GOOD MORNING EVERYBODY.
FOR THOSE WHO DON'T KNOW ME I
AM THE DIRECTOR OF THE HIV
DYNAMICS AND REPLICATION
PROGRAM.
IT IS AN HONOR AND A PRIVILEGE
TO HAVE THIS OPPORTUNITY TO
KICK OFF STEVE'S RETIREMENT
SYMPOSIUM AND ON BEHALF OF THE
ENTIRE DRP.
STARTING WITH A FEW OBLIGATORY
BIOGRAPHICAL DETAILS, STEVE
RECEIVED HIS PHD FROM HARVARD
1975.
ABOUT THAT TIME MOVING TO
UNIVERSITY OF UTAH AND AND
THEN MOVED TO UC SOUTH TO DO
POSTDOCTORAL FELLOWSHIP IN THE
LABORATORY.
1979 STEVENS RECRUITED TO
HARVARD AS A SENIOR STAFF
INVESTIGATOR.
AND IN 1984 RECRUITED TO THE
RESEARCH PROGRAM.
HE SERVED AS DEPUTY DIRECTOR
FOR OVER TEN YEARS AND BETWEEN
1995 AND 1999 DIRECTOR OF THE
LABORATORY AT EBL.
WITH THAT WAS ROLLED INTO THE
FEDERAL GOVERNMENT IN 1989
STEVE JOINED THE HIV DRP
PROGRAM FOR KEEPING THE
ACRONYM IN PLACE.
HE SERVED AS THE CHIEF FROM
1999 THROUGH HIS RETIREMENT
EARLY THIS YEAR ALSO SERVING
AS THE ACTING CHIEF FROM THE
DRP AND BETWEEN 2006 WAS
DIRECTOR OF THE HIV DRP.
HE HAS MADE A GROUNDBREAKING
CONTRIBUTION TO THOSE AREAS OF
VIROLOGY LIKE STRUCTURE AND
FUNCTION OF HIV-1 AND
MECHANISM SEPARATE RESISTANCE
TO THE INHIBITORS AND ALSO HIV
RESISTANCE JUST TO ELABORATE
BRIEFLY TOGETHER WITH EDDIE
ARNOLD WITH THE STRUCTURE WITH
THE ENZYME AND COMPLEX
DOUBLE-STRANDED DNA IT WAS
LANDMARK PAPER PUBLISHED 1983
AND DURING HIS TIME HE MADE
IMPORTANT CONTRIBUTIONS TO
UNDERSTAND THE RETROVIRAL
INTEGRATION BY DEFINING THE
STRUCTURES OF THE VIRUSES FROM
RSV INFECTED CELLS.
STEVE HAS MADE NUMEROUS
CONTRIBUTIONS TO THE
UNDERSTANDING TO THE
INHIBITORS WORKING IN
COLLABORATION WITH A NUMBER OF
GROUPS INCLUDING EDDIE ARNOLD
AND OTHERS.
THEN RECENTLY STEVE HAS TURNED
MUCH OF HIS ATTENTION TO THE
UNDERSTANDING OF MECHANISMS OF
PERSISTENCE TO ACHIEVE THIS
LANDMARK PAPER IN 2014 TO SHOW
THAT HIV-INFECTED CELLS ARE
CAPABLE OF EXPANSION IN
INFECTED INDIVIDUALS AND
RECENTLY JUST LAST YEAR
SHOWING THE PRO- VICE VIRUSES
OF THE T CELL.
SO I HAVE TO CONFESS THAT AS A
YOUNG STUDENT MANY YEARS AGO
WHEN I FIRST MET STEVE I FOUND
HIM TO BE SEMI- INTIMIDATING.
[LAUGHTER]
I'M SURE MANY OF THE AUDIENCE
HAD THE EXPERIENCE HAVING
STEVE IN THE FRONT ROW READY
TO POUNCE THE MOMENT ANYTHING
WAS NOT CLEAR ASKING QUESTIONS
I THINK WERE DESIGNED NOT TO
HAVE A SIMPLE ANSWER.
WHEN I WAS RECRUITED TO THE
DAP NEARLY 20 YEARS AGO AS A
NEWLY TENURED PI FROM THE
START THAT WAS MY LAP SHE AND
WE BECAME CLOSE FRIENDS.
I VALUED HIS MENTORSHIP IS
STRONG SUPPORT AND HIS KEEN
INTELLECT AND APPRECIATED HIS
KEEN FEEDBACK FOR MY
RESEARCH.
ADMINISTRATIVELY OUR ROLES
SPLIT IN 2015 WHEN I BECAME
DRP DIRECTOR HE PROVIDED
VALUABLE ADVICE AND FEEDBACK.
I HAVE MET FEW PEOPLE IN MY
LIFE AS DEDICATED IN MY LIFE
AS STEVE AND NO ONE IS DEVOTED
TO FISHING.
[LAUGHTER]
HIS ENTHUSIASM IS CONTAGIOUS I
MYSELF WAS AN AVID FISHERMAN
CO-OWNER OF THE DEMANDS OF TO
PUSH FISHING TO THE BACK
BURNER.
AS HE STARTED FISHING AGAIN WE
HAD MANY OUTINGS ON THE
CHESAPEAKE BAY AND ON STREAMS
LOCALLY IN WESTERN MARYLAND
AND PENNSYLVANIA CATCHING A
BEAUTIFUL RAINBOW TROUT IN THE
STREAM IN PENNSYLVANIA NEXT
THEM ON SCHEDULE TO ACCOMPANY
HIM ON A FLOAT TRIP TO ALASKA
THAT I THINK WE WILL HEAR MORE
ABOUT THAT IT WAS THE PICTURE
TAKEN FROM LAST SUMMER FROM A
DRONE.
WE CAN SEE THE CAMP AND THE
THREE VOTES.
I WILL END THIS PART OF
INTRODUCTION BY CONGRATULATING
STEPHEN HIS ILLUSTRIOUS CAREER
OF THE FORWARD TO MORE
SCIENTIFIC AND UNSCIENTIFIC
INTERACTION.
WE HOPE YOUR TIME IS UP WITH
SUNNY SKIES AND CALM WATERS
AND BEAUTIFUL SUNRISES.
HERE IS THE PROGRAM AT A
GLANCE TALKS FROM FORMER
MENTORS AND COLLEAGUES AND
COLLABORATORS.
AT THE END OF THE DAY WE WILL
RECONVENE FOR AN INFORMAL
GATHERING AT THE BREWERY AND I
HOPE YOU CAN ALL JOIN US FOR
THAT.
SO THAT IS MY PLEASURE TO
INTRODUCE THE FIRST SPEAKER.
FROM THE UNIVERSITY OF UTAH
HAS RECEIVED MANY AWARDS
INCLUDING 2007 NOBEL PRIZE IN
PHYSIOLOGY OR MEDICINE IN THE
STEVE'S ADVISORS I MENTIONED
THE UNIVERSITY OF UTAH.
>>IT IS A PLEASURE TO BE
HERE.
FIRST A FEW WORDS ABOUT STEVE.
I WAS REALLY FORTUNATE HE WAS
ONE OF THE VERY FIRST GRADUATE
STUDENTS IN MY LAB.
AND AS A MENTOR YOU'RE NOT
SUPPOSED TO HAVE FAVORITES BUT
STEVE WAS A FAVORITE HE WAS
NOT ONLY BRIGHT AND ENERGETIC
BUT LOVES TO ARGUE.
[LAUGHTER]
BUT THE NICE THING ABOUT THE
ARGUMENT IS THAT AFTERWARDS
SCIENCE HAS IMPROVED.
AND AS A GRADUATE STUDENT AND
A PLEASURE TO HAVE AS A
COLLEAGUE CONGRATULATIONS.
>>SO MANY TIMES I GET BORED
WITH WHAT I AM DOING AND I
MOVE ON.
BUT ANXIETY IS A ENORMOUS
PROBLEM IN TERMS OF HUMAN
HEALTH.
THE NATIONAL INSTITUTE OF
MENTAL HEALTH REPORTED THE
LIFETIME OCCURRENCE OF CHRONIC
ANXIETY IS AS HIGH AS
20 PERCENT EVEN HIGHER IN
WOMEN THAN MEN.
AND THIS IS EVEN WORSE FOR
YOUNG ADULTS AND ADOLESCENTS.
AND THEN WITH THE DISABLING
DISEASE IT IS A BIG PROBLEM.
SO THE DISRUPTION GIVES RISE
TO CHRONIC ANXIETY AND THESE
OVER GROOMING OF COMORBIDITY.
AND WITH THE TRANSCRIPTION
FACTOR AND THE PRIMARY
FUNCTION IS TO MAKE SURE YOUR
HEAD IS IN THE RIGHT PLACE AND
SOMEONE.
AND IT RESULTS IN PATHOLOGICAL
BEHAVIOR.
AND HERE WE SHOW THAT THEY
KEEP GROOMING THROUGH THE
LACERATIONS.
AND THEN TO EXHIBIT ALL STRONG
MALE SEX BIAS AND THIS IS A
MAP OF 20 DIFFERENT MICE
SHOWING THAT FEMALES SHOULD
HAVE MUCH MORE AGGRESSIVE
PATHOLOGY THAN MALES.
AND THOSE RELATED TO
PRODUCTION OF FEMALE HORMONES
AND THE ANSWER IS YES.
WOMEN AND FOR THE ENZYME
REQUIRED.
SO AGAIN WHAT YOU CAN SEE IF
YOU DO THAT TO A FEMALE MOUSE
THEN YOU GO DOWN TO THE MAIL
LEVELS.
SO IF WE SPEAK OF THE SAME
HORMONES SITE PROGESTERONE
INTO MEALS THEN YOU CAN
INCREASE THEIR LEVEL OF
ANXIETY CLOSE TO THAT OF
FEMALES.
THAT IS SHOWN HERE.
THAT OTHERWISE IT DOESN'T DO
ANYTHING TO IT.
DOESN'T TELL YOU HOW THAT
INTERACTS.
SO THE FIRST QUESTION IS ARE
THE CELLS RESPONSIBLE I'M
SORRY WHAT IS THE CAUSE OF THE
AGENT?
SO SIMPLY WE DO SOME ANALYSIS.
AND THEN DRIVE THAT EXPRESSION
AND BUT THEN PUT THE SIGNAL
WITH THE T SITES AND THEN
WHEREVER THAT IS EXPRESSED AND
THEN WHEN YOU TURN ON GFP.
BUT I WANT TO EMPHASIZE THIS
IS NOT A TRANSCRIPTION MARKER.
BUT HERE WE HAVE AN ENORMOUS
SURPRISE.
SO THAT YOU WILL KNOW EVEN
MORE.
AND THEN HERE YOU SEE THE
LABELS 30 PERCENT POPULATION
IN THE BRAIN.
SO THESE ARE THE CELLS OF THE
BRAIN AND CELLS THAT GO INTO
THE BRAIN AFTER THE RAIN
STARTS TO FORM AND THEN THEY
ARE THEY ARE TO CLEAR CELLULAR
DEBRIS AND THEN HAS A RESPONSE
TO INFECTION THIS IS WHAT THEY
SAID TO BE DOING WHEN WE
STARTED THIS WORK.
BUT WE SAY SOMETHING QUITE
DIFFERENT.
WE ARE SUGGESTING LABELED WITH
THE MARKER REGULATES THE
MIDDLE OF CIRCUITS THAT
DISRUPTION OF THAT REGULATION
LEADS TO THE PATHOLOGY I WAS
TALKING ABOUT.
SO IT HAS BEEN POSITIVE DURING
THE FIRST WEEK AND THEN
DIRECTLY INTO THE BRAIN AND
THEN ONCE THE SYSTEM HAS BEEN
DEVELOPED THAT CAN GET INTO
THE BRAIN AND THIS IS BEEN
REPEATED SO OFTEN BECAUSE THAT
WE COME ALONG AND SAY MAYBE
THERE IS MORE TO THE STORY
THAN WE THINK THE THEN READING
THE PAPER CAREFULLY FROM THEIR
OWN DESCRIPTION THEY CAN ONLY
ACCOUNT FOR ABOUT 60 PERCENT
OF REDUCING THEIR MARKER.
AND THEN TO LABEL THAT HAS A
STOP SIGN AND THEN THE CELLS
BECOME RED SO THOSE THAT ARE
LABELED WITH CR ONE AND HERE
BUT THEN USING THE TWO
PHOTON.
SO THE FIRST THING TO
INTEGRATE AND THEN I MENTIONED
THE OTHER POPULATION OF
70 PERCENT AND THEN THAT TO
CREATE AFTER BIRTH AND THAT IS
ABOUT 30 PERCENT SO THERE IS
TWO POSSIBILITIES AND ONE IS
AT THE GREEN CELLS TURN ON
WITH THE POSSIBILITY THERE IS
ANOTHER SEPARATE WAY AND
LOOKING TO SEE ONE IT IS
EXPRESSED IN OF THOSE SYSTEMS
AND THEN YOU SEE A STRONG
EXPRESSION THE EE SEVEN AND A
HALF THERE IS NOTHING.
AND THEN TO START SEEING THE
EXPRESSION THERE IS LITTLE BIT
LEFT YOU CAN EASILY DETECT BUT
THERE'S FAR LESS AND THAT
RULES OUT THE FIRST
HYPOTHESIS.
SO E-SEVEN AND A HALF HAS THE
YIELD THAT GOES DIRECTLY TO
THE BRAIN AND THEN THEY ARE
BORN AGAIN IN THE YOLK SAC
WITH THE GESTATION AND THEN
STARTING AT.
>>DR. PARKER: AND A HALF IN A
VERY DIFFERENT ROUTE OF ENTRY
AND OBVIOUSLY IF YOU DO THAT
TO HAVE THESE DIFFERENT
PROPERTIES.
>>SO WHAT ARE THE
SIMILARITIES BETWEEN THESE TWO
POPULATIONS?
BECAUSE I HAVE TO GO THROUGH
THIS THAT THERE IS NO
POPULATION.
SO THERE ARE THREE MARKERS
THAT ARE SPECIFIED IT WITH A
TRANSCRIPTION FACTOR WHICH IS
THE RECEPTOR AND THESE ARE
BONA FIDE AND IF ALL EXPRESSED
EQUALLY IN THE TWO POPULATIONS
SO THEREFORE WE CAN SAY WITH
CONFIDENCE IT IS BONA FIDE.
SO AS THESE POPULATIONS BEHAVE
WITH THE DIFFERENT HISTORIES
DO THEY BEHAVE SIMILARLY?
SO HOPEFULLY THIS WORKS.
THAT WAS QUICK.
NOW KEEP GOING.
THE MOVIE ISN'T WORKING BUT
WHAT YOU WOULD HAVE SEEN IS
SENDING THE PROJECTIONS YOU
CAN WATCH IT AND IT TAKES
ABOUT 20 SECONDS WE HAD IT
DOWN TO ABOUT THREE SECONDS
AND THEN IT COMES DOWN AND THE
FIELD IS SURROUNDED.
ANYWAY.
NOW THE OTHER PROPERTY IS THAT
THEY DO SYNAPTIC PRUNING AND
ABOUT TWICE AS MANY
CONNECTIONS THAT YOU NEED AND
THEN USE PRUNING TO PULL OUT
THE WEAKER CONNECTIONS AND
THEN REINFORCE THE STRONG
PROJECTIONS.
NOW SOMETHING IS GOING REALLY
WRONG.
ANYBODY?
[INAUDIBLE CONVERSATIONS]
THERE WE GO.
SO ANOTHER ASSAY FROM PRUNING
WAS DEVELOPED FROM HARVARD AND
WHAT SHE DID WAS TAKE THE UNIT
WITH A VERY HIGH AFFINITY FOR
THE SYNAPSE BUT THE LABEL ON
AND INJECTED INTO THE BRAIN
INTO THE EYE AND THEN IT GOES
TO THE THALAMUS AND THEN THOSE
WE CONNECTIONS ARE TAKEN UP SO
THEN YOU HAVE A LABEL INSIDE
THE CELLS.
SO THAT IS DOUBLY LABELED AND
THEN YOU CAN DO THIS FOR
HUNDREDS OF CELLS AND PRETTY
MUCH INDISTINGUISHABLE.
SO MOLECULARLY THEY ARE
SIMILAR.
THERE ARE 40 GENES ESSENTIALLY
SO THE POPULATIONS ARE QUITE
SIMILAR MOLECULARLY.
SO THEN THE NEXT QUESTION IS
THE CAUSE FOR THE ANXIETY WE
WILL ADJUST THIS.
SO YOU TAKE A MOUSE BUT THEN
YOU FILL IT UP HOWEVER YOU
WANT AND WE CAN PUT THAT IN
FROM A NORMAL MOUSE OR A
MUTANT MOUSE IT WITH THAT
EXPERIMENT THIS IS A GROWTH
FACTOR RECEPTOR AND THEY
REQUIRE A RECEPTOR FOR
SURVIVAL.
SO WE CAN USE THIS METHOD
SIMPLY TO ELIMINATE FROM THE
BRAIN AND THEN USE THE
PROCEDURES AND WITH THE
DEVELOPMENT AND THEN REINJECT
ON BOTH SIDES AND THEN WAIT A
FEW MONTHS AND WITH THE
POPULATION WE GO BACK.
AND THEN 100 PERCENT
ENGRAFTMENT.
SO THIS IS INJECTING NORMAL OF
HOW MUCH TIME THEY SPENT
GROOMING.
AND THEN HERE IS LOOKING AT
ANXIETY AND LOOKING AT THE TWO
PLATFORMS.
AND THIS IS WHAT WE CALL
CLOSED AND ENCLOSED AND
ACTUALLY SUPPORTS BOTH
PLATFORMS EQUALLY WHERE A
MUTANT MOUSE LOOKS DOWN AND IT
WAS SCARY.
SO MOST OF THE ASSAY IS
AVOIDANCE TYPE.
SO THAT SAYS THAT PURIFIED
CELLS ARE CAUSAL THAT ONLY BY
THEMSELVES WITH THE PHENOTYPE
THEN IT IS A NORMAL BEHAVIOR.
SO WITH THE UNDERSTANDING OF
THE PROCESS.
NOW WE REALLY NEED HELP.
[LAUGHTER]
SO I WILL TAKE YOU THROUGH
THESE.
AND A WAY OF ACTIVATING.
[INAUDIBLE CONVERSATIONS]
SO THE FIRST STEP IS WHAT WE
WANT TO DO IS MODULATE.
SO THE FIRST PERSON IS HOW IS
THAT POSSIBLE?
AND THEN WITH THOSE ACTION
POTENTIALS.
SO WHAT DOES THAT MEAN CRACKS
SIMPLY OF CELL FUNCTION
PUMPING INTO THE CELL AND THAT
IS ENOUGH ESSENTIALLY THROUGH
TRANSCRIPTION AND SO ON.
AND WITH GENERAL DOBSON AND
THEN WITH THE REGIONALS OF THE
BRAIN.
AND WITH THE WATTAGE OF THE
LASER AND SO SHOWING YOU 20
CONTROL SLIDES SO YOU JUST
HAVE TO TAKE MY WORD FOR IT IS
VERY SPECIFIC CUTS.
PERIOD.
SPACE AND SPECIFICITY IS
IMPORTANT.
BUT IF YOU DO THE RIGHT ONE
SOMETHING HAPPENS.
IS ALSO IMPORTANT ASTROCYTES
AND NOTHING HAPPENS FINALLY
HAS BEEN REPORTED OF NON-
EXPRESSING NEURONS BUT THE
LEAKAGE IS EXTREMELY LOVING
CANNOT BE DETECT ON —-
DETECTED IN THE RESPONSE.
THAT IT IS UNDETECTABLE.
BUT NONE THE LION NOT ON —-
NONETHELESS WE DON'T SEE THE
EFFECT IF WE LEAVE OUT THE
COMPONENT THAT WE TALKED ABOUT
THAT SHOULD NOT AFFECT FREE
EXPRESSION WHAT MAKES THIS
RECEPTIVE IN THE NEURON.
SO WE HAVE DONE ALL OF THOSE
CONTROLS THAT YOU WILL NOT SEE
ANY OF THEM SO NOW LET'S MOVE
ON TO THE ACTIVE EXPERIMENTS
THEMSELVES.
SO HERE WE PUT IT INTO THAT
PART OF THE BRAIN.
SO WHAT YOU DO IS TAKE MOVIES
FOR TWO MINUTES TURN ON THE
LASER KEEP IT ON FOR TWO
MINUTES THEN TURN OFF THE
LASER KEEP RECORDING.
SO HERE WE HAVE SEE 57 CONTROL
THAT'S FOR MANY GENERATIONS SO
IF YOU TURN ON THE LASER
NOTHING HAPPENS BUT THEN THEY
DEPENDED.
THAT HERE IS THAT CONTROLS IT
IS 100 FOR LESS SO IF YOU TURN
OFF THE LASER INDEPENDENT EASY
SOME BACKGROUND NOISE THAT HAS
NOTHING TO DO WITH THE LASER
BUT AN EXPERIMENT THIS IS PRE-
AND POST, OR DURING BUT THEN
IT IS VARIABLE THEN BETTER
THAN OTHERS.
>>AND THEN WITH THE
PREFRONTAL CORTEX AND THEN
THEY TURN ON THE LASER AND THE
RESULTS ARE QUITE ROBUST AND
MEASURABLE AND QUANTITATIVE
THAT TELLS US WE CAN TURN ON
THE SYSTEM WITH THE PREFRONTAL
CORTEX AND THEN WE ONLY GET
THE RESPONSE AND THEN THE
CENTRAL AMYGDALA.
SO HERE IS THE MAZE AGAIN
EXPLORING BOTH AND THEN IT
STARTS TO EXPLORE AGAIN AND
THEN TO QUANTITATE THIS IN
DIFFERENT WAYS.
AND THEN TO EXPLORE THE ENTIRE
SURFACE IF IT IS A MUTANT
MOUSE THEN IT STAYS OUT IN THE
CENTER.
SO WHAT YOU CAN SEE ANOTHER
REACTION IS FREEZING WHICH IS
A REACTION OF ANXIETY.
SO FOR EXAMPLE WITH PRE- AND
THEN POST DURING THE LASER
THAT PATTERN DOES NOT CHANGE
FOR THE BLA AND THEN YOU CAN
QUANTITATE THAT.
IF YOU GO INTO THE HIPPOCAMPUS
THEN YOU CAN INDUCE BOTH
BEHAVIORS AS WELL AS HIGHER
LEVELS OF ANXIETY THAT IS
SHOWN HERE FREE AND DURING AND
THEN AFTER THEN YOU CAN SEE
QUANTITATIVE AND THEN WITH THE
CONTROL AND THEN HERE WE CAN
SEE THAT CONTROL WITH THE OPEN
FIELD TEST AND THEN SO THAT
ESSENTIALLY SAYS WITH THAT
BEHAVIOR BUT THAT IS THE SAME
SO SO FAR IT IS ACTIVATING SO
WHAT HAPPENS IF YOU ACTIVATE
OTHERS?
SO UNFORTUNATELY WE CANNOT DO
THAT EXPERIMENT BUT WE CAN
ACTIVATE BOTH POPULATIONS AND
WE ARE EXPRESSING THIS IN BOTH
POPULATIONS AND HERE'S WHAT WE
WERE EXPECTING SO ONE
POSSIBILITY IS IT IS NO
DIFFERENCE WITH ALTERNATIVE
YOU WOULD SEE HIGHER LEVELS IN
THAT SAYS THAT BOTH ARE
PARTICIPATING SO WHAT WE HAVE
DONE IS COMPLETE THE
UNEXPECTED.
IT ACTUALLY INHIBITED AND NOW
IT'S BACK DOWN TO ZERO.
AND THEN TO THINK ABOUT THIS
SO THROW EVERYTHING AWAY.
YOU HAVE TO REVISE HOW YOU
LOOK AT IT.
SO AT THE GENETICS EXPERIMENT
EXIST IS THAT AS IT ACTS AS A
BREAK AND REDUCES ANXIETY AND
THEN WORKING AS AN ACCELERATOR
TO INCREASE ANXIETY AND
BEHAVIOR AND TOGETHER WITH ANY
STATUS THAT THEY WANT AND THEN
YOU CAN REGULATE HOW MUCH
ANXIETY.
BECAUSE REMEMBER A LITTLE BIT
OF ANXIETY IS GOOD.
GETS US EXCITED AND MOVING
THAT TOO MUCH IS PATHOLOGICAL.
SO NOW YOU HAVE A VERY FINE
SYSTEM CONTROLLING THAT
BEHAVIOR.
SO I DO HAVE MICROGLIA IN THE
FIRST PLACE?
WE ARE DOING THE SAME THING
THEY HEAR THEY ARE WORKING
OPPOSITE TO EACH OTHER.
SO THE MODEL MAKES A STRONG
PREDICTION OTHERWISE WE SHOULD
GET NOTHING OTHERWISE IT
SHOULD BE ELEVATED FOR ANXIETY
IN THAT SECTION WHICH YOU SEE.
NO ONE WOULD HAVE EVER
PREDICTED THIS.
SO WITH THAT I WILL STOP.
THANK YOU.
[LAUGHTER]
[APPLAUSE]
>>THANK YOU VERY MUCH.
WE HAVE TIME FOR A LITTLE BIT
OF QUESTIONS.
>>
[INAUDIBLE]
>>WE CAN MAKE MICE THAT HAVE
EITHER ONE.
>>AND AT DIFFERENT TIMES
QUICK.
>>CERTAINLY WE CAN USE THAT.
WE STILL DON'T HAVE A WAY OF
REGULATING OR ANY MARKERS
SPECIFIC WITH THE SEQUENCING.
BUT MY GUESS IS TO FIGURE OUT
A WAY TO MODULATE BOTH
POPULATIONS.
WHAT I SHOULD POINT OUT WE CAN
PUT ANYTHING WE WANTED SO WE
COULD CHANGE THAT TO ADDRESS
THE QUESTION.
>>SOME OF THOSE DIFFERENT
PROPORTIONS WITH THE MOUSE
STRAINS?
>>WE DON'T KNOW.
>>SO IF YOU HAVE LESS
EXPRESSION YOU SEE THAT THERE
AS WELL?
>>YOU MODULATE.
>>
[INAUDIBLE]
>>AND HETEROZYGOTES WE DON'T
SEE THAT.
BUT IN BETWEEN WE SHOULD SEE
THE PHENOTYPE.
AND THEN I THINK WE CAN DO
THAT.
WHAT I SHOULD POINT OUT BUT
THEN THEY KEEP DIVIDING AND
DIVIDING AND THEN WENT TO STOP
AND REPLICATE.
AND THEN TO KNOW WHAT THE
GROWTH RATES AND THEN GO A
LITTLE BIT FASTER.
SO THOSE EXPERIMENTS CAN BE
DONE.
>>ANY ASSOCIATED TO HUMANS?
>>VERY INTERESTING QUESTION.
THAT FOR SOME REASON THEY
CHOSE THE OTHER AS WHAT YOU
NORMALLY HAVE.
BUT IN MOST POPULATIONS IN THE
ROLE THAT IS AT 90 PERCENT.
SO IT COMES IN FROM HIGHER
PRIMATES THEN TO MAINTAIN A
VERY HIGH LEVEL IN HUMANS.
AND WE HAVE TAKEN THAT
POLYMORPHISM STICKING INTO
MICE.
IT FUNCTIONS AS A MUTATION SO
THE HIGHER LEVELS OF ANXIETY.
SO WITH THE SYSTEM IT IS
SIMILAR.
SO THERE IS ONLY ONE
POPULATION NOW AND THAT IS THE
AFRICA POPULATION.
ALL OTHERS ARE REALLY HIGH.
SO THERE IS POLYMORPHISM.
>>FASCINATING.
THANK YOU.
>>DO YOU SEE ANY EFFECT ON
GENDER WITH YOUR
TRANSPLANTATION EXPERIMENTS?
>>NOW.
ALL OF THOSE TOOK PLACE THEN
WITH THOSE HUNDREDS AND
HUNDREDS OF EXPERIMENTS.
>>THANK YOU VERY MUCH.
WE SHOULD MOVE ON.
[APPLAUSE]
THE NEXT SPEAKER FROM CORNELL
MEDICINE.
OUR SECOND OF THREE TODAY
SHARING THE NOBEL PRIZE IN
1989 FOR DISCOVERING THE
ORIGIN IN MANY OF US ALSO
KNOWN HER AS A FORMER AND THEN
GOING FROM 1976 THROUGH 1979.
THANK YOU.
>>THANK YOU VERY MUCH.
NICE TO BE HERE.
THERE ARE SEVERAL WAYS TO GET
THIS KIND OF TALK.
ONE OF COURSES TO SAY HI TO
STEVE AND WE LOVE YOU AND GO
ON TO TALK ABOUT YOUR GREAT
NEW WORK IN MY HEAD WORK IS ON
THIS BEAUTIFUL AS WHAT YOU
JUST SAW THEN I WOULD DO
THAT.
THE SECOND WAY IS TO SAY HI
STEVE.
WHAT YOU DOING RECENTLY AND
TALK ABOUT THAT BUT I KNOW
THERE IS PLENTY TO COME FROM
ALL OF YOU THAT HE IS BEEN
DOING THE LAST TEN YEARS.
THE THIRD AND MOST FUN AND
APPROPRIATE UNDER THE
CIRCUMSTANCES IS TO TALK ABOUT
THINGS WE DID TOGETHER BOTH AT
THE TIME WHEN HE WAS A POSTDOC
IN THE LATE SEVENTIES IN SAN
FRANCISCO AND ENCOUNTERS
THEREAFTER.
STILL COME IN DOING THIS IT'S
NICE TO HAVE A THEME AND THAT
CAME FROM A MOST UNUSUAL
JOURNALISTIC AND THEN CRUISING
THE INTERNET WHILE LOOKING FOR
SOME PICTURES BUT THE CAPTION
SAID STEVE USES TRUTH IN
TEAMWORK.
AS I THINK ABOUT MOST
IMPORTANT AND NOW APPROACHING
50 YEARS ALTHOUGH NOT QUITE
BUT I NOTICED THERE IS A
PATTERN EVERY CURRENT BEHAVIOR
THAT INVOLVES TRYING TO DO
IMPORTANT THINGS FOR THE
COMMUNITY AT THE SAME TIME
EXHIBITING HIS OWN EXCELLENCE
AS A SCIENTIST AND A NARRATOR.
I WILL SPEND SOME TIME TALKING
ABOUT THREE EPISODES OUT OF
SEVERAL THAT EXEMPLIFY WORKING
AS A COMMUNITY TO ACHIEVE THE
AMBITIONS OF INDIVIDUALS.
I WILL DO THAT STARTING WITH
POSTDOCTORAL WORK AND THEN TO
OTHER ASPECTS OF OUR LIFE
TOGETHER AS A RETROVIRAL
ADJUST TO EXEMPLIFY THE
QUALITIES IT IS A BIG DAY FOR
HIM.
SO THE FIRST THING I WANT TO
TALK ABOUT.
WHAT IS GOING ON?
[LAUGHTER]
DO I NEED TO CALL FOR HELP OR
JUST DISPLAY ANXIETY?
[LAUGHTER]
I ALWAYS USE A MATH.
[LAUGHTER]
YOU CAN AFFORD IT.
[LAUGHTER]
BUT I DO NEED TO SCRATCH AT
THE MOMENT.
ONE —- I DO NEED THE CRUTCH AT
THE MOMENT.
I'M TOLD PLEASE WAIT.
THAT IS NOT IN MY CHARACTER.
[LAUGHTER]
>>ANY QUESTIONS SO FAR?
[LAUGHTER]
WHAT ARE YOU DOING NOW?
OTHER USER?
FORTUNATELY MY TALK IS NOT
THAT SO WE WILL DEFINE.
SO TRUTH IN TEAMWORK AND I
WILL TALK ABOUT MOST
EXTENSIVELY ABOUT THE TRUTH AS
STEVE WAS IN OUR GROUP.
HE WAS NOT THERE UNTIL 81
THAT'S WHY THE DATES APPEAR TO
UNDERSTAND THE STRUCTURE OF
PROVIRAL DNA.
THAT WORK WAS DONE TO A VERY
LARGE EXTENT BY STEVE.
BUT THERE WAS A PROVIRAL TEAM
IN THE LAB AND PETER ALSO
RECENTLY RETIRED FROM BROWN
UNIVERSITY WHO WAS STEVE'S
PARTNER ON THIS WORK BUT IN
LOOKING AT THE INTEGRATED
FORMS OF DNA AND PETER IS A
KEY COMPONENT.
IN ADDITION TO PUT THIS IN OUR
OWN LAB BUT WE ALSO HAD THE
ADVANTAGE OF WORKING WITH THE
WEST COAST TUMOR VIRUS.
EVERYBODY WAVED TO PETER I SAW
HIM RECENTLY IN SAN DIEGO FOR
DINNER.
AND THEN TO HAVE DINNER IN SAN
DIEGO BUT MANY OTHERS THAT
MANY INSTITUTIONS GATHERED
REGULARLY TO TALK ABOUT THINK
THEMES OF TUMOR VIROLOGY AND
THERE'S MORE TO SAY ABOUT THAT
IN A MOMENT BUT IT'S IMPORTANT
TO REMEMBER WHAT I TELL YOU
ABOUT WHAT HE ACCOMPLISHED IN
POSTDOCTORAL TIMES WAS CARRIED
OUT IN THE CONTEXT OF THIS
VERY INTERACTIVE GROUP PLUS
THE MUCH LARGER GROUP THAT
SHARED RESULTS OPENLY WITH
COLLABORATIONS AND IMPORTANT
TIMES TOGETHER AND THOSE
INSIGHTS WERE WIDELY SHARED.
COME ON.
ARE YOU KIDDING?
SERIOUSLY?
I CANNOT ADVANCE THE SLIDES.
AND THEY HAD NO PROBLEM
BEFORE.
WHAT THE HELL IS GOING ON?
SO WE KNOW HOW THE VIRUS
REPLICATES MAYBE IT'S NOT SO
RECENT THAT WE DID NOT KNOW
THE ANSWERS TO SOME BIG
QUESTIONS HOW MANY SITES WERE
AVAILABLE AND THE VITAL ROLE
DNA AND THOSE CHROMOSOMES HOW
THEY ORGANIZED IS IT A
SUITABLE TEMPLATE FOR THEIR
GENOMES AND MESSENGER RNA IS
THAT VIRUS ACTUALLY INFECTIOUS
AND WHAT IS THAT INTEGRATION
ITSELF SO THINKING ABOUT THOSE
QUESTIONS AT THE TIME STEVE
ARRIVED AND WERE ANSWERED AT
THAT POINT IS WHAT TYPE OF
TECHNOLOGIES WERE REVEALED AND
I TALKED TO STUDENTS SO WHAT
IS THE BIG DEAL WE COULDN'T ON
THE CHICKEN CELLS FOR HERE IS
THE COLLABORATION CAME IN
LOOKING AT CLONE INFECTED
NONPERMISSIVE RAT CELLS AND WE
COULD CLONE CELLS AND NOT
MOLECULES BY LOOKING AT THE
CLONES THE CELLS AND P.M.
PRESCRIPTION ENZYMES WITH A
LOT OF EXTRA ENZYMES AND
THINGS TO PEOPLE IN THE
INTERNAL GROUP AND THOSE
PROBES THAT WERE VERY SPECIFIC
FOR THE VIRAL GENOME AND THEN
THE ENVELOPE GENE IS DEFINED
BY THOSE MUTATIONS THAT PETER
HAS SUPPLIED TO US THE DEAD
MIRACULOUSLY WITH THE STAGE
CAN DO THE MAPPING REQUIRED
BUT THEN THE OTHER THINGS WE
WOULD HAVE LIKED TO BE ABLE TO
DO AND THEN IN SOME WAYS TO
STUDY THAT TO BE AVAILABLE SO
WHAT WERE THE'S KEY FINDINGS
OF STEVE?
FIRST AS WE HAD DEDUCED ONLY
ONE OR TWO PRO VIRUSES THERE
WERE A FEW WITH ALL THE PRO
VIRUSES WERE THAT PETER SHANK
HAD DESCRIBED A VERY SIMILAR
TECHNIQUES BUT ALL OF THEM HAD
CELL DNA FOR THOSE SEQUENCES
AND DERIVED FROM BOTH ENDS OF
VIRAL RNA AND NOTING THAT THE
SEQUENCE IS UNIQUE TO THE DNA
AND OF COURSE THAT CREATED
SOME ISSUES OF THEIR OWN.
AND PERHAPS MOST IMPORTANTLY
ALL THOSE VIRUSES LIKE THAT
HAVE DIFFERENT RESTRICTION
MAPS SO THEY ALL SEEM TO BE IN
DIFFERENT SITES THERE WERE TWO
FEATURES ABOUT THIS BUT THERE
ARE SOME FEATURES HERE WORTH
RECALLING BECAUSE THEY EXUDE A
CERTAIN TONE OR FEELING THE
FIRST TO SHOW WITH THE
NONPERMISSIVE CELLS THAT THEY
WERE INDEED INFECTIOUS
SECONDLY WE DID SOMETHING IN
THE ACKNOWLEDGMENTS AS THEY
WERE PUBLISHED IN 1878 THAT
ILLUSTRATED THE SENSE OF
CAMARADERIE WE WERE TROUBLED
BY THE FACT THERE WERE
SEQUENCES AND WE DIDN'T KNOW
HOW YOU COULD POSSIBLY GET
THAT AT THE FAR END OF THE PRO
VIRUS AND THEN GOING ON WITH A
DIFFERENT FIELD ENTIRELY BUT
THEN TO PIPE UP AND SAY
JUMPING FROM THE FIRST
TEMPLATE TO THE SECOND WHAT
ABOUT TO THE THIRD AND THEN AS
FAR AS WE CAN TELL THAT IS THE
EXPLANATION AND REVERSE
TRANSCRIPTION INTO DID HOLD US
TO AN ACCOUNT WAS CREATED AND
GOING OUT OF THE DEEP PAST A
MATHEMATICAL ANALYSIS TO SAY
WE ONLY MAP 13 INTEGRATION
SITE BUT ONLY TEN VIGOROUSLY
SO THERE MUST BE A LOT MORE
THAN TEN OR 15 SITES IN THE
GENOME.
AND HE DID THAT BY USING THE
TRADITIONAL BIRTHDAY PROBLEM
BUT I WILL NOT TRY TO RECITE.
AND THAT THERE ARE AT LEAST 40
REGIONAL SITES WITH THAT
SPECIFICITY OF A LOCAL
SEQUENCE AND FOR THOSE THAT
MUST BE PERMISSIVE AND I WOULD
LIKE TO THINK ABOUT STEVE'S
ATTITUDE.
