>> WELCOME TO THE D.N.A.

REPAIR INTEREST GROUP.

VIDEOCONFERENCE NOW.

WE HAVE A SPECIAL

ANNOUNCEMENT COMING UP.

MINI SYMPOSIUM ON APRIL 10th

HERE AT THE NATCHER

CONFERENCE CENTER.

WE'LL HAVE A JAPANESE

SCIENTIST.

THIS WILL GO FROM 1:00 TO

5:00 P.M. HERE ON NATCHER.

UNNOT FORTUNATELY, IT WON'T

BE VIDEO CAST BY ANYONE IN

THE AREA WILL BE WELCOME TO

COME.

DR. SUEDASAURER FROM COBY

UNIVERSITY.

DR. TANCA FROM SAKA

UNIVERSITY.

DR. COR YOKO FROM ASAKA.

DR. MAC NAMMO.

FROM THE NATIONAL INSTITUTE

HEALTH SCIENCES IN TOKYO AND

DR. MAKI FROM NARA.

ALL WILL BE SPEAKING HERE AT

THE NATCHER CONFERENCE

CENTER, AND YOU'RE ALL

WELCOME TO ATTEND, IF YOU

CAN MAKE IT TO BETHESDA.

UPCOMING VIDEOCONFERENCES IN

APRIL DR. SANTOS FROM SUNNI

WILL SPEAK ABOUT RECOGNITION.

IN MAY JAN DRAKE AT NIHS

WILL SPEAK ABOUT THE HISTORY

OF D.N.A. REPAIR AND IN JUNE

WE WILL HAVE OUR THREE YOUNG

INVESTIGATOR TALKS,

DR. ELABORATEE AT NIEHS,

DR. BOTCH EVA FROM BROOK

HAVEN AND UNIVERSITY OF

NORTH CAROLINA.

AND I APOLOGIZE IF MY

PRONOUNCE DELIBERATIONS WERE

NOT CORRECT -- PRONOUNCE

PRONUNCIATIONS WERE NOT

CORRECT.

>> TODAY WE WILL BE HAVING

TALKS FROM OHAD U.S. IN

PORTLAND, AND THE WAY WE'LL

BE RUNNING IT WILL BE A

LITTLE DIFFERENT AND WE'LL

HAVE DR. BOR EXPLAIN WHAT

WE'LL BE DOING TODAY.

>> OKAY, THANK YOU.

CAN YOU HEAR ME?

>> YES, WE CAN.

>> WELL, I WANT TO THANK

STEPHEN FOR PICKING THIS UP

AND BEING THE FIRST TO GIVE

THIS MORE COMPREHENSIVE KIND

OF PRESENTATION AND THE IDEA

BEHIND IT WAS THAT WE'VE HAD

THIS INTEREST GROUP FOR MANY

PURPOSES AND OF COURSE IN

THE FIELD AND FROM

INDIVIDUAL INVESTIGATORS BUT

ALSO TO INSTIGATE

COLLABORATIONS AND

DISCUSSIONS AT CENTERS WHERE

WE ARE NOW.

AND SO THE IDEA WAS TO ONCE

IN A WHILE HAVE A MORE

COMPREHENSIVE TALK THAT WAS

NOT NECESSARILY COVERING ONE

INVESTIGATOR BUT THE KINDS

OF THINGS THEY ARE DOING IN

OH U.S. AND TO GIVE US ALL A

FEEL FOR ONGOING ACTIVITIES

THAT WOULD ENHANCE

DISCUSSION AND COLLABORATION

BETWEEN US IN THE FUTURE.

SO WE'LL SEE HOW THIS GOES.

I'M SURE IT WILL BE VERY

WELL-DONE BY STEVE AND

THANKS, STEPHEN, FOR PICKING

IT UP AND THAT'S THE IDEA.

>> OKAY, SO WE'LL HAVE

SEVERAL PRESENTATIONS TODAY.

INITIALLY IT WILL BE TALKING

ABOUT OH U.S. AND THEN

IDENTIFICATION OF SMALL

MOLECULE INHIBITORS OF

D.N.A. KAPPA AND

DECONSTRUCTING THE PATHI.

SO PLEASE, EVERYONE BE SURE

TO MUTE YOUR MICROPHONES

EXCEPT AT OHSU AND NOW YOU

CAN TAKE IT AWAY.

>> THANKS, KIM.

LET ME KNOW IF THE VOLUME IS

TOO SKOST AND WE'LL MAKE

ADJUSTMENTS.

IS THIS RIGHT?

JUST RIGHT?

OKAY.

SO THIS IS JUST AN AERIAL

PHOTO OF THE OHSU CAMPUS AND

THAT THE BUILDING THAT WE

ARE BROADCASTING FROM IS

INDICATED BY THE AERIAL AND

WE REALLY APPRECIATE THE

OPPORTUNITY TO LET YOU KNOW

A LITTLE BIT ABOUT WORK

THAT'S GOING ON AT OHSU IN

THE AREA OF D.N.A. REPAIR

REPLICATION AND MUTE

GENESIS.

AND SPECIFICALLY FOR THE

OPPORTUNITY TO BE ABLE TO BE

THE GUINEA PIGS IN THE NEW

SYSTEM.

I THINK THAT THIS HAS A LOT

OF POTENTIAL TO BE VERY

VALUABLE.

SO THIS OPENING SLIDE -- AND

I'LL GO INTO A LITTLE BIT

MORE DETAIL FROM SOME OF THE

LABORATORIES.

WE DON'T HAVE TIME TO BE

FEATURING EVERY ONE.

WE'RE STILL TRYING TO GIVE

TWO SCIENTIFIC PRESENTATIONS.

BUT THIS WILL JUST GIVE YOU

A BIT OF AN OVERVIEW OF THE

TYPES OF OPPORTUNITIES THAT

ARE HERE.

WE COVER A VARIETY OF

SUBJECTS IN THE AREA OF

D.N.A. REPAIR.

WE HAVE RESEARCH AND BASIC

REPAIR AND NUCLEOTIDESIC

REPAIR AND WHEN I'M DONE

WITH MY PORTION OF THE

PRESENTATION, DR. MAUREEN

WILL BE PRESENTING BOTH HER

RESEARCH AS WELL AS AN

OVERVIEW IN THE FAN CONI

ANEMIA AND WE HAVE A FEW

INVESTIGATORS WHO ARE

WORKING IN THAT AREA.

ALSO IN THE AREA OF D.N.A.

REPLICATION, WON'T HAVE

TIMING INTO INTO ANY DETAIL,

BUT DR. MATTHEW IS AN

INVESTIGATOR HERE WHO WORKS

IN THE VERY INTERESTING AREA

IN TERMS OF DELAYED

REPLICATION OF CHROMOSOMES

THAT ARE ASSOCIATED WITH

CANCER CELLS AND THAT

PARTICULAR WORK IS REALLY

QUITE SIGNIFICANT IN TERMS

OF THE IMPLICATIONS ON THE

IMBALANCE IN JET STREAMIC --

GENOMIC STABILITY.

I'LL TALK MORE ABOUT MUTE

MUTEOGENESIS AND TODAY WE

JUST WON'T HAVE TIME TO GET

INTO THE D.N.A. DAMAGE OF

RESPONSE PATHWAYS AT ANY

GREAT LEVEL.

FIRST LAB I'D LIKE TO FOCUS

ON IS THE LAB OF DR. MOLLCOM,

THAT THE MAJOR FOCUS OF HER

LAB IS TO LOOK AT THE

FUNDAMENTAL MECHANISMS BY

WHICH ENVIRONMENTAL TOXIC

AND EXPOSURES AS WELL AS

INDOXNUS KPALES AND

RADIATION LEAD TO CANCER AND

OTHER TYPES OF DISEASES,

SPECIFICALLY SHE HAS THREE

FUNDED PROJECTS.

THE FIRST IS IN THE AREA OF

LOOKING AT THE FORMATION,

REPAIR AND TOLERANCE OF

D.N.A. PROTEIN PROCESS

LINKS.

AND IN THIS WORK THEY USE A

VARIETY OF REACTIVE ALDEHYDES

AND MOST PREDOMINANTLY THEY

HAVE USED FORM ALDEHYDE TO

BE INVESTIGATING THIS.

THEY USED A SERIES OF

TECHNIQUES, INCLUDING GENOMEWIDE

SCREENS.

THEY LOOK AT CELLULAR

RESPONSE PATHWAYS, AS WELL

AS THE FUNDAMENTAL

BIOCHEMICAL MECHANISMS FOR

TOLERANCE OF THESE LESIONS.

THEY ALSO WORK IN THE AREA

OF BASIC INCISION REPAIR, OF

OXIDATIVE D.N.A. DAMAGE AND

THE CELLULAR PATHWAYS THAT

RESPOND TO THAT.