WE DID SOME SIGNIFICANTLY
LATER ONE PERSON PUBLISHED A
LOT EARLIER AND I WAS
DISTRESSED BY THIS I DID NOT
LIKE HAVING SOMEONE BEFORE
THEY HAD THE GOODS BUT IT WAS
PUBLISHED ANYWAY.
BUT THEN WE ARTFULLY PUT
TOGETHER THE PAPER WAS THE
STATEMENT THAT DISCUSSION
SESSION THAT SAID THEY ARE AS
CONSISTENT AS OUR FINDINGS BUT
THE FACT THAT STEVE WAS MY DAD
ON THAT OCCASION BUT STEVE WAS
CALLED AND SAID IN THE LONG
RUN THE GOOD THINGS OPEN UP
THAT'S IN A RELATED TO HAD THE
RIGHT PERSON WORKING FOR ME.
SO HERE IS THE DESCRIPTION OF
WHAT STEVE WAS DESCRIBING SO
IT'S NOT JUST THE
INTERPRETATION AT THE TIME
THAT THE DEFINITION OF WHAT
COULD BE DONE IN THE FUTURE SO
THAT'S WHY IT'S USEFUL TO LOOK
BACK TO THINK ABOUT WHERE IT
STANDS IN HISTORY OF
RETROVIROLOGY.
SO WE CONCLUDED AND PROVEN ITS
TO BE A TOTALLY APPROPRIATE
TEMPLATE FOR TRANSCRIPTION AND
THAT WITH VIRAL RNA THAT COULD
PROVIDE SO WITH THE MORE
EFFORTS FOR THOSE VIRUS TO SAY
THIS IS TRUE THAT WE DON'T
HAVE TIME TO REVIEW IN DETAIL
BUT THAT IS KNOWING THERE IS A
LOT OF POTENTIAL INTEGRATION
SITES BUT IT WAS POSSIBLE TO
THINK ABOUT THE PRO VIRUS AND
THEN TO ACTIVATE THE GAINS BUT
THIS IS AN IDEA THAT WE TALKED
ABOUT A LOT I AM ON SABBATICAL
IN 78 AND 79 TO UNDERTAKE THE
TASK TO SHOW IT COULD ACT
USING THE LEUKEMIA VIRUS TO
REVERT BY INTEGRATING INTO THE
PRO VIRUS AND WITH THE HARD
WORK AND THEN TO SHOW THE FACT
THAT HAPPEN.
AND THEN TO INSPIRE EFFORTS SO
THAT MIGHT CAUSE CANCER EVEN
THOUGH MORE SLOWLY BY
INSERTING TWO AND ACTIVATING
SO AT THAT POINT WE ARE
LOOKING FOR SIGNS OF THE
PRECURSORS TO OTHER GENES WITH
A SIMILAR POTENTIAL THAT COULD
BE IMPLICATED AS A RESULT OF
THE RETRO FILE A ASSERTION AND
THEN TO DO SIMILAR WORK AND
THAT IS THE GENESIS FOR THE
DISCOVERY OF GENES SO THE
THIRD POINT IS THAT THE PRO
VIRUS AND THAT MADE US THINK
HOW THE RETROVIRUS MAY SERVE
AS GENETIC FACTORS FROM
ARTIFICIALLY WHEN THEY ARE
MOLDED AND AS YOU KNOW IS A
REASONABLY COMMON PRACTICE BUT
WE ARE ALSO ABLE TO SAY THAT
THERE ARE SOME THINGS THAT WE
NEED TO KNOW THAT WE WILL AND
WITHOUT MORE DEVELOPMENTS AND
TECHNOLOGY IN ONE IS THAT
DETAILED MECHANISM TO CLONE
AND SEQUENCE OF THE PRO VIRUS
UNDERCURRENT RESTRICTIONS WE
KNOW A LOT ABOUT THAT NOW I
WILL NOT DESCRIBE IT THAT
STILL DIDN'T HAVE THE TOOLS
FOR DOING IN VITRO INTEGRATION
AND NOT UNTIL PAT BROWN CAME
ALONG TO BUILD THE BASE AND
HIS COLLEAGUES CAME ALONG AND
LEARNED HOW TO USE THE
PURIFIED REACTIONS AND THIS IS
AN AREA WHICH IS STILL GOING
STRONG AND YOU WILL HEAR MORE
ABOUT THIS.
BUT THEN STILL THERE WAS THE
QUESTION KNOWING WHAT IS THE
REAL PRECURSOR AND LEARNING
FROM MY GROUPS AND OTHERS THAT
THEY HAD TO FIND NOT JUST A
LINEAR FORM OF DNA BUT TO
CIRCULAR FORMS AND ANY ONE OF
THOSE BY SOME MODEL COULD HAVE
PROVIDE A SUITABLE PRECURSOR
BUT THAT WAS NOT INTEL IN
VITRO REACTIONS WERE SUITABLY
DEPLOYED IT WAS POSSIBLE TO BE
CLEAR THAT LEAVING OPEN SOME
QUESTIONS WHY THAT DNA IS
THERE BUT MAYBE IT IS THERE
BECAUSE IT CAN BE WITH THE
ENDS OF LINEAR DNA AND THEN
FINALLY INTEGRATES FROM THE
DRUG TARGET AND YOU'LL HEAR
ABOUT THAT FROM DIMITRI IN
JUST A MOMENT SO FIRST I WILL
SAY WORD OR TWO ABOUT THAT
COLLABORATION ABOUT A DECADE
AFTER HE LEFT TO SAN FRANCISCO
WITH JOHN YOUNG AND PAUL BATES
HAD OVERCOME THE PROBLEM OF
NOT KNOWING WHAT THE RECEPTOR
WAS OR SUBGROUP A WHICH WAS
THE MOST COMMON FORM AND THEY
HAD SOME DIFFICULTY WITH A
SMALL GENE WITH THE TVA
RECEPTOR AND THEN ADD GENE
WHICH INTERESTINGLY THEY WERE
NOT VERY SURE WHAT TO DO WITH
THIS BUT THEN THIS WAS A NICE
IDEA TO USE THIS AS A WAY TO
TARGET VACUUMS WITH CELL TYPES
TO CARRY THAT GENE INTO THE
CONTROL OF THE SPECIFIC
PROMOTER SO HIS POSTDOC STILL
WORKS ON THIS PROBLEM TO BUILD
A TRANSGENIC ANIMAL THAT
ALLOWED THE MUSCLE CELLS BUT
AS SHOWN BY ANOTHER COMMUNITY
WITH THAT GROUP OF
INVESTIGATORS WHO ARE WORKING
TO MAKES USE OF THAT
TRANSGENIC MICE FOR A VARIETY
OF PURPOSES AND WITH SOME
OTHER TISSUES TO START MEETING
FAIRLY REGULARLY AT THE NIH
AND ELSEWHERE IN THE SPIRIT
NOT DISSIMILAR FROM WHAT WE
EXPERIENCED IN THE WEST COAST
IN THE CONTRIBUTION TO THIS
DEVELOPMENT TRYING TO SEEK
TRUTH BY MORE MAKING MORE
USEFUL ANIMAL MODELS.
SO BRIEFLY I WANT TO TALK
ABOUT STEVE'S ROLES IN
CREATION WITH ALL OF THE
TEXTBOOKS THAT WERE PUBLISHED
THAT HAVE BEEN GUIDES FOR
EVERYBODY AROUND THE WORLD FOR
THE LAST 40 SOME YEARS AND IN
THIS CASE IT'S A LOT OF
EDITORS AND WRITERS AND
READERS AND OTHERS WHO MAKE
CONTRIBUTIONS SO NOW I DRAW
YOUR ATTENTION TO A COUPLE OF
ASPECTS THAT HAD GONE ON SONG.
SO MIRACULOUSLY PUBLISHED IN
1997 AND THEN STILL 25 YEARS
LATER WAR AND PEACE HAS
SURVIVED BUT THIS IS A PRETTY
DURABLE BOOK BUT PEOPLE FORGET
THE PREDECESSORS BUT FROM 1872
THOSE THAT HAVE AND THOSE THAT
HAVE COPIES OF THAT BUT WE
RECOGNIZE THINKING ABOUT HOW
TO PROCEED TO MAINTAIN
AUTHORITATIVE TEXT WERE AWARE
BY THE EARLY EIGHTIES THE
FOLKS IN THE TWO FIELDS JUST
BEING SEPARATED SO THE TEXT IS
ALSO BE SEPARATE.
SO IN THE FIRST VOLUME WHICH
JOHN AND I WERE BOTH EDITORS
IN APPENDIX WAS NEEDED YOU
HAVE TO ACCUMULATE A LOT MORE
INFORMATION THAN COULD BE
INCORPORATED INTO THE
CHAPTER.
STEVE VOLUNTEERED OR COERCED
INTO BEING THE EDITOR OF THE
APPENDIX WHICH PROVED TO BE
100 PAGES LONG.
HE DID GET SOME HELP BUT THAT
WAS 100 PAGES OF STUFF AND
THEN TO ASSEMBLE ALL THE
STUFF.
AND WITH THE RETROVIRUS
ESPECIALLY ABOUT THE SEQUENCE.
AND THEN TO PUT OUT A SECOND
ADDITION AND IN 1985 AND THAT
PAPER HAD THE APPENDICES THAT
STEVE WAS THE LEAD TO DEVELOP
THIS OF 650 PAGES.
SO IT IS LIKE THE APPENDIX TO
THE APPENDIX BUT THIS IS A
GARGANTUAN TASK SO THANK YOU
TO THE'S SLIDE DELAYS MY TIME
IS COMING TO AN END BUT MY
LIFE AND RETROVIROLOGY AS IT
WAS THAT FROM BACK IN MY TIME
STEVE IS A VERY LARGE PERSON
WHO MAKES A LARGE ENTRANCE
WITH A LARGE EFFECT IT IS
DIFFICULT FOR ME TO IMAGINE SO
I STILL DO NOT BELIEVE THAT
THIS IS A RETIREMENT DAY YOU
CAN BETTER EXPRESS THE
ENTHUSIASM.
THANK YOU.
[APPLAUSE]
I'M HAPPY TO TAKE QUESTIONS OR
COMMENTS.
THANK YOU VERY MUCH.
>>NOW GOING FROM THE PAST TO
THE PRESENT THE ASSISTANT
PROFESSOR IS A LEADER OF
STRUCTURAL BIOLOGY AND LOOKING
AT THE HIV-1 STRUCTURE.
>>THANK YOU VERY MUCH.
IT IS A DISTINCT PLEASURE TO
BE HERE.
AND THEN TO TELL YOU AND THEN
TO DISCUSS OR TO INTRODUCE
CONGRATULATIONS IT IS AN HONOR
TO WORK WITH YOU OVER THE LAST
FIVE YEARS I AM TALKING TODAY
TO UNDERSTAND MECHANISMS AND
THE DRUG-RESISTANT VARIANT.
SO BEGIN WITH A VIRAL
REPLICATION CYCLE WE DO KNOW
THAT THERE IS AN WITH THE
REVERSE TRANSCRIPTION PUTTING
IT INTO NUCLEAR IMPORT AND
THEN THE PROTEINS WILL GO TO
THE LINEAR AND WITH THE DNA TO
FORM BUT THAT REFER TO IT AS A
PRE- INTEGRATION COMPLEX AND
THEN TO CONDUCT INTEGRATION TO
ESTABLISH THE PRO VIRUS AND
THE TARGET CELL WITH A
PARTICULAR PREFERENCE AND IN
THE CASE OF HIV-1 SO ONE OF
PARTICULAR INTEREST AND IT'S
NOT A MEDICAL TO STRUCTURAL
BIOLOGY FOR WHAT YOU CAN DO IS
RECAPITULATE THE STRUCTURE OF
THE PIC SO BIOCHEMICALLY THE
WAY THAT YOU DO THIS IS THAT
WITH A VIRAL DNA THERE ARE 27
VERSIONS AND THOSE BIOCHEMICAL
CONDITIONS WITH THAT SYNAPTIC
COMPLEX.
AND THE NUCLEOTIDES.
BUT THEN WITH THAT TARGET DATA
AND THEN IT COMPLEX THAT DATA
AND THEN SOON THEREAFTER AS
THE CATALYTIC REFORM.
SO WITH THE TARGET DNA WE HAVE
STRUCTURAL BIOLOGIST AND NOW
THEIR STRUCTURES FROM MANY
DIFFERENT RETROVIRUSES.
AND FAIRLY DIVERSE
ARCHITECTURALLY.
AND THEN TO SHARE A CONCERN
WHICH IS SIMILAR TO THE PFC.
AND THROUGHOUT THE ENDS OF
VIRAL DNA.
SO THIS IS FROM THOSE SUMS SO
MY TALK IS FOCUSED HIV
SUSPICION ONE —- SPECIFICALLY
WITH THEIR UNDERSTANDING OF
RESISTANCE.
SO I JUST HAVE TO POINT OUT
THERE ARE FIVE MEDICALLY
APPROVED DRUGS USING TO TREAT
AND THEN THE FIRST-GENERATION
NOW SECOND-GENERATION DRUGS
THAT ARE BETTER BECAUSE THERE
IS LESS RESISTANCE.
THEN THE IMPORTANT COMPOUNDS
FROM OUR COLLEAGUES AT NCI.
TERRY AND STEVE HAVE HAD A
LONG COLLABORATION TO TEST ALL
OF THESE.
SO THEN ADDRESSING THAT BASIS
FOR INHIBITION OF INTEGRATION.
AND THEN OF THE CHEMICAL
FEATURES OF POTENCY ESPECIALLY
AGAINST CHEMICAL RESISTANT
VARIANTS.
SO THE FIRST QUESTION.
SO A NUMBER OF YEARS AGO THIS
WAS THE LEADING CLINICALLY
APPROVED SECOND GENERATION
DRUG.
BUT THEN TO SYNTHESIZE SO THEY
WERE CHEERED TO CAPTURE DNA
AND THEN YOU CAN WITH THE
COMPOUNDS FOUND.
SO HERE WE CAN SHOW THEY ARE
COMPETENT TO INHIBIT
INTEGRATION AND THEN TO START
DOING THAT STRUCTURAL
VIROLOGY.
AND NOW WE HAVE A WORKFLOW TO
DETERMINE THE STRUCTURES.
AND HERE THIS IS WORK FROM A
COUPLE OF YEARS AGO SO THESE
ARE THE AFTER SITES.
AND THEN YOU CAN SEE CLEAR
DENSITY.
THERE ARE THREE IMPORTANT
FEATURES OF THE COMPOUNDS.
THERE ARE THREE
ELECTRONEGATIVE ATOMS THAT SO
THEY UNLOCK THE D ACTIVE
SITES.
BUT ON THE RIGHT SIDE THIS TAX
ON THE VIRAL DNA.
IT MAKES UP THAT AT THE SITE
AND THAT'S WHY THEY ARE
COMPOUNDS.
ON THE LEFT SIDE TO THE
COMPOUNDS OF THOSE VARIANCE.
BUT THEY ARE TILTED AND THAT
IS SIMPLY BECAUSE THE NEGATIVE
ADAMS IS EMBEDDED INTO THE
CORE SO THE RIGHT SIDE IS
IDENTICAL AND THE LEFT SIDE IS
THE REGION OF FIRST POTENCY.
SO THAT IS GREAT.
WE DISCUSSED HOW THEY BIND TO
THAT STRUCTURAL BASIS.
AND OVER A DECADE.
AND OF COURSE A LOT OF THIS
WAS KNOWN FROM EARLY ON ALL
THOSE MECHANISMS.
BUT ALSO WHICH THE HIV
DEPENDENT HOW YOU ALIGN SO
THEY ARE NOT DISTINCT IT IS
THE ACTIVE SITE OF THE HIV AND
THEN IT IS COLORED BUT YOU CAN
APPRECIATE THAT THERE ARE
REGIONS THAT ARE SIMILAR BUT
OTHERS THAT ARE DISTINCT ALL
THESE MESSAGES ARE IMPORTANT
DRUG-RESISTANT PERIOD ON —-
VARIANCE THAT ARISE IN THE
POPULATIONS.
AND THOSE THAT ARE NOT
IDENTICAL.
SO ONE OF THE CONSEQUENCES IS
THAT IF YOU TAKE THE EXACT
SAME COMPOUND AND WITH THAT
AND THEN AND THAT IS RATHER
DISTINCT AND THEN THE SAME
THING HAPPENS AND THEN NOT TO
ASK IT WE ARE BEGINNING TO
FIND OUT SO THAT'S WITH THE
NATURE OF THE ACTIVE SITE
THERE IS A DISPARITY HERE SO
THEN THEY ARE PERFORMING WITH
THOSE PROPENSITIES FOR THE
DRUGS AND THE PRELIMINARY DATA
THEN WITH THIS COMPOUND TO
EXTEND OUT.
AND TO VALIDATE THIS.
SO THIS LEADS TO THE SECOND
PART THERE ARE MANY
DRUG-RESISTANT VARIANCE THIS
IS A SNAPSHOT FROM THE
PUBLICATION THERE ARE HUNDREDS
IF NOT THOUSANDS OF DRUG
RESISTANCE ANY WITH THE CLOSE
COLLABORATION SO WE SELECTED
THOSE THREE POSITIONS.
AND THEN WHAT STEVE SMITH DID
THE SINGLE AND DOUBLE AND
TRIPLE MUTANT AND THEN THE
POTENCIES OF THE
SECOND-GENERATION AND THEN TO
HIGHLIGHT TWO THINGS IT WAS
MORE POTENT SO WE FOUND SOME
WEAK SPOTS AND THAT IS WITH
THE MUTATIONS.
AND THEN WE WILL TALK ABOUT
THIS NOW.
SO COMPREHENSIVE VIROLOGY AND
THEN TO PRESENT WHAT WE ARE
FINDING FOR STRUCTURAL
VIROLOGY ALONG THE PATHWAY
THAT THIS WAS THE TRIPLE
MUTANT SO WHAT ARE WE FINDING
FROM THE STRUCTURE OF VIROLOGY
THERE ARE FOUR IMPORTANT
RESIDUES AND THEN HISTIDINE
THAT IS NOT A RESISTANT
VARIANT SO LET'S LOOK AT THE
SINGLE-USE SO THAT SINGLE
MUTANT TO MAKE THAT
INTERACTION AND THEN TO
COMPARE THAT SO WITH THE G1 40
MUTATION AND THEN THAT LEADS
TO THAT CONFIGURATION.
[INAUDIBLE]
2148 K MUTATION THERE IS A
SHIFT IN THE BACKBONE SO THOSE
THAT ARE SHIFTED OUTWARDS AND
HIM BEING 114 WITH THE EFFECTS
OF 138.
THIS IS A STRUCTURE OF
DETRIMENTAL MUTATION SUPPORTED
BY AND TO MARGIE AND VIROLOGY.
AND THEN AS A SINGLE MUTANT
WITH THE POSITION OF 148 AND
THAT DOUBLE MUTANT IS
INTERESTING BUT IF THE RESIDUE
IS THAT POSITION IT DOESN'T
HAVE THE DETRIMENTAL EFFECTS.
WE THINK THIS ONE THAT WE
CONFIGURATION OF THE
MOLECULES.
BUT THAT WE DON'T COMPLETELY
UNDERSTAND THIS THAT THE
ENTOMOLOGY AND VIROLOGY SEEMS
TO BEHAVE.
SO WITH Q1 48 K IT IS
INTERESTING IN TERMS OF
RESISTANCE SO THAT INFLUENCE
POSITION BUT NOW IT EXTENDS
OUT.
I FORGOT TO MENTION THESE ARE
THE TWO MAGNESIUM'S.
IT AFFECTS THE LOCAL CHARGE
DISTRIBUTION OF MAGNESIUM AND
THEN WE SEE THE SAME THING.
SO RESISTANCE ONLY ARISES WITH
THE MUTATION.
SO IF YOU HAVE THE SINGLE
MUTANT OR THE DOUBLE MUTANT.
SO NOW IF YOU HAVE THAT DOUBLE
MUTANT BUT THEN IT EXTENDS OUT
TO THE TWO MAGNESIUM IONS
THAT'S WE SEE SUBSTANTIAL
DIFFERENCE IN POTENCY.
WE THINK THE LOCAL CHARGE THEN
THE DISTRIBUTION IS AFFECTED.
SO NOTICE THAT FORWARD THE
REMAINS POTENT SO THEN IN
ADDITION WE NOW HAVE
STRUCTURES OF THE VARIANT AND
THE HISTIDINE VARIANT THAT
ARISES IN PATIENTS QUITE
FREQUENTLY BUT THIS ONE MOST
FREQUENTLY THAN OTHERS.
AND WHAT WE ARE FINDING IS
THAT PROXIMITY IN THE CHARGE
THAT MATTERS.
AND IT'S QUITE CLOSE AND THE
DRUG LOSES SIGNIFICANT
POTENCY.
AND IT REMAINS A LITTLE MORE
POTENT THAT IS THE FURTHEST
AWAY AND THE WEAKEST OF THE
THREE AND THE DRUG REMAINS
HIGHLY POTENT SO THEY CAN BE
PRIMARILY ATTRIBUTED Q1 48
THIS IS SIMILAR AND THERE ARE
SOME SIMILARITIES FROM A FEW
YEARS AGO.
AND WE HAVE MORE INSIGHTS NOW.
AND WITH THAT COMPREHENSIVE
NATURE OF THE STUDY.
AND THIS IS REORIENTING THE
POSITIVELY CHARGED POSITION.
IT IS A POSITIVE CHARGE TO
DRIVE RESISTANCE SO AS THE
LAST FEW SLIDES ESPECIALLY THE
DRUG-RESISTANT VARIANCE.
GETTING BACK TO THE PLOTS NOW
FOR THE REMAINS HIGHLY POTENT
AGAINST THE DEVIL AND THE
TRIPLE MUTANT.
SPECIFICALLY LOOKING AT THE
TRIPLE MUTANT AND THERE ARE
SOME INSIGHTS WE ARE JUST
LEARNING ABOUT AND WE JUST
DESCRIBED PREVIOUSLY WITH THE
CHEMICAL FEATURES OF THE
DRUGS.
SO I WILL JUST HIGHLIGHT ONE
NEW ASPECT AND THIS IS A
RECENT DEVELOPMENT.
SO I MENTIONED THAT VIRAL DNA
FORMS THE TERMINAL DNA COULD
EITHER HAVE ONE WITHOUT ONE
CONFIGURATION BUT THAT IS NOT
CONTRIBUTING SOMEWHAT WE ARE
FINDING THAT TRIPLE MUTANT
THAT ADAPTS MULTIPLE
CONFIRMATIONS BUT WHEN THAT IS
FOUND IS LARGELY IN THE STACK
THIS ISN'T JUST SPECIFIC TO
THESE TWO STRUCTURES IN FACT
WE SEE THIS ACROSS THE BOARD.
AND THEN IT MAKES SENSE FROM
THE PERSPECTIVE WHEN YOU HAVE
THAT STACKED CONFIGURATION SO
THE IDEA AND THEN AT TEMPLE
UNIVERSITY SO THIS IS AN
EXPLANATION FROM THOSE DYNAMIC
CONSIDERATIONS AND THEN YOU
CAN HAVE THE ENZYME TO HAVE
THE STACK CONFIGURATION IS A
THERMODYNAMIC SITE AND THEN
THEY TAKE THAT EXPERIMENTAL
OCCUPANCY AND THEN WITH THE
DELTA GEO.
OF NON- STACKING.
AND THAT'S TO SUPPORT THE IDEA
WITH A CONSTRUCTIVE INTERFACE
WITH THOSE WITH A RESISTANT
VARIANCE AND THEN TO IDENTIFY
WAYS OF COMBATING DRUG
RESISTANCE AND THEN HIV THE
THAT CAN PROBABLY TRANSLATE.
AND THE CENTRAL HYPOTHESIS IS
SEVERAL WELL-DEFINED
TREATMENTS AND THEN TO HAVE A
COMPREHENSIVE UNDERSTANDING
THEN THE WITH THOSE EFFICACIES
ACROSS BIBLE SUBTYPES.
SO THANK YOU THERE IS A LOT OF
PEOPLE TO THINK MANY ARE IN
THE AUDIENCE.
AND THEN AS NOW AS A POSTDOC
IN MY GROUP AND ALL THOSE THAT
THE NCI NOT JUST AS A
COLLEAGUE AND THEN TO CONVINCE
ME WHEN YOU THINK ABOUT THIS
WORK IN THE BEST POSSIBLE WAY
AND THEN SEE A MENTOR AND
COLLEAGUE OVER THE YEARS.
THANK YOU.
[APPLAUSE]
>>AND THAT LOW-KEY
INTEGRATION THOSE THAT
OCCURRED NATURALLY.
AND WITH THOSE RETROVIRUSES
AND MAYBE EVEN SOME THAT YOU
HAVE LOOKED AT.
>>THAT'S A GOOD QUESTION.
[LAUGHTER]
I WILL HAVE TO GET BACK TO YOU
ON THAT.
AND WITH THE HIV THAT'S A
LITTLE SURPRISING.
AND THEN BASED ON GENOMIC AND
IT SEEMS TO BE THERE IS
ANOTHER WAY TO SLICE THAT.
>>WE ARE FINDING THAT THERE
ARE SIMILARITIES OF COURSE AND
THOSE THAT ARE ABOUT
40 PERCENT CONCERNED OF COURSE
THERE IS IMPORTANT DIFFERENCES
OF THE Q1 86 IT IS A RESCUE
THAT HAS AN IMPORTANT AND
LARGE AFFECT.
SO I GUESS WE HAVE NOT THOUGHT
EXTENSIVELY WITH INHIBITION OF
OTHER RETROVIRAL'S BUT PERHAPS
WE SHOULD THINK MORE OF THOSE
IMPLICATIONS.
>>I WOULD BE ON THE LOOKOUT.
>>I AGREE THAT THIS IS OF
INTEREST.
>>
[INAUDIBLE]
THERE IS NOTHING FUNDAMENTAL
FOR THOSE REACTIONS.
>>SO IT IS QUITE CONCERNING.
WE THINK IT HAS TO DO WITH A
POSITIONING WITH THE PROXIMITY
TO CARRY OUT THE INTEGRATION.
SO THEN THE FILE HAS TO BE
POSITIONED IN THIS SPECIFIC
WAY IN A MEETING AFTER THE
STRUCTURE AND THE POSITION.
>>THANK YOU VERY MUCH.
GREAT WORK.
[APPLAUSE]
WE WILL NOW TAKE A 15 MINUTE
BREAK.
[INAUDIBLE CONVERSATIONS]
>>WE SHOULD START TO FILTER
IN AND TAKE OUR SEATS.
OKAY WE SHOULD GET STARTED
AGAIN SO WE STAY ON TIME.
THE NEXT SPEAKER FROM SCRIPPS
RESEARCH IN LA JOLLA
CALIFORNIA PETER HAS BEEN A
WORLD LEADER IN MOST RECENTLY
FOCUSING AND JOINING US
REMOTELY AS YOU CAN SEE THIS
WILL GO SMOOTHLY THANK YOU
VERY MUCH.
>>THANK YOU FOR INVITING ME
AND TO PARTICIPATE.
[INAUDIBLE CONVERSATIONS]
>>I WANT TO TALK ABOUT THE
COLLABORATIONS IN SCIENCE AND
THIS ECHOES SOME OF THE
THOUGHTS EXPRESSED TO ENGULFED
IN THE COLLABORATIONS AND IT
STARTED AND CONTINUES TO THIS
DAY.
ONE OF THE HIGH POINTS WAS THE
BISHOPS LAB IN SAN FRANCISCO
AND ALL OF THAT COOPERATIVE
AND THAT MARKS WITH MY TIME IN
SCIENCE AND OF COURSE I WAS
ABLE TO PARTICIPATE IN WORK ON
THE VIRUS THAT YOU LEFT ME
WITH A GIFT.
SO OVER THE PAST 20 PLUS YEARS
AND HAS BEEN AN IMPORTANT
FACTOR IN MY SCIENCE.
THANK YOU STEVE.
SO THEY ARE STARTED AND THEY
GROW AND FLOURISH AND THEY
BECOME DORMANT FACED WITH NEW
COLLABORATIONS SO WHEN I
JOINED SCRIPPS I WAS FORTUNATE
TO BE HOUSED AT THE ONLY
BIOLOGY LAB.
ANDERSON'S THE INTERACTION
THAT IS IMPORTANT SO NOW LET
ME TURN TO MORE SPECIFICS.
I WANT TO TALK ABOUT SOME OF
THE RECENT DATA.
AS YOU MAY KNOW EXISTING AS A
SUBUNIT AND P1 TEN AND WE
TALKED IN THE GENERATION THIS
IS FIRST LINKED TO CANCER THAT
WAS CARRIED OUT IN THE TUMOR
VIRUS THAT USED BY THE DNA
TUMORS AND LATER I DISCOVERED
THE GENE IN THE RETROVIRUS AND
THEN WE EXPRESS THAT BUT THEN
ALSO INDUCE TUMORS INTO THE
ANIMAL THEN FINALLY THAT IT IS
MUTATED FREQUENTLY AND THEY
CONFER TO THE PROTEIN AND ALSO
INDUCE TUMORS.
SO THESE FINDINGS MAKE THEM A
POPULAR AND PROMISING TARGET.
BUT AFTER MORE THAN 20 YEARS
AND MORE THAN INHIBITORS WE
HAVE NO BREAKTHROUGH.
WE HAVE FIVE FDA APPROVED
INHIBITORS.
FOUR OF THESE ARE EXCLUSIVELY
ONLY ONE WORKS WITH SOLID
TUMORS AND THEN TO USE THE
REFERENCE AND THE TERMINOLOGY.
>>I'M SORRY I HAVE TO STOP
YOU FOR A SECOND.
YOU ARE NOT SHARING YOUR
SLIDES.
>>ACTUALLY I AM STILL TALKING
ABOUT INTRODUCTORY STUFF.
>>I'M SORRY.
>>I AM NOT ON THE DATA SLIDE
YET.
THAT WILL COME IN THE SECOND.
>>OKAY.
SORRY.
>>SORRY ABOUT THE LONG
INTRODUCTION BUT I WANTED TO
COMMENT ON THE IMPORTANCE OF
COLLABORATION.
SO TALKING ABOUT INHIBITORS
TALK ABOUT ONE SPECIFIC
INHIBITOR THE BENEFITS FOR THE
PATIENT COMES AT THE COST OF
SIGNIFICANT RISK AND THE SIDE
EFFECTS REPRESENT INHIBITION.
SO THE INHIBITOR HAS NOT
DISCRIMINATED AND THAT IS
INJURIOUS BUT THE OBVIOUS
SOLUTION IS TO MAKE THOSE
SPECIFIC INHIBITORS THAT THERE
IS A PROBLEM USUALLY WE DON'T
HAVE THE CHEMICAL SPACE THAT
IS NEEDED.
SECOND WE DON'T HAVE THAT
STRUCTURAL INFORMATION THAT
DISTINGUISHES MUTANT.
WE HAVE PARTIAL INFORMATION
BUT NOT COMPLETE INFORMATION.
I WOULD LIKE TO TALK ABOUT THE
STRUCTURAL STUDY AND IT IS A
COLLABORATION WHICH IN THIS
CASE IS TRANSCONTINENTAL AND
AFTER FIVE YEARS WE HAD ZERO
RESULTS SO IN ORDER TO MAKE
PROGRESS WE NEEDED A LOT MORE
INSTRUMENT TIME THAN WE EVER
COULD GET LOCALLY AT SCRIPPS.
SO I TALKED ABOUT THIS WITH A
FRIEND IN SHANGHAI AND HE HAD
SEVERAL INSTRUMENTS SO WE
DECIDED TO COLLABORATE.
AND HAVING PERMISSION FOR THAT
COLLABORATION AND THE REQUEST
HAD TO GO ALL THE WAY TO THE
EMBASSY IN BEIJING BUT AT THE
BEGINNING WE HAD NO LUCK TO
THE PROTEIN AND THEN IF IT WAS
COLLECTED AT THE EDGE OF THE
SCREEN AND IT TOOK FIVE
MINUTES BEFORE THEY SAW
SOMETHING AND THEN THIS IS THE
FIRST PICTURE THAT WE SAW BUT
THEN WE HAD A MODEL.
SO THEN THE NEXT QUESTION IS
HOW CLOSE IS THAT MODEL TO THE
STRUCTURE?
IT IS VERY CLOSE BUT THERE ARE
SOME SIGNIFICANT DIFFERENCES
WE TRY TO IDENTIFY THE DOMAINS
HE WANTS TEN FOR THAT
CATALYTIC SUBUNIT AND THE
WELL-DEFINED STRUCTURALLY AND
FUNCTIONALLY.
P-LETTER 85 ALSO HAS FIVE
DOMAINS.
WE COULD IDENTIFY ALL OF THE
DOMAINS.
BUT ONLY TWO OF THE DOMAINS OF
P-85 SO THAT LEFT US AT A
POINT, AT THE SAME POINT THEY
COULD NOT SEE AND WE COULD NOT
SEE THAT EITHER AND THAT IS
EXTREMELY MOBILE AND FLEXIBLE
SO WE DECIDED TO USE A TRICK
THAT IS SOMETIMES SUCCESSFUL
TO BIND A SMALL MOLECULE TO
THE PROTEIN AND TO USE THIS
OFFICE SPECIFIC.
AND ON —- ALPHA SPECIFIC TO
SEE THE DENSITY OF ELECTRON
DENSITY AND WITH THESE
ADDITIONAL AREAS ARE ABOUT THE
EXPECTED SIZE OF THE MISSING
DOMAINS WE COULD NOT IDENTIFY
THOSE BECAUSE IT WAS SIMPLY
TOO LOW FOR WHAT WE DID SEE
HOWEVER WAS ONE OF THESE SAT
ON THE ACTIVE SIDE AND IN THAT
POSITION IT WOULD PREVENT FROM
APPROACHING THE CELL MEMBRANE
AND WOULD ALSO BLOCK GETTING
INTO THE ACTIVE SIDE.
SO THAT WAS THE INHIBITORY
EFFECT INDUCED BY BILL 719
THAT IS NOMINALLY A
COMPETITIVE INHIBITOR AND WHAT
WE HAVE SEEN HERE IS THAT IN
ADDITION FROM THAT COMPETITION
IT ALSO INDUCES HAS A SEPARATE
CONFIRMATION ALL CHANGE BY
BLOCKING THE ACTIVE SIDE.
SO THAT MADE IT VERY IMPORTANT
TO IDENTIFY THE IDENTITY OF
THIS PARTICULAR DOMAIN.
IN ORDER TO DO THIS WE HAVE
TWO APPROACHES AND WE USED
CHEMICALS WITH IT WAS
INTERESTING BECAUSE IT'S NOT
THE RESULTS THAT WE WANTED
WITH THE DOMAINS BUT TO
CROSS-LINK FOLLOWED BY
DIGESTION IS THE NUMBER OF
PEPTIDES THAT ARE ESSENTIALLY
HYBRIDS WITH PART OF THE
SEQUENCE DERIVED FROM ONE
GROUPING AND THE OTHER FROM
THE OTHER PROTEIN AND TO BE ON
THE IDENTITY OF THE CONTACT
PROTEINS.
AND IF WE DO THIS SO LET'S
SKIP THIS SLIDE WE DO FIND
THAT SITTING ON THE ACTIVE
SIDE IS THE DOMAIN THAT HAS
NEVER BEEN KNOWN TO BE
INVOLVED IN THE AUTOREGULATION.
AND OBVIOUSLY THAT HAPPENS AS
A SOLID POSITION ONLY IN THE
PRESENCE OF AN INHIBITOR
KNOWINGLY ON —- NORMALLY THIS
DOMAIN MOVES AROUND BUT THERE
IS AN ADDITIONAL BLOCK AND WE
ALSO IDENTIFIED AS AGE THREE
SO NOW WE WERE ENCOURAGED BY
THE DATA SO NOW WE CAN
ACTUALLY ATTACK THE MUTANTS.
THEY ARE MAPPED ONTO SIDES OF
THE CATALYTIC SUBUNIT TO
MUTATIONS ONE IN THE KINDNESS
DOMAIN.
SO THESE TWO ARE COMPLETELY
DIFFERENT SO WE LOOK AT THIS
BIOLOGICAL AND GENETIC METHODS
AND WE CAN SEE THAT KINDNESS
DOMAIN MUTATION DOESN'T
REQUIRE BINDINGS WE CAN MUTATE
AND THERE'S NO DIFFERENCE IN
ACTIVITY BUT IT DOES NEED PAD
FIVE SO IS NEEDED TO RECRUIT
THE PROTEIN SO THOSE ARE
EXACTLY THE OPPOSITE.
THEY ARE DEPENDENT ON
RESPONDING AND THAT MAY
MEDIATE THEY DO NOT NEED THAT
DOMAIN AND P-85.
SOME OF THOSE SUFFICIENT
AMOUNTS OF PROTEIN WITH THAT
GAIN OF FUNCTION.
WE HAVE TO GROW THE PROTEIN IN
THE PRESENCE AND THEN USE BILL
719 TO DO THAT WITH THAT
NEGATIVE FEEDBACK LOOP THAT
HAS HIGH CONCENTRATION FOR
GAIN OF FUNCTION'S YOU CAN
NEVER PRODUCE ENOUGH SO YOU
ALWAYS HAVE TO HAVE AN
INHIBITOR.
THIS BECOMES IMPORTANT LATER.
SO IF WE LOOK AT THE KINDNESS
WE RECEIVE A NUMBER OF CHANGES,E
CHANGES IN THE STRUCTURE OF
THE PROTEIN.
I WANT TO COMMENT ON SOME OF
IT.
THE MAIN CHANGES OCCUR AROUND
THE ORIGIN THAT GOES AWAY FROM
THE CORE OF THE ENZYME AND IT
ALSO ALLOWS THE KINDNESS
DOMAIN TO BECOME VERY
FLEXIBLE.
THAT IN TURN MENACES THE
LISTED BINDINGS SO THE
APPROACH IS ENHANCED AT THE
SAME TIME THE ACTIVATION LOOP
IS FROM WHAT IS ORIGINALLY
HERE AND THE ACTIVATION
CHANGING IS INCREASING ENZYME
ACTIVITY.
THE NEXT ITEM THAT WE SAW IS
THAT NOW HERE WE HAVE THE
PROTEIN AND BILL 1759.
SO THEN THIS TO THE KINDNESS
DOMAIN SO NOW AS THAT BECOMES
FLEXIBLE AGAIN WITH THE ACTIVE
SIDE OF THE KINDNESS DOMAIN IS
TOTALLY ACCESSIBLE.
AND HERE IS A DETAILED
ILLUSTRATION AND THAT BECOME
STREAM ON —- EXTREMELY
FLEXIBLE TO INTERACT WITH THE
CELL MEMBRANE.
AND THEN THAT AND WAS
TRANSLATED INTO FURTHER
CHANGES SO THOSE MULTIPLE
CHANGES AT ALL CAN BE
INTERPRETED TO THE GAIN OF
FUNCTION AND MY GUESS THAT ARE
INTRINSIC TO THE MOLECULE THE
INCREASE KINDNESS ACT WITH
IMPROVED RECRUITMENT.
BUT WE DON'T HAVE A UNIFIED
MOLECULAR MECHANISMS WE HAVE
CHANGES BUT WE DON'T HAVE THE
MECHANISM.
SO NOW LET'S SWITCH.
THERE IS A BIG SURPRISE SO THE
REGULATORY SUBUNIT AND THE ABD
SO THAT BRINGS THE QUESTION
ARE THEY STILL LINKED OR
COMPLETELY DISASSOCIATED WITH
THIS REGULATORY AND CATALYTIC
SUBUNIT AND THE ANSWER IS YES
THEY ARE.
AND YOU CAN SHOW THAT THEY ARE
TOGETHER SO THAT LINK BETWEEN
THE ABD IS NOT DISRUPTIVE.
BUT THE ABD BECOMES VERY
FLEXIBLE AND THAT FLEXIBILITY
IS INCREASED WITHIN P-85.
SO WE HAVE HERE A MECHANISM
THAT IS PART OF THE MOLECULE
AND TO BE DISENGAGED FROM
INTERACTION AND IN DETAIL THAT
DISENGAGEMENT IS TRIGGERED BY
THE MUTATION THAT DISRUPTS SO
WITH THAT DOMAIN AND THE
DISRUPTION DATES TO THE
DISENGAGEMENT.
ESSENTIALLY THIS IS ALL I WANT
TO SAY ABOUT THE EXPERIMENTAL
RESULTS.
IT IS CLEAR WE HAVE VERY
DETAILED INFORMATION WITH THE
STRUCTURAL DIFFERENCES BETWEEN
MUTANT.
SO WE LIKE TO COME BACK TO THE
QUESTION SO WE HAVE SHOWN THAT
INHIBITOR BY ITSELF SO THAT IS
DUE TO THE INHIBITOR TO GROW
THE MUTANT DOOR IS THAT TO THE
ORIENTATION?
AND THE BEST ANSWER WE CAN
GIVE IS WHEN WE DEFINE MUTANT
SPECIFIC DIFFERENCES USING
THAT AS A STANDARD BUT THE
PRESENCE OF THE INHIBITOR IS A
POSSIBLE COMPLICATING FACTOR
AND THEN THE NEXT STEP IS A
CHANGE OR IMPROVEMENT IN THE
ABSENCE.
SO ALREADY PHARMA AND STARTUPS
ARE STARTING TO MAKE MUTANT
SPECIFIC INHIBITORS WITH SOME
FOR SUCCESS BUT NONE OF THESE
CLAIMS HAVE BEEN SUBSTANTIATED
SO IT IS IMPORTANT TO LOOK IN
THE NEAR FUTURE TO HAVE SOUND
INHIBITORS BUT HERE ARE THE
PEOPLE WHO HAVE DONE THEIR
WORK SO THOSE DEVELOPING THAT
PURIFICATION PROCEDURE AND
THEN TAUGHT THAT PROCEDURE TO
THE COLLABORATORS IN SHANGHAI.
SO MOST OF THAT COMPUTATIONAL
WORK THAT IS NECESSARY AND TO
INTERPRET THE RAW DATA WITH
THE TEAM IN SHANGHAI INCLUDES
A GRADUATE STUDENT AND THEN
THE YOUNG SCIENTISTS TO SEND A
STATA AND THEN THAT PERSONAL
ACTION WITH HIM HE IS
EXTREMELY BRIGHT AND VERY
DEDICATED.
WE HAVE A VERY GOOD
RELATIONSHIP.
AND THEN WITH THE HEAD OF THE
INSTITUTE.
AND JAPAN DID MUCH.
SO WITH THIS I WOULD LIKE TO
AND AND BY ENDING AND WANT TO
WISH STEVE A WONDERFUL AND
SUNNY RETIREMENT.
THANK YOU I CANNOT HEAR YOU.
I THINK SOMEHOW WE ARE CUT
OFF.
I CANNOT HEAR.
I WILL STOP SHARE.
>>WE CAN STILL HEAR YOU
ONLINE.
>>WE CANNOT HEAR ERIC.
THE MICROPHONE AT THE
AUDITORIUM IS NOT WORKING.
[INAUDIBLE CONVERSATIONS]
IT IS REMARKABLY QUIET.
APPARENTLY THERE IS A
MICROPHONE IN THE AUDIENCE
THAT IS MUTED AND IT NEEDS TO
BE TURNED ON AGAIN.
>>YOU DID A WONDERFUL TALK.
I JUST THINK IT IS ODD THAT A
SYMPOSIUM IN DOCTOR HUGHES
HONOR IS KEPT INTENTIONALLY
SILENT FOR SO LONG.
[LAUGHTER]
SOMEHOW THAT SEEMS MAYBE THAT
WAS PLANNED.
>>HI PETER.
CAN YOU HEAR ME?
HOW MANY MUTATIONS ARE THERE
IF YOU WERE GOING TO MAKE
INHIBITORS TO SEPARATE
MUTATIONS?
>>ACTUALLY THAT ACCOUNTS FOR
80 PERCENT SO WITH THE
INHIBITORS AND ACTUALLY THERE
IS ONE INHIBITOR THAT EXPOSE
THE MUTATIONS THAT IT IS A
LITTLE HARD TO BELIEVE BECAUSE
THOSE ALSO STRUCTURALLY ARE
VERY DIFFERENT.
BUT ANYWAY IF YOU MAKE THE
INHIBITOR AND WITH 80 PERCENT
OF THE PATIENTS.
>>THANK YOU.
>>THEY ARE SILENT AGAIN BUT
TO FOLLOW UP ON THAT WHAT IS
THE LIKELIHOOD TO SELECTIVELY
TARGET THOSE DOMAINS?
IT SEEMS LIKE THAT IS
CHALLENGING.
>>THERE ARE ACTUALLY QUITE A
FEW STRUCTURES AND WITH THOSE
FIGURES.
>>IT SOUNDS LIKE WE HAVE THE
AUDITORIUM BACK.
>>IT SHOULD BE POSSIBLE TO
MAKE THE INHIBITOR.
>>THANK YOU.
>>'S OVER THE PAST 36 YEARS
WANT TO REFLECT ON THE
ADVENTURES WE HAVE HAD
TOGETHER.
BUT BASICALLY WE HAVE BEEN
ABLE TO LEVERAGE BASIC
RESEARCH INTO HELP TO CREATE
SOME OF THE DRUGS FOR TREATING
HIV INFECTION.
I HAPPEN TO HAVE GREAT FORTUNE
WORKING IN BOSTON AS A POSTDOC
THIS WAS IN THE MID- EIGHTIES
AND THIS LED TO SO MANY
INSIGHTS ABOUT VIRUSES HOW
THEY WORK AND FUNCTION AND
ASSEMBLE.
AND WHEN HIV WAS ALL IN THE
NEWS AND THEN TO HAVE A DEAF
SENTENCE AND CLEARLY SOMETHING
NEEDED TO BE DONE I WAS VERY
COMMITTED TO USE MY SKILLS
WITH THAT DESIGNED TO HELP
FIND A SOLUTION AND I MET
STEVE AT A VERY FORTUNATE TIME
HE WAS ALREADY EXTREME —-
EXCEEDINGLY TALENTED NATURAL
VIROLOGIST WITH SUCH WONDERFUL
ORIGINS.
AND HE HAD MUCH THE SAME
PASSION AS I DID.
WE MET INITIALLY WITH THAT
INFRASTRUCTURE OF THE
RHINOVIRUS 14 AND THEY ALSO ON
THE VISIT WITH STEVE HUGHES SO
WE TALK ABOUT THINGS WITH
RETROVIRUS AND EXPRESSION SO
THAT WAS BEYOND MY TIME SO
IT'S VERY PASSIONATE TO BE
EXTREMELY TALENTED SO THAT THE
RETREAT IN THE MOUNTAINS SO
THAT WAS A FUN TIME AND THAT
STEVE SAID THEY HAD JUST
SUCCESSFULLY EXPRESSED.
AND I SAID WAIT I REALLY
WANTED TO WORK ON THESE
PROTEINS AND THIS IS EXACTLY
HOW I WANT TO GET MY LAB
STARTED.
I DIDN'T HAVE A JOB YET BUT
FOR THE TENURE-TRACK POSITIONS
BECAUSE OF THIS COLLABORATION
AND FORTUNATELY I MOVED TO
WHERE I COULD IMMEDIATELY IN
FACT DROVE TO THE VERY FIRST
WEEKEND AFTER ARRIVING AND
THEN TO MAKE MESSES THAT THIS
WAS A VERY CHARGED BEGINNING
BUT THEN THERE WERE SOME DARK
YEARS WE SPENT TIME AND MONEY
TRYING TO GET CRYSTALS AND
SPENT FOUR YEARS WE ENDED UP
WE TRIED DNA.
WE TRIED TO WORK WITH P60 SIX
TO AVOID THE COMPLEXITIES BUT
AFTER A YEAR AND A HALF OF THE
CRYSTALS GROWING THE ONLY ONES
THAT DIFFRACTED AT ALL THIS
WAS IN 1989 AND WE WERE
WORKING WITH THE HETERO SO
DURING THIS ENTIRE PERIOD THE
PROTEIN WAS PRODUCED IN
STEVE'S LAB AND ALSO VERY
INVOLVED IN THIS AND IN TERMS
THAT A LOT OF THE GROUNDWORK
HAD BEEN MADE FOR THE
RETROVIRUS SOME OF THE FIGURES
HERE IT WAS A THERAPEUTIC
TOPIC STUDYING BASIC
BIOCHEMISTRY LIKE A GREAT
BASIS AND NERVOUS OF THE
COMPLEXITY AND WE WILL JUST GO
GET IT.
BUT NOT UNTIL 1992 THAT WE
START THIS WAS 180 WITH THAT
PART STRUCTURE AND IT WAS ONLY
PART OF IT BUT NATURE TOOK IT
BECAUSE IT DID NOT KNOW.
[LAUGHTER]
BUT THEN TO USE THAT AS A
YARDSTICK.
SORRY FOR ACTUALLY FIGURING
OUT AT THE SAME TIME WE HAVE
MUCH BETTER RESOLUTION DATA
BUT THIS WAS AFTER FOR A VERY
DARK YEARS AND THEY WERE
WORKING AND THREE OR FOUR HAD
THE SOLUTIONS BUT THIS WAS TOO
IMPORTANT.
SO ANYWAY WE SUCCEEDED TO HAVE
THE FULL STRUCTURE AND THAT
WAS PHENOMENAL.
THIS WAS FOLLOWING A
PUBLICATION WITH THE INHIBITOR
THAT THEY COULD DESCRIBE THE
STRUCTURE WITH COMPLETE
SIDECHAINS AND OF COURSE THIS
IS THE FIRST STRUCTURE WITH
ANY NUCLEIC FOUND ONE —- BOUND
SO BE LEARNED HOW THE RACES
WORK AND HOW TO ENGAGE IT WAS
VERY INFORMATIVE.
AND BRIEFLY WITH THAT AND
RANGE ON —- WITH THAT
ARRANGEMENT IT WAS P6 SIX ONLY
BUT THOSE STRUCTURES VERY
BEAUTIFUL INCLUDING THE RIGHT
HAND ARRANGEMENT THAT IS THE
MOST IMPORTANT PART BECAUSE
THAT IS THE CATALYTIC
INCORPORATION A BEAUTIFUL
ASPECT OF THOSE STRUCTURES
THAT WE PROCEEDED TO OBTAIN
ALSO IN COLLABORATION WITH THE
REST OF THE WORLD BY THAT TIME
AND THEN WE CAN GET IN
UNDERSTANDING OF THAT
MECHANISM AT THE ATOMIC LEVEL
AND PART OF WHAT WAS INVOLVED
WITH MASSIVE CHANGES IN THOSE
ARE THE MANY DIFFERENT TYPES
OF CRYSTALS IT HAS ALLOWED US
TO HAVE MANY DIFFERENT
INSIGHTS IT IS VERY SIMILAR
WITH A TWO DECADE EARLIER
LEVEL BUT THIS ALSO ESPECIALLY
ALLOWED US TO DESCRIBE ANY
FUNCTIONAL SPACE THAT WAS
ELUSIVE SUCH OF THE STRUCTURE
WITH THE INHIBITORS THAT TOOK
US TWO DECADES TO GET.
PEOPLE THOUGHT WE WERE LAZY OR
STUPID.
AT THIS POINT IN YOUR
ATTENTION WITH THE GOLD
MARBLES AND I ALSO WANT TO
POINT OUT SOME OF THIS
ESSENTIAL ASPECTS OF THE
CATALYTIC MECHANISM AND WITH
THAT ACID RESIDUE AND THEN
THAT INCOMING INCORPORATES AND
THAT WAS CATALYZED.
AS YOU SEE THE CHEMISTRY
OCCURS.
WITH THE LOSS WITH THE
PHOSPHATE AND THEN ONCE GETS
THAT TRANSLOCATION.
SO THIS IS RELEVANT.
SO STEVE AND I WERE VERY
INTERESTED TO TRY TO USE THIS
INFORMATION TO ACCELERATE THE
PROCESS OF DRUG DISCOVERY.
TO INCORPORATE EMPLOYEES IN
THE PROCESS BUT THEN VERY MUCH
LIKE THE NORMAL SUBSTRATE AND
WE WILL TELL YOU MORE ABOUT
THEM BUT THERE ARE SIX THAT
ARE FDA APPROVED GOING INTO A
HYDROPHOBIC POCKET.
AND THOSE THAT WERE APPROVED
AND THAT NONE INHIBITORS
ESPECIALLY SO ONE OF THE
THINGS THAT STEVE AND I
ENTERED THE PROJECT WITH IS
THE APPRECIATION THAT THE
DRUGS COULD BREAK DUE TO
RESISTANCE SO WE ANTICIPATED
IT WOULD BE A MAJOR PROBLEM
FOR A REVERSE TRANSCRIPTION.
IT WAS RELATIVELY CONSERVED
TARGET SO WE SOLVE THE
STRUCTURES OF ALL THE DRUG
RESISTANCE IS ONE —- MUTANTS
AND IF YOU WANT TO DEFEAT THAT
SO TO DETERMINE THE MECHANISM
OF INNOVATION MOST OF THOSE
ARE FAIRLY STRAIGHTFORWARD BUT
IN THE CASE OF THE NUCLEUS BUT
NOT JUST AT THE ACTIVE SITE
BUT AND TO HAVE THAT MECHANISM
OF RESISTANCE AND WITH THAT
ENHANCED ABILITY AND THAT
COULD OCCUR EITHER THROUGH
EXCLUSION BUT IN ONE OF THE
PAPERS THAT WE PUBLISH
TOGETHER IN 1989 SO THERE'S
RESISTANCE MOST SITES HAVE GO
BELOW THE LINE BUT THEN WITH
OPPOSITE STEREOCHEMISTRY.
AND THEN WITH THAT CASE THAT
WAS ACTUALLY THE EXACT SAME
PLACE WHERE IT WOULD BE
LOCATED EXPLAINING WHY TO HAVE
GREATER RESISTANCE SO IN TERMS
OF ONE OF THE MOST FASCINATING
ASPECTS WE ARE HOPING THAT
WALTER SCOTT WAS INVESTIGATING
THE POSSIBILITY THAT IT WOULD
ACTUALLY EXERCISE AFTER
INCORPORATED AND THEN TO
UNDERSTAND THE MUTATIONS THAT
WERE ADDED DISTANCE AND TO SEE
THAT AROMATIC RING CAN STACK
WITH THE MOLECULE AND THE
ENHANCED BONDING WITH THAT
SOCIAL MECHANISM.
SO WHAT THEY WILL SHOW YOU IN
THIS DIAGRAM IS BASICALLY WHAT
IS HAPPENING AND THAT
PHOSPHATE AND THEN IT ATTACHES
TO THE PEACE SITE BECAUSE THAT
IS AVAILABLE AND THEN THAT CAN
OCCUR FOLLOWED BY
TRANSLOCATION YOU CAN ACTUALLY
HAVE ONE WAY OF REDUCING THAT
EXTENSION AND ANOTHER CAN
OCCUR.
AND THE OTHER IS IN THIS CASE
SO WITH THAT PETRO PHOSPHATE
SO WE HAVE DETERMINED THIS IS
EXCISED BY THE ATTACHMENT AND
THAT SUCH AS THE CLOSING AND
THEN WITH THOSE MUTATIONS TO
PROMOTE THAT PROCESS AND TO
BACK UP THE MECHANISM.
[INAUDIBLE]
BUT THEN THEY SHOWED EXACTLY
HOW THAT AND THEN TO STABILIZE
THE BINDING.
AND HOW WE COULD LEVERAGE THE
WORK AND WITH THAT HIV
INFECTIONS SO WHILE STEVE AND
I WERE SCHEMING IT WAS UP TO
PAUL JANSEN IN BELGIUM AND HE
BEGAN THE SEARCH OF THE
COMPOUNDS OF THE NON-
NUCLEOSIDE INHIBITORS.
SO WE BEGAN CRYSTALLIZATION
AND PROTEIN THAT STEVE WAS
PRODUCING AND IN 1890 DOCTOR
PAUL PUBLISHED A PAPER AND
WITH THE TIVO COMPOUNDS WITH A
TWO PAGE PROPOSAL AND SO WHAT
ARE THE POSSIBILITIES THAT WE
DON'T HEAR BACK AND THE ANSWER
IS NO SO I WAS ASKING SO WHEN
I COULD COME TO MEET AND
INSTEAD WE HAVE GOT SOMETHING
AND DOCTOR PAUL TURNED OUT TO
BE THAT STEVE AND I HAD A
CHANCE TO MEET WITH HIM
MULTIPLE TIMES HE TURNED OUT
TO BE A VERY WARM PERSON.
WHAT IS THE PH OF THE
SOLUTION?
AND THEN TO DO RETRO
SYNTHESIS.
HE HAD DISCOVERED 75 DRUGS IN
HIS CAREER AND IS SO TOTALLY
COMMITTED SO THROUGH THESE
COLLABORATIONS THAT MAKE THESE
THINGS POSSIBLE.
SO THEN HAVING THE STRUCTURE
AND THEN WE COULD SEE THESE
COMPOUNDS HAVE VERY SIMILAR
SHAPE IN COMMON BINDING AND
ACTUALLY THAT EXPANDED.
SO THEN WE WERE CARRYING OUT
STUDIES DRIVING THE SYNTHESIS
OF THE INHIBITORS BUT AGAIN
THIS WAS WITH PROTEINS AND
EVENTUALLY WE CAME UP WITH
COMPOUNDS THAT ARE STILL IN
USE TREATMENT TODAY BUT AND
THE DESIGN ESPECIALLY PROVE
THOSE CONNECTIONS AND THE
TYPES THAT WERE INVOLVED TO
THE EXTENT THAT THE EVENTUAL
COMPOUNDS HAD PHENOMENAL
CHARACTERISTICS.
AND ESPECIALLY DURING THE
PROCESS SO TO DETERMINE THE
STRUCTURES OF MANY OF THE
DERIVATIVES AND WE HAVE SEEN
IN A SEPARATE PROJECT IN THE
LATE EIGHTIES A POSSIBLE ROUTE
WAS FOR THE COMPOUND TO OTHER
UNDERGO CONFORMATIONAL
CHANGES.
SO TO CRYSTALLIZE THESE IDEAS
IN THE 2004 PAPER WHEN WE
SHOWED THE INHIBITOR FOR
EXAMPLE FOR RESISTANCE IT IS A
FLEXIBLE AND HAVE BETTER COULD
ADAPT TO CHANGES IF IT IS
RELATIVELY COMPACT SO IT IS
ONE WAY THAT YOU CAN OVERCOME
RESISTANCE AND THEN YOU CAN
GET TO MORE ACTIVITY AND THOSE
RESISTANT VARIANTS BUT THE KEY
WAS ALL THE WAY THE COMPOUND
WAS SCREAMING AGAINST THE
DRUG-RESISTANT VARIANT AND
EVEN THEN THE CALCULATIONS ALL
OF THOSE REUSED IN THE DESIGN
CALCULATIONS.
AND THE OUTCOME IS THAT THESE
INHIBITORS FOR THOSE HIV
MEDICATIONS AND THEN TO FIX
THOSE COMBINATIONS THAT HE
MADE ROYALTY-FREE ALSO
COMBINED WITH ONE OF THE
INHIBITORS AND THEN IT KEEPS
ON GIVING AND IN 2001 IT WAS
LIKE ONCE EVERY TWO MONTHS
CAME OUT OF A COMBINATION OF
THE INHIBITOR SO I ALSO WANT
TO MENTION THIS BEAUTIFUL
STRUCTURE IN COLLABORATION
WITH STEVE HUGHES THE PAST
SEVERAL YEARS THE WAY THAT WE
HAVE BECOME WITH THE
PROCESSING AND THEN IN THIS
FORM THAT WE TRY TO UNDERSTAND
THE STRUCTURE AND TO
UNDERSTAND MORE ABOUT POLY
PROTEIN.
SO AT THIS STAGE I LIKE TO
THINK THE INCREDIBLE TEAMS OF
PEOPLE SO THE LATE PAT CLARK
AND ESPECIALLY I CAN'T IMAGINE
AND IT IS FUNDED THROUGH THE
PROJECTS FOR THOSE PERIODS AND
STEVE HAS GENEROUSLY FOUND
WAYS TO FUND THROUGH NCI BUT
SO WITH THOSE IMAGES OF STEVE
WHO ALL HAPPEN TO BE FROM
ITALY TWO OF MY GREATEST
FRIENDS AND MENTORS STEVE AND
MIKE HERE WE ARE HAVING FUN.
BUT ANYWAY.
THANK YOU SO MUCH.
STEVE, I WISH YOU ALL THE BEST
AND ALL THE BEST ENJOYMENT IN
LIFE.
[APPLAUSE]
>>OUR TYPING HAS SLIPPED A
LITTLE BIT SO WE HAVE TO SKIP
QUESTIONS BUT YOU CAN ASK
DURING THE LUNCH BREAK.
SO LIKE ALL OF THE OTHER
SPEAKERS TODAY, JOHN NEEDS NO
INTRODUCTION THE LEADING
EXPERT OF ALL THINGS
RETROVIROLOGY AND FOR TODAY'S
EVENT THE FOUNDING DIRECTOR
WEDNESDAY BECAME DIRECTOR AND
STEVE IN PARTICULAR AND WITH
THAT HIV PERSISTENCE.
THANK YOU ERIC IT IS AN HONOR
TO BE HERE AND THEN TO HEAR
ALL OF THESE WONDERFUL STORIES.
[LAUGHTER]
SO I PROGRAMMED THIS A LITTLE
BIT DIFFERENTLY.
I WON'T SHOW TOO MUCH DATA
EXCITING THOSE STUDIES AT THE
VERY END SO DESPITE THE VERY
LONG FRIENDSHIPS.
SO THEN COLLABORATING ON ANY
RESEARCH PROJECT OVER THE LAST
FEW YEARS.
AND THAT CAME OUT VERY WELL.
>>STEVE AND ME OUR FRIENDSHIP
GOES BACK A VERY LONG TIME.
SO IN PARTICULAR WITH THIS
PARTICULAR THING BUT IT WAS
GOING AROUND THE PHRASE THAT
AS I THOUGHT ABOUT IT MORE AND
MORE A VERY LARGE AMOUNT WHAT
WE HAVE ACCOMPLISHED OVER THE
YEARS WITH DRP THAT RELATES TO
GETTING RID OF THE FEELING
THAT IS EXPRESSED IN THIS RACE.
THAT IS THE SHINY DOGMA.
[LAUGHTER]
I WILL ILLUSTRATE THIS BY
POINTING OUT BECAUSE I COULD
NOT FIND ANYTHING.
AND THEN THE SHINY DOG
PICTURES APPEAR.
SO I AM QUITE SURE BY 1977
THAT THIS TALK NOW MEETING THE
RETROVIRUS AND YOU CAN TELL
THIS IS A STUDY THAT HAROLD
WAS REFERRING TO AS SOME
LENGTH.
AND IT WAS A VERY IMPORTANT
DESCRIPTION THAT ORIGINALLY
THE GENOME.
AND THEN THE FACT THAT COULD
BE INTEGRATED.
>>AND SO WITH THE INTEGRATION
SITE THE QUESTION TO
UNDERSTAND WHAT'S GOING ON
WITH THE HIV INFECTION.
SO SO IN FACT AFTER SOME YEARS
WE BECOME FAMOUS FOR ALWAYS
SITTING IN THE FRONT EACH ONE
WHO ALWAYS SHOWED UP FIRST
WOULD PUT A MARKER ON BOTH OF
THOSE SEATS TO CONSUME THE
TRADITION YEAR AFTER YEAR I
DON'T KNOW HOW FAR BACK IT
GOES BUT IT GOES BACK A LONG
TIME.
SO AS WE PLAY THAT OUT WE
BECOME FAIRLY WELL-KNOWN FOR
THIS.
AND THE OTHER THING THAT WHAT
HAROLD ALLUDED TO WAS THE
RETROVIRUS BOOK THAT THE THREE
OF US WORKED OUT FOR A LONG
TIME IN THE BOOK WAS PUBLISHED
IN 97 ABOUT THE TIME WITH NCI
TO FORM THE DRP.
STARTING WITH A MEETING IN
WASHINGTON DC AND HAROLD HAD
JUST BEEN APPOINTED THE ANY
AGE DIRECTOR AND SIMILAR TO
THE POT CIRCLE SO WE ACTUALLY
LAID OUT THE WHOLE STRUCTURE
OF THE BOOK AND WE PRETTY MUCH
ENDED UP BEING THE FINAL
CHAPTER WITH A LIST OF AUTHORS.
AND THE MEETING WAS QUITE
PRODUCTIVE BUT THEY HAD
FLIGHTS TO CATCH AND I HAD TO
GET HOME AND HAROLD I
RECOLLECT WOULD GIVE A BIG
LECTURE AND THEN TO BE GREETED
WITH THE RED CARPET.
BUT THEN IT RAN A LITTLE LONG.
BUT IT WAS ONLY IN VERY SHORT
TOPICS.
TO THE NATIONAL AIRPORT BUT
GOING IN THE WRONG DIRECTION
AND THEN GETTING MORE AND MORE
ANSI.
AND SAY HAVE A MAP IN THE
TRUCK.
AND THEN SAID HURRY DON'T
STOP.
>>AND THEN TURNING UP ON ROCK
CREEK PARKWAY EXCEPT THAT IT
TURNS AROUND AND GOES THE
WRONG WAY AT RUSH HOUR.
SO THEN FINALLY MAKING IT UP
TO THE RED CARPET.
AND THEN LENDING TO A QUESTION
TO RECOGNIZE THE THEME OF THIS
AND HAVE A MORE OR LESS LOST
ONE SIDE?
SO MANY WOULD AGREE IT IS
SUCCESSFUL.
THAT WAS THE STORY.
SO WHAT GOT ME INTO THE DRP.
SO AT THE TIME AFTER HIV WAS
DISCOVERED WITH THE VIRUS TO
SEQUENCE IT WAS MORE THAN
OTHER RETROVIRUSES THEN YOU
HAVE A LARGE AMOUNT OF GENETIC
DIVERSITY.
SO THAT WENT TO THE GENERAL
DOGMA.
AND THEN AT THE TIME IN THE
NINETIES AND PATIENTS WERE
CONSTANTLY FAILING THEM WITH
THE EXISTENCE AND WITH THE LOW
POTENCY SO I PUBLISHED THIS
REVIEW THIS IS THE SAME AS HIV
TO SHOW THE RATE AT WHICH IT
OCCURS IN THIS PARTICULAR CASE
I HAD DONE AN EXPERIMENT BUT
OF K BUT OF COURSE THE
PATIENTS WERE CHICKENS THERE
WAS VERY LITTLE DIVERSITY BUT
IF YOU TAKE THAT VIRUS THEN
YOU GET GENETIC DIVERSITY SO
THE CONCLUSION IS THAT HIV
DOES NOT HAVE THAT PATIENT
RATE AND THAT THE LIFESTYLE IS
CONSTANTLY REPLICATING OVER
AND OVER AGAIN.
SO THAT REPLICATION OVER AND
OVER IS THE PROPERTY OR THE
CONSEQUENCE THAT ALLOWS THE
ACCUMULATION IN THE POPULATION
AND DRUG RESISTANCE IS
SLIGHTLY DELETERIOUS AND WITH
THE VIRUS THAT THEY COME FROM
BECAUSE OTHERWISE THEY WOULD
BE SLIGHTLY FAVORABLE.
HE WOULD NEVER SEE AN
EFFECTIVE DRUG.
SO THE IDEA IS THAT THE
FAILURE OF HIV THERAPY IS NOT
DUE TO THE POTENCY OF THE
DRUGS THAT ALREADY RESISTANT
TO THE POPULATION.
SO THIS ATTRACTED THE
ATTENTION OF STEVE AND GEORGE
BECAUSE THE BUILDING HAD JUST
BEEN BUILT.
AND THEY WERE LOOKING TO FILL
IT.
THAT OCCUPIED A COUPLE OF
FLOORS BUT THERE WERE SEVERAL
FLOORS STILL TO BE FILLED.
SO DRUG RESISTANCE WOULD BE A
GOOD TEAM TO FOCUS ON AND THEN
OBVIOUSLY DEMONSTRATED TO
THINK ABOUT DRUG RESISTANCE
THE ONLY THING IS TO DO IT.
AND THEN MANAGED TO PERSUADE
ME THAT FOR PERSONAL REASONS
PART-TIME SUGGEST 25 YEARS AGO
WE GOT STARTED FROM 1997.
AND WITH THAT RESISTANCE
PROGRAM AND WITH THOSE CURRENT
PIS IN THE PROGRAM AND THEN IT
CAME TO BE OUR FAVORITE
RESTAURANT.
AND WITH THE SPEAKER AT THE
LECTURE A FEW YEARS AGO.
AND THEN THAT IS REMARKABLY
SUCCESSFUL AND THEN FOR THAT
TRANSFORMATIONAL BIOCHEMICAL
AND SO ON.
AND THEN AFTER HE RECRUITED ME
ONE OF MY FIRST RECRUITS.
[LAUGHTER]
BECAUSE THE PROGRAM THAT HE
WAS IN WAS CLOSED TO BRING
SOME OF THE STAFF INTO THE
NCI.
SO STEVE BECAME A VERY
IMPORTANT PART NOT ONLY
GETTING THE DRP GOING BUT A
VERY IMPORTANT MEMBER GOING
FORWARD AND THEN EVENTUALLY
INTEGRATES THE INHIBITORS AS
WELL SO BUNCH OF BASIC
STUDIES.
AND OVERALL WITH THE
DIRECTIONS OF THE PROGRAM I
WAS HELPING AND TO WORK MOSTLY
WITH THE CRITICAL GROUP AND
THEN WITH THE ORIGINAL GOAL TO
TAKE THIS POSITION IS WITH THE
PERSPECTIVE AND SO THAT WAS
ONE OF THE THINGS ALSO BETTER
THAN ON A MORE PERMANENT
BASIS.
BUT NOW IT WAS TRANSLATIONAL
RESEARCH SECTION AND THEN THE
FIRST GOAL WITH ADVANCED
TECHNOLOGY FOR HIGHLY
SENSITIVE ASSAYS.
AND THEN WITH THAT REPLICATION
AND SO ONE OF WHAT IS
HAPPENING.
AND ALSO AT THE TIME TO DISPEL
A FEW MORE SHAGGY DOGMAS.
FOR EXAMPLE A DOGMA THAT STILL
HAS LONG-TERM PERSISTENCE THAT
REPLICATION OF CELL TO CELL
TRANSFER.
AND WITH THOSE SANCTUARY SITES
AND THEN AS HAS BEEN SHOWN
REPEATEDLY WITH THE GROUP AND
COLLABORATORS AND IN THE FIRST
PLACE WITH A VERY LARGE LEVEL.
AND MOST INDIVIDUALS ON
THERAPY.
AND THEN THAT WAS SHOWN THAT
WASN'T SENSITIVE TO KEEP
THROWING DRUGS IT AND THEN TO
BRING THAT DOWN A LITTLE BIT.
AND THEN TO BE PUBLISHED IS
THE ATTORNEY GROUP.
SO IF YOU LOOK AT THE SYMBOLS
AT THE VIRUS SEQUENCE BEFORE
THEY STARTED A RETROVIRAL
THERAPY AND THAT WAYS IDENTITY
MY —- IDENTICAL AND TAKE THAT
OTHER PERSON THREE YEARS IN
THERAPY AND THE SEQUENCES AND
ALL OF THE INDIVIDUALS AND
THAT MEANS ABSENCE OF
APPLICATION TO ANY SIGNIFICANT
EXTENT.
THAT WE HAVE SUCH NICE DATA
SHOWING THIS.
THE PATIENTS HAD BEEN ON
THERAPY FOR A LONG TIME.
>>UP TO THIS POINT I HAVE
BEEN WORKING BASIC STUDIES ON
INTEGRATION BUT ALSO A LOT AND
THAT WAS INHIBITORS IN THE
RESISTANCE AND THAT WAS OVER
THE RTA F.
AND THEN WITH THAT INTEGRATION
FOR THE SITES AND THAT THIS IS
SENSITIVE ENOUGH AND THAT YOU
CAN STILL USE THAT WITH THAT
INTEGRATION SITES AND AT THE
SAME TIME FRANK BROUGHT THE
FAMOUS PATIENT.
I HAVE TO TELL YOU THIS PERIOD
ON —- PERSON WAS NAMED PATIENT
ONE NOW IT WOULD BE CALLED
PARTICIPANT ONE.
AND HE HAD A HISTORY AND WHERE
HE SHOWED UP FIRST SHOWING UP
AT THE CLINICAL STUDY THAT
FRANK WAS MANAGING AND
OBVIOUSLY HAVE BEEN INFECTED
FOR QUITE A LONG TIME SO HE
WAS NOT VERY HEALTHY AT THE
TIME HE SHOWED UP HE WAS PUT
ON A COMBINATION THERAPY AND
THEN RAPIDLY BROUGHT HIS LEVEL
DOWN AND THEN MAINTAINED WITH
CHANGING THERAPIES UNTIL HE
HAD A COUPLE OF PROBLEMS WITH
ACCESS TO DRUGS.
SO THEN THE VIRUS CAME BACK UP
AND WAS BACK DOWN AGAIN AND
THEN AFTER TEN YEARS DESPITE
THE FACT YOU ARE ON GOOD THERE
APPEARS STARTED TO GO UP.
SO FRANK RECOGNIZED THIS IS
BRILLIANT WITH THOSE OF
SEVERAL INTERESTING THINGS.
SO IT INVOLVES IN TO SHARE THE
DNA IN —- YOUR POINT WOULD BE
DIFFERENT.
AND THOSE THAT COME FROM THAT
ORIGINAL MOLECULE.
AND WITH THAT PRIMER.
AND THEN WITH THAT METHODS IF
YOU WANT AND WITH THAT
SEQUENCING YOU GET ONE
SEQUENCE AND THAT SHOWS YOU
ARE THAT DNA MOLECULE.
AND WITH THAT SAME INTEGRATION
SITE COMING FROM DIFFERENT
ORIGINALS CELLS.
AND FROM THAT ORIGINAL
INFECTED CELL.
AND WHAT WAS A BIG SURPRISE
AND THEN THE DAD CAME IN ON
THE IPHONE.
AND SAID AND WITH THOSE
DIFFERENT BREAKPOINTS I DON'T
THINK ANYBODY EVER THOUGHT OF
THE DOGMA AT THE TIME.
AND THEN TO BE BEFORE YOU
STARTED THERAPY.
AND THEN THE CLONES THE GRAY
ONES INDICATE YOU ONLY SAW IT
WANTS BUT THEN I YOU CANNOT
SAY THEY ARE CLONES.
I WOULD BE WILLING TO BET
EVERY SINGLE ONE OF THESE IS
OPERATING ON ITS OWN BECAUSE
THAT SENSITIVITY.
THAT LOOKING AT ONE IN
100,000.
AND SO THEY WOULD YOU GET DOWN
TO ONE YOU ARE SEEING 100,000
ARE GREATER.
SO THE ONE THAT IS MARK THERE.
SO WITH THESE DESIRES AND THEN
THAT VIRUS COMING UP SLOWLY IN
THE PATIENT THAT WE SAW AS THE
INCREASING PERSISTENCE.
AND THEN ADDING MORE DRUGS AND
THEN IT IS NOT A
DRUG-RESISTANT VIRUS.
BUT EVER THESE VERY LARGE
CLONES.
SO WHAT IS IT THAT ALLOWS
THEIR SURVIVAL AND ONLY VERY
QUITE A FEW FRACTIONS OF ANY
OTHER CLONES YOU COULD FIND
EVEN IN THE SMALL QUANTITIES.
AND THEN TO BE SUPPRESSING AND
THAT I VERY SKEPTICAL ABOUT
CELL CULTURE LOOKING AT THIS
FIREBALL LATENCY.
BECAUSE THOSE POPULAR CULTURE
LOOKING AT THE VIRAL DNA WHICH
IS A VERY COMMON METHODS OF
WORKING QUITE HARD.
AND THEN AND THEN TO MAKE THE
INFECTIOUS VIRUS SAID THAT CAN
SHOW VERY NICELY THAT YOU
COULD NOT SEE SO WHATEVER IT
IS HE STILL DON'T KNOW WHAT IT
IS WITH THE PRO VIRUS IS NOT
DNA METHYLATION AS IT IS THE
MOST POPULAR CELL CULTURE.
SO THIS IS STILL SOMETHING
THAT WE NEED TO WORK OUT.
THE FREQUENCY IS VERY VERY
SMALL ONE OUT OF 10000 AND A
GOOD ONE AND A GOOD
INDIVIDUAL.
SO THAT IS PRESENT WITH MORE
THAN 10 MILLION CELLS SO IF
YOU WANT TO GET RID OF THESE
THE CHALLENGES VERY LARGE.
AND THEN TO HAVE SPECIFIC
ANTIGENS BUT THEN THE OTHER
THING THAT CAME OUT OF THIS
AND THEN TO BRING OF THAT
IDEAL CONSTRUCTION.
AND THAT WAS NEVER THOUGHT OF
NOW PEOPLE ARE DOING GENE
THERAPY.
AND THEN WITH THAT COUPLE OF
GENES WITH THOSE DRAMATIC OF
CLUSTERS WITHIN A VERY SMALL
REGION AND THOSE THAT ARE
CLUSTERED.