AND A VERY LONG STANDING WE

HAVE IS IN THE AREA OF

INITIATION OF BASIC REPAIR

OF PHOTO PRODUCTS,

SPECIFICALLY THE DIMERS AND

CURRENT WORK IN THIS AREA IS

PREDOMINANTLY FOCUSED ON THE

DELIVERY OF THESE NEW REPAIR

SYSTEMS INTO HUMAN SKIN.

THE SECOND LAB I WILL

MENTION IS THAT OF DR. MITCH

TURKER.

HE HAS LONG STANDING

RESEARCH INTEREST IN THE

AREA OF THE MECHANISMS OF

FORMATION OF MUTATIONS AND

ALSO A BIT MORE RECENTLY BUT

REALLY NOW WELL ESTABLISHED

IN THE AREA OF HOW

ENVIRONMENTAL TOXIC AND

EXPOSURES MAY MODULATE

EPOGENIC SILENCING.

AND MITCH HAS HAD LONG

STANDING FUNDING IN THE AREA

OF THE EFFECTS OF DEEP SPACE

RADIATION ASSOCIATED WITH

DEEP SPACE TRAVEL.

THEY USE A VARIETY OF

MODELS.

THIS IS JUST ILLUSTRATING AN

ATRT MUTANT IN A KIDNEY CELL,

IN WHICH THERE HAS BEEN AN

EXPOSURE TO PROTONS IN WHICH

THERE HAS BEEN CHROMOSOME

DAMAGE AND REARRANGEMENT

LEADING TO INACTIVATION OF

APRT.

AND IN THE OTHER AREAS -- SO

THEY HAVE A NUMBER OF BOTH

MOUSE MODELS, AS WELL AS

CELL MODELS FOR THIS.

THE SECOND AREA THAT MITCH

HAS REALLY BEEN WORKING

TOWARD PINE EITHER HAS TO DO

WITH THE -- PINEERING HAS TO

DO WITH THE ENVIRONMENTAL

TOX CANTS THAT WILL BE

MODULATING EPEGENETICS AND

SPECIFICALLY THEY HAVE BEEN

ABLE TO TAKE THE BRACA ONE

PROMOTER AND THEN LINK THIS

TO HPRT AND THEN LOOK AT THE

EFFECTS OF BOTH UNDER NORM

OXIC AS WELL AS HIGH POX

YIGSIC EFFECTS IN TERMS OF

THE ACCUMULATION OF THE

RESISTANCE IN THESE AS

EVIDENCE OF MODULATION OF

EPEGENETICS SILENCING.

SO I'D LIKE TO JUST GIVE A

QUICK OVERVIEW ALSO OF MY

GROUP, IN WHICH WE HAVE

AREAS OF RESEARCH AND REPAIR

TOLERANCE AND REPLICATION IN

GENIS.

FOR QUITE SOMETIME WE'VE

WORKED ON THE AREA OF

INITIATION OF BASIC INCISION

REPAIR BY UV DAMAGE BY A

CATEGORY COSLEASES AND WE

HAVE THIS LONG STANDING

INTEREST IN D.N.A.

REPLICATION BYPASS WITH

SPECIFICALLY MODIFIED

NUKETIDES.

AND THE FOCUS IS PRIMARILY

CANCER TO A LESSER EXTENT

AGING BUT WE HAVE A NUMBER

OF STUDIES LOOKING AT HOW

DEFICIENCIES IN D.N.A.

REPAIR CAN LEAD TO METABOLIC

DISEASE.

AND THE OTHER END POINTS

THAT WE CURRENTLY DON'T HAVE

MODELS FOR, THOUGH, ARE IN

THE AREA OF HEART DISEASE AS

WELL AS NEURODEGENERATION.

SO I'D LIKE TO JUST TURN NOW

TO A FAIRLY SHORT STORY THAT

HAS BEEN DEVELOPED IN MY LAB

OVER THE LAST COUPLE OF

YEARS.

THIS WORK HAS BEEN A MAJOR

COLLABORATION BETWEEN OUR

COLLEAGUES AT VANDERBILT

UNIVERSITY, GROUPS AT THE

NIH CHEMICAL JENOMICS CENTER

AND WORK -- THAT HAS ITS

FOUNDATIONS IN WORK BY

DR. MINUTECO IN OUR GROUP AS

WELL AS KENRA KINRIN

YAMANAKA.

AND AS I'M SURE YOU'RE AWARE

THAT D.N.A. POLL KAPPA IS A

MEMBER OF THE Y-FAMILY

POLYMERASE.

IT HAS A LOW-FIDEST

REPLICATION OF NON-DAMAGE

D.N.A. BUT IN WORK BOTH OUT

OF OUR LAB AS WELL AS A

NUMBER OF OTHER LABS,

ESPECIALLY THAT OF NICK,

THAT IT'S BEEN SHOWN TO BE

CAPABLE OF BYPASSING VERY

BULKY LESIONS IN THE MINOR

GROUPS, SPECIFICALLY THE

GROUP OF QUANINE.

AND IN THIS SLIDE IS JUST

SIMPLY ILLUSTRATING THAT THE

TWO QUANINE ADDUCTS OF A

HYDRO CARBONS CAN BE

BYPASSED AND THAT THESE ARE

PRIMARILY THE ONES THAT ARE

BYPASSED ARE LOCATED IN THE

MINOR GROUP.

OUR LAB HAD ALSO LOOKED,

AGAIN W OUR COLLEAGUES AT

VANDERBILT IN TERMS OF

D.N.A. DAMAGE THAT RESULTS

FROM BY PRODUCTS LIPID

OXIDATION.

WE'VE USED FORMED SKMOID OUR

COLLEAGUES HAVE BEEN ABLE TO

MAKE D.N.A.S THAT ARE

SPECIFICALLY MODIFIED WITH

THESE AND WE'VE SHOWED A

WHILE BACK THAT THE POLL

KAPPA WAS CAPABLE OF

BYPASSING THIS.

THAT ARENA THEN EXTENDED

THESE STUDIES TO LOOK AT

LARGER AND LARGER TYPES OF

CROSS LINKS, AND IN THIS

PARTICULAR STUDY, WE BEGAN

WITH D.N.A.S THAT WERE

MODIFIED BY GAMMA

HYDROXYPROPANA D ADDUCT.

BUT THIS PARTICULAR LESION

IS IN A DINEIC EQUILIBRIUM

IN WHICH IT CAN OPEN WITH

REACTIVE HALED HYDE AND YOU

CAN THEN IN THE PRESENCE OF

VARIOUS PEPTIDES, WE HAVE

USED A SMALL PEPTIDES SUCH

AS THIS LI STRIP SLICE, AS

WELL AS LARGER PEPTIDES THAT

IN THE PRESENCE OF SODIUM

SANOBORAHIGH DROID THAT YOU

CAN THEN FORM VERY STABLE

D.N.A. PEPTIDE CROSS LINKS

WITH THESE ACROLEIN BRIDGE.

AND WHEN IRENA CARRIED OUT A

SERIES OF STUDIES LOOKING AT

THE REPLICATION OF POLL

KAPPA, THAT THIS IS A

NON-DAMAGE D.N.A. AND YOU

SEE THAT IT HAS A RELATIVELY

ROBUST PROCESSIST AND

EXTENTS TO FULL LENGTH.

BUT IF YOU NOW LOOK AT THESE

D.N.A. THAT'S CONTAIN VERY

BULKY D.N.A. PEPTIDE CROSS

LINKS, THIS IS THE LOCATION

OF WHERE THE ADDUCT WOULD

BE, AND YOU CAN SEE THAT

THERE IS ESSENTIALLY ALMOST

NO PAUSE AT THAT SITE AND

THAT YOU GET VERY EFFICIENT

REPLICATION BYPASS, WHETHER

THIS IS SIMPLY A GAMMA

HYDROXYPROPANO, WHICH IS NOT

SHOWN, OR THESE D.N.A.

PEPTIDE CROSS LINKS.

AND THE IF I DELT IS QUITE

REMARKABLE.

WHEN YOU LOOK AT SINGLE

NUCLEOTIDES -- SORRY.

WHEN YOU ARE COMING SINGLE

NUCLEOTIDES INCORPORATION

THAT ONE SEES THAT FOR THE

NON-DAMAGED, OBVIOUSLY YOU

HAVE A CORRECT INCORPORATED

BUT YOU ALSO SEE THE SAME

HIGH-FIDELITY INSERTION

OPPOSITION THESE BULKY

ADDUCTS.

SHE ALSO WAS THE PIONEER IN

THE STUDIES IN WHICH WE WERE

LOOKING AT REPLICATION

BYPASS OF INNER STRAND

D.N.A. CROSS LINKS AND THIS

AGAIN WAS WORK THAT WAS

PINEERED -- PIONEERED IN

COLLABORATION WITH OUR

COLLEAGUES AT VANDERBILT IN

WHICH WE WERE USING ACROLEIN

TO BE ABLE TO MEDIATE THE

INNER STRING CROSS LINKS OF

QUANINE, SPECIFICALLY IN THE

CPG SEQUENCE CONTEXT WE

THINK IS MOST BIOLOGICALLY

RELEVANT.