SO IN A REGION IF YOU THINK
ABOUT IT A COUPLE OF TINY
AXONS IN THERE SO YOU CAN SEE
THAT IS NOT TO SURPRISING VERY
EASILY MISSED EVEN IF THEY
WOULD BE THERE THEY ARE ALL
POINTING IN THE SAME
DIRECTION.
SO THEN YOU GET INTEGRATIONS
AND THOSE DIRECTIONS AND NO
PARTICULAR BIAS MAY BE A
SLIGHT INTEGRATION BIAS BUT
THAT IS A DIFFERENT STORY.
SO WHEN YOU LOOK AT A BUNCH OF
THESE THOSE THAT HAVE THE SAME
PROPERTY OF CLUSTERED
INTEGRATIONS THESE ARE
EXPANDED LIKE EVERYTHING ELSE
BUT WITH THE EXPANSION AS I
WILL SHOW YOU IN A SECOND.
SO THIS IS RELATIVE TO THE
PROTEIN AND BASICALLY THEY ARE
DEREGULATING THE EXPRESSION SO
ACTUALLY IT IS INSIDE THE GENE
IT IS WELL-PLACED TO BE
REMOVING A NEGATIVE REGULATORY
DOMAIN AND THEN THAT ALL TIME
RETRO- VIROLOGY.
THAT'S CANCER FROM BEFORE.
AND THESE ARE CANCERS.
AND THOSE THAT YOU CAN SEE.
AND WITH THE PW H.
THEY ARE NOT CANCERS WITH
UNIMPEDED GROWTH AND DOING
SOMETHING ELSE.
SO ONE OF THE THINGS TO KEEP
IN MIND SO THIS IS A LIST OF
GENES THAT WE CAME UP WITH
FROM THE PAPER THAT WE
PUBLISHED A YEAR AGO.
SO WITH THAT INTEGRATION CAN
BE QUITE DRAMATIC IT'S ONLY A
COUPLE OF PERCENT SO IT'S NOT
A HUGE PART OF WHAT'S GOING ON
THE CHEST THE MOST STRIKING
PART THAT PROBABLY DOING
SOMETHING INTERESTING ONE OF
THE THINGS THAT WE DID WAS TO
LOOK AT THE PATIENTS ON
THERAPY WHEN IT'S A LITTLE
MORE COMPLICATED BUT SO WE
LOOK AT THAT AMPLIFICATION
RATIO SO TO HAVE A SITE THAT
IS AMPLIFIED YOU SEE THAT
TWICE IN YOUR SAMPLE THAT
WOULD HAVE THE AMPLIFICATION
RATIO.
SO ALL OF THOSE ARE AMPLIFIED
TO EXACTLY THE SAME EXTENT IT
IS NOT DRIVING THE
AMPLIFICATION IT'S BEING
DRIVEN BY CELL BIOLOGY AND THE
ENERGY RECOGNITION AND WITH
THE INFECTED CELL AND EVEN THE
GENES BECAUSE THE NUMBERS ARE
SMALL EVEN THOSE THAT WE
IDENTIFIED IF YOU LOOK AT THE
AVERAGE IN THE GENES THAT WE
IDENTIFIED TO HAVE THESE
CLUSTERS AND INTEGRATION SITES
IT'S NOT SIGNIFICANTLY GREATER
THAN FOR THE GENES OVERALL
THIS EXPERIMENT WAS NOT DONE
BY US BUT IT WAS DONE BY ONE
OF OUR FAVORITE ALUMNI AND
WHAT HE DID WAS TO ISOLATE THE
CELLS FROM THOSE PATIENTS BY
THEIR SPECIFICITY INCLUDING
HIV SO WE TAKE THOSE CLONE
CELLS IN THE TWO WHERE THE
INTEGRATION SITES WERE SO THAT
ALMOST ALL OF THESE ARE VERY
HIGHLY EFFECTIVE WITH ONE
EXCEPTION BUT IF YOU LOOK AT
THE RED ONE SO LOOK AT THOSE
IN THE GENES SO IN THIS CASE
OVER ONE THIRD OF THEM MORE
THAN ONE THIRD OF THESE HAVE
INTEGRATION SITES WITH THESE
GENES THAT THERE IS A SYNERGY
BETWEEN THE STIMULATION OF
THESE GENES AND ONE GUEST THAT
I HAVE IS NOT JUST THE GROWTH
OF THE CELLS BUT THE EXTENT TO
WHICH THEY ARE LOST DURING THE
DK PHASE BECAUSE THE GROWTH OF
THE CELLS FROM WHAT THESE
REPRESENT HAVE THE UP-AND-DOWN
PATTERN IF THE EXPRESSION OF
THE GENES IS INHIBITORY THEN
THEY WILL BE SELECTIVE BECAUSE
OF THAT SO THAT IS A GUESS
RIGHT NOW.
>>SO WE LEARNED SURVIVAL BY
THE CELLS THESE ARE FOR
IMPORTANT CONTRIBUTIONS AND
THEY WILL SHOW YOU THIS BUT A
BUNCH OF OTHER PEOPLE HERE
CO-AUTHORED A PAPER YOU COULD
ALREADY SEE WHEN THEY STARTED
THE ANTIRETROVIRAL THERAPY
WITHIN A FEW WEEKS IT MUST BE
UP TO 100,000 OR MAYBE
1 MILLION CELLS SO THE IDEA
THAT IT JUST DOESN'T WORK THEY
CANNOT HAVE BEEN RESTING VERY
LONG BECAUSE THEY ARE
PROVIDING CONSTANTLY AND ALSO
TO BE MODIFIED I HAVE NOT
SHOWED YOU THIS EITHER BUT THE
PATTERN OF INTEGRATION CHANGES
AFTER YOU START THERAPY
BECAUSE OF THE SELECTION THOSE
OF THE SAME ORIENTATION.
TALKING ABOUT CLUSTERS OF
INTEGRATION SO WITH THAT I
WILL CLOSE.
THANK YOU TO THOSE OF YOU WHO
ARE HERE AND HAVE DONE SO MUCH
WONDERFUL WORK OVER THE YEARS
AND TO SEVERAL PEOPLE AND OF
COURSE THANK YOU TO STEVE FOR
ALL OF HIS YEARS WITH US.
[APPLAUSE]
>>SORRY TO SLIP A LITTLE BIT
YOU CAN ASK QUESTIONS AT LUNCH
SO THE NEXT SPEAKER IS LARRY
WHO HAD THAT DISTINGUISHED
CAREER AND ALSO SERVED
ADMINISTRATIVE GOALS HERE IN
FREDERICK INCLUDING THE
PRESIDENT THERE ON THAT SHE'LL
BE GIVING US THE FIRST OF
SEVERAL MAD SCIENCE AND THERE
WOULD BE SOME FISHING SCIENCE
AS PART OF THE STOCK ONE —-
PART OF HIS TALK.
>>I MAY TALK A LITTLE BIT
ABOUT STEVE'S FISHING
CAPABILITY FIRST OF ALL
CONGRATULATIONS ON YOUR
RETIREMENT AND ALL OF THE
SCIENTIFIC ACCOMPLISHMENTS YOU
HAVE HAD DURING YOUR CAREER.
I WILL CONCENTRATE TODAY ON
STAGE PREPARATION FOR
POSTRETIREMENT.
HE HAS WORKED REALLY HARD AT
THAT.
[LAUGHTER]
AND IS QUESTIONING WHETHER OR
NOT HE IS DEDICATED.
SO WE MOSTLY HAVE FISH IN
WESTERN MARYLAND BUT LET ME
TELL YOU HE SENDS HIS
APOLOGIES TO NOT MAKE THE
TRIP.
WHEN BOB RETIRED HE TALKS HE
BY THE HOUSE IMPORTANT SHORT
ON —- PORT CHARLOTTE THE LAST
HURRICANE THE I WENT RIGHT
OVER THERE HOUSE.
I AM GLAD TO REPORT THEY
SUFFERED MINOR DAMAGE.
THEY NOW HAVE ELECTRICITY AND
WATER IN THEIR HOUSE.
BUT UNFORTUNATELY BOB CANNOT
ATTEND BUT HE SENT ME SOME
SLIDES AND I USE THOSE IN MY
PRESENTATION AS WELL.
SO LIKE I MENTIONED WE DECIDED
IN 2061 OF HIS FRIENDS JOINED
US AS WELL SO THE FOUR OF US
MONTHS BEFORE I HAD BEEN NOT
THAT WE HAD NOT BEEN ON THE
RIVER.
SO WE FLEW TO BATTLE WITH THE
MATERIAL THAT WE NEEDED AND HE
FLEW WAS UP TO THE TOP OF THE
LAKE AND THEN TO PUMP UP THE
RAFT.
AND NONE OF US HAVE EVER WROTE
A RAFT LIKE THIS BEFORE.
SOMETIMES WE WORRY ABOUT THE
DECISION-MAKING.
SOMETIMES WE WORRY ABOUT THAT
EVEN MORE.
BUT WE FOUND OUT QUICKLY BY
SAYING GOODBYE TO THE FLOOD
PLANE BUT THEN WE REALIZED WE
WOULD NOT SEE ANOTHER PERSON
FOR FIVE OR SEVEN DAYS OF OUR
TRIP.
WE DID NOT EVEN SEE A CAMPSITE
THIS ENTIRE TIME THAT WE DID
FIND ONE MORE ELEMENT.
HE WAS QUITE GOOD AND IS
IMPORTANT BECAUSE THE SNOWFALL
THAT THEY GET NOTES IN THE
SPRING AND THEN IT BRINGS NEW
CHANNELS AND THEY HAVE TREES
HANGING OVER THE BRAIDS.
THEY ARE CALLED SWEEPERS FOR A
GOOD REASON.
HE HAS FOUND WAYS WITH AN
UNSCHEDULED SWIM YOU WILL
NOTICE HIS CLOSE ARE HANGING
AND DRYING OUT ANY QUESTIONS
JUST ASK STEVE.
[LAUGHTER]
THE OTHER THING WE FOUND OUT
RAPIDLY ON THE TRIP IS THAT
STEVE IS A CHEF EXTRAORDINAIRE.
ONCE WE FINISHED OUR DAY AND
SET UP CAMP TO BUILD A FIRE
GOT EVERYTHING READY PULL UP A
CHAIR STEVE OWEN SIT IN IT AND
TELL US WHAT HE NEEDED BE
WOULD BRING HIM EVERY SINGLE
THING HE NEEDED.
NOW YOU CAN SEE HE HAS GROWN
SOME SALMON BUT IT WAS PRETTY
EASY TO KEEP THE MASTER CHEF
AT WORK SINCE WE DIDN'T HAVE
TO CARRY ANYTHING ONCE WE
BROUGHT INTO THE RAFT WE HAD
STEAK AND BAKED POTATOES AND
SALADS AND HE MADE SPAGHETTI
AND THEN WE DID NOT SUFFER
FROM A LACK OF FOOD.
I SHOW HERE AND HERE WITH A
RAINBOW TROUT THAT HE GOT AND
ALTHOUGH FISHERMEN ARE KNOWN
TO STRETCH THE TRUTH THAT HE
GOT OVER 100 FISH ON THE TRIP
DOES NOT STRETCH THE TRUTH.
AND THEN AND THEN WE DECIDED
THAT WE WERE CHECKING OUT THE
FOLLOWING AND ALSO WE HAD A
CLOSER LOOK AT IT SO EVEN WITH
STEVE WE CANNOT RUN IT.
SO OUR LAST RAFTING TRIP TO
ALASKA THAT TURNED OUT TO BE
ONE OF THE BEST DECISIONS WE
HAD ADDITIONAL PEOPLE JOIN
US.
YOU ARE RECOGNIZED.
AND OF COURSE FUTURE SATELLITE
JOINS US AS WELL.
AND I HAD TO PUT THIS IN
BECAUSE IT WAS QUITE AN
ADVENTURE.
THAT WE HAD TO TAKE A SHORT
CUT AND TAKE EVERYTHING OUT OF
THE PLANE BECAUSE THEY HAVE TO
LAND IN THE TUNDRA AND THEN
THEY HAVE TO TAKE OFF AGAIN.
AS SOON AS THE PILOT LOOKED AT
STEVE SO THEY NEVER HAD ANYONE
LIKE THIS BEFORE THEN THE
CHILLS RAN DOWN MY SPINE.
[LAUGHTER]
BUT THE FLIGHT WAS ONLY A 30
MINUTE FLIGHT BUT 15 MINUTES
OF THOSE GETTING STEVE INTO
THE PLANE.
[LAUGHTER]
SO HE HAD TO CRAWL THROUGH
WHERE THE PILOT SEAT WAS OVER
THE CONTROLS AND GOT INTO THE
PASSENGER SEAT.
AND THE FRAME OF THE PLANE WAS
SO LOW HE COULD NOT GET HIS
HEAD AND I WAS REALLY
CONCERNED.
BUT HE WENT UP AND AROUND AND
EXTENDED HIS HEAD ABOVE THE
FRAME THE PILOT SAID PLEASE
DON'T YOUR HEAD TO THE
FABRIC.
[LAUGHTER]
SO WE LANDED NO PROBLEM.
THIS SHOWS THE TROUT CLOSE TO
CAMP.
I DON'T KNOW IF JEFF TOOK THIS
OR WHO BUT WE HAVE SOME
ABSOLUTELY WONDERFUL DRONE
SHOTS.
SO KNOWING THAT BOB HAD MOVED
TO FLORIDA THIS IS ANOTHER
OPPORTUNITY AND THAT 100-POUND
TARPON HE BROUGHT UP TO THE
BOAT.
AND DID I SAY STEVE IS
COMMITTED TO FISHING?
HE WOULD BE TAKING A NAP
ALTHOUGH HE SAID HE WAS JUST
RESTING HIS BACK YOU KNOW HE
HAS A FISHING ROUTE RIGHT
HERE.
[LAUGHTER]
HE WILL NOT NOT GIVE UP THE
OPPORTUNITY TO CATCH MORE
FISH.
SO STEVE HAS JOINED US AT A
FLY FISH ON —- FLY FISHING
CLUB NEAR GETTYSBURG.
IT'S A NICE FLY FISHING AREA
WE HAVE A NUMBER OF PEOPLE.
I NEED TO SHOW HIS FISH.
THERE IS A TIGER THERE IS A
RAINBOW.
CLICK ON HIS NAME.
SO THEY HAVE A NICE THING THAT
SO THEN AN OLD FISHING BUDDY
FROM THE UNIVERSITY OF
MINNESOTA WE HAD PEOPLE IN THE
AUDIENCE AND THEN TO BE UP
THERE THAT IS A NICE PLACE TO
VISIT SO JUST BECOME FRIENDS
WITH STEVE AND HE WILL FIX YOU
UP.
THE OTHER THING WE DO THAT IS
REWARDING TO PARTICIPATE WITH
PROJECT HEALING WATERS TO
BRING OUT FROM WALTER REED AND
TO SERVE AS GUIDES WHEN IN
FACT STEVE WAS THE GUY TWO
WEEKS AGO FOR ONE OF THESE
OUTINGS.
SO BRADY HAS BEEN FISHING WITH
HIM FOR A NUMBER OF YEARS AND
NOW LAST WEEK THIS IS A
PICTURE LAST WEEK LOOKING
FURTHER TO GETTING OUT TO THE
FISHING AREA IT WAS
3:00 O'CLOCK IN THE MORNING SO
LET ME TELL YOU THAT IF ANYONE
IS WORRIED ABOUT STEVE WRITING
OUT TO THE SUNSET WITH HIS
RETIREMENT I THINK HE IS
HEADED INTO THE SUNRISE.
[APPLAUSE]
>>THANK YOU VERY MUCH.
SO I WILL BE GOING ON A TRIP
TO ALASKA NEXT SUMMER SO I
WILL REPORT BACK HOW THAT
GOES.
WE WILL BREAK FOR LUNCH WE ARE
RUNNING A LITTLE BIT BEHIND
THE LET'S RECONVENE AT 1:30
WE'LL GET STARTED.
NEXT SPEAKER IS JEFF, RECRUITED
WITH STEVE BY GEORGE VANDER WOOD
IN THE 1980s, SENIOR
INVESTIGATOR, DEPUTY DIRECTOR AT
NCI CANCER RESEARCH, A LONGTIME
FISHING BUDDY AND I EXPECT WE'LL
HEAR ABOUT THAT IN THIS TALK.
>>WELL, THANK YOU.
IT'S A REAL HONOR TO BE IN
POSITION TO BRAG ABOUT STEVE,
AND STAY A LITTLE BIT ABOUT THE
RELATIONSHIP I'VE HAD WITH HIM
THAT SPANS THE TOPIC OF SCIENCE,
AN IMPORTANT FEATURE THAT
RELATES TO OUR FAMILIES, AND
FISHING.
STEVE IS A BIG BY, OKAY?
AND HIS HANDSHAKE IS FIRM AND
WELCOMING.
HE'S STRONG ENOUGH TO BREAK A
SLEDGEHAMMER WHILE HITTING THE
ROCK IN FAIR FASHION, NOT
OVERREACHING A SPIKE OR
SOMETHING.
BUT IN ADDITION, HE'S GOT A HUGE
PERSONALITY, A BIG INTELLECT,
HELPED LOTS OF PROJECTS WHEN HE
WAS INVOLVED.
STEVE MOVED IN NEXT DOOR TO MY
FAMILY, MY WIFE ANN AND MY
DAUGHTER LESLIE.
IN THE TOP TWO FLOORS OF THE
BUILDING BEHIND US IN COLD
SPRING HARBOR.
WE SHARED A WALL.
YOU COULD WALK THROUGH THIS, I
DON'T KNOW WHAT THAT WAS ABOUT.
A FEW MONTHS AFTER STEVE MOVED
IN, MY SON RUSSELL WAS BORN.
SO, HAVING THAT EXTENDED FAMILY
WAS AN ESSENTIAL COMPONENT OF
DEALING WITH THE 24/7 RESEARCH
ENVIRONMENT AT COLD SPRING
HARBOR.
IT IS A MARVELOUSLY INTENSE
PLACE TO WORK.
BUT OUR FAMILIES SORT OF
FUNCTIONED AS ONE EXTENDED
FAMILY.
WE ATE DINNER TOGETHER THREE OR
FOUR NIGHTS A WEEK FOR FOUR
YEARS, OKAY?
AND THE KIDS, MY DAUGHTER,
LESLIE, RUSSELL, MY SON, PHILIP
AND CHRISTINA, WHO IS IN THE
AUDIENCE OVER HERE, WERE RAISED
PRACTICALLY AS SIBLINGS.
AND THEY CELEBRATED BIRTHDAYS
TOGETHER AND I THINK THAT'S
PROBABLY EASTER.
SOMETIMES THEY WERE LOOKED AFTER
BY STEVE, THE TWO DAUGHTERS, AND
SOMETIMES BY ME, THE TWO
DAUGHTERS.
BUT OF COURSE -- LET'S GET THIS
RIGHT.
MOST FREQUENTLY, BY MY WIFE,
ANN, AND ALL THE KIDS, OR
VICTORIA AND ALL THE KIDS.
I DON'T THINK WE COULD HAVE
SURVIVED WITHOUT HIM.
WELL, STEVE AND I WORKED IN THE
LAB, REALLY 24/7.
AND THERE WERE LATE, LATE
NIGHTS.
WE FINALLY RECOGNIZED YOU DON'T
MAKE A DAY ANY LONGER BY CUTTING
HOURS OFF OF THE END OF THE DAY
AND SEWING THEM TO THE MORNING.
SO WE MADE A PACT, THE PACT WAS
THE FIRST PERSON DONE IN THE LAB
AFTER MIDNIGHT WOULD GO AND HAUL
THE OTHER PERSON OUT OF THE
LABORATORY.
INDEED THAT HAPPENED MANY TIMES
IN THE YEARS WE WORKED THERE
TOGETHER.
SO MY WORK WAS ON THE CONTROL OF
SACCHAROMYCES CEREVISIAE,
STEVE'S WORK ON RETROVIRAL
VECTORS, ISOLATION OF
CYTOSKELETAL GENES.
LET'S SEE.
RIGHT HERE.
THERE WASN'T MUCH TIME FOR
RECREATION.
BUT WE DID FINALLY DISCOVER THAT
WE HAD A LOVE OF FISHING.
AND I HAD TO DIG THROUGH MY
PICTURES TO FIND ONE IN WHICH I
HAD THE BIGGER FISH.
[LAUGHTER]
MOST OF WHAT WE DID WAS FROM A
CANOE WITH A LIGHT WEIGHT FLY
RODS CATCHING FISH ABOUT 6
INCHES LONG.
BUT THAT SHARED LOVE WAS
IMPORTANT COMPONENT OF EARLY
RELATIONSHIP.
NOW, AS WE MOVED TO FREDERICK IN
1984 GEORGE RETHE VIEWED FIRST
STEVE, THEN ME, ON THE BASIS OF
STEVE'S RECOMMENDATION, GEORGE
AND ME OR MY WORK AT THE TIME,
AND GEORGE CHEATED.
WE TOOK THE FAMILIES, HE BROUGHT
THEM TO HIS FARM IN VIRGINIA,
RIGHT, TOLD THE KIDS THEY DO
COULD HAVE A FARM IF STEVE AND I
ACCEPTED HIS INVITATION TO WORK
IN FREDERICK.
[LAUGHTER]
SO, THE END RESULT WAS STEVE HAS
A 63-ACRE FARM IN WEST
FREDERICK, I HAVE 28-ACRE FARM
EAST OF FREDERICK.
NOW, LET'S SEE.
THE CONVERSATIONS RESULTED IN
FORMAL COLLABORATIONS.
THIS IS ONE OF OUR PAPERS,
ANALYSIS OF THE CLEAVAGE SITE
FOR ENZYME THAT CUTS AT A SINGLE
SPOT IN THE YEAST GENOME, TO
INITIATE SWITCHING.
THAT ENZYME, SINGLE-BASED
RESOLUTION, THAT ENZYME WAS
USEFUL IN MY SUBSEQUENT WORK
BECAUSE WE USED IT TO INITIATE
COMBINATION AND WENT ON LOOK AT
FIDELITY OF RECOMBINATION AND
GENES INVOLVED, BUNCH OF OTHER
ASSAYS.
KEY WORK.
IDENTIFICATION OF A GENE THAT
WAS A CANDIDATE FOR A GENE
IDENTIFIED IN STEVE'S LAB, AS
MENTIONED EARLY, SEQUENCING
SOMETHING WAS A BIG DEAL.
THEY DID SEQUENCE IT.
WE HAD MAPPED INTO THE YEAST
GENOME, KNOCKED IT OUT,
DEMONSTRATED IT WAS AN ESSENTIAL
GENE.
AND FUN12 MORE RECENTLY
IDENTIFIED INITIATION FACTOR
ELF5B.
THE BEST COLLABORATION WAS WHEN
WE PUT THE HIV REVERSE
TRANSSCRIPTASE INTO THE YEAST
RETROTRANSPOSON TI1.
THIS IS THE ASSAYS BASED ON
YEAST TRANSPOSON, IN WHICH RT
DOMAIN WAS SUBSTITUTED, MARKED
WITH A METABOLIC SCREEN, WHEN
THE RNA WAS MADE IT WAS REVERSE
TRANSCRIBED, AFTER THE INTRON
WAS SPLICED OUT, GENERATED
FUNCTIONAL HISTORY GENE, BEING
ABLE TO MAKE HISTIDINE, EACH OF
THESE IS RESULT OF TRANSPOSITION
EVENT.
THAT TURNED OUT TO BE USEFUL IN
A LOT OF PROJECTS.
FIRST OF WHICH WAS TY ELEMENTS
ARE RESISTANT TO NON-NUCLEOTIDE
REVERSE TRANSCRIPTATION
INHIBITORS.
HIV BASED ELEMENTS WERE
SENSITIVE, IF YOU ISOLATED HIV
RT DOMAIN FROM RESISTANT
VIRUSES, THEY WERE IN TURN IN
YEAST RESISTANT TO NNRTIs,
BASIS OF SEVERAL EXPERIMENTS
FROM THE LAB.
HERE BEGINS THE INITIAL ASSAY.
HERE IS WILD TYPE THAT IN THE --
WHEN INDUCED MADE
(INDISCERNIBLE) BUT IN THE
PRESENCE RARELY MADE
TRANSPOSITION EVENT, WE WERE
ABLE TO ISOLATE HIV RT DOMAIN
FROM PATIENTS, THE VECTOR MADE A
LIBRARY, SCREENED FOR ONES
RESISTANT TO THE DRUG,
IDENTIFIED FROM THE PATIENTS
RESISTANT, SLIGHTLY RESISTANT
ALLELES.
SO RESULTS OF THE BASIS OF THE
SCREEN FOR NEW NNRTIs WE
DEMONSTRATED, COMPOUNDS AND
EXTRACTS THAT HAD -- WERE ABLE
TO INHIBIT, WE VERSE
TRANSCRIPTASE IN YEAST FOR HIV.
THE IMPACT ON MY WORK, STEVE HAD
IMPACTS THROUGHOUT THE COMMUNITY
AS YOU'VE SEEN TODAY.
ONE OF THE FEATURES THAT STEVE
PROVIDED WAS INPUT ON PEOPLE'S
SITE VISIT.
HE WROTE A BOOKLET HOW TO
SURVIVE SITE VISITS.
IT HAD SEVERAL IMPORTANT
LICENSES LIKE THE TITLE SHOULD
BE THE CONCLUSION.
YOU'RE GOING TO HAND OUT COPIES
OF THE SLIDES, AND THEN THEY ARE
GOING TO WALK AWAY WITH A PIECE
OF PAPER THAT HAS EVERY
CONCLUSION FROM EVERY SLIDE ON
IT.
A SLIDE, IT'S NECESSARY,
MINIMIZE WHITE SPACE, BLOW
THINGS UP, AND MAYBE MOST
IMPORTANTLY IF THEY DON'T
BELIEVE YOU CAN DO THE
EXPERIMENT YOU'RE SCREWED.
SO UNDERSTAND CONTROLS.
THEN DON'T SHOW ALL THE DATA
THAT YOU EVER PRODUCED.
ALSO APPRECIATED STEVE'S ROLE IN
MY EFFORTS AS DEPUTY, TO
INCREASE COMMUNICATION BETWEEN
THE LABORATORIES, SO I INITIATED
WHAT WAS CALLED A LAB CHIEF
LUNCH, ALL LAB CHIEFS COULD
COME, AN OPPORTUNITY TO EXCHANGE
BETWEEN THE LABS ONGOING
RESEARCH, WHAT WAS HOT THIS WEEK
OR LAST.
FURTHERMORE STEVE'S CONTRIBUTION
ALONG WITH ME IN THE SITE VISIT
OF ALL THE LABORATORIES UP IN
HERE, FREDERICK, AGAIN CREATED A
NEXUS POINT FOR THE DISPERSAL OF
INFORMATION THAT WOULD SAY YOU
TWO GO HAVE A BEER TOGETHER
BECAUSE YOUR EXPERIMENTS ARE
FOCUSED ON SIMILAR OUTCOME.
NOW, AGAIN, THE FIRST SEVERAL
YEARS WE WERE HERE WE DIDN'T
FISH AT ALL.
I KNOW THAT'S SURPRISING.
BUT FINALLY IT GOT TO THE POINT
STEVE AND I AGREED THAT IF WE
WERE TO EVER DO THIS, YOU KNOW,
AGAIN, WE HAD TO GET BACK INTO
IT.
SO WE DID A LITTLE BIT OF
FISHING.
I HAVE A POND ON MY PROPERTY,
NICE RIVERS AROUND HERE TO TAKE
A CANOE ON.
BUT THIS IS ONE OF THE FIRST
FISH STEVE CAUGHT ON A FLY ROD,
STRIPED BASS, I KNOW THAT
BECAUSE THAT'S MY FLY ROD FROM
MY FISHING DAYS IN OREGON FOR
STEEL HEAD TROUT.
THE CAPTAIN THOUGHT WE WERE
CRAZY BUT THE SOLUTION IS GET A
NEW CAPTAIN, RIGHT?
AND SO WE DID.
WE GOT BRADY, REFERRED TO
EARLIER, AND WE MADE OUR OWN FLY
RODS, TIED OUR FLIES.
HERE OFF NORTH CAROLINA, SOUTH
CAROLINA, CATCHING FALSE ALBA
DOOR, TUNA ON A FLY ROD, ON
FLIES YOU TIED, DOESN'T GET A
LOT BETTER THAN THAT.
STEVE WAS, AS I SAID, A BIG
HELP, LOTS OF PROJECTS.
HERE HE IS HELPING ME PUT
TOGETHER THE FOURTH OF MY
ADDITIONS TO MY HOUSE, WHICH WAS
A THREE-CAR GARAGE WITH A
TRACTOR BAY.
AND SOMEWHERE I COULD NOT FIND
THE APPROPRIATE PICTURES,
THERE'S A FOUR-HOUR VIDEO OF 24
SCIENTISTS AND 12 AMISH MEN
RAISING A BARN ON STEVE'S
PROPERTY AFTER HIS BIG BARN
BURNED DOWN.
IT WAS ONE OF THE BEST DAYS OF
MY LIFE.
SO INTERESTING AND SOMEDAY MAYBE
YOU'LL TALK STEVE INTO SHOWING
THAT YOU VIDEO.
IT WAS FANTASTIC.
OUR FAMILIES HAVE BEEN TOGETHER
SINCE 1979.
OKAY?
AND WE'VE HAD THANKSGIVING
DINNER AND CHRISTMAS DINNER
EVERY YEAR SINCE THEN, OKAY?
WITH AN EXCEPTION FOR COVID, WE
EVEN DID THAT BY SKYPE.
HERE'S ONE OF THE RECENT ONES.
YOU'LL NOTICE IN IT A SCOTTISH
TOAST, I HOPE IT WILL PLAY.
MY DAUGHTER, LESLIE.
MY GRANDDAUGHTER, KAYLY ANN.
MY SON, RUSSELL.
PHILIP.
VICKIE AND STEVE.
WE CAN'T HEAR IT.
I'LL TELL WHAT YOU THE SCOTTISH
TOAST IS.
IT'S HERE'S TO US ... MORE'S THE
PITY.
PEOPLE LIKE STEVE ARE DAMN FEW.
HIS INFLUENCE ON EVERYBODY WAS
INCREDIBLE.
OUR DEAR DEPARTED FRIEND, AMAR,
WITH MY WIFE, ANN, THE IMPACT WE
ALL HAD.
FISHING PICTURES, OF COURSE, MY
FRIEND STEVE.
GOOD LUCK.
[APPLAUSE]
TIGHT LINES.
>>THANKS VERY MUCH.
NEXT SPEAKER IS CUED UP, BOB
SILICIANO JOINS US REMOTELY FROM
JOHNS HOPKINS, PIONEER IN THE
STUDY OF ANTIRETROVIRAL THERAPY
AND HIV PERSISTENCE, PROFOUNDLY
INFLUENCED RESEARCH DONE
PARTICULARLY WORK ON HIV
PERSISTENCE.
>>THANKS.
IT'S A PLEASURE TO BE HERE FOR
THIS SYMPOSIUM HONORING STEVE.
I'D LIKE TO TALK ABOUT
PROLIFERATION OF LATENTLY
INFECTED CELLS, A BARRIER TO
CURE.
THIS IS A TOPIC THAT STEVE TOOK
UP RELATIVELY LATE IN HIS
REMARKABLE CAREER, AND MADE
PROFOUND CONTRIBUTIONS THAT I'LL
TRY TO DESCRIBE, CONTRIBUTIONS
THAT WENT TO THE HEART OF THE
PROBLEM OF TRYING TO FIND A CURE
FOR HIV INFECTION.
WHEN PATIENTS STARTED ON
COMBINATION THERAPY THERE WAS A
RAPID DECREASE IN VIREMIA BELOW
LIMIT OF DETECTION, REFLECTING
SHORT HALF-LIFE OF CELLS THAT
PRODUCE MOST OF THE PLASMA
VIRUS.
NO MATTER HOW LONG PEOPLE HAVE
BEEN ON ART, VIREMIA REBOUNDS
RAPIDLY BECAUSE OF LATENTLY
INFECTED CDPOSITIVE 4 CELLS,
ARISE WHEN ACTIVATED CELLS
BECOME INFECTED TRANSITIONING
BACK TO MEMORY STATE
NON-PERMISSIVE FOR VIRAL GENE
EXPRESSION.
AND THESE CELLS CAN PERSIST
THROUGH THE NORMAL MECHANISMS
THAT MAINTAIN MEMORY.
SUBSEQUENT ACTIVATION THROUGH
THE ANTIGEN RECEPTOR CAN LEAD TO
REVERSAL OF LATENCY, AND VIRAL
REBOUND, IF TREATMENT HAS BEEN
INTERRUPTED, BUT CAN ALSO AS
I'LL SHOW YOU LEAD TO
PROLIFERATION OF INFECTED CELLS
WITHOUT VIRUS PRODUCTION
GENERATING NEW LATENTLY INFECTED
CELLS.
THESE ARE PRESENT IN EVERYBODY
WITH HIV INFECTION BECAUSE FROM
A SIMPLE BLOOD SAMPLE YOU CAN
PURIFY RESTING CD4 CELLS, THEY
ARE NOT PRODUCING ANY VIRUS, BUT
YOU PLATE THEM IN DILUTION AND
ACTIVATE WITH MITOGEN AND CAN
GROW OUT ENOUGH VIRUS TO MEASURE
BY ELISA ASSAY, VIREMIAS COMING
FROM THE LATENTLY INFECTED CELLS
THAT'S BEEN ACTIVATED UNDERGOES
EXPONENTIAL GROWTH.
THERE ARE LIKELY OTHER
RESERVOIRS, THIS ASSAY WE THINK
IS THE MOST CHALLENGING BARRIER
TO CURE.
NOW, WE KNOW VIRUS IN ACTIVATED
CELLS AND PRODUCTIVELY INFECTED
CELLS, LIKELY PROXIMAL SOURCE OF
VIRAL REBOUND BUT MAY HAVE BEEN
GENERATED THROUGH ACTIVATION AND
TEND TO HAVE OTHER SHORT
HALF-LIFE DUE TO VIRAL
CYTOPATHIC EFFECT OR SAMPLING
NORMAL CONTRACTION PHASE OF THE
IMMUNE RESPONSE WHICH ELIMINATES
MOST ACTIVATED CELLS.
LEAVING BEHIND RESTING MEMORY
CELLS TO PROVIDE IMMUNOLOGIC
MEMORY.
IN THESE HIGHLY QUIESCENT CELLS,
VIRUS REMAINS LATENT, BECAUSE
KEY HOST TRANSCRIPTION FACTORS
INCLUDING NF-kappaB ARE
SEQUESTERED, SO THAT THE VIRUS
REMAINS LATENT AND AVOIDS
CYTOPATHIC EFFECT AND CLEARANCE.
USING THIS MEASURE OF THIS
LATENT FORM OF THE VIRUS, SHOWN
MANY YEARS AGO, THE HALF-LIFE OF
THIS RESERVOIR IS EXTREMELY
LONG, 3.7 YEARS.
THIS WORK WAS PUBLISHED IN 1999
AND 2003, AND IN 2015 DAVID
MARGOLIS REPEATED THIS STUDY ON
PATIENTS ON A INTEGRASE
INHIBITORS AND FOUND EXACTLY THE
SAME HALF-LIFE.
NOW IN 2022 WE CAN ASK WHETHER
THIS SLOW DECAY CONTINUES IN
PEOPLE WHO HAVE BEEN ON VERY
LONG-TERM ART, AND WE FIND IT
DOES NOT.
THERE'S A HIGHER THAN EXPECTED
FREQUENCY OF LATENTLY EFFECTED
CELLS ON PEOPLE ON VERY
LONG-TERM ART, NOT DUE TO ANY
KIND OF FAILURE TO SUPPRESS
REPLICATION, WE SHOWED IN A
SERIES OF PAPERS THAT AS FAR AS
I CAN TELL NEVER READ BY ANYONE
THAT THERE IS UNEXPECTED
COOPERATIVITY IN DOSE-RESPONSE
CURVES OF SOME ANTIRETROVIRAL
DRUGS AND ENERGIES BETWEEN DRUG
CLASS THAT ALLOW 5 TO 10 LOG
SUPPRESSION OF SINGLE ROUND
INFECTION BY ART, SUFFICIENT TO
PREVENT ANY REPLICATION AND
PREVENT VIRAL EVOLUTION IN
ADHERENT PATIENTS, SHOWN IN MANY
OTHER STUDIES INCLUDING BY MARY
KEARNEY.
SO, WE WANTED TO CONFIRM THIS
HIGHER THAN EXPECTED FREQUENCY
OF LATENTLY INFECTED CELLS ON
PEOPLE ON LONG-TERM ART WAS REAL
AND SO WE USED INDEPENDENT
ASSAY, THIS IS A DIGITAL DROPLET
THAT MEASURES LACKING DEFECTS
AND GOT EXACTLY THE SAME RESULT.
SO, IT'S NOT DUE TO DE NOVO
INFECTION EVENTS, DOESN'T
REPRESENT THE PROLIFERATION OF
LATENTLY INFECTED CELLS.
SO MY FIRST INDICATION THAT THIS
KIND OF SITUATION MIGHT BE
TAKING PLACE CAME FROM AN
ANALYSIS OF RESIDUAL VIREMIA.
THIS IS THE TRACE LEVEL OF FREE
VIRUS IN THE PLASMA OF PEOPLE
WHO ARE DOING WELL ON ART.
STUDIES BY JOHN COFFIN'S GROUP
SHOWED THAT IN MOST PEOPLE YOU
CAN DETECT RESIDUAL VIREMIA
OFTEN AT LEVEL AROUND 1 COPY PER
MIL, WE SPENT A LONG TIME
EXPENSING AND SHOWED IT IS
ARCHIVAL IN CARE, DRUG SENSITIVE
THAT CONTINUES TO BE RELEASED
WITHOUT ANY EVOLUTION, FOR YEARS
VIRUS COMING FROM LATENT
AFFECTED CELLS ACTIVATED, CAN'T
REPLICATE BUT CAN BE DETECTED
WITH SENSITIVE ASSAY.
THE MOST INTERESTING THING ABOUT
THIS VIRUS, I SHOULD POINT OUT
IN STUDIES WE SHOWED THAT
ADDITION OF ANOTHER DRUG TO
THREE DRUG REGIMEN COULD NOT
SUPPRESS RESIDUAL VIREMIA, IT'S
COMING FROM CELLS.
DESPITE EXTENSIVE AMOUNT OF
DIVERSIFICATION IN INDIVIDUAL
PATIENTS DURING UNTREATED
INFECTION, THE RESIDUAL VIREMIA
YOU SEE IN THE SETTING OF ART
WAS OFTEN DOMINATED BY A SINGLE
SEQUENCE, THAT IS A A SINGLE
SEQUENCE, SINGLE NUCLEOTIDE
DIFFERENCE OVER 2 TO 3-YEAR
PERIOD, WE THOUGHT THIS MUST
REPRESENT EXTENSIVE
PROLIFERATION COPYING WITHOUT
AREA INTO PROJECTY ELSE IN.
BUT PROOF OF INFECTED CELL
PROLIFERATION REQUIRES
INTEGRATION SITE ANALYSIS.
MY WIFE JANET IN 2004 PROVIDEDED
FIRST MAPPING USING INVERSE PCR
TECHNIQUE AND MOST OF
INTEGRATION SITES THAT SHE FOUND
WERE IN TRANSCRIPTIONAL UNITS
CONSISTENT WITH EARLIER STUDY
USING IN VITRO INFECTION.
ALL THE SITES WE FOUND WERE
UNIQUE.
AND THEN STEVE DEVELOPED A
HIGH-THROUGHPUT METHOD FOR HIV
INTEGRATION SITE ANALYSIS.
AS HAS BEEN DESCRIBED.
AND FOUND THAT THE SITES WERE IN
GENES, SOMETIMES GENES
ASSOCIATED WITH CELL
PROLIFERATION BUT MOST
IMPORTANTLY OVER A THIRD OF THE
SITES WERE IN GENES, SORRY, OVER
A THIRD OF THE INTEGRATION SITES
WERE IN EXPANDED CLONES.
IN OTHER WORDS, THE SAME SITE
WAS FOUND IN MULTIPLE CELLS,
PRESENT IN THE SAME SAMPLE,
SOMETHING THAT COULD ONLY BE
EXPLAINED BY PROLIFERATION OF
INFECTED CELLS.
I THINK IT'S REALLY HARD TO
OVERSTATE THE IMPORTANCE OF THIS
FINDING.
AFTER ALL, THE STIMULI THAT
DRIVE T CELL PROLIFERATION ALSO
INDUCE VIRAL GENE EXPRESSION AND
MOST PRODUCTIVELY INFECTED CELLS
HAVE A SHORT HALF-LIFE.
SO, IT WASN'T AT ALL CLEAR THAT
INFECTED CELLS COULD
PROLIFERATE.
THIS STUDY BY STEVE AND
COLLEAGUES REALLY DEMONSTRATED
THAT THIS COULD HAPPEN IN VIVO,
AND NOT ONLY DOES IT HAPPEN IN
VIVO BUT TO A REMARKABLE EXTENT
THAT YOU COULD CAPTURE PROGENY
CELLS DERIVED FROM THE SAME
INITIAL INFECTED CELL, IN CELL,
IN A SINGLE BLOOD SAMPLE.
AROUND THIS TIME USED WHOLE
GENOME SEQUENCING TO SHOW HIGHLY
DEFECTIVE DUE TO LARGE DELETION,
ANALYSIS CAPTURES ONLY THE END
OF THE VIRAL GENOME SO IT'S
POSSIBLE MANY SITES IDENTIFIED
WITH INTEGRATION SITE ANALYSIS
ARE PROVIRUSES THAT ARE
DEFECTIVE.
BUT THIS CONCEPT OF INFECTED
CELL PROLIFERATION WAS EXTENDED
TO CELLS CARRYING REPLICATION
COMPETENT VIRUS IN STUDIES, AND
IN FACT USING VIRAL ASSAY YOU
CAN SHOW MANY OF THE INDEPENDENT
ISOLATES FROM A SINGLE BLOOD
SAMPLE ACTUALLY CARRY THE SAME
PROVIRAL SEQUENCE, 57% WITH
EXTRAORDINARY DEGREE OF IN VIVO
PROLIFERATION OF INFECTED CELLS.
AND IF YOU FOLLOW CLONES OF
INFECTED CELLS OVER A PERIOD OF
ONE TO FOUR YEARS, YOU SEE SOME
PERSIST, DETECTED AS FRACTION OF
ISOLATES BUT OTHERS APPEAR AND
DISAPPEAR DURING THIS TIME
FRAME.
SO THAT THESE CLONES ARE
ESSENTIALLY WAXING AND WANING, A
FINDING THAT'S INCONSISTENT WITH
A CONTINUOUS CELL AUTONOMOUS
STIMULUS AS THE SOLE DRIVER OF
PROLIFERATION, AND RATHER IT
APPEARS THAT ANOTHER MAJOR
FACTOR IS ACTUALLY ANTIGEN
DRIVEN PROLIFERATION SHOWN BY
SIMONETTI IN THE AUDIENCE,
ISOLATED CELLS RESPONDING, AND
SHOWED AMONG CMV RESPONDING
CELLS, VERY LARGE CLONES OF
INFECTED CELLS HAD SAME
INTEGRATION SITE, SAME PROVIRAL
SEQUENCE, SOMETIMES COMPRISING
OVER 80% OF ALL OF THE
CMV-RESPONSIVE HIV INFECTED
CELLS, SOME CLONES ARE LARGE
ENOUGH TO BE SEEN AMONG CELLS
RESPONDING NON-SPECIFICALLY TO
CD3, CD28.
SO, EVEN THOUGH IMMUNE SYSTEM
CAREFULLY CONTROLS TC
PROLIFERATION, REPEATED EXPOSURE
TO THE SAME ANTIGEN CAN GENERATE
LARGE CLONES TRANSFORMING THIS
RESERVOIR FROM COLLECTION OF
QUIESCENT CELLS TO ONE THAT'S
DOMINATED BY LARGE POPULATIONS
OF INFECTED CELLS ALL DERIVED
FROM A SINGLE CELL, LARGE
CLONES.
AND THE THING WE HAVEN'T YET
EXPLAINED IS HOW DOES THIS
HAPPEN, GIVEN THAT THE STIMULI
THAT DRIVE T CELL PROLIFERATION
ALSO INDUCE VIRAL GENE
EXPRESSION, AND MOST
PRODUCTIVELY INFECTED CELLS DIE
VERY QUICKLY SO HOW DOES THIS
HAPPEN?
TURNS OUT THERE'S UNCOUPLING
PREEN T CELL PROLIFERATION AND
PROVIRAL INDUCTION, WE CAN SHOW
THIS IN ASSAYS, SPLIT THE WELLS
AND TAKE HALF OF EACH WELL,
SUBJECT CELLS TO ADDITIONAL
ROUND OF STIMULATION, AND YOU
CAN SHOW THAT EVEN THOUGH ALL OF
THE CELLS HAVE PROLIFERATED,
SOME OF THEM DON'T PRODUCE VIRUS
UNTIL AFTER A SECOND OR THIRD OR
EVEN A FOURTH ROUND OF T CELL
ACTIVATION.
SO THERE'S SOME STOCHASTIC
FEATURE THAT DETERMINES WHETHER
A PROVIRUS IS INDUCED FOLLOWING
AN INITIAL STIMULUS OR
SUBSEQUENT STIMULUS SO THAT IT'S
A T CELL ACTIVATION DOES NOT
ALWAYS CAUSE PROVIRAL INDUCTION
SO WE THINK THIS IS HOW VIRUS
CLONES CAN BE GENERATED.
BUT SO WHAT ARE THE CLINICAL
CONSEQUENCES OF THIS
PROLIFERATION OF INFECTED CELLS?
WELL, IT REALLY ALL COMES DOWN
TO REBOUND.
THIS IS THE WHOLE ROLE OF HIV
CURE RESEARCH IS TO PREVENT
VIRAL REBOUND.
AND SO ONE QUESTION WE CAN ASK
IS DO THESE EXPANDED CLONES OF
LATENTLY EXPECTED CELLS
CONTRIBUTE TO VIRAL REBOUND, WE
CAN DETECT CLONES WITH OUR IN
VITRO SAMPLING, IN VIVO SAMPLING
THE RESERVOIR CURVES
CONTINUOUSLY AND ANTIGEN WILL
INDUCE T-CELL ACTIVATION,
PRODUCTION OF VIRUS, THIS MAY
EXPLAIN THE CLONAL NATURE OF
RESIDUAL VIREMIA BUT WHEN
REMOVED DO THEY GIVE RISE TO
REBOUND?
REBOUND MAY NOT ALWAYS COME FROM
THESE LARGE CLONES.
THIS WAS FIRST SHOWN BY KATIE
BARR AND MICHELE, IN STUDIES
USING ASSAY TO SHOW EVEN IN
PATIENTS WHO HAD A RESERVOIR
THAT WAS DOMINATED BY A SINGLE
LARGE CLONE VIRAL REBOUND
FOLLOWING TREATMENT AND ERUPTION
OFTEN CAME FROM MINOR VARIANTS,
PHYLOGENETICALLY NOT PREVIOUSLY
DETECTED, A NUMBER OF
EXPLANATIONS OFFERED FOR THIS
SURPRISING FINDING, THIS IS WHY
IT WAS SUGGESTED IT MIGHT BE DUE
TO RECOMBINATION DURING REBOUND.
BUT WE HYPOTHESIZE A SIMPLER
EXPLANATION, AUTOLOGOUS COULD
BLOCK RESIDUE, RELATIVELY LITTLE
ATTENTION PAID TO AUTOLOGOUS
NEUTRALIZING ANTIBODIES, BUT IT
IS THE EFFICACY OF THE
AUTOLOGOUS NEUTRALIZING ANTIBODY
RESPONSE THAT DRIVES VIRAL
EVOLUTION AND THAT FUNDAMENTALLY
IS RESPONSIBLE FOR THE FACT WE
DON'T HAVE AN HIV VACCINE.
THESE ANTIBODIES ARE EFFECTING
BUT ESCAPE OCCURS QUICKLY.
THE ABSENCE OF ANTIBODIES AND
PRESENCE OF CONTROL ITT FROM HIV
NEGATIVE OR AUTOLOGOUS, LOOK FOR
REDUCTION IN VIRAL OUTGROWTH AND
COMPARED TO CULTURES WITHOUT
ANTIBODY OR WITH CONTROL IgG
WE FOUND THAT IN MANY CASES
OUTGROWTH IN THE PRESENCE OF
AUTOLOGOUS IgG WAS GREATLY
REDUCED, AND THIS IS LIKELY A
DRAMATIC UNDERESTIMATE OF IN
VIVO EFFECT BECAUSE 50 MICROGRAM
PER mL READINGS IN THESE
EXPERIMENTS IS OVER TWO LOGS
LESS THAN IN VIVO CONCENTRATION
OF IgG.
SO, ON AVERAGE WE GOT A 4.6-FOLD
REDUCTION IN FREQUENCY OF
OUTGROWTH, IN 6 OF 15 DONORS
OUTGROWTH OF OVER 80% OF THE
RESERVOIR VIRUS WAS BLOCKED BY
LOW CONCENTRATION OF AUTOLOGOUS
IgG.
WE SEQUENCED OUTGROWTH VIRUSES,
TO UNDERSTAND THIS PHENOMENA,
AND STRUCTURE OF THE RESERVOIR
IS APPRECIATED BY CONSIDERING
FIRST THE VIRUSES THAT GREW
WITHOUT ANTIBODIES, TWO BRANCHES
OF THE PHYLOGENETIC TREE
INCLUDING ONE LARGE EXPANDED
GROWN.
OUTSIDE THE PRESENCE THEY ARE
CONFINED TO UPPER BRANCH OF THE
TREE, WHERE THERE ARE MUTATIONS
IN THE B 3 AND B 4 REGIONS OF
ENVELOPE GENE CONSISTENT WITH
ESCAPE FROM NEUTRALIZING
ANTIBODY RESPONSE, WE CAN PROVE
THIS IN NEUTRALIZATION ASSAYS,
SHOWING VIRUS IN UPPER PART OF
THE TREE ARE HIGHLY RESISTANT
SHOWN BY RED COLORS, OTHER
ISOLATES INCLUDING THIS LARGE
CLONE ARE HIGHLY SENSITIVE,
READILY NEUTRALIZED.
PRESENCE OF LARGE CLONES
PROVIDES A SIMPLE EXPLANATION
FOR DISCREPANCY BETWEEN VIRAL
OUTGROWTH ASSAY AND VIRUSES THAT
APPEAR UPON REBOUND.
WE'VE GONE ON TO SHOW THERE'S A
PHYLOGENETIC RELATIONSHIP
BETWEEN IN VITRO AND VIRUSES
THAT CAUSE REBOUND IN VIVO.
IN OTHER WORDS, REBOUND IS A
SELECTION PROCESS.
AND THE RESERVOIR IS ACTUALLY
CONTINUOUSLY SORT OF SAMPLED IN
VIVO THROUGH EXPOSURE TO ANTIGEN
AND THEN WE CAN SEE WHETHER THE
VIRUSES INDUCED CAN NOW
REPLICATE ONCE ART IS REMOVED
AND IT TURNS OUT SOME OF THEM
CANNOT BECAUSE THEY ARE
NEUTRALIZED BY EXISTING ANTIBODY
RESPONSE.
HOWEVER, BECAUSE THIS SAMPLING
PROCESS OF THE RESERVOIR THAT
OCCURS IN VIVO IS CONTINUOUS, IT
CAN FIND MINOR VARIANTS THAT ARE
NOT NEUTRALIZED, THIS IS WHAT
LEADS TO VIRAL REBOUND.
I THINK IT'S ENCOURAGING THAT
THE SIZE OF WHAT JEFF LIFSON
CALLS REBOUND COMPETENT
RESERVOIR IS SMALLER THAN WE
THOUGHT.
BUT I SHOULD POINT OUT THAT IN
SOME INDIVIDUALS, THESE LARGE
EXPANDED CLONES CARRY
NEUTRALIZATION RESISTANT VIRUS.
THE PROCESS BY WHICH THE VIRUS
ESCAPES FROM AUTOLOGOUS
NEUTRALIZING ANTIBODIES SORT OF
PREDATES AND IS ORTHOGONAL TO
THE PROCESS OF ANTIGEN-DRIVEN
CLONAL EXPANSION.
SO THINKING ABOUT REBOUND, IT'S
USEFUL TO CONSIDER THE RESERVOIR
SORT OF ON THE LOG SCALE.
AND IT'S VERY CLEAR THAT THE
CLONAL EXPANSION PROCESS THAT
STEVE PROVIDED EVIDENCE FOR IS
REALLY VERY IMPORTANT IN SORT OF
THE MAINTENANCE OF THE LATENT
RESERVOIR.
WE ALSO KNOW RAPID REBOUND CAN
COME FROM MINOR VARIANTS PRESENT
AT FREQUENCY LET'S SAY LESS THAN
1%.
AND THE CONTINUOUS SORT OF
SAMPLING OF THE RESERVOIR THAT
OCCURS IN VIVO CAN FIND THESE
MINOR VARIANTS.
THIS GIVES RISE TO SOBERING VIEW
OF THE PROSPECT OF ACHIEVING
ERADICATION.
SOME YEARS AGO ALLISON HEAL
DEVELOPED A MODEL FOR THE DEGREE
OF RESERVOIR REDUCTION REQUIRED
TO ACHIEVE A GIVEN DEGREE OF
DELAY IN VIRAL REBOUND, AND SHE
BASED THIS MODEL ON LEVEL OF
RESIDUAL VIREMIA, THIS AGREES
VERY WELL WITH KNOWN CASES OF
VERY LONG TIME TO REBOUND, WHAT
THE MODEL SHOWS IS FOR A SORT OF
TWO LOG REDUCTION IN SIZE OF THE
RESERVOIR, YOU ONLY GET A FEW
WEEKS DELAY IN REBOUND.
I THINK THIS IS A VERY SOBERING
VIEW OF POSSIBILITY OF ACHIEVING
CURE THROUGH RESERVOIR
REDUCTION.
99% REDUCTION IN RESERVOIR.
ONLY REALLY CAUSING MINOR DELAY,
NOWHERE NEAR ACHIEVING THAT
DEGREE OF REDUCTION.
IN FACT I WOULD SAY IN CLINICAL
STUDIES THERE REALLY ISN'T ANY
CONVINCING EVIDENCE IN STUDIES
THAT DON'T INVOLVE BONE MARROW
TRANSPLANTATION.
I DON'T WANT TO END ON THIS
DEPRESSING NOTE SO I'LL SHOW
THIS PHYLOGENETIC TREE FROM AN
SIV-INFECTED MACAQUE, AND THIS
SHOWS ALL THE EVOLUTION THAT
OCCURS FROM INITIAL STOCK
SEQUENCE OVER THE COURSE OF ONE
YEAR OF UNTREATED INFECTION,
ANIMAL IS PUT ON ART AND
FOLLOWED FOR FOUR YEARS,
REPRESENTATIVE OF TEN ANIMALS IN
COMBINATION WITH DAN'S GROUP.
VIRUSES REPLICATING AT THE TIME
OF ART INITIATION FOUND IN
PLASMA SHOWN IN BLACK, AND THESE
VIRUSES HAVE ACCUMULATED
MUTATIONS IN VARIABLE REGIONS OF
THE ENVELOPE GENE CONSISTENT
WITH ESCAPE FROM AUTOLOGOUS
NEUTRALIZING ANTIBODIES.
OTHER VARIANTS THAT ARE
ANCESTRAL AND THAT HAVE FEWER
MUTATIONS ARE ALSO PRESENT AND
ARE PRESERVED IN THE RESERVOIR,
AND IN FACT ACTUALLY BECOME MORE
PROMINENT WITH TIME, DECREASING
TIME, FOR REASONS I DON'T HAVE
TIME TO GO INTO.
BUT WHAT'S REALLY INTERESTING IS
WHY AREN'T ALL OF THESE VARIANTS
REPLICATING AT THE TIME OF ART
INITIATION?
OF COURSE ONE HYPOTHESIS IS THAT
THEY CAN'T REPLICATE.
THEY ARE ALREADY NEUTRALIZED BY
YOU A TOLL -- AUTOLOGOUS
NEUTRALIZING ANTIBODIES, A
DIFFERENT WAY TO LOOK AT THE
PROBLEM INSTEAD OF THINKING
ABOUT HOW MANY LOGS OF REDUCTION
IN THE RES WE -- RESERVOIR WE
HAVE TO ACHIEVE WE CAN THINK
ABOUT SIMPLY FINDING A WAY TO
PREVENT OUTGROWTH OF THE SUBSET
OF VIRUSES THAT ARE NOT ALREADY
CONTROLLED BY THE IMMUNE
RESPONSES.
I'M GOING TO STOP THERE.
I WANT TO, AGAIN, CONGRATULATE
STEVE ON HIS REMARKABLE
CONTRIBUTIONS NOT ONLY TO THE
CURE FIELD BUT TO RETROVIROLOGY
IN GENERAL, AND I WANT TO THANK
THE PEOPLE FROM OUR GROUP
INVOLVED IN THIS WORK, WORKING
WITH MY WIFE JANET FOR MANY
YEARS, HO DID THE ORIGINAL WORK,
AND SHE NOW HAS A LAB AT YALE,
FRANCESCO HAS A LAB AT JOHNS
HOPKINS, ALLISON HEEL IS NOW
ASSISTANT PROFESSOR AT JOHNS
HOPKINS, AND DANIEL ROSENBLUM,
WE HAVE A TERRIFIC TEAM,
LONG-TERM ART PROJECT, EMILY AND
KUMAR.
I WANT TO THANK DAN, STEVE,
LUIS, PEOPLE WHO INSPIRED OUR
WORK AND FUNDING AGENCIES.
I'LL STOP THERE TO ANSWER ANY
QUESTIONS.
-789D.
[APPLAUSE]
>>THANKS, BOB.
INSPIRING AS ALWAYS.
>>WE HAVE TIME FOR ONE OR TWO
QUICK QUESTIONS, IF THERE ARE
ANY.
>>VERY NICE TALK.
IT FOLLOWS ALONG WITH SOMETHING
I DON'T LIKE TO SAY BUT IT'S
(INDISCERNIBLE).
BUT YOU TALK ABOUT PERSISTENT
VIREMIA, HAVE YOU LOOKED TO SEE
PERSISTENT VIRUS STILL BEING
SHAPED BY THE SAME ANTIBODY
COURSES?
>>SO, WE HAVEN'T -- GOOD
QUESTION.
WE HAVEN'T HAD A CHANCE TO DO
THAT YET.
WE'RE PARTICULARLY INTERESTED IN
LOOKING AT THIS, PARTICULARLY IN
THESE PATIENTS, BUT LONG-TERM
ART TO SEE WHAT THE NATURE OF
THE VIREMIA IS, NATURE OF LARGE
CLONES, TURNS OUT IN THE
PATIENT -- IN LONG-TERM ART
STUDY I SHOWED YOU, THE
RESERVOIR WAS ACTUALLY DOMINATED
BY -- AT LEAST IN PEOPLE WHO
STUDIED SO FAR BY SINGLE CLONES
AND VARIATIONS TO EXTENT WHICH
THOSE ARE AFFECTED BY AUTOLOGOUS
NEUTRALIZING ANTIBODIES.
>>ANYBODY ELSE?
ALL RIGHT.
THANK YOU VERY MUCH.
[APPLAUSE]
NEXT IS RON SWANSTROM,
UNC-CHAPEL HILL, LEADER IN HIV
RESEARCH, UNDERSTANDING OF
DIVERSITY, RESISTANCE, TROPISM,
I'M HAPPY RON COULD BE HERE, HE
ALMOST WASN'T FOR A REASON MAYBE
HE WILL TELL US.
>>A THIRD OF MY SLIDES HAVE
BEEN USED.
[LAUGHTER]
[ INDISCERNIBLE CONVERSATION ]
>>MY TALK IS DIFFERENT.
A LITTLE MORE IRREVERENT.
I'M REMINDED OF THE TIME WHEN WE
WENT TO COLD SPRING HARBOR AS A
YOUNG POSTDOC, STUDYING
(INDISCERNIBLE) AND THE
MICROPHONE WAS OUT.
I WAS A SECOND-YEAR POSTDOC,
TERRIFIED WITH NO MICROPHONE TO
ENTERTAIN THE CROWD FOR WHAT
SEEMED LIKE AN ETERNITY.
I WANT TO SAY THIS TALK WOULD BE
DIFFERENT, I GUESS THAT'S
BECAUSE MY RELATIONSHIP WITH
STEVE HAS BEEN DIFFERENT THAN
MANY OF YOURS.
STILL GOOD.
[LAUGHTER]
ANYONE WANT TO LOOK AT MY MUSIC
LIST?
[LAUGHTER]
IT IS KIND OF WEIRD, YOU GET TO
SEE OTHER PEOPLE'S COMPUTERS.
[LAUGHTER]
NOT NECESSARILY THE PRETTIEST
THING.
THIS IS WHERE WE WERE FIVE
MINUTES AGO.
>>WELL DONE.
>>OKAY.
THANK YOU.
SO, YEAH, JOHN STOLE MY TITLE.
I TRIED TO BE MORE THEATRICAL.
I WANT EVERYONE TO NOTE THE
SYDNEY POWELL DISCLAIMER DOWN
THERE THAT I WILL USE AS PART OF
MY TALK.
IF ANYBODY I SAY IS NOT TRUE,
LET'S BLAME SYDNEY POWELL.
45 YEARS IS A REAL NUMBER.
I SUSPECT HAROLD MAY HAVE MET
STEVE BEFORE I DID.
OTHER THAN THAT, IT'S US, BUDDY.
SO WE'RE THE ONES THAT MADE IT
THROUGH UCSF, AND WE'RE NOT
GOING TO MAKE IT TO THE NEXT
SLIDE.
[LAUGHTER]
SO, YEAH, THIS IS WHERE WE
WORKED AT UCSF, I WAS THERE
BEFORE STEVE, STILL THERE A
LITTLE AFTER STEVE TOO.
WE BOTH HAD THIS COMMON
INTEREST.
RETROVIRAL DNA.
I CAN'T FULLY DISENTANGLE MY OWN
CAREER FROM HAVING STEVE AROUND.
SO THIS TALK IS REALLY
INTERSECTIONS WHICH WERE IN MANY
WAYS LESS OBVIOUS, LESS DIRECT,
BUT STILL THERE.
SO STEVE OF COURSE CAME FROM
HARVARD, WHILE I CAME FROM
UC-IRVINE WHICH WAS KNOWN FOR
(INDISCERNIBLE).
WE HAD REALLY DIFFERENT
BACKGROUNDS WHEN WE ARRIVED AT
UCSF.
THIS IS A FAMOUS PICTURE
EVERYONE IMPORTANT AT UCSF --
YOU'LL SEE STEVE IN THE MIDDLE.
I'M ON THE SIDE BY THE OTHER
POSTDOCS IN THE LAB OFF TO THE
EDGE OVER THERE.
THAT WAS MY INTRODUCTION.
AND PETE SAME UP HERE, I THINK
ON AN EXTENDED OR RETIREMENT
VACATION IN FLORENCE WHICH IS
THE ONLY THING THAT KEPT HIM
FROM BEING HERE AND I PROMISED
WOULD BRING HIM UP.
HE CAME TO UCSF FROM UNC, AND
WHICH IS WHERE I WOUND UP.
BUT WE REMAIN FANS.
YOU'VE SEEN THIS PAPER BEFORE.
I WANT TO POINT OUT PETER AND
STEVE WORKED TOGETHER TO PUBLISH
THESE PAPERS BACK TO BACK IN
"SCIENCE," AND THE FIRST ONE,
THIS IMPORTANT CONCEPTUAL
ORGANIZATION, VIRAL GENOME, AND
STEVE AND THEN PETER'S PAPER
LOOKING AT THE INTEGRATION, AS
YOU HEARD.
I WAS IMPRESSED LOOKING BACK TO
GO SO FAR AS TO ORGANIZE GENETIC
MAP OF THE VIRUS WITH BIOLOGICAL
ABILITY TO RESCUE THESE VIRUSES,
WHETHER OR NOT THEY WERE
DEFECTIVE OR NOT, INTELLECTUAL
LEAPS AT THE TIME.
ONE OF THE THINGS I NOTICED IN
THE SAME ISSUE OF "CELL,"
MICHELE AND JEFF PUBLISHED THAT
TM9 TRANSPOSITION RESULTED IN
REPEAT DURING INSERTION IN
GENOME, WE CAN'T BE SURPRISED
STEVE PICKED UP ON THAT AND
MONTHS LATER WAS ABLE TO SHOW
RETROVIRUSES ALSO GENERATED
DIRECT REPEAT IN CELLULAR DNA
WHEN THEY INTEGRATE.
SO MY, AS I SAID, MY OWN LIFE IN
ALL OF THIS AT THE SAME TIME, IN
THAT MILIEU WHEN CLAUDIA BECAME
AVAILABLE, HOFFMAN, FAR RIGHT,
KIND ENOUGH TO GIVE ME THE
CHANCE TO SEQUENCE DNA HE
CLONED, I WAS ABLE TO SEQUENCE
THE CIRCLE JUNCTION AND GET SOME
NUCLEOTIDE SEQUENCES TO LIKE
PPT, THINGS LIKE THAT.
IT WAS AS HAROLD MENTIONED AN
ENVIRONMENT THAT WAS FUN TO BE
PART OF.
SOMETIMES, MOST OF THE TIME, I'M
ENVIOUS OF STEVE FOR THE WORK HE
DOES.
SOMETIMES I'M DISMAYED.
SO, CORRECT ME IF I'M WRONG.
A POSTDOC IS GOING TO TELL
EVERYBODY WHAT ENDOGENOUS
VIRUSES LOOK AT IN CHICKENS, AND
SHOWS ONE SLIDE.
NO TALK.
AND THIS IS ACTUALLY -- THIS IS
FROM THE PAPER, I DON'T KNOW IF
IT'S EXACTLY THE SAME AS THE
SLIDE, BUT CASUAL GLANCE THAT'S
WHAT IT LOOKED LIKE.
SO, I'M FORTUNATE ENOUGH TO
PARTICIPATE IN THIS BOOK.
I'M GRATEFUL TO HAROLD FOR
GIVING ME THIS OPPORTUNITY TO BE
CO-AUTHOR, YOUNG POSTDOCS,
REALLY GRATEFUL HE PICKED ME.
IT'S A WAY A YOUNG PERSON GETS
TO BE PART OF A COMMUNITY.
HAROLD WAS VERY COMPLIMENTARY OF
STEVE FOR DOING APPENDICES.
I HAD A DIFFERENT VIEW.
I WAS HAPPY TO HAVE MY FACE AND
MY NAME ON THE TABLE OF
CONTENTS.
STEVE MANAGED TO ENGINEER HIS
NAME SIX TIMES.
[LAUGHTER]
SO THREE LESSONS I LEARNED EARLY
ON.
I -- ONE-UPMANSHIP CAN BE AN
ART FORM.
I CAN HANG OUT WITH BIG PEOPLE.
THEN I LEARNED THAT IF YOU
DIDN'T GET ASKED A QUESTION
AFTER YOUR TALK, YOU HAVE TO
ANSWER ONE OF THREE WAYS.
PROBABLY PARAPHRASE, IF YOU
LISTEN TO STEVE ANSWER
QUESTIONS, IT'S ONE ONE OF
THESE THREE ANSWERS.
YOU TOLD YOU THAT BUT YOU
WEREN'T LISTENING.
I TOLD YOU THAT BUT I GUESS YOU
AREN'T SMART ENOUGH TO
UNDERSTAND.
YOU'RE TRYING TO REINTERPRET MY
DATA, YOU'RE WRONG.
STEVE AND I LEFT UCSF, WENT ON
OUR OWN.
I CAN THINK OF MYSELF AS A
SMALLER PART, TRAVELING TOO
FAST, NOT STAYING AT ONE PLACE
LONG ENOUGH.
STEVE LARGER PLANET ON THE
OUTSIDE, MOVING SLOWLY, THINKING
MORE DEEPLY ABOUT PROBLEMS,
MAKING NICE CONTRIBUTIONS.
AND WE HAVE THIS KIND OF COMMON
SET OF INTERESTS, WHICH ARE
IMPORTANT IN THE FIELD OF LAW,
BUT WE SEEM TO COVER A LOT OF
THE SAME TERRITORY OVER THE
YEARS.
I'LL NOT GO INTO STEVE'S
GENETICALLY MODIFIED ORGANISMS.
[LAUGHTER]
ONE OF THE THINGS THAT HAPPENED
IN THE LAB WHEN WE WERE
POSTDOCS, THE TECHNIQUE OF
CLONING VIRAL DNA BY CLONING
CIRCLES AFTER THIS TECHNIQUE OF
PHENOL EXTRACTION.
WE WERE ABLE TO CLONE CIRCLES,
STUDY CIRCLE JUNCTIONS, WHICH IN
A MATERIAL WAY LOOKED AT, THAT'S
NICE, THEN LOOKED ON, BUT THE
NEXT VERSION OF THIS REALLY
BENEFITED FROM STEVE BECAUSE AS
YOU KNOW STEVE WAS MAKING
COMPETENT VECTORS.
WE MADE THIS REPLICATION
COMPETENT, AFFECTING CELLS WITH
T ANTIGEN.
YOU COULD SEE ACCUMULATION OF
CIRCULAR DNA, CLONE THINGS OUT,
ACTUALLY FIGURE OUT IF YOU --
ABOUT HALF THE DNA IS DEFECTIVE.
THAT WAS SOMETHING I HAD TO
ADMIT STRUCTURES LOOK MORE
INTERESTED TODAY, MORE
UNDERSTANDABLE THAN WE WERE ABLE
TO FIGURE OUT AT THE TIME.
FUN TO LOOK BACK AT THEM.
SO STEVE REALLY TOOK TO ASKING
QUESTIONS ABOUT THE REPLICATION
FUNCTIONS AROUND THE LTR, IT
STARTED SLOW.
YOU CAN SIGH ONE PAPER, IN TEN
YEARS TO START WITH.
THEN NOTICE A PERIOD OF THREE
YEARS WHERE HE PUBLISHED TEN
PAPERS, ALL ABOUT U 5, U3,
POLYPURINE TRACK, CIRCLE
JUNCTIONS, ALL OVER.
BUT HUGE AMOUNT OF ENERGY.
AND I HAVE TO ADMIT, SIMILAR,
NICE PAPERS, BUT TEN IN THREE
YEARS, SOMETIMES IT WAS TEDIOUS,
I MEAN READING THOSE.
[LAUGHTER]
SO THIS IS JUST A RECENT PAPER
WHERE STORIES ABOUT MUTATIONS
AND POLYPURINE TRACT,
RESISTANCE, NOT SENT TO THE DOG
HOUSE.
NO CHANGES IN SENSITIVITY TO A
VARIETY OF INTEGRASE INHIBITORS.
ONE OF THE MANY THINGS HE WILL
BE REMEMBERED FOR.
STEVE AND I HAD THIS BIFURCATION
IN OUR CAREERS.
LOOKING BACK I CAN SEE IT REALLY
CLEARLY.
HIV SHOWS UP, STEVE AND I ON OUR
OWN, REVERSE TRANSCRIPTION.
INDEPENDENTLY DECIDE IT WOULD BE
IMPORTANT TO MAKE AMOUNTS OF HIV
RT AND E. COLI.
SPOILER, YOU CAN SEE, IT'S NOT
FROM MY PAPER.
OUR EFFORTS, WE WERE PUTTING IN
FRAGMENTS OF HIV FRAGMENTS.
AND GETTING SOME LITTLE
ACTIVITY, ONE DAY, NOT THE NEXT
DAY.
A COLLEAGUE SAID WHY DON'T YOU
GET ANTIBODIES AND DO A WESTERN
BLOT.
WE DID THAT.
WE SAW IN THE SECOND LANE OVER,
PROCESSED RT.
SO IT WAS THERE, WE DIDN'T MAKE
VERY MUCH OF IT.
WE DECIDED GOSH, IF WE CAN'T
MAKE VERY MUCH BUT HAVE NICE
ASSAY FOR PROTEASE, LET'S STUDY
THE PROTEASE.
A BIG BIFURCATION.
I USED A CRAPPY EXPRESSION
SYSTEMS.
COULDN'T MAKE BIOCHEMICAL
ANALYSIS BUT ENOUGH TO DO A
WESTERN, AND THAT WAS MY START
IN PROTEASE.
WELL, STEVE FIGURED OUT HOW TO
MAKE CHEMICAL AMOUNTS, IT HAS
SET THE STAGE FOR A NUMBER OF
IMPORTANT STUDIES.
YOU HEARD FROM EDDY ARNOLD
TODAY.
I SHOW THIS SLIDE BECAUSE I SHOW
IT TWICE A YEAR IN CLASSES THAT
I TEACH, THAT YOU CAN SEE,
BARELY SEE THE N 3, AZT BLOCKING
THE NUCLEOTIDE POSITION, ATP
GETTING ORGANIZED TO GO IN THERE
AND BE PRIMORDIAL EXONUCLEUS
ACTIVITY, ELEGANT STUDY, I THINK
IF I HAD JUST PICKED A DIFFERENT
EXPRESSION VECTOR, COULD HAVE
BEEN A DIFFERENT RESPONSE ALL
THESE YEARS.
[LAUGHTER]
WE DID AN EXPERIMENT IN
HUTCHINSON'S LAB, EXPRESSING
PROTEINS IN E. COLI, IF THEY
FOLD NICE THEY LOOK NICE IN
WESTERN, IF YOU DON'T FOLD RIGHT
YOU GET THIS DEGRADED.
YOU CAN USE THIS ASSAY, ENZYME
ACTIVITY, INTERESTING THINGS
ABOUT POSITIONS.
AND STEVE ALSO USED THIS ASSAY.
HE ACTUALLY FIGURED OUT THAT YOU
COULD DO IT IN A VARIANT, SAME
THING HAPPENED IN THE VARIANT,
TEMPERATURE SENSITIVITY AT 37,
BUT NOT AT 42.
SO YOU CAN DO IT IN THE VIRUS
ITSELF.
NOW, STEVE DID DO A PROTEASE
EXPERIMENT ONCE, ANOTHER FRIEND
OF HIS, LOOKING AT FUSION
PROTEIN IN BACTERIA, SIMILAR
TYPE OF ASSAY SO WE DID CROSS
OVER ONCE IN A WHILE.
WE BOTH DEVELOPED AN INTEREST IN
DUG RESISTANCE.
I'LL POINT OUT I'M SURE PAUL
BOYER FEELS LIKE HE SENT YEARS
PURIFYING RT AND DOING ASSAYS.
IT'S LED TO MANY STUDIES.
THIS IS IN PREPARATION FOR
RECENT DRUG RESISTANCE PAPER
SHOWING VARIANTS, CHEMICALLY
ACTIVE IN PRESENCE OF ATPs.
WE SPENT TWO YEARS SELECTING
RESISTANCE TO PROTEASE
INHIBITORS.
DON'T LIKE THE PROFILES.
SPEND ANOTHER TWO YEARS
SELECTING FOR PROTEASE INHIBITOR
RESISTANCE, FIFTH YEAR GET TO
SEQUENCE THEM AND SEE THIS DRUG
RESISTANCE IN CELL CULTURE.
THIS CAME IN PLAY WITH
OBSERVATION WITH FRANCOIS MADE,
HE NOTED THAT VIRUSES WITH
RESISTANCE TO PROTEASE
INHIBITORS BECAME MORE SENSITIVE
TO AZT.
STEVE AND ALEXANDRIA FIGURED
THAT OUT.
YOU CAN SEE WHAT THEY DID WAS
THEY TITRATED -- INTERPRETATION
IS THAT PROTEASE IS BAD,
CLEAVAGE IS BAD, LESS SRT IN THE
VARIANT, THEY TESTED DIRECTLY
AND GOT INTERESTING RESULT, WE
REPRODUCED PROTEASE.
YOU'RE CAPPING THE N, IT'S A
TERMINAL THING, SO AC 50 DOESN'T
CHAIN.
IF YOU TITRATE WITH A MUTANT YOU
MAKE IT -- RESIDUAL RT IS MORE
SENSITIVE BECAUSE THERE'S NOT
VERY MUCH, YOU CAN TITRATE OUT.
AZT, YOU WITH SEE, IT'S MORE
LIKE NAVAPANE THAN 3TC, WE DID
THE SAME THING, TITRATING.
WE DID THE SAME THING, TITRATING
PROTEASE WHICH HAS THIS INDIRECT
EFFECT ON RT, YOU BECOME MORE
SENSITIVE TO PROTEASE INHIBITORS
BECAUSE PROTEASE INHIBITORS ARE
TITRATING ACTIVITY.
YOU REQUEST SEE TENOFOVIR
SLIGHTLY AFFECTED.
OTHERS LOOK JUST LIKE EXPERIMENT
STEVE DID.
TO ME, FASCINATING THING, IT
TELLS YOU THE DIFFERENCE IS AZT
CAN BE REMOVED.