BUT IN COLLABORATION WITH

CARAMELA RISO THAT HE WAS

ABLE TO CINT THIGHS A

VARIETY OF D.N.A.S IN WHICH

THERE WOULD BE A MIMIC OF A

-- SYNTHESIZE --

>> I AM GOING TO PROBABLY

STOP USING THE POINTER.

THE ICL 1 IS A REFLECTION OF

WHAT WOULD HAVE OCCURRED

FOLLOWING AN INITIATION BY A

PORTION OF THE NUCLEOTIDES

EXCISION REPAIR IN WHICH YOU

HAVE BOTH FIVE AND THREE

PRIME OVERHANGS AND BLOCKED

BY BID OXIDES.

AND INNER STRENGTH CROSS

LINK TWO IS IMAGINING IF YOU

HAVE AN EXO CHEWING DOWN BUT

LEAVING IN TACT THE THREE

PRIME SIDES AND THEN THE ICL

3 IS A SCENARIO IN WHICH YOU

WOULD DIGEST THE PRIME

TOWARD THE CROSS LINK AND

THE IC L 4 WOULD BE

CONSIDERED PROBABLY A

MINIMAL CROSS LINK.

IT'S PROBABLY MORE LIKELY

THAT IT WOULD JUST BE TWO GS

CROSS LINKED TOGETHER.

BUT TO GET TO THE DATA, YOU

CAN AGAIN SEE THAT ARE

NON-DAMAGED D.N.A., WHETHER

THERE IS A DOWNSTREAM PRIMER

OR NOT.

THE POLL KAPPA CARRIES OUT

EFFICIENT SIN THEYSIS.

HOWEVER IN THE CASE OF THE ICL

OF ONE AND TWO, ONE SEES

THAT THERE IS A VERY

SIGNIFICANT BLOCK AND SO THE

COMMON FEATURE OF THIS IS

THAT IF YOU HAVE A

SIGNIFICANT AMOUNT OF BASE

PAIRINGS STILL DOWNSTREAM

THAT HAVE CROSS LINK KAPPA

IS VERY INEFFICIENT AT

BYPASSING.

HOWEVER, WHEN YOU NOW MAKE A

MINIMAL CROSS LINK SUCH AS

I'VE SEEN IN THE ICL 4, THAT

ONE IS THEN -- THEN SEES

THAT THE BLOCK IS

SIGNIFICANTLY REDUCED AND

THAT YOU CAN GET REPLICATION

BYPASS AND THE SINGLE

NUCLEOTIDES INCORPORATIONS

INDICATE THAT THIS IS A

HIGHLY FAITHFUL REACTION.

NOW, POLL KAPPA IS NOT ABLE

TO PANEL TO -- BE ABLE TO

BYPASS MAJOR GROUP LESIONS

AND THESE ARE COMPLEMENTARY

LESIONS.

INSTEAD OF PUTTING THESE ON

THE AMINO GROUP OF QUANNINE

THAT THESE ARE ATTACHED BY

WAY OF THE ADENNINE IN SIX.

AND SO WE'VE LOOKED AT THE

REPLICATION BYPASS OF

LESIONS IN WHICH THERE IS A

LINKAGE TO THE ADENINE AND

WHAT WE SEE IS THAT POLL

CAPA IS VERY SIGNIFICANTLY

BLOCKED BY THESE.

WE'VE LOOKED AT BOTH INNER

STRAND D.N.A. CROSS LINKS,

AS WELL AS PEPTIDE CROSS

LINKS AND THESE WERE IN

STRONG AGREEMENT WITH WORK

THAT WAS PUBLISHED QUITE

SOMETIME AGO ON POLL KAPPA

OUT OF NICK'S LAB LOOKING

THAT IF YOU NOW PLACE THE

POLY PSYCHE COLOR MEDIC

CARBONS IN A MAJOR GROUP

THAT, IT'S UNABLE TO BYPASS.

SO THE QUESTION THAT WE THEN

RAISED WAS THERE WHETHER

THERE WAS A BIOLOGICAL ROLE

FOR POLL KAPPA THAT WE MIGHT

BE ABLE TO TAKE A LOOK AT

AND SPECIFICALLY IS THERE

ANY POTENTIAL ROLE FOR POLL

KAPPA IN MEDIATING THE

CELLULAR RESPONSES TO D.N.A.

DAMAGING AGENTS AND ALSO IS

THERE ANY EVIDENCE TO

ASSOCIATE DISREGULATION OF

POLL KAPPA WITH CANCER?

AND SO IN WORK THAT WE

CARRIED OUT WITH ROB MOSES

AND SUSAN KNOLLSONE AND OUR

GROUP THAT THESE WERE

STUDIES IN WHICH THE LEVEL

OF POLL KAPPA WAS THEN

MODULATED BY SIRNAS AND WHEN

WE LOOKED TO SEE WHETHER

THAT MODULATED THE RESISTANCE

OF CELLS TO A TREATMENT WITH

C, WHICH IS GOING TO PRODUCE

MINOR GROUP CROSS LINKS THAT,

WE SEE A VERY MODEST BY

SIGNIFICANT DECREASE IN THAT

SURVIVAL.

THAT, THIS WORK ALSO WAS

EXTENDED TO LOOK AT THE

NUMBER OF RADIALS THAT ARE

BEING FORMED IN CELLS IN THE

PRESENCE OR ABSENCE OF A

KNOCKDOWN OF POLL KAPPA.

AND WHAT YOU SEE ON THE

RIGHT IS THAT FOLLOWING A

TREATMENT OF CELLS THAT HAVE

BEEN SUPPRESSED FOR POLL

KAPPA, THAT THERE IS A MAJOR

INCREASE IN THAT RESISTANCE.

AND SO WE THEN TURN TO TAKE

A LOOK IN THE LITERATURE,

WHETHER OR NOT THERE IS AN

ASSOCIATION WITH POLL KAPPA

BEING MODULATED IN TUMORS.

AND PROBABLY THE BEST

EXAMPLE OF THIS IS WORK THAT

WAS REPORTED BACK IN 2010,

IN WHICH IT WAS DEMONSTRATED

THAT BOTH AT AN MRNA AND

PROTEIN LEVELS THAT POLL

KAPPA WAS UPREGULATED IN

CLEOMAS AND IN FACT, NOW

POLL KAPPA IS SOMETIMES USED

AS A BIOMARKER FOR THE GRADE

OF LE OMAS AND YOU CAN SEE

IN THE BOTTOM PANEL THAT AS

THE GRAY NUKE LOMA IS

INCREASING THE POLL KAPPA IS

GENERALLY GOING UP.

THIS IS BIOLOGICALLY

SIGNIFICANT BECAUSE WHAT ONE

SEES IS THAT IN PATIENTS

THAT APPEAR TO BE NEGATIVE

OR NORMAL LEVELS OF D.N.A.

POLYMERASE KAPPA AT THE TIME

OF DIAGNOSIS OF NUKE OMAS

AND FOLLOWING OUT SURVIVAL

OF THOSE PATIENTS, THAT IN

CASES IN WHICH YOU HAVE A

POLL KAPPA-POSITIVE GLIOMA

THAT THE MEAN SURVIVAL

LONGEVITY IS ONLY RIGHT

AROUND TEN TO 12 MONTHS.

WHILE IT IS MUCH, MUCH

SIGNIFICANTLY LONGER IN

PATIENTS THAT ARE POLL KAPPA

KAPPA-NEGATIVE.

AND SO WE THEN ASK THE

QUESTION WHETHER OR NOT IN

THE HYPOTHESIS WHETHER SMALL

MOLECULE INHIBITORS CAN BE

USEFUL IN THE TREATMENT OF

CANCERS WITH DISREGULATED

POLL KAPPA EXPRESSION AND

WHETHER SPECIFICALLY YOU

COULD RENDER TUMOR CELLS

MORE SUSCEPTIBLE TO THE CITE

OH TOXIC EFFECTS WHEN USED

IN COMBINATION WITH SMALL

MOLECULE INHIBITORS.

SO THIS IS A STUDY THAT WE

HAD INITIATED WITH A GROUP

AT THE NIH, THE NCGC.

OUR PRIMARY CONTACT THERE

HAS BEEN DR. ANTON SEMINOFF

AND THEY HAVE BEEN FABULOUS

COLLABORATORS.

PREVIOUSLY THEY HAD WORKED

WITH SAM WILSON AND ROGER

WOOD GATE TO DEVELOP A

FLUORESCENT-BASED STRAND

DISPLACEMENT ASSAY THAT

COULD BE USED FOR SCREENING.

AND THIS ASSAY IS EXTREMELY

EFFICIENT IN ITS DESIGN.

THERE IS A TRIPAR DIET

SUBSTRAIT WHICH CONTAINS A

QUENCHING MOLECULE TO BLACK

HOLE QUENCHER TWO.

IN THE BLUE IS THE PRIMER

STRAND ON WHICH ONE WOULD BE

CARRYING OUT D.N.A.

SYNTHESIZE AND DOWNSTREAM OF

THIS IS THERE IS A REPORTER

STRAND THAT AT THREE PRIMEIN

CONTAINS TAMA THAT AS LONG

AS THE REPORTER STRAND IS

BOUND TO THE TEMPLATE

STRAND, THE PHYSICAL

PROXIMITY OF THE TAMARA WITH

THE BLACK HOLE QUENCHER

ESSENTIALLY LIMITS TO A

VERY, VERY GREAT EXTENT THE

AMOUNT OF FLUORESCENCE THAT

CAN BE MEASURED.

HOWEVER, UPON ADDITION OF DNTPS,

ONCE ONE HAS BEEN ABLE TO

CARRY OUT STRAND INVASION OF

THE REPORTER STRAND, THEN

THE MOUNTING TEMPERATURE OF

THE REPORTER IS SUCH THAT AS

SOON AS YOU'VE BEGUN STRAND

DISPLACEMENT, IT THEN GOES

INTO SOLUTION AND YOU CAN

MONITOR THE FLUORESCENCE.

AND THIS IS QUITE AMENABLE

TO SCREENING ASSAYS.

SO KEN IN MY LAB, A GRADUATE

STUDENT, QEPD THIS SYSTEM

FOR US USING D.N.A.

POLYMERASE KAPPA.

SHE FIRST WORKED ON THIS

USING JUST THE 384 WELL

PLATES WITH 10 MICROLITER

REACTIONS AND WHAT YOU SEE

ON THE LEFT IS THAT UPON

ADDITION OF POLL KAPPA THAT

THERE IS A SIGNIFICANT

INCREASE IN THE FLUORESCENCE

THAT IS MEASURED.

THE REASON THAT THE FIRST

DATA POINT DOES NOT START

OUT AT ZERO IS SIMPLY THE

LENGTH OF TIME THAT IT

REQUIRES TO DO THE CUTTING

AND TO BE ABLE TO GET THIS

INTO OUR INSTRUMENT WAS

RIGHT ABOUT A MINUTE, MAYBE

A LITTLE BIT LONGER.

AND SO WE DON'T CAPTURE THAT

FIRST MINUTE'S WORTH OF SIN

THEY SIS.

BUT YOU CAN SEE THAT YOU

HAVE AN EXTRAORDINAIRELY

LARGE SIGNAL-TO-NOISE RATIO

BETWEEN NO POLAR MEARSS AND

WITH THESE DATA, THEN THE

GROUP AT THE NCGC THEN

REDUCED THIS TO THEIR

SYSTEM, WHICH IS THE 1536

WELL PLATES FOR MICROLITER

REACTIONS AND ONE CAN SEE

THAT THIS WORKED

EXTRAORDINARILY WELL ALSO

AND THAT SYSTEM WHERE THE

MAERGS YOU HAVE NO SIGNAL

AND WITHOUT THE ADDITION OF

POLL KAPPA, YOU GET A VERY,

VERY ROBUST SIGNAL.

AND SO THE IDEA THEN IS THEN

TO SCREEN FOR INSIBTORS OF

-- INHIBITORS OF POLL KAPPA

THEN BY LOOKING FOR THE LACK

OF FLUORESCENT SIGNALS

COMING THROUGH.

SO WE DID A SERIES OF PILOT

SCREENS WITH THEM.

IN THE PILOT STUDY THIS

TOTALED ABOUT 15,000

MOLECULES THAT WERE SCREENED

AT SEVEN CONCENTRATIONS

EACH.

THIS ALSO INCLUDED THE

LIBRARY FORM COLOGICALLY

ACTIVE COMPOUNDS.

THIS IS THE VERY FIRST

TIME-STEP THAT ONE TAKES,

WHICH IS 12820 MEMBER

LIBRARY.

AND JUST TO GIVE YOU AN IDEA

OF WHAT THESE LOOK LIKE.

THESE ARE A COUPLE OF THE

1536 PLATES IN WHICH ON THE

FAR LEFT YOU CAN SEE WHAT

WOULD BE AN ADDITION POLL

KAPPA AND THAT'S IN THE RED

BOX.

IN THE BLUE BOX IS WHERE

THERE WAS NO POLL KAPPA

BEING ADDED.

PLATE ON THE LEFT WOULD

INDICATE THE LOWEST

CONCENTRATION OF THE

INHIBITORS THAT WOULD HAVE

BEEN PUT INTO THESE AND YOU

CAN SEE THAT EXCEPT FOR

POSSIBLY ONE PARTICULAR WELL

THAT THERE WAS NO INHIBITION.

WHILE AT THE HIGHEST

CONCENTRATIONS YOU CAN SEE

THAT THERE WERE A VARIETY OF

HITS.

SO THE GREAT THING ABOUT

THEIR PARTICULAR SYSTEM IS

THAT YOU ARE LOOKING AT

SEVEN DIFFERENT

CONCENTRATIONS OF EVERY

COMPOUND IS SO YOU CAN

GENERATE IC 350 CURVES AND

THIS IS JUST A

TYPICAL-LOOKING DATA COMING

FROM THAT.

OBVIOUSLY, YOU WANT THINGS

THAT ARE INHIBITING AT THE

LOWEST CONCENTRATIONS.

AND SO THE RESULTS OF THIS

INITIAL PILOT SCREEN THAT WE

CARRIED OUT WAS THAT THEY

IDENTIFIED APPROXIMATELY 500

HITS AND THAT THIS WAS

RAPIDLY TRIAGED DOWN TO

AROUND OF 0, IN WHICH THOSE

60 COMPOUNDS WERE THEN GIVEN

TO OUR LAB TO LOOK AT

SECONDARY ASSAYS.

KINRIN CARRIED OUT A

RADIOACTIVE GEL-BASED PRIMER

EXTENSION ARRAY SAY AND THE

RESULTS OF THIS WERE WE WERE

VERY PLEASED THAT OUT OF THE

60 THEY IDENTIFIED, WE ONLY

HAD THREE FALSE-POSITIVES BY

EVIDENCE THE BY INABILITY TO

REPRODUCE THAT IN A PRIMARY

EXTENSION ASSAY.

IT TURNED OUT THAT FIVE OUT

OF THE 60 WERE VERY, VERY

DIFFICULT TO WORK WITH

BECAUSE THESE DID NOT ALLOW

ANY OF OUR D.N.A.S TO EVER

MIGRATE INTO THE GEL.

BUT WE WENT AHEAD AND

CONTINUED WORKING UP 52 OF

THESE AND THE CRITERIA FOR

PRIORITYIZATION WAS ONE THAT

THE NCGC PUTS IN PLACE, IN

WHICH THERE ARE A NUMBER OF

CRITERIA THAT HAVE TO BE

ABLE TO PASS.

ONE, WHETHER OR NOT THERE

ARE PROBLEMATIC FUNCTIONAL

GROUPS, WHETHER YOU GET A

REALLY GOOD DOSE-RESPONSE

CURVE, IS IT F.D.A.-APPROVED

AND CAN THIS BE OPTIMIZE BID

MEDICINEAL AND CHEMISTRY?

JUST TO GIVE YOU AN IDEA OF

WHAT THIS IS LOOKING LIKE,

USING THOSE STRINGENT

CRITERIA, WE REDUCED THIS

DOWN TO REALLY CONCENTRATING

ON THREE INHIBITORS.

WE WERE ABLE TO CARRY OUT

PRIMARY EXTENSION ASSAYS AND

YOU CAN SEE THAT WE

DEVELOPED REASONABLE-LOOKING

IC 50'S FOR ALL OF THESE.

AND WHEN WE LOOKED AT THE

CONCENTRATION THAT WE GOT

COMPARED TO THE SCREEN,

GENERALLY OURS WERE

INDICATING AN IC 50 OF

APPROXIMATELY TWO TO

THREEFOLD IN THE GROUP --

[INDISCERNABLE]

 POLL KAPPA IS RESPONSIBLE

FOR REPLICATION BYPASS OF

DAMAGE D.N.A.S.

WE LOOKED AT THE GAMMA

HYDROXYDG TO SEE IF IT WAS

ALSO INHIBITING REPLICATION

BYPASS OF THESE.

AND DATA TURNED OUT QUITE

NICE AND IN AGREEMENT WITH

WHAT WE SAW IN THE

REPLICATION OF THE UNDAMAGED

D.N.A..

AND SO WHAT THIS IS

INDICATING IS THAT WE ARE

ABLE TO USE NON-DAMAGED

D.N.A. AS A VERY GOOD

SURROGATE OF REPLICATION ON

DAMAGE TEMPLATES BUT WE'LL

CONTINUE TO USE THAT AS ONE

OF THE CRITERIA.