WHAT THIS TELLS YOU IS THAT THE
RT THAT PUTS IT ON IS NOT THE
SAME THAT TAKES IT OFF.
AND SO I THOUGHT THAT WAS A
WONDERFUL INSIGHT TO GET TO
PARTICIPATE IN.
SO, STEVE HAS BEEN GENEROUS TO
ME OVER THE YEARS.
ONE PLACE, WHEN YOU HOST THE
RETROVIRUS MEETING SOMEONE GETS
ASKED, AND THEY GET TOLD FIND
SOMEONE ELSE YOU THINK YOU CAN
WORK WITH. STEVE WAS ASKED,
INVITED ME TO DO IT, I GREATLY
ENJOY, PICK THE COVERS.
YOU SAW THIS.
THIS IS 1993, YOU SAW THIS
EARLIER IN EDDY'S TALK, HARD TO
TAKE THE EXCITING WORK AND PUT
IT ON THE COVER, THE OTHER THING
WAS, IT WAS HARD TO FIGURE OUT
THE FIRST PAPER PEER REVIEWED WE
PUBLISHED, I THINK ABOUT THE
PROVIRUS ID, I THINK THAT'S IT,
BUT WE PUT THAT ON THE BACK.
A WAY OF MAKING SURE HOWARD KNEW
WE WERE THINKING ABOUT HIM.
THIS IS MY COPY OF THE
RETROVIRUS BOOK.
IT'S AN OLD COPY.
IT WAS WONDERFUL TO BE INVITED
TO WRITE A CHAPTER, AND I'M
GRATEFUL FOR THAT.
YOU'LL SEE BECAUSE I USED THE
WRONG EXPRESSION SYSTEM, I HAD
TO WRITE THE PROCESSING OF VIRAL
PROTEINS.
I WONDERED IF IT WAS A SPLIT
VOTE WHETHER OR NOT I WOULD GET
TO BE AN AUTHOR.
I WAS TOLD I HAD TO TELL A
FISHING STORY.
HOW MANY TIMES A YEAR TO DATE,
HOW MANY PEOPLE HAS STEVE TAKEN
FISHING?
IT'S LIKE EVERYBODY, RIGHT?
HALF THE PEOPLE IN THE ROOM.
STEVE, HOW MANY TIMES HAVE WE
BEEN FISHING?
>>HOW MANY TIMES HAVE I ASKED
YOU?
>>I'M AVAILABLE.
[LAUGHTER]
SO THE ANSWER IS ZERO.
HOW DO I FEEL?
HE TAKES STRANGERS FISHING.
[LAUGHTER]
ED SAID I HAD TO TELL A FISHING
STORY.
I GREW UP FISHING IN THE
SIERRAS, AND HERE'S A COUPLE
YEARS AGO, A LONG TRIP.
THERE'S THE BACKGROUND, CRABTREE
MEADOW, YOU CAN FISH FOR GOLDEN
TROUT, GORGEOUS BUT THE SIDES OF
YOUR HAND.
YOU WOULD CATCH AS MANY ON
VELVEETA CHEESE AS A FLY.
NOT HARD TO CATCH.
THAT ONE GOT RIGHT BACK IN THE
WATER.
I FOUND MY LOVE FOR FISHING.
IT'S NOT -- YOU CATCH ALSO TROUT
BUT IN WESTERN MONTANA YOU CAN
CATCH REAL TROUT.
THERE'S A FAMOUS GUIDE OUT
THERE, FOR THE RICH.
THE RICH AND FAMOUS SCIENTISTS.
JACK MEYER TAKES THEM FISHING.
THAT WAS THE WAY -- THERE ARE
TWO WAYS THAT I THINK THAT I
REACHED MY HEAVEN FOR FISHING.
ONE, THESE FISH ARE KIND OF BIG
FOR ME TO HANDLE WITH MY HANDS.
YOU TRY TO GET THE FISH BACK IN
THE WATER SO IT DOESN'T DO GOOD
TO HAVE ME DROP THEM IN THE
BOAT.
THE OTHER, YOU BORROW A LINE,
JACK, WE'RE GOING TO NEED A
BIGGER NET!
[LAUGHTER]
THE OTHER THING THAT WAS
INTERESTING ABOUT THE TRIP, JACK
TURNS AROUND, THERE'S A FLY IN
THE BACK OF HIS SHIRT.
JACK, YOU'VE GOT A FLY BACK
THERE.
SO, THAT'S DR. X.
I SAID, I KNOW A LOT OF PEOPLE
WHO KNOW DR. X, I'M GOING TO
TELL THAT STORY.
HE MADE ME PROMISE I COULDN'T
TELL IT.
[LAUGHTER]
AT LEAST I CAN'T TELL YOU WHO
DR. X IS BUT EVERY ONE OF YOU
KNOWS DR. X.
[LAUGHTER]
SO GOING OUT ON TOP, IT'S GREAT
TO SEE THE WORK, YOU'VE SEEN
THIS SEVERAL TIMES.
REALLY WONDERFUL TO SEE THIS
BECAME AN INSTANT CLASSIC.
WHAT YOU CAN IMMEDIATELY INFER
FROM IT.
YOU TALK ABOUT INTEGRATION
SITES, THERE'S A GAZILLION.
IF YOU WANT TO TALK ABOUT
ANYPLACE IN THE GENOME, THEY
HAVE A LOT TO SAY ABOUT IT.
AND SO MANY OF YOU KNOW I HAVE
AN INTEREST IN MACROPHAGE
VIRUSES, THE WHOLE IDEA MOVING
FROM ACTIVATED T CELL TO
MACROPHAGE IS PROBABLY NOT AS
PROFOUND AS INSECT TO MAMMALIAN
CELL BUT MUST BE SOMETHING ALONG
THOSE LINES, RAISING THE
POSSIBILITY, IT TOOK A LONG TIME
BUT HE DID AGREE.
I NEED FIVE MILLION INTEGRANTS,
I WANT 10% EFFICIENCY TO DO A
COMPARISON.
MY HOPE IS WE'LL SEE CERTAIN
GENES DROP OUT AND WE CAN FIGURE
OUT WHAT CELLS ARE BEING
AFFECTED BY THE PATTERN OF
INTEGRATION SITES NOW.
THERE'S SOME -- TURNS OUT TO BE
ERP, WE'LL SEE WHAT HAPPENS.
WE'VE BEEN WORKING ON GETTING
FIVE BILL INTEGRANTS.
I GOT ANEMIC AND HAD TO STOP FOR
A WHILE.
I'M FEELING BETTER NOW AND WE'LL
GET BACK TO IT.
[LAUGHTER]
MY CONFIDENCE IS SHAKEN TODAY,
WHETHER OR NOT HE'S RETIRING.
I STOPPED BY THE HOUSE AND GOT A
LETTER, NICE LETTER.
FIRST I WANT TO THANK YOU FOR
MANY YEARS OF SERVICE TO OUR
COUNTRY THROUGH YOUR EFFORTS AS
A RESEARCHER, FROM THE WHITE
HOUSE.
THE MOST IMPORTANT
CONTRIBUTIONS, HIV SPECIFICALLY.
YOUR WORK WILL FOREVER BE PHONE
FOR QUALITY.
THAT'S REALLY NICE, COMING FROM
THE PRESIDENT.
THEN HE SAID MORE.
ONE OF YOUR COLLEAGUES HAS
CONVINCED ME THAT YOUR UNFUND
PROJECT MAPPING INTEGRATION
SITES IN MACROPHAGES IS
ESSENTIAL FOR OUR GOAL OF
CONQUERING HIV.
[LAUGHTER]
THEREFORE USING THE POWERS
INVESTED IN ME, IN THE
CONSTITUTION, I AM SUSPENDED ALL
YOUR FISHING LICENSES UNTIL THAT
PROJECT IS COMPLETE.
LET DR. COLLINS KNOW SO THE
ACTION CAN BE REVERSED, SIGNED
BY THE PRESIDENT.
[APPLAUSE]
WHEN YOU GO TO PubMed, STEVE
AND I APPEAR TOGETHER ONCE.
IT SURPRISES ME, BUT THAT'S THE
WAY IT TURNED OUT.
IT'S NICE THAT THAT'S THE SAME
NUMBER OF PAPERS HE'S PUBLISHED
WITH MY DAUGHTER.
[LAUGHTER]
MANY OF YOU KNOW ADRIAN, SHE
JUST HAD A BABY GIRL, ELEANOR.
WE'RE IN GOOD SHAPE.
SHE'LL BE BACK SOON.
WHAT CAN I SAY, ALL THE BODIES
IN THE SOLAR SYSTEM, IT'S TRUE
FOR STEVE.
JUPITER'S GRAVITY AFFECTS ALL
BODIES IN THE SOLAR SYSTEM.
IT'S TRUE FOR STEVE.
SUNSET, MAYBE IN FREDERICK, BUT
CERTAIN DAYS THE SUN IS SETTING
IN FREDERICK, IT'S RISING IN THE
WESTERN SIDE OF ALASKA.
HAVE FUN.
[APPLAUSE]
I DON'T KNOW IF THAT'S AMENABLE
TO QUESTIONS.
>>ANY COMMENTS, QUESTIONS OR
OTHERWISE?
IF NOT, THANK YOU VERY MUCH.
>>THAT'S A TOUGH ACT TO FOLLOW.
IT'S A PLEASURE TO INTRODUCE THE
FIRST OF OUR TWO DRP SPEAKERS,
FIRST A MEMBER SINCE FOUNDING,
CURRENTLY DEPUTY DIRECTOR, WITH
STEVE AND I HAVE BEEN CLOSE
FRIENDS FOR 35 YEARS, SINCE WE
WERE IN MADISON TOGETHER.
>>THANK YOU SO MUCH.
RON, THANKS A LOT.
THIS IS HARD TO FOLLOW.
SO I WOULD LIKE TO ECHO WHAT
EVERYBODY SAID.
THIS IS TRULY A WONDERFUL
OPPORTUNITY TO CONGRATULATE
STEVE AND ALSO I WOULD LIKE TO
FIRST TALK A LITTLE BIT ABOUT
THE TRIBUTE TO STEVE.
I FIRST MET STEVE IN COLD SPRING
HARBOR IN 1989, I WAS A POSTDOC
FROM HOWARD'S LAB, MY FIRST
ROTATION.
AND HE WAS CHAIRING THE SESSION.
I WAS PRESENTING SO, FIRST TIME,
I WAS A LITTLE INTIMIDATED.
VERY MUCH INTIMIDATED.
ESPECIALLY NOTICED HOW WHEN
PEOPLE WENT OVER TIME, HOW STEVE
WOULD SNEAK UP BEHIND THEM AND
PEOPLE LOOKED AT THEM AND
JUMPED, WAS REALLY INTIMIDATED.
I ALWAYS RESPECTED HIM ABOUT HIS
SCIENTIFIC INSIGHT BUT REALLY
GOT TO KNOW STEVE AFTER I JOINED
DRP HERE IN 1999.
AS MY COLLEAGUE HAS IMPACTED ME
IN DIFFERENT WAYS.
ONE OF THE WAYS HE IMPACTED MY
SCIENTIFIC CAREER HE'S ONE OF
THE LUMINARIES IN OUR FIELD,
INSPIRED ME TO EXPLORE MORE
INTERESTING TOPIC, TOOK LONGER
VIEWS OF THINGS, TAKE MORE
CHANCES.
AND ALSO HIS QUESTIONS, ALSO NOT
JUST ME BUT A LOT OF PEOPLE IN
LINE, IF YOU WANT TO SHOW DATA
YOU BETTER BE PREPARED, HE WILL
ASK IF YOU'VE DONE CONTROL TO DO
XYZ, AND IF YOU WANT TO PRESENT
YOUR FAVORITE HYPOTHESIS YOU
HAVE TO THINK ABOUT OTHER
POSSIBILITIES, IS HE GOING TO
ASK IF IT'S POLICY, EXPLANATION
FOR THAT, ALSO IN ADDITION ALSO
HELPED US EDUCATE A LOT OF
JUNIOR COLLEAGUES, AND HE ASKED
A QUESTION GENTLE, VERY
INSIGHTFUL QUESTIONS IN
SCIENTIFIC WORKING PROGRESS
MEETINGS, SOME OF MIGHT HAVE
STUDENTS HAD COMMENTED THAT
BASICALLY WE HAVE COLD SPRING
HARBOR FIRST ROLE IN WORK IN
PROGRESS SO I'M NOT NERVOUS WHEN
I PRESENT AT COLD SPRING HARBOR.
SECOND THING I WANT TO MENTION
IS THAT WE HAVE AT NIH OUR
ANNUAL ETHICS REVIEW THAT WE
HAVE TALKED ABOUT RESEARCH
ETHICS.
STEVE HAS GENEROUSLY OPENED HIS
DISCUSSION TO EVERYBODY IN DRP,
SO FOR ONE DAY MOSTLY IN
DECEMBER WE GET TOGETHER AND
DISCUSS THE ONE CASE WE HAVE TO
DISCUSS, STEVE ALWAYS MANAGED TO
MAKE IT VERY ENTERTAINING, VERY
INTERESTING, AND WE ALWAYS HAVE
A VERY NICE DISCUSSION, SO THAT
I ALWAYS REALLY APPRECIATE.
SO, I HAVE TO SAY, STEVE TOOK ME
FISHING ONCE AND TURNED OUT TO
BE I WAS NOT COORDINATED ENOUGH
TO GET THE FISH AND PUT IT BACK
IN THE WATER WITHOUT MAKING A
BIG MESS.
SO, BUT WE FOUND SOMETHING IN
ADDITION TO SCIENCE THAT BOTH OF
US ENJOYED VERY MUCH, GOOD FOOD.
SO, THIS IS ONE PICTURE AFTER A
SYMPOSIUM WE HAD THE SPEAKERS IN
A RESTAURANT, AND THIS IS
ANOTHER PICTURE OF US IN OUR
FAVORITE THAI RESTAURANT BETWEEN
BETHESDA AND FREDERICK THAT
AFTER WE GO TO A MEETING IN
BETHESDA WE OFTEN STOP AT THIS
THAI RESTAURANT, REASON BEING
THE TRAFFIC IS REALLY BACKED UP
ON 270, WE MAY AS WELL HAVE
DINNER FIRST.
THIS IS MANY A GREAT MEAL, I
SHARED WITH STEVE, I ENJOY
STEVE'S FRIENDSHIP AND
SCIENTIFIC IMPACT, LOOKING FOR
MANY MORE INTERACTIONS TO COME.
SO TODAY I'LL TELL YOU ABOUT OUR
WORK, HOW HIV SELECT VIRAL
GENOME.
SO, TO GENERATE INFECTIOUS VIRUS
HIV MUST MINE RNA AND PACKAGE
INTO PARTICLES.
IN INFECTED CELLS HIV RNA,
THERE'S LITTLE RNA, SO IT HAS A
TOUGH, TOUGH JOB.
BUT IN ADDITION TO THAT MAKE
THINGS MORE DIFFICULT, MORE
RECENTLY SHOWN THAT NOT ALL HIV
RNA ARE CREATED EQUAL, AND ONLY
A SUBSET OF RNA IS
PREFERENTIALLY PACKAGED, STUDIES
FROM MASUDA'S GROUP AND THEN
ALICE, BASICALLY THREE IN A ROW,
IN HIV IN OUR REGION.
SOMETIMES POLYMERASE STARTS AT
FIRST, SECOND, OR THIRD.
SOMETIMES FURTHER DOWN.
AND GENERATE IF YOU USE ONE OF
THESE TO GENERATE RNA WITH THREE
GUANOCINES, THE SAME EXCEPT THIS
BEGINNING NUCLEOTIDE.
HOWEVER, IT'S BEEN SHOWN THAT
THREEG RNA IS THE MOST ABUNDANT,
BUT 1 GNA IS PROFESSIONAL
DAMAGED.
WE GO HUH, HAVE TO TRY IT
OURSELVES.
WE DID THE EXPERIMENT, WHAT YOU
SEE IS THE INITIATION POINTS OF
ALL HIV WITH RNA IN CELL VERSUS
THE VIRUS.
AND IS SHOWN IN DIFFERENT COLOR
AS YOU CAN SEE, 3G BLUE MOST
ABUNDANT IN THE CELL, 1G IS MOST
ABUNDANT IN THE VIRUS.
SO WHAT THIS SAYS IS THAT HIV
MANAGED TO DISTINGUISH TWO RNA,
THAT'S 9 KB LONG, ONLY DIFFERENT
BY 2 NUCLEOTIDES.
TO DO THIS, WE COLLABORATED WITH
KARIN'S LAB.
HYPOTHESIS, MIKE SUMMERS AS WELL
AS OUR GROUP, VERY LIMB --
SIMILAR WE THINK IT MUST RELEASE
RNA INTO STRUCTURES AND THAT'S
HOW THEY CAN TELL THE
DIFFERENCE.
SO, THE HIV HAS A COMPLEX
STRUCTURE, WHAT PEOPLE THINK
THEY LOOK LIKE.
TO TELL YOU THE PART WE WANT TO
NOTE, I WILL DISCUSS THE POLY-A
STEM RIGHT HERE AND SL1 AND DIS.
DIS IS SIGNAL, THIS IS THE
SEQUENCE WE THINK RNA
DIMERIZATION IS.
PREVAILING HYPOTHESIS EARLIER
WAS RNA FOLDING TO DIFFERENT
CONFIRMATION, DIMERIZATION
SIGNALS OPEN SO.
IN THIS DIS SEQUENCE, NO LONGER
AVAILABLE TO MAKE DIMER, AND SO
NOW PACKAGABLE.
IT'S THOUGHT THIS STRUCTURE WAS
EXTENDED TO BE LONGER.
HOW DOES THE TWO STRUCTURES TELL
US HOW VIRUS MANAGES TO RNA, THE
IDEA IS THAT IN THIS STRUCTURE
IT'S DIFFERENT IN 1G.
IN 1G RNA IT'S SHORTER, 3B RNA
IT'S LONGER.
MAKES NON-PACKAGABLE FORM MORE
OFTEN, WE TESTED THIS HYPOTHESIS
BY MUTATING DIS AND YOU FIND TO
MAKE THE SEQUENCE MORE OR LESS
STABLE, OUR DATA DOES NOT
SUPPORT THIS PARTICULAR
HYPOTHESIS.
IF THAT'S THE CASE WHAT COULD BE
THE POSSIBLE MECHANISM?
SO, WE ACTUALLY THINK THAT A
LARGE RNA FORMS MULTIPLE
STRUCTURES, AND THEY HAVE
ENSEMBLE STRUCTURES, WE THINK 3G
RNA TEND TO FLOAT INTO
CONFIRMATIONS, MULTIPLE
CONFIRMATIONS, MOST NOT SUITABLE
FOR PACKAGING.
1G CAN FOLD SUITABLE FOR
PACKAGING.
IN VITRO TRANSCRIPTION WAS DONE,
AND THESE MADE A PAGE TO
SEPARATE THEM.
YOU CAN SEE SOMETHING DIFFERENT
RIGHT AWAY, IN THE 1G RNA YOU
CAN SEE BOTH DIMER AND MONOMER.
IN 3G RNA, MOSTLY MONOMERS.
SO HOW ARE THESE STRUCTURES
DIFFERENT?
KARIN'S LAB PERFORMED CHEMICAL
PROTEIN LOOKING AT THESE TWO,
MOST OF THEM STRUCTURE TENDS TO
BE WHAT THEY CALL EXPOSED, THAT
IS THEY FORM BOTH POLY- A AND
SL1.
MONOMERIC FORMS THAT ARE
SEQUESTERED.
THERE ARE MULTIPLE STRUCTURES,
TWO DOMINANT STRUCTURES I'M
GOING TO SHOW YOU.
THIS IS WHAT WE CALL EXPOSED
STRUCTURE.
IN THIS CASE, POLY-A IS FORMED.
HERE IS DIS.
READY TO BASE PAIR.
NC BINDING SIDES ARE EXPOSED,
READY TO BE BOUND BY PROTEIN.
ON THE OTHER HAND 3G RNA, POLY-A
IS MELTED.
YOU CAN SEE THE DIS IS
SEQUESTERED.
AND TWO NC BINDING SITES ARE NO
LONGER AVAILABLE.
WE THINK THAT'S THE DIFFERENCE.
TO BETTER PROBE THIS AND TO FIND
OUT MORE, WE HAVE TO ASK CAN YOU
CHANGE IT?
WE IDENTIFIED TWO TYPES OF
MUTANT THAT 2G RNA CAN BE
PACKAGED.
THESE MUTANT TURN OUT TO BE
MUTANT AROUND POLY-A REGION.
FIRST TYPE OF MUTANT WE CHANGED
STABILITY OF POLY-A, A LITTLE
SHORTER AND LESS STABLE.
AS YOU CAN SEE IN THIS CASE IN
THE CELL LINING THE DISTRIBUTION
IS SIMILAR TO THE WILD TYPE.
HOWEVER, IN THE VIRUS RNA YOU
CAN SEE NOW IN THE PACKAGE ALLOW
MORE 3G RNA, IT CAN ALSO BE
PACKAGED.
THE SECOND TYPE OF MUTANT WE
MADE, WE ADDED TWO NUCLEOTIDE
AND ADD ADDED, CG RNA CAN ALSO
BE PACKAGED.
SO WE HAVE ALSO LOOKED AT RNA
STRUCTURE WITH THESE.
AND WE TAKE ALL MUTANTS AND PUT
THEM IN THE SHELL LIKE I SHOWED
EARLIER.
SO WHAT YOU CAN SEE, IF YOU ADD
NUCLEOTIDES BETWEEN THE TAR AND
POLY-A, WHETHER IT'S 3G OR 1G
RNA NOW YOU CAN SEE MOSTLY
DIMERIC RNA.
ON THE OTHER HAND IF YOU
DESTABILIZE POLY-A STEM, MOST OF
THE RNA REGARDLESS OF 3G OR 1G
RNA ARE MONOMERIC IRAN. -- RNA.
HIV HAS A DYNAMIC STRUCTURE, IF
YOU CAN FOLD 5-PRIME POLY-A
HAIRPIN RNA CAN FOLD INTO
PACKAGABLE FORM.
AND THEY ARE DIMER READY.
EXPOSED, HAVE EXPOSED GAG
BINDING SITE, READY TO BE
PACKAGED AS GENOME.
ON THE OTHER HAND, IF THESE RNA
ARE NOT FOLDING TO POLY-A HAIR
HAIRPIN, THEY GET SEQUESTERED
AND CANNOT BE PACKAGED AS
GENOME.
AND THIS IS THE DIFFERENCE, THIS
IS HOW HIV DISTINGUISH BETWEEN
1G AND 3G RNA, LIKE THEIR
GENOME.
THEN WE ASK IS THIS UNIQUE?
IS THIS UNIQUE HIV OR DOES IT
EXTEND TO OTHER VIRUSES?
AND THIS IS DONE IN
COLLABORATION WITH BRENDA'S LAB,
ALSO RENEE'S LAB.
THE FIRST THING WE WANT TO DO IS
ASK ABOUT HIV.
OUR STUDY AS WELL AS OTHER
STUDIES ARE BASED ON NL4-3, DO
THEY ALSO USE TRANSCRIPTION
START SITES AND DO THE SAME
THING?
THIS IS EXAMPLE IN ALPHA 3, AS
EXAMPLE.
FIRST THING WE WANT TO KNOW
OTHER TYPE OF VIRUSES, WE TOOK
THREE ISOLATES AND ASKED TO SEE
WHAT HAPPENED.
ONE STRAIN TURNED OUT TO BE VERY
SIMILAR TO NL4-3.
THE SURPRISE SAME WITH WHO OTHER
STRAINS OF ISOLATE, TURNED OUT
TO BE THEY SHIFTED SIGNAL
TRANSCRIPTION A LITTLE BIT,
STARTED THE TRANSCRIPTION, 1G,
ALSO HAD VERY LOW 3G, SHIFTED
DOWN TO USE THE FIFTH XYLAZINE
AS A START SITE.
IF YOU LOOK AT CELLULAR RNA
STILL USED MULTIPLE
TRANSCRIPTION SITE, SECOND IF
YOU LOOK, STILL PACKAGED, THOSE
TWO ARE STILL TRUE.
WE THEN LOOKED AT HEPATITIS C
VIRUS, VERY PREVALENT IN THE
WORLD, USING MJ 4 AND CDH 185.
IN SIMILARLY, CAN YOU SEE THEY
HAVE MULTIPLE TRANSCRIPTION, 1G
IS PREFERENTIALLY PACKAGED.
THESE TWO ARE MORE 1G RNA IN THE
CYTOPLASM TO BEGIN WITH.
SO WE'RE FAIRLY CONFIDENT HIV
GENERALLY USE NEIGHBORING SITES
AND MAKE MULTIPLE FULL LENGTH
RNA, AND SUBPOPULATION IS
PACKAGED.
BUT WHERE DO THESE COME FROM?
FROM PROGENITOR VIRUSES OR
SOMEHOW HIV GOT INTO THE HUMAN
POPULATION AND THEY DO THESE
THINGS?
WE LOOK AT PROGENITOR VIRUSES,
HIV ORIGINATED FROM SIV CTC,
THESE TWO ARE THE ISOLATES
THOUGHT TO BE SIMILAR TO THE
VIRUS THAT GOT INTO THE HUMAN
POPULATION.
WE TALKED TO BRENDAN AND HAVE
GOTTEN SOME VIRUS FROM HIM AND
WE LOOK AT WHAT HAPPENED TO THE
VIRUSES SIMILAR TO PROGENITOR OF
GROUPM AND N, THEY BEHAVE
SIMILAR IN L4, 3.
THEY PACKAGE 1G RNA.
WE LOOK AT WHAT HAPPENED TO
GROUP O VIRUS, SIV GORILLA, THEY
EXHIBIT SIMILAR THINGS EXCEPT IN
THEIR CASE IN GORILLA THEY SEEM
TO HAVE A LITTLE BIT MORE 1G TO
BEGIN WITH.
SO, WE WOULD LIKE TO PUSH THIS A
LITTLE FURTHER AND ASK HOW FAR
DOES THIS CONSERVATION GO?
SIV CPC IS THOUGHT TO BE
RECOMBINANT.
THERE'S NOT A LOT OF REAGENT
AVAILABLE SO WE WERE ABLE TO
OBTAIN ONE STRAIN OF RCM
INFECTIOUS VIRUS, THIS GUY WAS
NOT LIKE ANYBODY ELSE.
MOSTLY PRODUCED 3G RNA AND
CELLULAR AND VIRUS ARE NOT TOO
DIFFERENT.
THIS GUY IS ACTUALLY THE
EXCEPTION TO THE RULE.
SO WE STILL CANNOT CONCLUDE AT
THIS POINT BEYOND DIRECTLY
RELATED VIRUS HOW FAR DOES THIS
CONSERVATION GO.
HOWEVER, WE CAN LOOK AT ANOTHER
VIRUS, AND SO WE LOOK AT HIV-2
AND RELATED VIRUSES.
AND BECAUSE OF TIME CONSTRAINT I
WON'T SHOW YOU THE DETAIL,
BOTTOM LINE IS THAT HIV-2 CAUSES
AIDS, AND CAME FROM ZOONOTIC
TRANSMISSION OF DIFFERENT VIRUS
SIV SMM, SO WE LOOK AT THESE
VIRUSES AND FIND THAT THEY ALSO
USE MULTIPLE TRANSCRIPTION START
SITES, BOTH HIV 2 AND SIVMAC
SELECTIVELY PACKAGE A
SUBPOPULATION OF RNA.
TO BRIEFLY SUMMARIZE, I SHOWED
YOU TODAY HIV ACTUALLY USES
NEIGHBORING SEQUENCES TO
GENERATE MULTIPLE FULL LENGTH
RNA, AND THE CONTEXT AFFECT THE
RNA STRUCTURE AND FUNCTIONS.
ONLY A SUBSET OF THEM ARE
PREFERENTIALLY PACKAGED.
THESE FEATURES LARGELY CONSERVED
IN MANY PRIMATE LENTIVIRUSES
INCLUDING THE PROGENITOR OF
HIV-1.
THESE FEATURES PREEXIST PRIOR TO
INTRODUCTION INTO HUMAN
POPULATION, AND HIV 2 AND HIV-2
RELATED ALSO HAVE SIMILAR
FEATURES.
I WOULD LIKE TO ACKNOWLEDGE THE
PEOPLE IN MY LAB.
A LOT OF PEOPLE IN THE LAB,
MOSTLY EVERYBODY CONTRIBUTED TO
SOME CURRENT OR FUTURE WORK, AND
THESE ARE THE FORMER MEMBERS,
AND THESE STUDIES ARE DONE IN
COLLABORATION WITH KARIM'S LAB,
JONATHAN WAS A GRADUATE STUDENT
IN HIS LAB, NOW HE'S IN MY LAB
LOOKING AT RNA BIOLOGY.
AND BRENDAN KEEL, AND WE WANT TO
THANK ERIC AND UNIVERSITY OF
ALABAMA FOR REAGENTS.
THANK YOU SO MUCH.
[APPLAUSE]
>>QUESTIONS?
>>WONDERFUL WORK.
ENJOY THE SPECIALIZED PACKAGING.
IS THERE SPECIALIZATION RNAs
GET TRANSLATED?
>>GREAT QUESTION!
I WOULD LIKE TO TELL YOU ABOUT
THAT ANSWER IN A COUPLE OF
MONTHS.
WE'RE IN THE PROCESS OF DOING
THAT.
ONE THING I DO WANT TO SAY IS
THERE ARE SPECIALIZED BUT
DIVISION IS NOT ABSOLUTE.
SOME 3G IS STILL PACKAGED.
SO WE ARE CURRENTLY LOOKING AT
THAT AND IT'S COMPLICATED.
IT'S NOT ONE OR THE OTHER.
THERE'S REALLY INTERESTING
MECHANISTIC THINGS THERE.
>>I'M WONDERING WHY, WHY DOES
IT HAVE SUCH A COMPLICATED
SYSTEM.
>>AH, GREAT QUESTION.
WHY DO THEY EVEN BOTHER WITH
THIS, RIGHT?
HIV IS A SERIOUS VIRUS, THEY
DON'T DO SOMETHING THIS
COMPLICATED FOR NO REASON AT
ALL.
THERE'S A GOOD REASON FOR THAT.
SO THAT'S ANOTHER QUESTION WE'RE
CURIOUS ABOUT.
WHAT DO YOU REALLY NEED THE
DIVERSITY FOR THE VIRUS TO WORK?
YEAH.
YES, RON?
>>SIMILAR QUESTION, SPLICING,
WHERE THE DECISION TO NOT BE
AVAILABLE FOR SPLICING IN TERMS
OF THESE STRUCTURES.
>>YEAH, I KNOW.
I HATE TO SOUND SO REPETITIVE
BUT WE'RE TRYING TO DO THIS.
AS YOU KNOW, BEAUTIFUL STUDIES
LOOKING TO FIGURE OUT USING NGS
TO LOOK AT SPLICING CURRENTLY
DEVELOPING SIMILAR METHODS TO
LOOK AT 5 PRIME END.
I THINK WE'RE GETTING THERE BUT
HOPEFULLY WE'LL HAVE SOME
ANSWERS FOR YOU SOON.
YES?
>>IS THERE A WAY TO SUPPRESS
ONE OR THE OTHER RNA TO SEE WHAT
THAT DOES?
>>YES.
THERE IS A WAY.
SO WE MAKE MUTATIONS NEAR THE
PROMOTER REGION AND YOU CAN'T
SHIFT TRANSCRIPTION, OBVIOUSLY
WHAT HAPPENS WHEN YOU SHIFT THE
TRANSCRIPTION, YEAH, HOPEFULLY
WE'LL FIND OUT SOON.
>>THANKS VERY MUCH.
[APPLAUSE]
WE'LL TAKE A 15-MINUTE BREAK AND
RESUME.
>>IT'S A GREAT HONOR TO
INTRODUCE THE NEXT SPEAKER, MIKE
BISHOP FROM SAN FRANCISCO,
PROFESSOR EMERITUS AS UCSF
SHARED THE NOBEL PRIZE FOR
HAROLD VARMUS, MIKE ALSO SERVED
AS CHANCELLOR OF UCSF, A BIG
ADMINISTRATIVE JOB.
IT'S AN HONOR FOR YOU TO JOIN US
TODAY
>>THANK YOU VERY MUCH.
THE NOBEL PRIZE, AN ANECDOTE
ABOUT THAT.
I WAS SHARING A ROOM AT COLD
SPRING HARBOR, LET'S SAY RATHER
GRIM LIGHT, A DOUBLE-DECKER BED,
I HURRIED OVER TO GET THE OTHER
BUNK.
WORRIED THE BUILDING MIGHT
COLLAPSE ON ME.
[LAUGHTER]
THERE WE ARE, BOTH HAD GOTTEN IN
AFTER A LATE NIGHT.
AND FROM BELOW, STEVE GROWLS AT
ME, HE SAYS, DO YOU REALIZE,
BISHOP, YOU'RE GOING TO
STOCKHOLM!
I SAID WHAT?
SO, THE FIRST ANYONE RAISED THE
POSSIBILITY, AND IT DIDN'T
HAPPEN AGAIN UNTIL THE LEGENDARY
3 A.M. BUNKO EVERYBODY TALKS
ABOUT.
OKAY.
LET ME JUST SAY I'M REALLY
DISAPPOINTED I CAN'T BE THERE
BECAUSE OF PERSONAL
CIRCUMSTANCES.
STEVE WAS A WONDERFUL COLLEAGUE
DURING HIS TIME WITH HAROLD AND
ME, GREAT SCIENTIST,
ENTERTAINING INDIVIDUAL, AND
HE'S REMAINED A FRIEND OVER THE
YEARS EVEN THOUGH WE'VE BEEN
SEPARATED BY, WELL, THE WHOLE
CONTINENT.
WHEN STEVE WAS WITH US, I HAD A
NICKNAME WITH HIM WHICH NO ONE
ELSE ADOPTED FOR REASONS THAT
MIGHT BE OBVIOUS.
MY NICKNAME FOR STEVE WAS
"HUGE."
IT'S EASY TO EXTRACT FROM HIS
FAMILY NAME, HUGHES, AND IT
EMBODIES THE SORT OF PRODIGIOUS
HEFT OF BODY, MIND AND SOUL THAT
IS THIS MAN.
HE'S QUITE A PACKAGE.
AND I'M GOING TO OPEN THAT
PACKAGE BY MEANS OF ANECDOTES.
I WAS A PARTICIPANT OR WITNESS
IN MOST OF THESE.
BUT IN A FEW, I'VE HAD TO RELY
UPON SECONDHAND ACCOUNTS,
SUPPLEMENTED BY MY OWN
IMAGINATION.
TO BEGIN, STEVE NEVER HESITATES
TO SPEAK TRUTH TO POWER.
I'M GOING TO GIVE TWO EXAMPLES.
THE FIRST INVOLVED A TATTERED
T-SHIRT THAT STEVE WORE ALMOST
CONTINUOUSLY TO ME.
THE IMAGE WAS A BARBED WIRE
FENCE.
THE TEXT READ "STEVE LUBBOCK."
STEVE WAS ACQUAINTED WITH
LUBBOCK, HE SPENT SOME TIME
THERE, BECAUSE HIS MOTHER TAUGHT
AT THE STATE COLLEGE IN LUBBOCK.
AT SOME POINT, HAROLD TOOK
NOTICE OF THE T-SHIRT.
AND REMARKED, THAT I TOOK AT A
JEST.
STEVE, OBLIVIOUS TO THE JEST, IN
THAT TONE HE GENERALLY USES TO
DELIVER COUP DE GRAS, YOU HAVE
NOT SKIED LUBBOCK.
A POSTDOC MIGHT HAVE ASKED ABOUT
VERTICAL DROP.
NOT STEVE.
A POSTDOCAL FELLOW WHO I SHALL
CALL OMICRON.
OMICRON WAS A VIRTUOSO
EXPERIMENTALIST WITH A MAIN
FAULT.
HE LEFT BEHIND HIM WORKBENCHES
THAT RESEMBLED THE AFTERMATH OF
THE BATTLE OF VERDUNNES.
AND HE NEVER CLEANED THEM UP.
SO, ONE EVENING STEVE
ENCOUNTERED THE AFTERMATH IN A
ROOM DIVERTED WORK TO
RADIOISOTOPES, STEPHEN HAD
ENOUGH.
IT WAS IN SHAMBLES.
AT 10 P.M. OR SO HE PHONED ME AT
HOME.
OUR CONVERSATION WENT SOMETHING
LIKE THIS.
STEVE SPEAKING.
BISHOP
ALWAYS BISHOP, NEVER MIND, NOT
EVEN J MICHAEL, JUST BISHOP.
BISHOP, YOU HAVE TO DO SOMETHING
ABOUT THIS!
I AGREED AND ASSURED STEVE I
WOULD TAKE IT ON IMMEDIATELY THE
FOLLOWING MORNING.
STEVE.
NO, BISHOP, NOW!
THAT TONE AGAIN.
SO, THIS TIME IT WAS I WHO
PICKED UP THE PHONE AND CALLED
OMICRON.
WITHIN THE HOUR, THE RADIO
ISOTOPE ROOM LOOKED LIKE IT
MIGHT BE OF MIGHT BE ON THE
MARKET FOR SALE, AND I CREDIT
THE SUCCESS OF THE PHONE CALL TO
THE FACT OMICRON KNEW STEVE
STARTED THE WHOLE THING AND
THAT'S WHAT OMICRON MOST FEARED.
STEVE'S HEFT.
ON OCCASION, HOWEVER, HE SPENT
TIME IN SALT LAKE CITY WITH HIS
DOCTORAL MENTOR, AND HOW CAN I
PUT THIS?
STEVE DID NOT TAKE WELL TO THE
PROMINENCE THERE OF MORMON
THEOLOGY, WHICH HE DENIGRATED BY
TELLING MORMON BACK YARDS,
NOMROM.
HE USED THE TERM IN OCCASIONAL
DIATRIBES ABOUT THE MORES OF
MORMONISM, ENTERTAINING FELLOW
WORKERS AROUND HIM.
EVENTUALLY, I DON'T HEAR
ANYBODY, IS THAT OKAY, JOE?
>>YES.
>>WE HEAR YOU LOUD AND CLEAR.
>>WE MUTED THE AUDITORIUM SO WE
DIDN'T DISTURB YOUR TALK.
>>OKAY.
SO NOMROM.
QUITE AMUSING BUT THERE WAS A
SUBDUED POSTDOC BEHIND STEVE WHO
SPOKE UP.
STEVE, YOU MIGHT WANT TO KNOW
THAT I AM A BISHOP IN THE MORMON
CHURCH.
[LAUGHTER]
WE HEARD NO MORE DIATRIBES ABOUT
THE NOMROM.