WHAT I'D LIKE TO THEN

TRANSITION TO IS THEN SOME

OF THE WORK THAT KINRIN HAS

BEEN DOING TRYING TO NOW

WORK OUT BIOLOGICAL ASSAYS

TO DETERMINE IF THESE

INHIBITORS ARE CAPABLE OF

MODULATING THE BIOLOGICAL

ACTIVITY OF POLL KAPPA AND

THE FIRST ASSAY THAT SHE WAS

WORKING UP WAS IN LOOKING AT

HUMAN FIBROBLAST CELLS WHERE

YOU WOULD BE ADDING OR NOT

THE INHIBITOR AND THEN

CHALLENGING THESE CELLS WITH

MITOMYCIN C AND WHAT ONE

SEES IN THE LEFT HAND PANEL

IS THAT THE EYE BEING THE

INHIBITOR ALONE, HAD NO

CYTOTOXIC EFFECT AND THEN IN

THIS EXPERIMENT SHE HAD USED

TWO DIFFERENT CONCENTRATIONS

OF MITOMYCIN SKPRMS IN ONE

CASE WE HAVE ABOUT AN 85%

SURVIVAL.

BUT WHEN THAT IS INCLUDED IN

THE INHIBITOR PLUS THAT,

SAME CONCENTRATION THAT

THERE IS A SIGNIFICANT

INCREASE IN THE CYTOTOXICITY

THAT'S OBSERVED AND YOU SEE

THE SAME THING WHEN THIS IS

LOOKED AT WITH THE HIGHER

CONCENTRATIONS OF MITOMYCIN

C.

WE ALSO HAD, BECAUSE THE

ROBUSTNESS OF THAT ASSAY IS

ONLY IN THE RANGE OF ABOUT

TWO TO THREEFOLD, SHE

SCOURED THE LITERATURE,

LOOKING FOR ADDITIONAL WAYS

TO BE ABLE TO MEASURE THE

POTEENCY OF THESE INHIBITORS

IN A CELL BIOLOGY SYSTEM.

AND SPECIFICALLY, THERE WAS

A REPORT IN WHICH THEY WERE

DOING SIRNA KNOCKDOWNS IN

CELLS THAT WERE ALREADY

VARIATES SO THEY WERE

DEFICIENT.

AND I DON'T HAVE TIME TO GO

THROUGH ALL THE DETAIL IN

THIS SLIDE, BUT SIMPLY TO

ILLUSTRATE THAT IN A POLL

KAPPA NKDOWN THAT THERE WAS

ABOUT A FIVE TO EIGHTFOLD

INCREASE IN THIS CYTOTOXICITY.

SO WHAT WE DECIDED TO DO AND

JIM CLEAVER WAS KIND ENOUGH

TO SEND US XPV CELLS.

SO THE HYPOTHESIS WAS THAT

IF WE START WITH AN XPV CELL

AND ADD INHIBITOR OR NOT AND

THEN CHALLENGED THOSE HAVE-W

VARIOUS DOSES OF UV, THEN

COULD WE SEE ADDITIONAL

CYTOTOXICITY?

AND SO SHE LOOKED AT A

VARIETY OF UV TREATMENTS,

AND WHAT YOU SEE IS THAT IN

THE CASE OF INHIBITOR ALONE

THAT THERE IS NO REDUCTION

IN THE SURVIVAL OF THESE

CELLS, BUT WHEN YOU USE A

COMBINATION OF UV-PLUS

INHIBITOR THAT WE ARE SEEING

SIGNIFICANT INCREASES IN

THAT CYTOTOXICITY.

SO THE CURRENT STATUS OF OUR

WORK WITH LOOKING AT POLL

KAPPA INHIBITORS IS THAT WE

HAVE NOW EXTENDED THIS TO A

SCREEN OF APPROXIMATELY

400,000 MOLECULES.

THIS WAS DONE AT FIVE

CONCENTRATIONS RATHER THAN

THE SEVEN IN THE PILOT.

BUT THE REALLY NICE THING

ABOUT THIS PARTICULAR

LIBRARY THAT THE NCGCS

SCREEN IS ROGER WOOD GATE

AND SAM WILSON HAVE ALSO

SCREENED THE SAME LIBRARY

FOR INHIBITORS OF POLL IOTA

AND BETA.

AND AT THIS POINT WE ARE

DOING COMPARATIVE ANALYSES

OF THOSE COMPOUNDS, WHICH

ARE INHIBITORY TO PAUL KAPPA

VERSUS SOME OF THE OTHER

TRANSLESION OR REPAIR

POLYMERASES.

WE ARE WORKING TOWARD A

COCRYSTALIZATION WITH PAUL

KAPPA AND D PO4 AND WE ARE

CURRENTLY DESIGNING WHAT WE

WANT TO HAVE IS A SCREEN FOR

INHIBITORS OF POLL KAPPA

USING A CELL BIOLOGY,

SYNTHETIC RELATIONSHIP.

SO I DON'T WANT TO -- I WANT

TO MAKE SURE MAUREEN HAS

PLENTY OF TIME.

THIS WORK HAS ONLY BEEN

POSSIBLE BECAUSE OF A

TREMENDOUS GROUP OF PEOPLE

WORKING ON THIS, BOTH ARENA

AND KINRIN HAVE LAID THE

FOUNDATIONS AND REALLY MOVED

THIS FORWARD.

WE'VE HAD FANTASTIC

COLLABORATIONS HERE AT OHSU,

AT THE NCGC, VANDERBILT AND

I THINK I'VE ACKNOWLEDGED

THE OTHER FOLKS THAT HAVE

BEEN HELPING US.

SO I'D PREFER TO BE ABLE TO

TAKE QUESTIONS UNTIL AFTER

MAUREEN'S HAD A CHANCE AND

WE WILL SEAMLESSLY SFLID MY

PRESENTATION OVER TO HERS ON

DECONSTRUCTING THE FAN KONI

ANEMIA DAMAGE RESPONSE

NETWORK.

>> THANKS, STEVE.

AGAIN, LET ME KNOW ON YOUR

END IF THE VOLUME IS OKAY.

>> IT'S FINE.

>> OKAY.

THANK YOU.

GOOD MORNING.

WELL, STEVE STARTED HIS

PRESENTATION WITH A PICTURE

OF HOOD, WHICH IS THE POINTY

ONE THAT WE CAN SEE FROM OUR

LABORATORIES.

AND I AM GOING TO START MY

PRESENTATION WITH THE

VOLCANO MOUNT ST. HELENS.

THIS IS A VIEW FROM THE

NORTH LOOKING INTO THE

GROWING TONE.

WE CAN ALSO SEE THIS VOLCANO

FROM OHSU, TOO.

SO WE HAVE KIND OF A NICE

GEOLOGICAL TAB GOING ON

HERE.

>> BUT DON'T LET ANY OF THAT

DISTURB YOU FROM COMING TO

PORTLAND.

>> CERTAINLY NOT.

IT'S JUST GEOLOGY.

>> HAVE YOU BEEN UP THERE

FOR JUST WATCHING?

>> OH, YEAH.

THAT'S A PREREQUISITE OF

WORKING HERE IS THAT YOU

HAVE TO MOUNTAIN CLIMB AND

SKI AND SNOWBOARD.

I'M PRETTY SURE.

THANKS FOR THE QUESTION.

SO MY LAB IS FOCUSED ON

FRANK KONI ANEMIA AND WE

HAVE SEPARATE AREAS OF

INVESTIGATION, AS YOU CAN

SEE HERE.

WE'RE INTERESTED IN

DISCOVERING MUPROTEINS IN

THE FRANCONI PATHIS ARE

INTERESTED --

[INDISCERNABLE]

 PROTEINS THAT HAVE AN NUNC

ON REPLICATIONS.

AND ALSO USE HUMAN CELLS AS

WELL.

BUT I AM NOT GOING TO TALK

ABOUT THOSE STUDIES TODAY.

I THOUGHT I'D FOLLOW UP

STEVE'S TALK ON SMALL

MOLECULE SCREENING WITH WHAT

WE'RE DOING TO ASSAY FOR

SMALL MOLECULE INHIBITORS OF

THE FRANCONI PATHI --

PATHWAY.

BUT I WANTED YOU TO BE AWARE

OF OTHER AREAS OF THE

INVESTIGATION IN THE LAB.

SO I WANTED TO START OFF

WITH A GENERAL SLIDE ABOUT

FRANCONI ANEMIA.

IT'S A VERY RARE MULTIGENE

DISORDER, AND IT HAS

CLINICAL FEATURES THAT

INCLUDE VERY SPECIFIC

DEVELOPMENTAL DEFECTS,

TYPICALLY BONE MARROW

FAILURE OCCURS BEFORE SEVEN

YEARS OF AGE, AND THERE IS A

DECREASED SURVIVAL.

THERE IS ALSO CANCER

PREDISPOSITION, WHICH IS

MORE EVIDENT NOW THAT THE

PATIENTS ARE TRANSPLANTED

AND LIVE LONG ENOUGH TO

EXPRESS THIS CANCER

SUSCEPTIBLE FENE ZPOIP

THAT'S USUALLY EXPRESSED AS

SOLID TUMORS AND SQUAMOUS

CELL CARCINOMAS.

THE FRANCIONI IS

DEMONSTRATED BY SPONTANEOUS

ABNORMALITIES THAT INCLUDE

CHROMOSOME BREAKS IN

RAIDIALS.

THE CELLS ARE HYPERSENSITIVE

TO D.N.A. INNER STRAND CROSS

LINKING AGENTS SUCH AS

MITOMYCIN C AND SIS PLAITIN.

THIS IS AN UPDATED VERSION

OF THE FRANCONI ANEMIA

PATHWAY.

IN 2012, WHEN I FIRST

STARTED WORKING ON THIS --

THIS IS GOING TO AGE ME --

BUT THERE WAS ONLY ONE GENE

THAT HAD BEEN IDENTIFIED AND

THAT WAS C, WHICH YOU SEE UP

IN THE CORE COMPLEX, ONE OF

THE BLUE BALLS UP THERE IN

THE CORE COMPLEX.

THE CORE PLEX ACTS AS AN E 3

LYINGASE AND HAS OTHER

FUNCTIONS, AND IN RESPONSE

TO D.N.A. DAMAGE AND DURING

S PHASE, THIS CORE COMPLEX

PARTICIPATES IN A PROCESS

THAT MON UBIQUITOUSLY --

THESE ARE PAR LOGS THAT FORM

A COMPLEX AND THIS COMPLEX

IS THEN RECRUITED TO CHROMATIN

AND AS YOU CAN SEE IN THE

IMMUNOFLUORESCENCE PICTURE

THERE, YOU END UP GETTING

VERY BRIGHT SUBNUCLEAR FOSSI

AND REPAIR VIA THE FRANCONI

HOMULGUS COMBINATION IN

TRANSLESION.

THIS IS STILL UNDERGOING

INTENSE INVESTIGATION AT

THIS POINT.

THE MOLECULAR DETAILS OF

THAT PROCESS, AND THE

CHALLENGES HERE ARE THAT THE

PATHWAY IS VERY COMPLEX.

THERE IS FUNCTIONAL

INTERDEPENDENCE, IN

PARTICULAR AMONG THE CORE

COMPLEX PROTEINS.

MANY OF THESE PROTEINS ARE

ORPHANS WITH THE EXCEPTION

OF THE HIGHLY CONSERVED E 3

LIGASE L AND THE DEAH M.

SO THAT ALSO RESTRICTS SOME

OF THE STUDIES THAT ONE CAN

IMAGINE DOING.

IN ADDITION, THERE ARE MORE

FA PROTEINS TO BE DISCOVERED

AND PROTEINS IN THE CORE

COMPLEX AND THAT'S ONE OF

THE OTHER SUBJECTS WE'RE

WORKING ON IN THE LAB.

GENERALLY ONE WOULD LIKE TO

TARGET THE D.N.A. DAMAGE

RESPONSE.

AND THIS AGAIN, EMPHASIZES

WHAT STEVE ALREADY SAYS SAID

EARLIER, THIS MAY IMPROVE

THERAPEUTIC INDEX BECAUSE

CANCER CELLS OFTEN HAVE

DEFECTIVE D.N.A. REPAIR PATHWAYS 

PATHWAYS.

ANOTHER ASPECT IS THIS IS TO

TAKE ADVANTAGE AND EXPLOIT

SYNTHETIC RELATIONSHIPS AND

PROBABLY THE BEST EXAMPLE OF

THAT ARE PARP INHIBITORS

RIGHT NOW.

AND TO ME WHAT'S INTENSELY

INTERESTING IS TO ME IT'S AN

UNEXPLORED AREA OF

INVESTIGATION IN THE D.N.A.

DAMAGE RESPONSE AND THAT'S

LOTS OF DIFFERENT TARGETS

THAT ONE CAN EXPLORE IN THIS

RECORD.

-- REGARD.

SO I WANT TO SPEND JUST A

COUPLE OF MINUTES TALKING

ABOUT OUR EXPERIMENTAL

STRATEGY AND THIS IS

RESTRICTED TO JUST THE

EXPERIMENTS WE DO IN ZETPUS.

ACE MENTION, WE ALSO DO LOTS

OF EXPERIMENTS USING MAMMALIAN

CELLS AS WELL.

BUT BECAUSE THIS IS AN

UNUSUAL MODEL, I THOUGHT I'D

SPEND SOMETIME ON IT.

WHAT THE POWER OF THIS

SYSTEM IS THAT THE ZEB HAS

LAID VERY LARGE EGGS, WHICH

THEN CAN BE EASILY

MANIPULATED TO COME UP WITH

AN EXTRACT THAT IS FULLY

CAPABLE OF INVITRO

REPLICATION.

ONCE YOU HAVE D.N.A. AND IT

CAN BE ANY KIND OF D.N.A.,

DEPENDING ON THE TYPE OF

EXTRACT, THERE ARE LOTS OF

DIFFERENT EXTRACTS ONE CAN

MAKE.

WE MAKE A LOW-SPEED EXTRACT

THAT IS INCAPABLE OF THIS

INVITROREPLICATION.

AND YOU CAN SEE THERE ON THE

RIGHT IS DECONDENSED SPERM

CHROMATIN THAT HAVE THEN

BECOME DECONDENSED AND

THEY'VE GONE THROUGH ONE

FULL CYCLE OF REPLICATION.

THE POWER OF THIS SYSTEM IS

THAT ONE CAN ADD D.N.A.

SUBSTRAITS.

YOU CAN ADD CHEMICALS AND

YOU CAN ALSO DEPLETE

PROTEINS USING SPECIFIC

ANTIBODIES.

YOU CAN ALSO ADD BACK

INTERFERING PROTEINS AS WELL.

AND SO THIS IS ONE OF THE

EXPERIMENTAL PLATFORM THAT'S

WE USE TO DECONSTRUCT THE

FRANCONI PATHWAY.

WHAT WE FOUND THAT STIMULATED

OUR INTEREST IN SCREENING

WAS THAT WE COULD TAKE

D.N.A. AND ADD D.N.A. TO THE

EXTRACTS, WHICH WERE D.N.A.-FREE 

D.N.A.-FREE, BECAUSE THIS,

THE PROCESS THAT I SHOWED

HERE EXTRACTS D.N.A., SO

THIS IS NOW A D.N.A.-FREE

EXTRACT.

SO NOW YOU ADD D.N.A. TO

THIS.

AND WHAT WE FOUND IS UNDER

THOSE CIRCUMSTANCES D 2,

WHICH IS HERE THE SHORT FORM.

THIS IS NON-UBLATED SAINT D

2.

NOW WHEN D.N.A. IS ADDED, IT

BECOMES UBQUINNATED AND THIS

IS DPENGT DEPENDENT ON THE

AMOUNT OF D.N.A..

THIS WAS A SURPRISE TO US

AND IT WAS ALSO A WAY TO

THINK ABOUT PERHAPS DOING

SCREENS USING THIS ASSAY,

WHERE ALL THE COMPONENTS

THAT ARE REQUIRED, EVEN THE

ONES WE DON'T KNOW ABOUT YET

-- ARE PRESENT AND ACTING IN

A NATURALLY SELF-SYNCHRONIZED

MANNER.

SO WE KNOW THAT THE D.N.A.

THAT WE ADD TO THE EXTRACT

STIMULATES D 2 BECAUSE WE

DID AN EXPERIMENT WHERE WE

TAGGED UBQUITIN.

IT WAS A HISS TAG AND RAN UP

THROUGH A NICKEL COLUMN AND

WHEN WE DID THAT, WE FIND,

AS YOU CAN SEE BELOW, THAT

THE EXTRACT WAS STIMULATED

WITH D.N.A..

IT HAS A VERY -- WAS ABLE TO

CAPTURE THE HISS TAG UB

QUINNATED D 2.

WHAT WE'VE DONE NOW IS WE'VE

BEGUN TO DO A LIBRARY

SCREENING IN EXTRACT.

AND THE WAY THIS WORKS IS WE

STIMULATE THE EXTRACT, ADD

THE CANDIDATE DRUG AND IN A

MULTIPLEX WAY, SO JUST A

PLATE, AND I'VE GOT DOWN

HERE -- THIS IS AN ACADEMIC

THROUGH PUT.

SO THIS ISN'T A HIGH THROUGH

PUT AT ALL BECAUSE WE'RE

ANALYZING THIS BY IMMUNOBLOG

AND YOU CAN SEE AT THE

BOTTOM OF THE SLIDE THE

POSSIBLE OUTCOMES.

SO IT'S POSSIBLE THAT THE

COMPOUND MIGHT BLOCK

UBQUINNATION OF SAINT D 2

AND IT'S ALSO POSSIBLE THAT

WE MIGHT FIND COMPOUNDS THAT

BLOCK DEUBQUINNATION OF D 2.