IT CAME TO PASS STEVE HAD TO
MOVE ON.
HIS SEARCH FOR LANDING PLACE HAD
ONE REQUIREMENT.
OUT OF THE FEAR OF EARTHQUAKES,
HE WOULD NOT SETTLE ON THE WEST
COAST.
HE WAS ADAMANT ABOUT THIS OF .
I TALKED TO HIM REPEATEDLY.
I THOUGHT HE WAS A DAMN FOOL,
KNOWING HIS LOVE OF THE
OUTDOORS.
AND HE WOULD NOT, NO
STATISTICAL -- NO AMOUNT OF
STATISTICAL ARGUMENT WOULD
PERSUADE HIM.
SO WE HAD TO SURRENDER HIM TO
THE EAST COAST.
OFF HE WENT.
AS YOU'VE HEARD BEFORE, TO THE
COLD SPRING HARBOR LABORATORY
WHERE I SUSPECTED THAT THE
PRINCIPAL ATTRACTION OF THE
PLACE, TO HIM, WAS EASY ACCESS
TO THE STRIPED BASS OF LONG
ISLAND SOUND.
IT WAS ALSO EXPECTED AN
EARTHQUAKE OF SORTS TO OCCUR THE
FIRST TIME STEVE CROSSED WITH
J.D. WATSON, PRESIDENT OF THE
LAB AT THE TIME, OF COURSE.
TO MY KNOWLEDGE, THAT EARTHQUAKE
NEVER HAPPENED.
INSTEAD, STEVE KEPT HIS HEAD
DOWN, MADE GOOD USE OF THE
RESOURCES AT HIS DISPOSAL, AND
EARNED A PROMINENT APPOINTMENT
AT NCI FREDERICK, THAT
EVENTUALLY BROUGHT HIM BACK INTO
MY ORBIT, A BIT BECAUSE HAROLD
WAS DIRECTOR OF NIH AT THE TIME,
AND HAD RECRUITED ME TO CHAIR
REVIEW OF THE INTRAMURAL PROGRAM
OF THE NATIONAL CANCER
INSTITUTE, WHICH WAS LOSING A
BIT OF ITS LUSTER.
SO, STEVE HAD BEEN AT FREDERICK
BY THAT TIME LONG ENOUGH TO BE
DISSATISFIED.
NOW HE SAW AN OPPORTUNITY AND
POUNCED.
HE PRESENTED ME WITH A
NON-NEGOTIATABLE PROPOSAL, TRUTH
TO POWER, I WOULD BOLSTER THE
RESEARCH PROGRAM AT FREDERICK TO
THE POINT WHERE IT WAS EQUAL TO
WHAT WAS GOING ON IN BETHESDA,
OR MOVE THE ELIGIBLE SCIENTISTS
AT NCI FREDERICK TO THE MAIN
CAMPUS.
DO IT, BISHOP.
NOW OF COURSE I COULDN'T DO IT,
I COULD ONLY TRY TO PERSUADE MY
COMMITTEE TO TRY TO DO IT, IF I
WAS SO DISPOSED.
STEVE'S STILL IN FREDERICK.
DRAW WHATEVER CONCLUSION YOU
WISH.
I SUSPECT THERE ARE QUITE A FEW
PEOPLE WHO ARE GLAD STEVE STAYED
IN FREDERICK.
STEVE'S MOVE TO FREDERICK
ALLOWED HIM TO MIMIC MARIO
CAPECCHI, BEING OFF THE BEATING
PATH.
HE BOUGHT A FORM AT THE FOOT OF
A MOUNTAIN, ALONG A NARROW AND
POORLY PAVED ROAD, NEAR WHERE
THE UNION AND CONFEDERATE ARMY
SKIRMISHED BEFORE THEY MOVED TO
ANTIETAM WHERE THEY WOULD FIGHT
ONE OF THE BLOODIEST BATTLES IN
THE HISTORY OF WARFARE.
IT WAS IN CONVERSATION ABOUT
THAT LOCALE THAT I DISCOVERED
STEVE IS, AMONG OTHER THINGS, AN
AMATEUR HISTORIAN.
PROBABLY NOT UP TO HIS
CAPABILITIES BUT NEVERTHELESS
ENTERTAINING.
THIS FARM, YOU'VE HEARD ALLUSION
TO THIS, WAS REPLETE WITH A
BARN.
AND I SAW THAT BARN AT THE TIME,
RATHER BEDRAGELLED, WITH
SOLIDIFIED RESIDUE OF PREVIOUS
OCCUPANTS AND LITTLE ELSE IN IT.
IN DUE COURSE THE BARN DID WHAT
BARNS OFTEN DO.
IT BURNED TO THE GROUND.
AH, ANOTHER OPPORTUNITY.
STEVE RESOLVED TO REBUILD AND TO
DO IT IN STYLE. IN STYLE.
HE EVEN HAD THE REQUISITE
INSURANCE.
I CAN'T ACQUIESCE TO YOUR
SUSPICIONS.
YOU HEARD STEVE RECRUITED SOME
AMISH, KNOWN FOR TRADITIONAL
BARN RAISINGS, AND THEY WERE
HANDY IN HIS NEIGHBORHOOD.
SO HE GOT HIS BARN RAISING,
START TO FINISH, IN ONE DAY.
THE BARN IS A SPLENDID EDIFICE,
DEVOID OF ANYTHING RESEMBLING A
FARM ANIMAL OR HAY, AT LEAST
LAST TIME I VISITED, STEVE WILL
TELL YOU GROUCHILY, SOME YEARS
AGO NOW.
NOW, THE BARN RAISED STEVE, TO
OCCUPY HIM DURING RETIREMENT.
I DON'T KNOW WHAT STEVE PLANS TO
DO WITH ALL OF HIS NEWFOUND FREE
TIME, BUT I'M CERTAIN THAT SOME
OF IT WILL BE DEVOTED TO THE ROD
AND REEL.
NOW, YOU'VE SEEN EVIDENCE OF
THIS ALREADY, SO I TOOK OUT MY
SLIDES TO DOCUMENT IT.
BUT STEVE, AND THIS IS NO
SURPRISE, STEVE PREFERS BIG
FISH, REALLY BIG FISH.
AND HE'S TAKEN SOME OUT OF THE
CHESAPEAKE AND THE RIVER IN THE
HEART OF THE ALASKA WHICH HE
VISITED REPEATEDLY OVER THE
YEARS AND SENT ME A PHOTOGRAPH
OF THE TERRAIN, WITH THE RIVER
RUNNING THROUGH IT, SO TO SPEAK.
AND IT'S JUST VAST, AS FAR AS
YOU CAN SEE, EMPTY WILDERNESS.
AND IT'S AWE INSPIRING.
AND I THINK IT SUGGESTED TO ME
HOW STEVE MAY VIEW HIS
RETIREMENT, UNBOUNDED
OPPORTUNITY.
WHERE IT WILL LEAD HIM, I CANNOT
IMAGINE.
I CAN ONLY PREDICT HE WILL NOT
LIE IDLE.
SO, HERE'S HUGE CONGRATULATIONS
ON THE COMPLETION OF A SPLENDID
CAREER, I WISH YOU THE BEST IN
WHAT IS TO COME.
THANK YOU.
[APPLAUSE]
>>THANK YOU VERY MUCH.
ANOTHER TOUGH ACT TO FOLLOW.
SO, IT'S A PLEASURE TO WELCOME
MARY KEARNEY AS OUR SPEAKER,
CURRENTLY DRP DEPUTY DIRECTOR
WITH WEI SHAU HU WHO YOU HEARD
FROM, PIONEERING ASSAYS,
COLLABORATED WITH STEVE ON WORK
ON HIV RESERVOIRS.
>>THANK YOU.
OKAY.
WELL, I DIDN'T GET THE MEMO THAT
WE HAD TO INCLUDE FISHING
STORIES, LIKE JEFF DID, I THINK
IT'S BECAUSE EVERYONE KNOWS I'VE
NOT BEEN FISHING WITH STEVE.
I HAVE HEARD MANY FISHING
STORIES, HOWEVER, AT OUR MANY
DINNERS TOGETHER, OVER THE YEARS
AT OUR FAVORITE RESTAURANT WITH
ALL OF THE P.I.s AND INVITED
SPEAKERS, AND THE ONLY ONE I
RECALL IS ONE THAT INCLUDED A
NEW FISHING BOAT THAT SOMEHOW
INCLUDED USING A PINK OR PURPLE
CONDOM IN SOME WAY.
THE STORY I HEARD WAS THAT WHEN
HIS WIFE FOUND A CONDOM IN HIS
PANTS POCKET, IT DIDN'T OCCUR TO
HER MAYBE HER HUSBAND WAS HAVING
AN AFFAIR.
INSTEAD SHE THOUGHT WHAT KIND
OF NEW-FANGLED FISHING THING IS
HE DOING NOW.
[LAUGHTER]
WE WOULD ALL COME TO THE SAME
CONCLUSION IF WE FOUND THAT IN
THE PANT POCKET OR OFFICE OR
SOMEWHERE.
I'M GOING TO TELL YOU ABOUT MY
COLLABORATIONS WITH STEVE AND HE
HAS SUPPORTED ME SO MUCH IN MY
CAREER HERE AT THE DRP OVER THE
LAST 20 YEARS.
AND I TITLED IT CLONAL
EXPANSIONS WITH HIV DRP BECAUSE
THEY ARE NOT JUST WITH STEVE BUT
THEY INCLUDE COLLABORATIONS WITH
JOHN AND FRANK AND MANY OTHERS
IN THE HIV DRP.
SO, WHILE STEVE WAS STUDYING
TRANSCRIPTASE AND INTEGRASE, I
STARTED IN 2001 IN JOHN
HAWKINS'S LAB, PERSISTENCE
DURING THERAPY.
WHEN INDIVIDUALS INITIATE ART,
LEVELS OF PLASMA VIREMIA DROP
PRECIPITOUSLY AND DECLINED TO
LEVELS THAT WERE BELOW DETECTION
WITH CLINICAL ASSAYS.
SO, IN JOHN COFFIN'S LAB, PALMER
DEVELOPED AN ASSAY WHERE THEY
COULD MEASURE LOW LEVELS OF
PLASMA VIREMIA, AND PLASMA
VIREMIA PERSISTS IN MOST
INDIVIDUALS AROUND A SINGLE
VIRUS PARTICLE PER mL OF
PLASMA, FOLLOWED UP WITH STUDIES
IN JOHN'S LAB I COLLABORATED ON
WHERE THE THEY MEASURED DNA
COPIES, MUCH HIGHER THAN NUMBER
OF RNA COPIES, SO DNA COPIES IN
INDIVIDUALS ON ART WILL REFLECT
NUMBER OF INFECTED CELLS THAT
PERSIST IN THESE INDIVIDUALS.
SO, WHERE THE COLLABORATION WITH
STEVE BEGAN WAS WHERE WE STARTED
TO STUDY GENETICS OF THE DNA, OR
PROVIRUSES THAT PERSISTED IN
INDIVIDUALS IN THERAPY, I'LL
START WITH SHOWING SLIDES
REPRESENTED AT THE THINK TANK IN
2012, BECAUSE THIS IS WHEN THE
COLLABORATION, MY COLLABORATIONS
WITH STEVE, BEGAN.
SO, IN THIS TALK I SHOWED
GENETICS OF SINGLE PROVIRAL
SEQUENCES IN INDIVIDUALS WHO HAD
BEEN ON ART FOR MANY YEARS, AND
COLLECTED LONGITUDINAL SAMPLES,
IN COLLABORATION WITH FANG AND
-- FRANK AND HIS LAB.
WE CAME TO EIGHT FINDINGS.
THE FIRST IS WE FOUND THAT
IDENTICAL SEQUENCES WERE IN
PBMCs, IDENTICAL PROVIRAL
SEQUENCES IN DONORS ON LONG-TERM
ART, YOU SAW SOME FROM A
PRESENTATION EARLIER TODAY
WORKING ON SIMILAR STUDIES AT
THE TIME.
THESE WERE LONGITUDINAL SAMPLES
IN THIS INDIVIDUAL, SAMPLE
COLLECTED 27 DAYS ON ART, FOUR
YEARS AND NINE YEARS ON ART.
YOU CAN SEE POPULATIONS, ONE
CIRCLES DOWN AT THE BOTTOM,
WHERE I HAD CAL PRO -- WITH
IDENTICAL PROVIRAL SEQUENCES,
SOME CONTAINED HYPERMUTANTS, WE
KNEW SOME THAT PERSISTED WERE
NOT REPLICATION COMPETENT, AND
WE FOUND NO EVIDENCE FOR
EVOLUTION, VIRAL EVOLUTION, ON
ART.
WHEN YOU SEE EVOLUTION, BRANCHES
GET LONGER AND LONGER,
LONGITUDINAL SAMPLES, HERE YOU
CAN SEE THAT SEQUENCES COLLECTED
NINE YEARS ON ART IN THE
DIAMONDS NOT ON LONGER BRANCHES
THAN THOSE COLLECTED 27 DAYS ON
ART IN THE CIRCLES.
SO, WE ALSO FOUND THERE WAS NO
CLUSTERING OF SEQUENCES BY
SOURCE, SO THERE WERE THE
SEQUENCES THAT WE COLLECTED IN
THE PLASMA, RNA SEQUENCES IN
PLASMA, AND SEQUENCES WE
COLLECTED IN CELLS, DNA
SEQUENCES, WERE NOT SEGREGATED
SUGGESTING PLASMA VIRUSES WAS
COMING FROM INFECTED CELLS IN
THE BLOOD WITH YEARS ON ART, AND
IDENTICAL SEQUENCE ON ART
MATCHED PRESENCE SHORTLY AFTER
ART, ONE OF THESE A CIRCLE 14
DAYS AFTER ART SO POPULATIONS OF
IDENTICAL SEQUENCES SEEM TO
PERSIST FOR THE DURATION OF
TREATMENT.
WE ALSO FOUND THAT IF WE
SAMPLED, OPEN CIRCLES, AT EARLY
TIME POINT IN PRE-THERAPY AND
YEARS LATER BEFORE STARTING ART
WE SAW CLEAR EVOLUTION, SO THREE
SAMPLES BEFORE STARTING ART IN
OPEN CIRCLES SO WE CAN SEE
DISTINCT POPULATIONS, SO TWO
SAMPLES THREE YEARS APART
WITHOUT ART CLEAR EVOLUTION, BUT
WHEN COLLECTED LONGITUDINALLY ON
ART THERE WAS NO DIVERGENT OVER
3 YEARS WITH ART AND SEQUENCES
THAT PERSISTED MANY YEARS ON ART
IN THE CELLS UP TO 9 YEARS WERE
THE SAME AS SEQUENCES THERE JUST
BEFORE INITIATING ART, THREE
SEQUENCES THAT REALLY DOMINATE
THE POPULATION FOR YEARS AND
YEARS ON ART.
IDENTICAL SEQUENCES THAT WERE
FOUND IN THE PLASMA DID NOT
ALWAYS MATCH IDENTICAL SEQUENCES
THAT PERSISTED IN THE CELLS,
SUGGESTING THAT NOT ALL OF THE
INFECTED CELLS CONTRIBUTED TO
PERSISTENT VIREMIA YOU SEE
DURING TREATMENT.
BUT WHAT WE FOUND TO BE MOST
INTERESTING IS THAT SOME OF
THESE POPULATIONS OF IDENTICAL
SEQUENCES WERE DUE TO
HYPERMUTANTS STOP CODONS LEADING
TO COLLABORATION WITH STEVE WHO
DEVELOPED ASSAY WITH SHAU HU,
THE ONLY WAY THEY COULD HAVE
ARISEN TO BE IDENTICAL AFTER
MANY YEARS ON ART WOULD BE IF A
SINGLE CELL WAS INFECTED WITH
DEFECTIVE PROVIRUS AND CLONALLY
EXPANDED.
AND SO WHAT WE HAD SUMMARIZED AT
THIS MEETING IN 2012 IN OUR
CONCLUSIONS LED TO REALLY THIS
COLLABORATION WITH STEVE HUGHES
THAT LED TO SEMINAL PAPER THAT
CHANGED OUR UNDERSTANDING OF HIV
PERSISTENCE, PUBLISHED IN
"SCIENCE" A COUPLE YEARS LATER.
SO WE FOUND WHAT WE SUMMARIZED
THAT THERE'S A RESTRICTED GROUP
OF HIV VARIANTS, DETECTED FOR
YEARS ON SUPPRESSIVE ART IN
PLASMA AND BLOOD CELLS,
SUGGESTING THAT RESULT FROM
EXPAND OF ONE OR MORE INFECTED
CELLS, THE IDENTICAL SEQUENCES
WERE NOT ON LONGER BRANCHES
WHICH SUGGESTED THAT CELLS WERE
INFECTED BEFORE INITIATING
THERAPY AND THAT THESE CELLS ARE
THE SOURCE OF THESE IDENTICAL
SEQUENCES SO THEY PERSISTED FOR
ALL OF THESE YEARS ON ART LIKE
CLONAL EXPANSION.
IDENTICAL SEQUENCES MATCH
VARIANTS THAT WERE PRESENT
SHORTLY AFTER INITIATING ART AND
SO THIS LED TO A LOT OF STUDIES
WHERE WE DEMONSTRATED THAT ART
WAS FULLY SUPPRESSIVE, FULLY
POTENT, THERE WAS NO ONGOING
REPLICATION DURING TREATMENT.
SO, THOSE STUDIES AND ONE OF
THOSE PATIENTS WAS ACTUALLY
PATIENT 1.
AND SO THAT PATIENT AND FOUR
OTHERS WERE USED TO MAP
INTEGRATION SITES AND FOUND
IDENTICAL SEQUENCES HAD
IDENTICAL INTEGRATION SITES, SO
YOU SEE THIS PIECHART MANY TIMES
TODAY, THIS IS THE DATA FROM ALL
OF THE PATIENT 1 AND ALL OF THE
PATIENTS COLLECTIVELY TOGETHER
SHOWING THAT SOME OF THE SITES
DETECTED ONLY ONCE, MANY OF THEM
ARE DETECTED OVER AND OVER
AGAIN, AND SOME OF THEM ARE
VERY, VERY LARGE CLONES IN THESE
INDIVIDUALS AND THIS IS WHAT LED
TO MY FIRST COLLABORATION WITH
STEVE, WITH THIS INTEGRATION
SITE ANALYSIS, SOME FOLKS ARE
MISSING IN THIS PICTURE
INCLUDING FRANZISKA SIMONETTI.
THE OBVIOUS NEXT QUESTION WAS
CAN THESE CLONES CARRY
REPLICATION CONFIDENT VIRUS?
"SCIENCE" PAPER SHOWED HIV
PERSISTED BECAUSE CELLS INFECTED
BEFORE INITIATING ART COULD
UNDERGO CLONAL EXPANSION AND
RESULTED IN RENEWABLE --
CONTINUOUS RENEWABLE SOURCE OF
INFECTED CELLS DURING TREATMENT
DESPITE THE FACT THERE WAS NO
ONGOING VIRAL REPLICATION BUT
THE OBVIOUS QUESTION WAS, CAN
THESE CLONES CARRY REPLICATION
COMPETENT PROVIRUSES, AND I WILL
SAY THAT I THINK -- IF I HAD TO
GUESS THE BULK OF THE PEOPLE IN
THE ROOM, CONVERSATIONS I HAD,
DID NOT SEEM TO THINK THEY COULD
CARRY REPLICATION COMPETENT
PROVIRUSES BUT FRANZISKA IN
COLLABORATION DID SHOW THAT
IN -- FIRST TO SHOW CLONES EVEN
VERY LARGE CLONES WERE
REPLICATION COMPETENT AND SOURCE
OF PLASMA VIREMIA IN THIS
INDIVIDUAL.
THIS PAPER WAS FOLLOWED BY
ANOTHER PAPER IN JOHN MELLOR'S
LAB, FOUND THE SAME IN FIVE
PATIENTS, IN THESE INDIVIDUALS
REPLICATION COMPETENT PROVIRUSES
WERE THE SOURCE OF PERSISTENT
VIREMIA DURING ART AND BOB TOLD
YOU TODAY SOMETIMES IT APPEARS,
OFTENTIMES, THESE CELLS MAY NOT
BE THE SOURCE OF VIRAL REBOUND
BACKUP IT'S NOT KNOWN IN THESE
INDIVIDUALS WHETHER THE CLONES
WOULD HAVE BEEN A SOURCE OF
PERSISTENT VIREMIA HAD THEY
STOPPED ART SO MORE STUDIES
SHOULD BE DONE ON THAT.
STEVE SAID HOW DO INFECTED CELL
CLONES PERSIST WITH REPLICATION
COMPETENT VIRUS, WE KNOW THEY
ARE THE SOURCE OF PERSISTENT
VIREMIA BECAUSE WE CAN DETECT
VIREMIA IN THESE INDIVIDUALS, WE
CAN SEQUENCE THE VIRUS, IT
MATCHES THE PROVIRUS IN THE
LARGE EXPANDED CLONES, SO WE
KNOW THAT THEY ARE NOT BLATANTLY
INFECTED, WE KNOW THEY ARE
PRODUCING VIRUS PARTICLES, WE
CAN DETECT THEM IN THE BLOOD AND
IN SOME CASES CAN DETECT THEM AT
FAIRLY HIGH LEVELS IN THE BLOOD.
SO, WE FORMED HYPOTHESIS AND
MADE THIS SLIDE, IT WAS STEVE'S
IDEA.
AND USING THESE EVIL GUYS TO
SHOW CELL CLONE WITH A
REPLICATION COMPETENT VIRUS, IN
THIS CASE EMB1 VIRUS, WHICH YOU
HEARD ABOUT, IT MUST BE A SMALL
FRACTION OF THE CELLS IN THE
CLONE THAT HAVE ACTIVE
PROVIRUSES, MAKING VIRAL RNA,
THERE ARE VIRUS PARTICLES WE CAN
DETECT IN THE BLOOD, CELLS
LIKELY DIE, THE VIRUS THEY ARE
PRODUCING IS GOING TO KILL THE
CELLS BUT WHEN THEY DIE OTHERS
CELLS MUST BE REACTIVATE,
PROVIRUS ANYWAY, TO PRODUCE MORE
VIRAL, IT PERSISTS OVER
LONGITUDINAL SAMPLES, THOSE
CELLS THAT ARE STILL EXISTING
THAT ARE NOT PRODUCING VIRAL RNA
BECAUSE THEY HAVE LATENT
PROVIRUSES CONTINUE TO DIVIDE.
WE SET OUT TO ADDRESS THIS
HYPOTHESIS, WE DEVELOPED AN
ASSAY WHERE WE COULD DETECT
POPULATIONS OF EXPANDED CELL
CLONES WITH IDENTICAL PROVIRUS,
COULD ASSAY CELLS DIRECTLY FROM
THE INDIVIDUALS WITHOUT
CULTURING THEM, WITHOUT
MANIPULATING THEM IN ANY WAY,
DIRECTLY FROM THE DONORS, AND
DETERMINE WHETHER OR NOT THEY
HAD VIRAL RNA.
AND THEN THESE CLONES ATTACKED
PROVIRUSES, VIRAL RNA IS
PRODUCING VIRUS PARTICLES THE
SOURCE OF PERSISTENT VIREMIA IN
THESE INDIVIDUALS, AND WITH THIS
ASSAY COULD REPORT THE FRACTION
OF CELLS WITH PROVIRUSES
TRANSCRIPTIONAL ACTIVE AND
FRACTION WITH PROVIRUSES THAT
ARE LATENT AND WE SET OUT TO DO
THIS.
THIS ASSAY WAS USED IN PATIENT
1, IDENTIFIED 34 INFECTED CELL
CLONES, WHAT FACTION HAD
DETECTABLE LEVELS OF VIRAL RNA?
THREE HAD INTERACT PROVIRUS, ONE
CHARACTERIZED BY SIMONETTI, TWO
OTHERS SHORTLY AFTER THIS WERE
ALSO IDENTIFIED THAT HAD INTACT
PROVIRUSES, SO THREE OF THE CELL
CLONES HAD REPLICATION COMPETENT
PROVIRUSES AND WHEN WE MEASURED
THE FRACTION OF CELLS WITHIN
THESE INDIVIDUAL CELLS THAT HAD
VIRAL RNA IT WAS ABOUT 2% OF
THEM AND BY THE WAY ANDREW MUSIC
IN MY LAB TWO DID THIS STUDY
ASSAY AND JOHN SDREMLE, 1200
SINGLE CELLS CARRIED, 2% HAD
DETECTABLE LEVELS OF VIRAL RNA
USING A SENSITIVE ASSAY.
AND SO THEY THEN LOOKED AT
ANOTHER ABOUT 30-SOME CLONES, 5
OTHERS HAD PROVIRUSES, KNOWN TO
BE EFFECTIVE.
FULL LENGTH PROVIRUSES WEREN'T
CHARACTERIZED, DON'T FLOW IF
THEY WERE INTACT OR DEFECTIVE
BUT IT WAS JUST AROUND 2 TO 5%
OF THE CELLS WITHIN THESE LARGE
EXPANDED CLONES THAT HAD VIRAL
RNA.
SO MOST OF THE CELLS WITHIN
THESE CLONES HAVE PRO VIRUS,
SOME OF THESE CLONES DID HAVE A
HIGH FRACTION OF CELLS THAT HAD
PROVIRUSES TRANSCRIPTIONALLY
ACTIVE, FURTHER WILL DETERMINE
IF INTEGRATION SITES OR OTHER
FACTORS PLAY A ROLE , WHY DOES
IT VARY CLONE TO CLONE?
WE CONCLUDED EXPANDED CELL
CLONES PERSIST BECAUSE MOST OF
THE MEMBERS HAVE LATENT
PROVIRUSES.
SO FROM THERE, STEVE AND I AND
OTHERS IN THE DRP WENT ON TO
PUBLISH MANY PAPERS, ONE IS THE
PAPER, I JUST TOLD YOU ABOUT, WE
SHOWED IT'S JUST A SMALL
FRACTION OF CELLS, BUT SINCE
THAT TIME, I WAS SURPRISED TO
SEE, STEVE, HOW PRODUCTIVE WE'VE
BEEN OVER THE PANDEMIC AND HOW
MANY STUDIES WE'VE PUBLISHED
SINCE 2019 WHEN WE PUBLISHED
THIS STUDY, WE'VE PUBLISHED A
LOT OF STUDIES, LOOKED IN LYMPH
NODES, SHOWN THE SAME EXPANDED
CELL CLONES ARE IN LYMPH NODES,
LOOKED AT ACUTE INFECTION,
CLONES ARISE EARLY, THIS PAPER
HERE.
WE TRACKED SIZE OF CLONES
LONGITUDINALLY, NOW WE HAVE BIG
PLANS MOVING FORWARD BUT I'M
SURE STEVE, ALTHOUGH RETIRED,
ABOUT CONSULT WITH US, ADVISE US
AS HE ALWAYS HAS SO WE CAN
CONTINUE TO DO AMAZING SCIENCE
IN THE DRP.
STEVE, THANK YOU SO MUCH FOR
EVERYTHING I HAVE LEARNED FROM
YOU, AND YOUR SUPPORT OF MY
CAREER.
I MUST SAY AS DEPUTY DIRECTOR OF
DRP WITH WEI SU, STEVE WAS
BRANCH CHIEF, SOMETHING LIKE
DEPUTY DIRECTOR AT THE TIME, I
NEVER IMAGINED THIS DAY THAT I
WOULD BE IN A POSITION THAT WITH
ONCE BEEN FILLED BY STEVE
HUGHES.
IT'S REMARKABLE THAT IT'S
HAPPENED, AND HAPPENED BECAUSE
OF THE SUPPORT I'VE HAD FROM
STEVE AND FROM JOHN COFFIN AND
OTHERS IN THE DRP.
STEVE, THANK YOU SO MUCH FROM
THE BOTTOM OF MY HEART.
THANK YOU.
I'LL JUST LEAVE IT THERE.
[APPLAUSE]
>>ALSO HAPPENED BECAUSE OF YOUR
GREAT WORK.
QUESTIONS?
>>SEQUENCES THAT WE FIND IN RNA
VIREMIA ARE NOT ALWAYS EASY TO
FIND IN DNA, THERE'S A VERY
SMALL FRACTION OF THE CELLS
PRODUCING THIS VIRUS, SOMETIMES
PEOPLE COULD HAVE THOUSANDS AND
TENS OF THOUSANDS OF COPIES OF
RNA.
HAVE YOU TRIED TO WORK THAT
BACKWARD, OH, HERE'S THE VIRAL
LOAD IN THIS PERSON ON THERAPY,
OH, THEY HAVE THIS MANY CELLS
THAT ARE PRODUCING VIRUS?
>>WE HAVE NOT DONE THAT.
I HAVEN'T THOUGHT OF DOING THAT.
I DON'T THINK THERE WOULD BE
CORRELATION BECAUSE IN THE
FAMOUS PATIENT NUMBER 1, THE
AMB1 CLONE WAS A VERY LARGE
CLONE, I THINK THE LARGEST
INFECTED CELL CLONE IN THAT
INDIVIDUAL, LEVEL OF VIREMIA
STILL QUITE LOW IN THAT
INDIVIDUAL COMPARED TO THE
LEVELS OF VIREMIA IN THE
SUBSEQUENT PAPER FROM JOHN
MELLOR'S LAB, AND COLLABORATIONS
WITH MY LAB AT WELL, THOSE
INDIVIDUALS HAD HIGHER LEVELS OF
PERSISTENT VIREMIA THAN PATIENT
1, BUT THE CLONES WERE SMALLER,
YEAH, QUITE SURE, AND SOME WERE
I THINK VERY SMALL, THE SOURCE
OF PERSISTENT VIREMIA DESPITE
HIGHER LEVELS THAN IN PATIENT
NUMBER 1.
>>VERY BEAUTIFUL WORK.
WHAT DISTINGUISHES CELLS THAT
PRODUCE RNA FROM THOSE THAT
DON'T.
WHEN YOU GET THIS RNA, FROM A
CELL THAT YOU CAN DO SINGLE CELL
RNAseq?
>>SO, I THINK WE HAVE NOT DONE
SINGLE CELL RNAseq.
I THINK THERE ARE PROBABLY
MULTIPLE THINGS THAT LEAD TO
LATENCY.
VERSE HAVE DATA THAT SUGGEST
INTEGRATION SITE COULD BE
IMPORTANT.
I THINK THERE'S A LOT MORE WORK
TO BE DONE IN THAT AREA, IF IT'S
A SMALL N, SO THERE'S MORE WORK
TO BE DONE, IF THE INTEGRATION
SITE HAS ANYTHING TO DO WITH IT.
IT COULD BE THE STATE OF THE
CELL, MAYBE THE DIFFERENTIATION
STATE OF THE CELL, LEVEL OF
ACTIVATION OF THE CELL, IT COULD
BE THE CELL CYCLE, COULD EVEN BE
SOMETHING TO DO WITH CIRCADIAN
RHYTHMS.
THERE'S A LOT OF DATA COMING OUT
NOW, VARYING OVER THE DAY.
THERE'S MORE WORK TO BE DONE TO
DETERMINE THAT.
>>OKAY.
THANKS VERY MUCH.
[APPLAUSE]
NEXT SPEAKERS, NEIGHBOR JEFF
LIFTON, IN THE FREDERICK
NATIONAL LAB FOR CANCER
RESEARCH, SPENT HIS CAREER AT
THE FOREFRONT OF THE FIELD OF
RETROVIRAL PATHOGENESIS,
PARTICULARLY KNOWN FOR WORK ON
SIV, COLLABORATED WITH STEVE ON
SEVERAL PROJECTS, WE'LL LIKELY
HEAR MORE DURING HIS TALK TODAY.
>>THANKS.
IT'S NOT THE SAME WITHOUT THE
SIZE 14s ON THE STAGE.
I'M GLAD I WAS ABLE TO FIND THIS
PICTURE.
MOST OF THE PICTURES TODAY HAVE
BEEN FROM WEBSITES AND THINGS,
HE LOOKS WAY MORE DOUR THAN IS
TYPICAL OF HIM.
I SURROUNDED HIM WITH FOUR OF
HIS FAVORITE THINGS, A SMILING
FACE TO CAPTURE SOMETHING THAT'S
MORE REFLECTIVE OF MY EXPERIENCE
WITH STEVE.
TO HAROLD'S INTRODUCTION, MY
PRESENTATION WILL FALL INTO THAT
CATEGORY, BASICALLY PRAISING
STEVE.
I WANT TO PRESENT IT FROM SORT
OF TWO SYNERGISTIC PERSPECTIVES.
ONE, TO TALK ABOUT THE
INTERACTIONS I'VE HAD WITH HIM
SCIENTIFICALLY, BUT ALSO ABOUT
THE INTERACTIONS I'VE HAD WITH
HIM OUTSIDE THE LAB, REINFORCING
THINGS THAT WE'VE HEARD ABOUT
TODAY THAT REINFORCE WHO STEVE
IS AS A PERSON, WHOSE DEEP
KNOWLEDGE OF MANY FIELDS HAS
CRITICAL ASSESSMENT OF THAT
KNOWLEDGE, HIS CLEAR, DIRECT,
BUT ALWAYS GRACEFUL EFFECTIVE
COMMUNICATION OF HIS INSIGHTS,
AND ALONG WITH THAT A GREAT
SENSE OF HUMOR, BOUNDLESS
ENTHUSIASM FOR THE THINGS HE'S
INTERESTED IN, AND AT THE SAME
TIME ENTHUSIASM FOR
COMMUNICATING AND SHARING THAT
ENTHUSIASM, THAT KNOWLEDGE, AND
SEEMINGLY INEXHAUSTIBLE PATIENCE
FOR DOING SO, TAKING GREAT PRIDE
AND PLEASURE WHEN HE'S ABLE TO
GET SOMETHING ACROSS IN A WAY
THAT BENEFITS SOMEONE ELSE.
SO FROM THAT PERSPECTIVE I'LL
START OUT BY SAYING I HAVEN'T
KNOWN STEVE AS LONG AS MANY
OTHER SPEAKERS.
I WAS RECRUITED TO FREDERICK IN
199 5, I CAME TO UNDERSTAND THE
SIGNIFICANCE OF THE SPREAD-HANDS
GESTURE.
[LAUGHTER]
BUT WHEN I FIRST CAME, AS A
NATIVE CALIFORNIAN, MY MODE OF
TRANSPORTATION WAS LESS DRAMATIC
THAN DOROTHY'S TRIP OVER THE
RAINBOW BUT WHEN I GOT TO
FREDERICK IT WAS SORT OF
EQUIVALENTLY STRANGE AND
DIFFERENT WORLD RELATIVE TO WHAT
I WAS USED TO AS A LONGTIME BAY
AREA RESIDENT, NEVER LIVED ON
THE EAST COAST, IF YOU ASKED ME
BEFORE I CAME HERE WILL YOU
SPEND THE NEXT 25 YEARS OF YOUR
CAREER WORKING ON AN ARMY BASE,
I WOULD HAVE LOOKED LIKE YOU
SHOULD BE COMMITTED.
BUT NEVERTHELESS THAT WAS THE
SITUATION.
WHAT WAS REMARKABLE IS THAT
SOMEONE WHO STARTED IN SCIENCE
AS A CELLULAR IMMUNOLOGIST, SORT
OF GOT INTO RETROVIROLOGY
THROUGH THE BACK DOOR BECAUSE OF
HIV AND AIDS, I REALIZED THAT I
HAD LANDED DOWN NOT ON TOP OF
THE WISHED WITCH OF THE WEST BUT
THE BIGGEST GROUP OF
RETROVIROLOGYISTS ANY, A GREAT
OPPORTUNITY BUT INTIMIDATING
BECAUSE THESE WERE THE PEOPLE
THAT WE'VE SEEN LITERALLY WROTE
AND EDITED THE BOOK ON THE
SUBJECT.
I WAS ENTHUSIASTIC ABOUT TRYING
TO FIND OPPORTUNITIES TO WORK
WITH SOME OF THESE FOLKS, IN
SPITE OF NOT HAVING
COLLABORATIVE PROJECTS FOR A
LONG TIME, IT WAS A GREAT
OPPORTUNITY TO LEARN AN AWFUL
LOT.
I THINK ONE OF THE REASONS WHY I
STRUGGLED TO FIND COLLABORATIVE
PROJECTS IS THAT MUCH OF MY WORK
AFTER I CAME TO FREDERICK
FOCUSED ON TRYING TO USE --
DEVELOP AND USE NON-HUMAN
PRIMATE MODELS FOR QUESTIONS
THAT COULDN'T AS READILY BE
APPROACHED IN THE CLINICAL
CONTEXT.
SO A LOT OF THE WORK WAS SORT OF
A LITTLE BIT OFF TO THE SIDE TO
SOME OF THE THINGS THAT STEVE
WAS INTERESTED IN.
NEVERTHELESS RETAINED A GREAT
INTEREST IN TRYING TO FIND A
PROJECT UNSUCCESSFULLY FOR MANY
YEARS, FORTUNATELY IN SPITE OF
THE LACK OF OVERLAP ON NON-HUMAN
PRIMATE, THERE WAS ANOTHER
SPECIES THAT PROVIDED
(INDISCERNIBLE) LARRY AND STEVE
SHARED PASSION FOR FISHING AND
MANAGED TO GET ME TO THE TROUT
STREAM IN PENNSYLVANIA, AND WITH
THE PASSION FOR FISHING WAS
THEIR PASSION FOR SHARING LOVE
OF FISHING.
HERE IS LARRY INSTRUCTING A
COLLEAGUE, JIM HOXY, AS A
NEOPHPYTE FLY FISHERMAN.
PARDON THE BAD PUN BUT I WAS
HOOKED.
EVEN TO THE POINT OF TRYING TO
FAMILIARIZE MYSELF WITH SORT OF
A FOREIGN AND UNAPPROACHABLE
ETHOS AND CULTURE FOR SOME PARTS
OF THE ANGLING COMMUNITY I
DIDN'T QUITE FULLY UNDERSTAND
WHERE THESE FOLKS WERE COMING
FROM BUT OVER TIME BEGAN TO GET
A FEEL FOR IT.
[LAUGHTER]
AND IT'S INTERESTING.
WE HAVE A LOT OF THINGS GOING
ON, A LOT OF DISTRACTIONS IN
LIFE, THE IDEA OF FINDING
YOURSELF IN THE BEAUTIFUL SPOT
WADING IN THE STREAM, AND ASKING
YOURSELF, NOW, AT THIS TIME OF
YEAR WITH THESE LIKE CONDITIONS
AND THIS TEMPERATURE AND THIS
WEATHER, IF I WERE A RAINBOW
TROUT WHERE IN THE POND WOULD I
BE?