SO WHAT WE DID INITIALLY,

BECAUSE ELLEN'S LAB FOUND

OUT THAT PER CUR-MINUTE, A

NATURAL PRODUCT, WE GOT

TOGETHER WITH JIM SCHNEIDER

AT EMORY AND TESTED PER

CUMIN IN MANY OF ITS

ORTHOLOGS.

-- I'M SORRY.

ANALOGUES.

HE MADE LOTS OF DIFFERENT

ANALOGUES TO KER CUMIN IN

ORDER TO DO STRUCTURAL

ACTIVITY RELATIONSHIPS.

AND WE DID IDENTIFY A VERY

STRONG INHIBITOR OF UBQUINNATION

IN THE ANALOGUE ES THIS

RIGHT HERE.

AND YOU CAN SEE COMPARED TO

THE OTHER ONES, THIS IS

LOOKING PRETTY INHIBITTRY.

SO WE -- WHAT WE DID THEN

WAS TRIED THE SAME

EXPERIMENT IN HELIX CELLS

AND INTERESTINGLY NOT JUST

YOU HAVE THIS BUT FOR ALL --

24 BUT FOR ALL THE COMPOUNDS

THAT WE HAVE IDENTIFIED SO

FAR, THE INHIBITION THAT WE

SEE IN EXTRACTS IS MIRRORED

IN HUMAN CELLS.

SO WHAT YOU CAN SEE HERE IS

THE TREATMENT OF HELIX CELLS

AFTER DAMAGE WITH HUMSU

RESULTS IN -- HU RESULTS IN

DIMINISHING APPEARANCE OF

UBQUINNATED ISOFORM OF C D

2.

THIS IS A LESS ACTIVE

ANALOGUE.

AND SO THE PREDICTION HERE

IS BY INHIBITING THE PATHWAY

IN CELLS THAT HAVE DEFECTS

IN GENES THAT ARE SYNTHETIC

WITH FA, THIS TREATMENT WILL

END UP BEING SELECTIVELY

TOXIC.

SO THIS CONCEPT IS

ILLUSTRATED HERE, WHERE IF

THERE IS A MUTATION IN GNA

AND THE PHENOTYPE WOULD BE

PREDICTED TO BE ALIVE,

MUTATION IN B ALIVE.

BUT IF YOU HAVE A MUTATION

IN BOTH THESE SPECIFIC A AND

B, THIS IS A LETHAL EFFECT.

AND SO THERE IS A SYNTHETIC

LETHAL SCREENING OF LIMITED

SYNTHETIC SCREENING

PERFORMED AS SHOWN HERE BY

PAPER BY KENNEDY AND OLIN

2007.

AND I WANT TO DRAW YOUR

ATTENTION ON THIS LIST OF

SYNTHETIC LETHAL TARGETS IS

HEM.

HEM IS DEFICIENT IN MANY

HEMATOLOGIC MALIGNANCIES AND

COL AND AOL HAS DIFICIENCY

MEASURED ALL DIFFERENT KINDS

OF WAYS, WHICH IS ANOTHER

ISSUE YOU NEED TO FACE WHEN

YOU ACTUALLY TRY TO USE

THESE COMPOUNDS.

BUT IN ANY CASE, THEIR

A.T.M. DIFICIENCY IS COMMON

IN CANCER.

AND SO WE COLLABORATED WITH

MITCH TURKER, WHO IS HERE AT

OHSU, AND THE PREDICTION

WOULD BE THEN IF WE INHIBITED

THE FRANC ONI PATHWAYS IN

CELLS THAT ARE DEFICIENT IN

A.T.M., WE WOULD HAVE A

SELECTIVELY TOXIC EFFECT.

AND AS YOU CAN SEE HERE, THE

ANALOGUE THAT I SHOWED

EARLIER THAT WE IDENTIFIED

AS THE VERY STRONG INHIBITOR

OF D 2.

THIS IS THE 24.

YOU CAN SEE HERE THE

KNOCKOUT VERSUS THE WILD

TYPE.

THERE IS A MORE TOXIC IN

EFFECT A.T.M. KNOCKOUT CELLS

AND THIS IS CONSISTENT WITH

A SYNTHETIC LETHAL EFFECT.

AND HERE IS THE -- AGAIN,

THE LESS ACTIVE ANALOGUE

HERE.

AND SO WHAT WE'RE DOING

RIGHT NOW IS WE'RE

CONVERTING THIS TO A

HIGH-THROUGH PUT SCREEN.

AS STEVE ALREADY MENTIONED,

THE FLUORESCENT-SPACE

SCREENS ARE SUFFICIENT TO DO

PROPER HIGH-THROUGH PUT.

YOU THINK OF THE COMPOUND

WE'VE IDENTIFIED SO FAR HAVE

PROOF OF CONCEPT BUT WHAT WE

REALLY NEED IS A VERY

SPECIFIC INHIBITOR OF SAINT

D 2 UBQUINNATION AND WE

THINK THAT THIS BIOLUM HE ISENCE

ENERGY TRANSFER METHOD WOULD

WORK.

WE'VE GOTTEN FUNDING FROM

THE INNOVATION -- AN

INNOVATION AWARD FROM THE

ORGAN TRANSLATIONAL CENTER

HERE IN OREGON.

SO I JUST WANT TO

ACKNOWLEDGE SOME OF THE

PEOPLE THAT HAVE CONTRIBUTED

TO THIS WORK.

CHELSEA MICHAEL JENKINS WHO

IS HERE AND WORKED ON THE

SCREEN FOR THE LAST YEAR TO

DEVELOP THE HIGH-THROUGH PUT.

MICHAEL WALLACE HAS REJOINED

THE LAB.

I BROUGHT HIM BACK

SPECIFICALLY TO WORK ON THIS

PROJECT.

AND OTHERS ARE ALSO

CONTRIBUTING AND THE PATH

MEMBERS IGOR IN PARTICULAR,

IGOR LANDIS, WHO IS NOW A

PROFESSOR, ASSISTANT

PROFESSOR HERE AT THE

THERAPY INSTITUTE, WORKED

VERY HARD ON THIS PROJECT,

AND COINVENTORS IGOR, ALEX,

AND STACEY -- THOSE HAVE ALL

MOVED ON TO BIGGER AND

BETTER LABORATORIES,

ESPECIALLY ALEX, WHO HAS HER

OWN LAB NOW AT THE

UNIVERSITY OF MINNESOTA.

I WANTED TO POINT OUT OUR

LONGTIME COLLABORATOR UDALL

LONG AND NIH OF THE

UNIVERSITY OF AM STER DAM.

JEN CHRISTIAN, WHO HELPED US

BEFORE SHE WENT TO THE

UNIVERSITY OF UTAH AND IN

PARTICULAR JIM SCHNEIDER AND

EMERY FOR THE ANALOGUES OF

PER CUMINY AND AS STEVE

ALLUDED TO, I WANTED TO

SPEND JUST A COUPLE MORE

MINUTES ON TELLING YOU ABOUT

THE WHAT I THINK IS -- MIGHT

BE INTERESTING FOR EVERYONE

TO HEAR ABOUT, WHICH IS THE

OHSU RARE DISORDERS RESEARCH

CONSORTIUM.

THIS IS ABOUT 150 STAFF AND

FACULTY THAT ARE INVOLVED

WITH THIS CONSORTIUM

STUDYING RARE DISORDERS HERE

AT OHSU, AND WE HAVE UNIQUE

RESOURCES, AND I KNOW PART

OF THE GOAL OF THIS NEW

STYLE OF TALK IS TO THINK

ABOUT COLLABORATIONS AND

PROBABLY PROGRAM PROJECT

GRANTS.

BUT WE DO HAVE UNIQUE

RESOURCES HERE.

ANIMAL MODELS, INTERNATIONAL

PATIENT REFERRALS, CELLWISE

DATABASES AND SPECIALIZED

TECHNIQUES.

THE CONSORTIUM IS COCHAIRED

BY UNION HAS LICK AND

MYSELF.

SO WE HAVE A DISTRIBUTED

STEERING COMMITTEE AS WELL.

AND I JUST WANTED TO POINT

OUT THE STRENGTHS OF JUST TO

HIGHLIGHT SOME OF THE

PROGRAMS WE HAVE HERE AND

THE RARE DISORDERS RESEARCH

CONSORTIUM.

THIS IS A LITTLE DAS6<„

-- OHSU RESEARCHERS DO STUDY

MORE THAN 90 RARE DISORDERS

BUT THERE ARE EIGHT DISORDER

IN PARTICULAR THAT OHSU

INVESTIGATORS SPAN THE WHOLE

PIPELINE FROM DISEASE GENE

DISCOVERY TO UNDERSTANDING

MECHANISMS, HAVING ANIMAL

CELL HUMAN RESEARCH

RESOURCES, INNOVATIVE

DIAGNOSTICS, PRECLINICAL AND

CLINICAL TRIALS AND YOU CAN

SEE THAT THERE ARE SOME GAPS

HERE THAT WE'RE FILLING IN

RIGHT NOW AND WE'RE ALWAYS

INTERESTED IN HEARING ABOUT

COLLABORATIONS IN THESE

AREAS.