THINKING ABOUT THE MINDS OF FISH
IS A WONDERFUL DIVERSION,
ESPECIALLY IN BEAUTIFUL OUTDOOR
PLACES, TO ME IT BETS THE HELL
OUT OF WHACKING A WHITE BALL
OVER PESTICIDE TREATED GRASS.
WITH STEVE'S TUTELAGE AND
SHARING OF EQUIPMENT AND
GUIDANCE, EVEN A TOTAL BEGINNER
LIKE ME ACTUALLY BEGAN TO HAVE A
LITTLE BIT OF SUCCESS, STEVE
MANAGED TO CONVEY A FEEL FOR THE
PROCESS OF FISHING AND BEING IN
THE MOMENT AND IT WASN'T
NECESSARILY ABOUT CATCHING THE
FISH, WHAT FISH YOU CAUGHT, AND
WHAT I REMEMBER IN PARTICULAR,
ONE TRIP THAT BROUGHT THAT HOME,
ONE STEVE WAS PROUD OF, THIS ONE
HE MANAGED TO LAND.
BUT STEVE ALSO CONVEYED TO ME
THERE'S NOTHING WRONG WITH
LANDING A BIG BEAUTIFUL FISH
EITHER.
THAT HAS BEEN A BOND.
THEY CONTINUED TO INSTRUCT ME.
I WAS A TOTAL NEWBIE, AS YOU CAN
OBSERVE FROM THIS PICTURE.
AND THEY SOON INSTRUCTED ME IN
THE LARRY ARTHUR TECHNIQUE OF
PRESENTING THE FISH TO THE
CAMERAMAN WITH THE ARMS FULLY
EXTENDED TO EXAGGERATE THE
EXPERIENCE.
I'VE GOT THAT DOWN NOW TOO.
I DIDN'T REALLY DEVELOP A
COLLABORATIVE PROJECT WITH STEVE
FOR THE FIRST TIME BUT MY FIRST
PROFESSIONAL INTERACTION WITH
STEVE WAS REALLY WITH REGARD TO
OUR SITE VISIT.
YOU CAN'T IMAGINE COMING FROM
THE OUTSIDE, IT'S TAKEN
SERIOUSLY BUT FOR GRANTED, BUT
UNFAMILIAR IF YOU'RE NOT
ACCUSTOMED.
JEFF ALERTED TO STEVE'S SURVIVAL
GUIDE FOR SITE VISITS,
INVALUABLE PIECE OF GUIDANCE AND
DOCUMENT.
WHAT WAS AMAZING TO ME WAS
STEVE'S WILLINGNESS TO SIT IN ON
OUR REHEARSAL SESSIONS AND
PROVIDE INCREDIBLE CONSTRUCTIVE
FEEDBACK IN HIS DIRECT,
SOMETIMES US HUMOROUS BUT COGENT
WAY, TO EXPRESS GRATIFICATION
WHEN YOU TOOK HIS GOOD ADVICE
AND THAT WORKED OUT WELL FOR
YOU.
JUST THE LAST FEW YEARS, MANAGED
TO FIND A COLLABORATIVE PROJECT
TO WORK ON WITH STEVE, AND I'M
NOT GOING TO BELABOR THE
BACKGROUND INFORMATION ABOUT
WORK A CLONAL PROLIFERATION IN
RELATION TO PERSISTENCE OF AID
VIRUSES SINCE IT WAS WELL
COVERED BY OTHER SPEAKERS THIS
AFTERNOON, BUT GOING FROM THIS
LANDMARK PAPER FROM STEVE AND
COLLEAGUES, DRP AND SEVERAL
OTHER CONCOMITANT PUBLICATIONS,
THE QUESTION ARISES WHAT IS
MECHANISM, WHAT IS IT DRIVING
THE PERSISTENCE IN CLONAL
PROLIFERATION OF CELLS HAVING
IDENTICAL PROVIRAL INTEGRATION
SITES?
AND IT BECAME MUTUAL INTEREST
FOR STEVE AND FOR ME, WHETHER OR
NOT SOME OF THE WORK THAT WE'VE
BEEN DOING IN DEVELOPING
NON-HUMAN PRIMATE MODELS MIGHT
BE APPLICABLE TO ADDRESSING SOME
RELEVANT QUESTIONS.
NOW, THEY HAVE ADVANTAGES IN
TERMS OF EXPERIMENTAL CONTROL.
YOU CAN CONTROL IDENTITY OF THE
INOCULATING VIRUS, EXACT TIMING
OF THE INOCULATION, ROUTE AND
DOSE OF INFECTION.
AND IN DEVELOPMENT FROM A
PROGRAM DEVELOPED BARCODED
VIRUSES TO TRACK DISTINCT
BARCODE CLONAL TYPES ON ISOGENIC
BACKGROUNDS IN INFECTED HOST.
YOU CAN CONTROL TIMING OF
INITIATION, DURATION,
INTERACTION.
ONE THING THAT WOULD BE
INTERESTING TO UTILIZE
IMMUNIZATION WITH EXOGENOUS
ANTIGEN, TO EXPLORE THIS
QUESTION IN MACAQUES.
WE DID MORE TISSUE SAMPLING
INCLUDING UP TO ELECTIVE
NECROPSY, BUT THERE ARE
LIMITATIONS.
THE RHESUS MACAQUE SEQUENCE AND
PATIENT ARE NOT AS WELL
DEVELOPED FOR HUMANS, TISSUE
SAMPLING IS AVAILABLE BUT
MACAQUES HAVE LIMITED BLOOD
LINE, WE OVERCAME BY DEVELOPED
PROCEDURES FOR LEUKAPHERESIS IN
THESE ANIMALS.
LET ME ASK A QUESTION, CAN WE
USE THIS MODEL TO FIRST ASSESS
SIV INTEGRATION BIOLOGY,
CAPITULATE FEATURES OF HIV IN
VITRO AND IN VIVO.
THE SECRET, I TOLD YOU THERE ARE
MANY ADVANTAGES TO WORKING WITH
NON-HUMAN PRIMATE PRODUCT ALSO,
THROUGH BITTER EXPERIENCE OVER
MANY YEARS WE'VE LEARNED THAT
BASED ON REAGENTS AND
INFORMATION SORT OF HIERARCHY OF
DIFFICULTY OF EXPERIMENTS FROM
MOUSE TO HUMAN TO MACAQUES,
SEEMS TO BE HARDER TO DO THINGS
IN NON-HUMAN PRIMATE MODELS BUT
IN THE RIGHT CONTEXT CAN BE WELL
WORTH THE EFFORT.
WITH STEVE'S APPROACH FOR RIGOR
WE SET OUT TO SEE, LET'S ASSIST
SIMILARITY OR DIFFERENCES IN
INTEGRATION PROCESS FOR HIV AND
SIV IN MACAQUE AND HUMAN PBMCs
USING VARIATION ON INTEGRATE
SITE ANALYSIS YOU HEARD ABOUT
EARLIER THROUGH THE PROGRAM.
AND INITIALLY WE CONDUCTED
COMPARATIVE SITE ANALYSIS FOR IN
VITRO INFECTION OF HIV VERSUS
SIV IN PRIMARY HUMAN CELLS CELL
TO VIRUSES, EXPLORING THE ROLE
OF THE VIRUS IN CONTRIBUTING
DIFFERENCES, AND COMPARED
INTEGRATION SITE ANALYSIS OF SIV
IN PRIMARY HUMAN PBMCS AND
MACAQUE PBMC TO EXPLORE HOST
CELLS, HIV AND MACAQUE CELLS WAS
A NON-STARTER, IN
SPECIES-SPECIFIC RESTRICTION
FACTORS.
IN ANY EVENT THAT WORK WENT
REASONABLY WELL, WE PUBLISHED A
PAPER TOGETHER, MY FIRST AND SO
FAR ONLY ONE WITH STEVE.
GRATIFYING PROJECT NONETHELESS.
A I LONG THE WAY I GOT EXPOSURE
TO NOT ONLY LEARNING NEW
TECHNIQUES BUT IN THE PROCESS
GETTING A FEEL FOR HOW STEVE
APPROACHES SCIENCE AND THINKS
ABOUT THINGS AND TELLINGLY THE
SENSE OF RIGOR HE BROUGHT TO THE
PROJECT, PARTICULARLY IMPRESSIVE
FOR ME AS I WAS GETTING EXPOSURE
AND LEARNING ENTIRELY NEW FIELD
IN TERMS OF SOME OF THE ASPECTS
OF THE INTEGRATION SITE
ANALYSIS, THE BIOLOGY AND
INFORMATICS OF IT.
AND THIS QUOTE SAYS IT ALL.
THE LEVEL OF RIGOR HE BROUGHT IN
GOING THROUGH THE ANALYSIS,
WHICH AGAIN STEVE SHARED WITH ME
THE CONSIDERATIONS AND MAKING ME
REALIZE, OH, I HADN'T THOUGHT
ABOUT THAT.
WE NEED TO TAKE THAT INTO
CONSIDERATION HOW WE LOOK AT
INTEGRATION SITES, BUT YOU'RE
ABSOLUTELY RIGHT.
REPEATEDLY GOING THROUGH THINGS,
WITH ALL THAT SAID, APPLYING
RIGOROUS THOUGHTS, WE'RE ABLE TO
SHOW BROAD SIMILARITIES BETWEEN
SIV AND HIV INTEGRATION AND
HUMAN AND MACAQUE CELLS BY
LOOKING AT SITES OF INTEGRATION,
AND WHEEL THERE WERE MODEST
DIFFERENCES IN BOTH THE VIRUS
AND THE HOST CELL SEEMED TO
CONTRIBUTE TO DIFFERENCES WHEN
WE DID APPROPRIATE ANALYSES, ON
BALANCE THINGS TO BE QUITE
SIMILAR.
AND THIS FIGURE FROM THE PAPER
SUMMARIZED THE MAPPING OF
INTEGRATION SITES, HIV INTO
HUMAN PBMCs, SIV INTEGRATION,
HIV INTO RESEARCH, YOU CAN SEE
CONCORDANCE BETWEEN INTEGRATION
PLOT AS YOU LOOK AT THE CIRCOS
PLOTTING ON THE HUMAN GENOME
ALTERNATIVE MAPPING TO HUMAN OR
RHESUS MACAQUE GENOME SYNTHETIC
LESIONS ON THE ALTERNATIVE
CIRCOS PLOT.
ESTABLISHING IN VITRO ANALYSIS
SUPPORTED GENERAL CONTENTION
SYSTEMS WERE SIMILAR ENOUGH TO
POTENTIALLY BE INFORMATIVE WAS
OUR INITIAL IN VIVO ANALYSIS
LOOKING AT GROUP OF ANIMALS THAT
WERE INFECTED, STARTING ON ART
AT 4 WEEKS INTO INFECTION, AND
MAINTAINED FOR ABOUT A YEAR
BEFORE ANALYSIS, A LITTLE OVER A
YEAR.
AND WHAT WE OBSERVED WAS THAT
EVEN IN SOME PRE-ART SAMPLES
WERE ABLE TO IDENTIFY AS EARLY
AS 2 OR 4 WEEKS POST-INFECTION
EVIDENCE OF CLONALLY EXPANDED
CELLS, SOME PERSISTENT INTO A
YEAR ON ART.
AND THIS WAS CONSISTENT WITH
SOME OF THE DATA THAT HAS BEEN
ALLUDED TO EARLIER, ON
HIV-INFECTED PEOPLE STARTED ON
EARLY ART.
WE WERE ABLE TO IDENTIFY UPWARDS
OF 100 CLONES SEEN AFTER A YEAR
ON ART, 20% OF THE INTEGRATION
SITES AFTER A YEAR ON ART WERE
FROM EXTENDED CLONES INCLUDING
CLONES WE SHOWED IN MULTIPLE
TISSUES.
TO BE CLEAR, THIS IS NOT A MODEL
FOR LOOKING AT EXTENDED CLONES
AND INTEGRATION SITES AND SO ON,
SUBJECT OF THE MULTIPLE
SELECTIVE PROCESSES THAT
OCCURRED OVER DECADES OF ART BUT
IT IS INTERESTING THAT A LOT OF
BASIC BIOLOGY THAT LEADS TO
THOSE THINGS IS ESTABLISHED AND
READILY DEMONSTRABLE IN THE
FIRST YEAR.
WE DID SEE ALSO INTEGRATIONS IN
SOME OF THESE OVERREPRESENTED
SITES, ASSOCIATED WITH POTENTIAL
INCREASED SURVIVAL OF CELLS
HARBORING INTEGRATION, JUST TO
SHOW THAT MOTHER NATURE HAS A
SENSE OF HUMOR, WHEN WE -- AT
THE TIME WHEN YOU'RE FIRST
ANALYZING DATA, WE WERE THINKING
THAT INTEGRATION SITE MIGHT PLAY
MORE COMMON OR FREQUENT ROLE
THAN WE CURRENTLY UNDERSTAND,
THE VERY FIRST CLONE WE
IDENTIFIED WAS ACTUALLY A
BLASTOMA LOCUS, INTERESTING TO
SAY THE LEAST.
ON ART WE DID EVEN JUST WITHIN
THE YEAR SEE EVIDENCE OF
SELECTION FOR INTEGRATION
OPPOSITE ORIENTATION OF
TRANSCRIPTION AS SEEN WITH
HIV-INFECTED PEOPLE AND OVERALL
CLONAL EXPANSION IN MACAQUES WAS
SIMILAR.
THIS MIGHT BE A USEFUL MODEL WE
COULD TRY TO USE TO STUDY
BIOLOGY POTENTIALLY PASS
INTERVENTIONS AND IN PARTICULAR
SETTING OUT TO TRY AND
UNDERSTAND ROLE OF
ANTIGEN-DRIVEN THROUGH
IMMUNIZATION PROLIFERATION OR
CYTOKINE DRIVEN PROLIFERATION,
TRYING TO TEST WHO LEADING
HYPOTHESES OF ANTIGEN DRIVEN
PROLIFERATION AND CYTOKINE
DRIVEN PROLIFERATION.
A LOT OF CHALLENGES, WE DID A
LOT OF CALCULATIONS TO DECIDE IF
WE WERE SPITTING INTO A GALE
FORCE WIND, DIFFICULT BECAUSE
OBTAINING SUFFICIENT IN
VIVO-DERIVED INTEGRATION SITES
TO ANALYZE FROM ANTIGEN-SPECIFIC
CELLS IN MACAQUES WAS GOING TO
BE DIFFICULT BECAUSE CURRENT
PROCESSES ARE ONLY ABOUT 10%
DEFICIENT, WE NEED TO SEE --
NEEDS TO BE AT LEAST 10 TO THE
6th CELLS TO IDENTIFY EXPANDED
CLONE, ONLY 100 TO 1,000 CELLS
ARE INFECTED.
THE SIGNS OF CD4 IN RESPONSES IS
OFTEN LOWER THAN CD8 T-CELL
RESPONSES AND SO THAT WAS A
LIMITATION.
AND INFECTED CELLS CAN BE
ELIMINATED EITHER THROUGH VIRAL
PROCESSES, MEDIATED CLEARANCE,
AND/OR ANTIGEN INDUCED CELL
DEATH PROCESSES.
ALSO WHILE WE HAD ADVANTAGES FOR
SAMPLING WERE THERE WERE
LIMITATIONS, AND FINALLY NEEDED
TO DEVELOP APPROACH FOR
SEPARATING IMMUNIZING ANTIGEN
SPECIFIC VERSUS OTHER CD4 CELLS
FOR INTEGRATION ANALYSIS.
I WANT TO CALL OUT OUR GROUP FOR
DEVELOPING THE APPROACH FOR
DOING THAT, INITIALLY SCREENING
MANY ANTIGENS, FINALLY
IDENTIFYING A HUMAN B CELL LINE
PRODUCING THE MOST ROBUST CD4
RESPONSES UP TO 3 TO 4% OF TOTAL
CD4 CELLS.
DEVELOPED A SCHEME FOR PRIME
BOOST IMMUNIZATION PRIOR TO
INFECTION, REBOOST AFTER 6
MONTHS ON ART, FOUR WEEKS POST
INFECTION, WE KNEW SOME CLONES
HAD BEEN ESTABLISHED BY THAT
TIME.
INFECTED ANIMALS WITH THE
BARCODED VIRUS, STARTED ON 4
WEEK SAMPLED BEFORE EACH
INTERVENTION, DEVELOPED AND
APPLIED PROCESS LOOKING AT BOTH
BLOOD CELLS AND LYMPH NODE CELLS
TO ENRICH FOR CD4 POSITIVE CELLS
USING A SHORT-TERM ANTIGEN
STIMULATION TO IDENTIFY CELLS
THAT SHOWED SPECIFIC RECOGNITION
OF THAT ANTIGEN BASED ON
INDUCTION OF ACTIVATION MARKER
BY FLOW CYTOMETRY, USING
INTEGRATION SITE ANALYSIS
APPROACH TO ASK A QUESTION, IS
THERE EVIDENCE FOR INCREASED
RATIO OF CLONAL INTEGRATION
SITES RELATIVE TO TOTAL
INTEGRATION SITES IN ANTIGEN
REACTIVE CD4 CELLS, FROM THE
IMMUNIZING ANTIGEN VERSUS TOTAL
CD4s, SORT OF A FALSE SETUP
BECAUSE IT'S BEEN ALLUDED TO ALL
REPRESENT CLONES BUT IN THE
LIMIT OF SENSITIVITY IS
OVERREPRESENTING, WE'RE TRYING
TO LOOK AT FOR VISUAL LEARNERS
SUMMARIZING THE SCHEME BEFORE
INFECTION, IMMUNIZING TWICE, AND
THEN INFECTING THE ANIMALS
STARTING THEM ON ART AT 4 WEEKS,
AND BOOSTING AGAIN AT 6 MONTHS
ON ART, WITH LEUKAPHERESIS,
BECAUSE OF COVID-RELATED DELAYS
HAS GONE ON LONGER, LOOKING AT
SITE ANALYSIS INCLUDING AFTER
IL-15 ADMINISTRATION.
REALLY WHEN I FOUND OUT ABOUT
THIS SYMPOSIUM I WAS HOPING WE
WOULD HAVE THIS PROJECT FURTHER
ALONG, THIS IS ONE OF THE
PROJECTS WHERE NOTHING HAS GONE
AS QUICKLY AS WE WERE HOPING IT
WOULD.
SO I DON'T HAVE THE DATA TO
PRESENT TODAY, WHAT I MOSTLY
HAVE IS JUST TEASER DATA, A LOT
OF HARD WORK IN OPTIMIZING
PROCEDURES TRYING TO MAKE THEM
RIGOROUS TO STANDARDS THAT STEVE
WOULD BE COMFORTABLE WITH ON
BOTH WET LAB SIDE OF THINGS AND
INFORMATICS SIDE AGAIN WITH A
LOT OF ADAPTATIONS OF THINGS
WELL WORKED OUT FOR HIV BUT
ADAPTING THEM TO MILIEU OF SIV
INFECTION IN MACAQUES.
ANALYSES ARE IN PROGRESS.
AND JUST AS A TEASER, WE DO HAVE
A PRELIMINARY SUGGESTS OF
INCREASED FREQUENCY, INCREASED
PROPORTION OF CLONAL INTEGRATION
SITES TO TOTAL INTEGRATION SITES
AND ALSO A LARGER SIZE OF THE
CLONES IN TERMS OF THE NUMBER OF
DIFFERENT REPRESENTATIVES FOR
IMMUNIZING ANTIGEN, LIMITED
NUMBER OF SPECIMENS WE'VE BEEN
ABLE TO ANALYZE TO DATE, AND
WE'RE WORKING ON SOME OF THE
COMPLETED ANALYSIS ALSO LOOKING
FORWARD TO LONGITUDINAL WITHIN
ANIMAL AND ALSO COMPARISONS,
JUST A BRIEF SUMMARY TO GIVE YOU
A FEEL FOR THE KINDS OF DATA
THAT WE'RE GENERATING, ON THE
LEFT SIDE CIRCOS PLOT, WITH
REASONS IN RED, ORANGE, UNIQUE
WITHIN THE ANALYSIS, PRE AND
POST THIRD BOOSTER, WE GOT A TON
OF DATA AND A LOT MORE TO
GENERATION BUT WE'RE EXCITED
THAT AFTER A LOT OF HARD WORK WE
THINK WE MAY HAVE AN INFORMATIVE
EXPERIMENT HERE TO ADDRESS THIS.
SO NEXT STAGE OF THINGS IS TO
COMPLETE THE IN VIVO PHASE OF
THE EXPERIMENT, THESE ARE VERY
LABOR INTENSIVE EXPERIMENTS ON
THE IN VIVO PART, AND SO THEY
ARE STAGGERED IN PAIRS OF
ANIMALS BECAUSE THAT'S ALL THE
ANIMAL FACILITY CAN HANDLE AT
ANY ONE TIME BUT IT'S GOING
WELL, COMPLETING ALL THE
INTEGRATION SITE ANALYSIS, AND
WE'RE IN THE PROCESS OF TRYING
TO ADAPT TECHNIQUES TO DO NOT
JUST INTEGRATE CELL ANALYSIS BUT
FOR BARCODE ANALYSIS AND
ASPIRATIONALLY HOPE TO DO SOME
OF THE FANCIER THINGS PEOPLE
LIKE OTHERS HAVE DONE, SINGLE
CELL RNA SEQUENCING AS ALLUDED
TO IN A PROFESSIONAL QUESTION TO
GET A FEEL FOR TRANSCRIPTOMICS,
TCR SEQUENCING ON SOME OF THESE
EXPANDED CLONES, LOOKING AT THE
SECOND HYPOTHESIS, THE ROAM OF
CYTOKINES, HOMEOSTATIC CYTOKINES
CONTRIBUTING TO CLONAL
PERSISTENCE OR EXPANSION, THIS
IS IMPORTANT BECAUSE SOME OF THE
WORK THAT WE'RE DOING AND MANY
IN THE FIELD ARE DOING TO LOOK
AT APPROACHES FOR REDUCING OR
ELIMINATING BIORESERVOIRS,
INCORPORATING CYTOKINES AS PART
OF IMMUNOTHERAPY APPROACH,
IMPORTANT TO UNDERSTAND IMPACT.
FINALLY IN ANOTHER DELAYED
PROJECT, SEEMS IRONIC IT'S
TAKING OFF AS STEVE IS
OFFICIALLY DEPARTING BUT LIKE
OTHERS I HOPE IT'S MORE
FIGURATIVE THAN LITERAL, WORKING
WITH HARRY BURKE AND COLLEAGUES
TO EVALUATE INHIBITOR THAT IS
ACTIVE AGAINST RESISTANT
VARIANTS THAT DMITRY PRESENTED
LOOKING AT OPTIMIZATION OF
FORMULATION AND
PHARMACOKINETICS, HOPING TO MOVE
TO EVALUATE ACTIVITY IN VIVO AND
NON-HUMAN PRIMATE.
I WANT TO ACKNOWLEDGE SOME FOLKS
THAT MADE KEY CONTRIBUTIONS TO
THE WORKS THAT I PRESENTED, YOU
PLAYED A KEY ROLE IN DEVELOPING
PROCEDURES FOR PROCESSING THE
ORIGINAL SPECIMEN FROM ANIMALS,
DEVELOPING TECHNIQUES FOR
ISOLATING CELLS, ADRIAN
SWANSTROM WHO HAS WRITTEN ON
COMPLICATED EXPERIMENTS, MAKING
SURE EVERYTHING GOT DONE AT THE
TIME IT WAS SUPPOSED TO,
QUANTITATIVE MOLECULAR
DIAGNOSTIC CORES THAT DID
MONITORING, NON-HUMAN PRIMATE
SUPPORT CORE FOLKS WHO PROCESSED
ANIMALS INCLUDING GEORGE AT THE
BETHESDA NON-HUMAN PRIMATE
FACILITY.
VAN AND CHRISTINE RESPONSIBLE
FOR OVERSEEING THE SEQUENCING
INVOLVED IN INTEGRATE SITE
ANALYSIS, CHARLIE GOODMAN FOR
INFORMATICS AND OPTIMIZATION OF
THE PIPELINE FOR LOOKING AT THIS
ALONG WITH HELPERS OF
STATISTICAL ANALYSIS TO MAKE
SURE WE CAN BELIEVE THE THINGS
WE'RE SEEING.
CELLULAR IMMUNOLOGY CORE HELPED
WITH FLOW CYTOMETRY SORTING, THE
CHEMISTRY CORE WORKED WITH
BIOLOGICAL PROTEIN PRODUCTS
CORE, AND GEORGE ON PREPARING,
ISOLATING, PURIFYING AND
CHARACTERIZING L15 WE'RE USING
AND VETERINARY STAFF IN BETHESDA
FACILITY, CRITICAL TO THESE
STUDIES, WITHIN THE DRP ANDREA
MADE KEY CONTRIBUTIONS TO THE
FIRST PUBLICATION, JOHN AND
STEVE HAVE BEEN INVALUABLE AND A
PLEASURE TO WORK WITH,
SUPPLEMENTED WITH WORK FROM SOME
COLLEAGUES AND DRUGS FOR
SUPPRESSION OF VIRUS IN THE
ANIMALS, I'D LIKE TO RECOGNIZE
OUR COLLEAGUES AT GILEAD.
I'LL STOP THERE SO IT'S BEEN AN
ABSOLUTE PLEASURE TO GET A
CHANCE TO FINALLY AFTER ALL
THESE YEARS WORK TOGETHER ON A
PROJECT, I'M REGRETFUL THAT, YOU
KNOW, THINGS ARE FINALLY
STARTING TO HEAT UP AND GET
EXCITING AT THE TIME THAT YOU'RE
AT LEAST PARTIALLY DEPARTING BUT
AT THE SAME TIME I WANT TO
CONCLUDE BY SAYING THANK YOU FOR
EVERYTHING.
IT'S BEEN A JOY TO HAVE A CHANCE
TO WORK WITH YOU AND GET TO KNOW
YOU.
I WISH YOU MUCH HAPPINESS AND
FULFILLMENT IN RETIREMENT AND
HOPE I CONTINUE TO CONSULT YOU
IN SCIENCE AND TRYING TO FIGURE
OUT THE MINDS OF FISH.
THANK YOU, STEVE.
[APPLAUSE]
>>THANKS VERY MUCH.
QUESTIONS?
>>THIS IS A QUESTION ABOUT HOW
LONG WE HAVE TO WAIT TO
(INDISCERNIBLE) AND WONDERING
IF -- YOU MENTIONED IN A RATIO
OF INTEGRANT (INDISCERNIBLE) TO
CHANGE OVER TIME.
AFTER A YEAR WHERE ARE YOU ON
THE RATIO RELATIVE TO
(INDISCERNIBLE)?
>>GOOD QUESTION.
I CAN'T ANSWER OFF THE CUFF.
BUT I WILL DREDGE UP NUMBERS.
WE CAN LOOK AT ONE YEAR, AND
COMPARE THAT TO SOME OF THE HIV
LITERATURE I USE AT VARIOUS
TIMES AND TRY AND GET BACK TO
YOU.
>>THANKS.
[APPLAUSE]
>>WE'RE JOINED BY NITA MAIHLE,
ON SIGNAL TRANSDUCTION, ONE OF
THE FIRST POSTDOCS, WILL KNOW
DOUBT SHARE WITH US RECOLLECTION
OF THE EARLY DAYS OF STEVE'S
CAREER.
>>A PLEASURE TO BE HERE WITH
STEVE TO HEAR HIM ROASTED AND
ADMIRED BY SO MANY.
WE WOULD BE REMISS IF WE DIDN'T
THINK ABOUT THE LARGE NUMBER OF
TRAINEES INCLUDING MYSELF STEVE
HAD THROUGH THE YEARS.
WE'VE HEARD OF HIS PROFESSIONAL
ATTRIBUTES INCLUDING KEEN
INTELLECT, AMBITION, CURIOSITY,
BOUNDLESS ENERGY AND GENEROUS
SPIRIT, THERE ARE AT LEAST THREE
I CAN THINK OF THAT CONTRIBUTED
A LOT TO OUR FIELD.
BIOMEDICAL RESEARCH AND THE
LIVES OF MANY OF THE FOLKS HE'S
MENTORED INCLUDING MYSELF.
I WAS STEVE'S THIRD FELLOW,
FIRST WAS FOUNDER OF STRATEGY,
THIRD WAS WELL KNOWN WATCHER
BIOLOGIST OF THE ACTIN MYOSIN
FAMILY, AND MYSELFA CANCER
BIOLOGIEST AT THE MAYO CLINIC
FOR 15 YEARS, AND YALE SCHOOL OF
MEDICINE FOR TEN YEARS, AND MORE
RECENTLY I'VE BEEN WORKING A
COUPLE SMALLER MEDICAL SCHOOLS
IN THE SOUTH HELPING THEM BUILD
RESEARCH AND EDUCATION PROGRAMS,
STILL ACTIVELY SERVE AS DEAN FOR
VIRTUAL ACADEMY FOR OVARIAN
CANCER.
COMING BACK TO STEVE'S
ATTRIBUTES, I THINK ONE OF THE
THINGS FOLKS DON'T OFTEN
R EALIZE, HE'S BEEN A TREMENDOUS
CHAMPION FOR WOMEN IN SCIENCE
AND WOMEN IN MEDICINE INCLUDING
MY OWN CAREER.
HE'S BEEN A STALWART ADVOCATE.
I RANDOMLY SPOKE WITH A WOMAN
WHO SAT DOWN AT LUNCH TODAY, LIZ
FROM UTAH, FIRST THING SHE SAID,
STEVE BEEN A CHEERLEADER FOR ME
AND MY WORK.
NO SURPRISE TO ME AT ALL.
AND I THINK IT'S VERY IMPORTANT
THAT FOLKS OF STEVE'S CALIBER
HAVE BEEN ABLE TO MOVE THE
NEEDLE IN THIS REGARD, WE HEAR A
LOT OF RHETORIC ABOUT THAT
TOPIC, PARTICULARLY TRENDY RIGHT
NOW, BUT THESE ARE VALUES STEVE
HAD FROM EARLY DAYS OF HIS
CAREER AND HE'S ACTED ON THEM
ACCORDINGLY.
SECOND, AGENDA RHETORIC IN
ACADEMIC MEDICAL CENTERS ABOUT
INTEGRITY AND CHARACTER RUNS
DEEP BUT WE HAVE VERY FEW ROLE
MODELS WHO HAVE STRONG PERSONAL
VALUES, PROFESSIONAL VALUES,
THAT THEY LIVE BY EVERY DAY.
IT WAS MY BISHOP EARLIER WHO
SPOKE ABOUT STEVE HAVING A
PENCHANT FOR SPEAKING TRUTH TO
POWER, AND CERTAINLY EVERYONE
WHO KNOWS HIM KNOWS THAT THAT IS
A PENCHANT THAT HE HAS.
HE'S REALLY SERVED AS TREMENDOUS
ROLE MODEL FOR ALL OF US IN
LIVING BY THOSE STANDARDS, AND
FINALLY THE LAST COMMENT IN
TERMS OF PERSONAL ATTRIBUTES,
STEVE IS EXCEPTIONAL IN MAKING
TIME TO BUILD RELATIONSHIP WITH
ANYONE HE CONSIDERS IMPORTANT
AND HE CONSIDERS EVERYONE HERE
IMPORTANT AND LOVES PEOPLE.
AS HIS MASTERY AT DOING THAT
WHEN I WAS A POSTDOC, I HELPED
HIM MOVE HIS LAB IN A U-HAUL
TRUCK FROM COLD SPRING HARBOR TO
THIS CAMPUS IN 1984.
AND ELAINE AND I USED TO JOKE
ABOUT STEVE ALWAYS BEING ON THE
THRONE, WE -- PHONE, AND WE
WOULD LIKE TO BE ON THE OTHER
END OF THE PHONE FOR FIVE
MINUTES BUT NOW HAVING BEEN ON
THE OTHER END OF THE PHONE OR
E-MAIL OR WHATEVER FOR MANY
YEARS I CAN TELL YOU ONE OF THE
THINGS ABOUT HIM, THE IMPORTANCE
THAT HE'S PUT IN OUR
RELATIONSHIP AND HIS
RELATIONSHIP WITH OTHERS, I
THINK IT'S A REALLY IMPORTANT
PERSONAL ATTRIBUTE TO HAVE,
SERVES AS A ROLE MODEL FOR US
ALL, DESERVINGS A ROUND OF
APPLAUSE FOR NOT ONLY THESE
ATTRIBUTES BUT MANY
CONTRIBUTIONS HE'S MADE IN OUR
CAREERS AND SIMILAR THINGS AS
WELL, STEVE.
[APPLAUSE]
>>THANK YOU.
EXTREMELY WELL SAID.
THIS BRINGS US ALMOST TO THE END
OF THE VERY REMARKABLE DAY.
I'D LIKE TO THANK ALL OF THE
SPEAKERS FOR TALKS BOTH EXCITING
AND ENTERTAINING.
I'D LIKE TO THANK YOU FOR
ATTENDING.
ALSO WOULD LIKE TO THANK THE
ONLINE AUDIENCE, WHICH HAS BEEN
QUITE LARGE, 200 PEOPLE, AND I
FEEL BAD THEY WERE NEGLECTED IN
TERMS OF CONTRIBUTING QUESTIONS
AND DISCUSSION BUT WE JUST HAD
TO BE ON A TIGHT SCHEDULE.
ALL THE TALKS TODAY HAVE BEEN
RECORDED.
THEY WILL BE AVAILABLE WITHIN
ABOUT A WEEK, I THINK ALL THE
REGISTERED PARTICIPANTS WILL
RECEIVE AN E-MAIL WITH A LINK TO
THAT.
IF YOU CAN'T FIND THAT OR DON'T
GET THAT, YOU KNOW ME OR TERRY
BURDETTE.
I'D LIKE TO THANK TERRY AND VAL
TURNQUIST FOR THE HARD WORK
PUTTING LOGISTICS TOGETHER, A
LOT OF THINGS CHANGED SINCE
COVID INCLUDING HOW YOU GET ON
THE BASE, HOW WE RUN THINGS HERE
IN THIS NEW HYBRID FORMAT, SO A
SHOUT OUT TO THEM.
I HOPE YOU CAN JOIN US THIS
EVENING FOR AN INFORMAL
GATHERING AT STEINHART'S BREWERY
IN DOWNTOWN FREDERICK, FLYERS
HAVE BEEN DISTRIBUTED TO FIND
YOUR WAY DOWN THERE.
AND I THOUGHT IT WOULD BE MOST
FITTING FOR STEVE TO HAVE THE
LAST WORD TODAY.
STEVE?
[APPLAUSE]
>>I REALLY WANT TO SAY THREE
THINGS QUICKLY.
I'M HUMBLED, HONORED, TOUCHED BY
ALL OF YOU FOR HAVING COME AND
FOR ALL THE KIND WORDS THAT HAVE
BEEN SAID.
BUT I THINK ONE OF THE THINGS
THAT I'D LIKE TO EMPHASIZE
DERIVES FROM WHAT SEVERAL
SPEAKERS SAID.
HAROLD BEGAN BY POINTING OUT WE
REALLY DON'T WORK AS
INDIVIDUALS.
YOU MAY SEE ON LATE NIGHT TV,
THE MAD SCIENTIST IS BY
THEMSELVES MAKING GREAT DISCOVE.
THAT'S COMPLETELY UNTRUE.
FOR ME TODAY I GET TO RELIVE
SOME OF THE REALLY WONDERFUL
COLLABORATIONS AND INTERACTIONS
AND HEAR FROM SOME FRIENDS AND
THINK ABOUT ALL THE THINGS THAT
WE ACCOMPLISHED, NOT AS
INDIVIDUALS BUT AS
COLLABORATORS, COLLEAGUES, I'VE
BENEFITED AS I HOPE YOU CAN
TELL, IMMENSELY, FROM HAVING
BOTH FABULOUS MENTORS AND ALSO
FROM HAVING REALLY WONDERFUL
COLLEAGUES.
AND THE THINGS YOU HEARD TODAY
NONE OF THOSE WERE THINGS I
EITHER DID OR COULD HAVE DREAMED
OF ACCOMPLISHING BY MYSELF.
THEY WERE ALL ACCOMPLISHED AS
PART OF LARGER COLLABORATIONS.
IT'S THROUGH THE STRENGTH OF
THOSE COLLABORATIONS AND THE
HELP AND HARD WORK AND INSIGHT
OF MY COLLEAGUES THAT I WAS ANY
KIND OF A SUCCESS AT ALL.
I WOULD LIKE TO SAY IN CLOSING,
THERE ARE UNFORTUNATELY A FEW
PEOPLE WHO REALLY LITERALLY
CAN'T BE WITH US, EITHER IN
PERSON OR VIRTUALLY.
AND I'M NOT GOING TO TRY AND
GIVE YOU A COMPREHENSIVE LIST
BUT WANT TO CALL OUT THREE
INDIVIDUALS I'M PARTICULARLY
SORRY ARE NOT HERE.
GEORGE OF COURSE, GEORGE WAS THE
PERSON WHO SET UP THE PROGRAM I
JOINED HERE IN 1984, BROUGHT MY
COLLEAGUE JEFF AND OUR GREAT
COLLEAGUE MARK DOWN FROM COLD
SPRING HARBOR.
GEORGE WAS A TRUE -- PEOPLE SAY
I'M LARGER THAN LIFE, GEORGE WAS
REALLY LARGER THAN LIFE IN THE
MOST WONDERFUL POSSIBLE WAY.
I MISS HIM INTENSELY TO THIS
DAY.
THE OTHER NAME FAMILIAR, I'M
PLEASED TO SEE MENTIONED BY
SEVERAL, CLARK, GRAND QUANTITIES
OF PURIFIED HIV REVERSE
TRANSCRIPTASE ENABLING ORIGINAL
STRUCTURAL WORK FROM EDDY AND
COLLEAGUES AND CHEMISTRY BY PAUL
BOYER AND OTHERS ALTHOUGH PAUL
WAS THE ONE WHO MADE IMMENSE
CONTRIBUTIONS TO THAT PROGRAM.
AND PAT, WHO WE LOST A COUPLE
YEARS AGO, IN HEARTBREAKING
FASHION.
WE DID COUPLE THINGS TOGETHER, I
MISSED DORSEY, AND WE'LL HAVE
THE DORSEY MEMORIAL LECTURE AT
SOME POINT, I'LL INVITE YOU
BACK.
VIRTUALLY, IF YOU CAN'T COME IN
PERSON, HEAR A WONDERFUL TALK IN
MEMORY OF JOE DORSEY, A VALUED
COLLEAGUE WE LOST MUCH TOO SOON.
I'LL THANK YOU ALL AND HOPE TO
SEE YOU A LITTLE LATER FOR A
BEER.