SO JUST TO RUN DOWN THE LIST

-- THIS IS AN IRON

ACCUMULATION DISORDER NUMBER

ONE.

AND OF COURSE, FRANCONI.

STERAL DEFECTS, OPHTHALMIC

DISORDERS, CHRONIC IS A

MAJOR FOCUS AND TARGETED

CANCER THERAPEUTICS.

CONNECTIVE TISSUE DISORDERS,

FATTY ACID OXIDATION

DISORDERS AND HUNTINGTON'S

DISEASE.

AND I WANTED TO FINISH BY

AGAIN HIGHLIGHTING THE OHSU

RESEARCH REVEL TO RARE

CANCER SUSCEPTIBILITY AND

MANY OF THESE INDIVIDUALS

ARE HERE IN THIS ROOM.

BUT IN ADDITION THERE ARE

MANY INDIVIDUALS AND THEIR

MENTEES WORKING ON D.N.A.

REPAIR DISORDERS IN OHSU.

SO IF YOU WANT MORE

INFORMATION ON THE RARE

DISORDERS RESEARCH CONSORTIUM,

WE HAVE AN ACTIVE WEBSITE

THAT IS KEPT UP TO DATE.

SO I'D DIRECT YOU THERE FOR

MORE INFORMATION.

AND THANK YOU VERY MUCH.

[APPLAUSE]

>> VERY NICE PRESENTATION.

AND VERY COMPREHENSIVE.

AND WE'RE HOPING THAT THAT

WILL BE POSTED ON THE

ARCHIVES SO PEOPLE CAN VIEW

IT AT THEIR LEISURE.

AND WE'RE GOING AROUND TO

DIFFERENT SITES NOW AND HAVE

ONE QUESTION PER SITE, IN

THE INTEREST OF TIME.

AND IF THERE IS STILL TIME

-- LET'S GO TO STONY BROOK

FIRST.

ARE YOU THERE IN STONY BROOK?

>> YES, WE'RE HERE AND I

THINK WE HAVE A QUESTION.

WHICH IS APPARENTLY FROM ME

BECAUSE I COULDN'T GET

ANYONE ELSE TO ASK ONE,

WHICH IS THIS IS FOR STEVE.

STEVE, DO YOU STILL SEE

INHIBITORY ENHANCING EFFECTS

ON UV SENSITIVITY, FOR

EXAMPLE, OR OTHER PHENOTYPES

IN POLL KAPPA KNOCKDOWN

CELLS FOR YOUR INHIBITOR?

>> THANKS.

WE ARE CURRENTLY WORKING ON

MAKING SOME STABLE KNOCKDOWNS

OF THE POLL KAPPA IN ORDER

TO BE ABLE TO REALLY PUT THE

VARIOUS INHIBITORS THROUGH

THEIR PHASES.

I THINK THAT THE USE OF THE

EITHER THE S.I.S IS SOMEWHAT

LIMITED BECAUSE OF THE

SIGNAL-TO-NOISE RATIO THAT

YOU HAVE TO WORK WITH.

AND SO WHAT WE'RE LOOKING

FOR IS TO BE ABLE TO DEVELOP

THESE THAT ARE GOING TO BE

VERY STABLE IN TERMS OF THE

VERY STRONG KNOCKDOWNS.

BUT THE DIRECTION, BRUCE,

YOU WERE GOING WITH?

>> THE QUESTION WAS WHETHER

THE PHENOTYPE IS ACTUALLY

DEPENDENT ON POLL KAPPA.

>> IT'S UNDER INVESTIGATION.

>> OKAY, THANKS.

>> OKAY, THANK YOU.

LET'S GO TO BALTIMORE NOW.

PLEASE MUTE AT STONY BROOK.

>> OKAY, I WANT TO THANK

YOU, STEVE AND BOTH OF YOU

FOR A VERY NICE PRESENTATION,

AND I THINK YOU JUST

FOLLOWED UP ON THIS IDEA

THAT WE GOT TO KNOW MORE

ABOUT WHAT'S GOING ON.

THERE ARE SEVERAL THINGS

THAT ARE OF INTEREST TO US

AND FOR EXAMPLE, WE ALSO ARE

WORKING WITH INHIBITORS HERE,

DAVID WILSON, BOB BAR SHH

AND MYSELF AND USING THE

INHIBIT AND STUDYING THE

FUNCTIONS AND WE HAVE A

NUMBER OF PEOPLE WORKING

WITH FRANC ONIS AND MICHAEL

SIDEMAN, BOB RUSH.

SO IT ALL FIT IN VERY WELL

TO ENHANCE FURTHER

DISCUSSION, COLLABORATION.

I KNOW YOU ARE DOING

PROJECTS AS WELL.

I GUESS I'VE TAKEN MY TIME

BUT A QUICK QUESTION OF

INTEREST WOULD BE THESE

PROTEIN D.N.A. CROSS LINKS,

THE PEPTIDE CROSS LINKS THAT

YOU MENTIONED TO D.N.A..

WHAT IS THE BIOLOGICAL

SIGNIFICANCE OF THAT?

-- OF THOSE?

STEPHEN?

>> I THINK THAT THERE HAS

BEEN A FAIR BI¤ OF WORK THAT

HAS LOOKED AT -- IF YOU HAVE

TREATMENT WITH AGENTS WHICH

ARE CAPABLE OF FORMING

D.N.A. PROTEIN CROSS LINKS,

THAT THERE IS A SIGNIFICANT

LITERATURE, ESPECIALLY FROM

ZACHOVICH AT BROWN

UNIVERSITY, IN WHICH THEN

YOU CAN DETERMINE IF THERE

IS A SEQUENTIAL DEGRADATION

OF THOSE PROTEINS DOWN TO

PEPTIDES, THAT THAT THEN

BRINGS THAT INTO THE SIZE OF

THE LESION THAT WOULD BE

CAPABLE OF BEING RECOGNIZED

BY NUCLEOTIDES EXCISION

REPAIR.

AND SO THE ROLE OF BEING

ABLE TO BREAK THESE PROTEINS

DOWN INTO MUCH SMALLER UNITS

IS STILL AN AREA OF ACTIVE

INVESTIGATION, BUT AS SLAB

HAS PUBLISHED SOME VERY NICE

DATA SHOWING THAT THE -- IF

YOU HAVE THE LARGER PROTEINS

THAT THESE ARING IF TO BE

REFRACTORY TO NER.

BUT ONCE YOU'VE DIE SWRESTED

THEM DOWN INTO LESS THAN

20,000 DALTONS, THEN THEY

BECOME VERY GOOD SUBSTRAITS.

AND ARENA HAS DONE SOME VERY

NICE WORK IN THE UV RAB

SYSTEM.

ALSO DEMONSTRATING THAT YOU

CAN HAVE QUITE ROBUST

ACTIVITY IN NER TO BE ABLE

TO EXCISE THOSE.

>> IT IS LIMITED TO NER?

IS IT LIMITED TO NER?

>> YOU MEAN OTHER THAN --

WELL, I'D LIKE TO THINK THAT

THE TRANSLESION.

BUT YOU MEAN IN TERMS OF A

REPAIR PATHWAY?

>> YES.

>> I THINK CERTAINLY

DR. MCCULA'S DATA ON LOOKING

AT BOTH ACUTE AND CHRONIC

FORMALED HYDE IN THE Y

SYSTEM AND LOOKING INTO THE

MAM ALIAN SYSTEM PROBABLY IT

IS GOING TO BE THE DOMINANT

PATHWAY THAT IS GOING TO END

UP LEADING TO A TOLERANCE

MECHANISM, WHILE UNDER

CIRCUMSTANCES, WHERE YOU

HAVE A MORE ACUTE HIGH-DOSE

EXPOSURE OF SOMETHING LIKE

FORM ALDEHYDE THAT THE

NUCLEOTIDES EXCISION REPAIR

SEEMS TO BE DOMINATING THAT

LANDSCAPE.

SO I THINK IT'S GOING TO

DEPEND UPON THE RELATIVE

AMOUNT OF DAMAGE AND THE

REPLICATION STATE OF THE

CELLS AND I THINK THAT

PROBABLY IN THE SAC SYSTEM

THAT POTENTIALLY THE HR

PATHWAY IS GOING TO BE MORE

DOMINATING THAN PENTAGON

OFFICIAL -- POTENTIALLY THE

NER.

BUT MIKE SITTING ACROSS THE

TABLE, COULD ALSO

POTENTIALLY COMMENT ON THE

RELATIVE USE OF NER VERSUS

H.R..

>> FOR THOSE I WOULD DEFER

TO YOU AND AMANDA.

>> OKAY.

>> OKAY, THANK YOU.

>> I HAVE TO SAY HI TO WIDON.

>> IS THAT SIDEMAN IN THE

BACKGROUND THERE?

HI, MIKE.

MIKE.

HOW ARE YOU DOING?

>> HI, MIKE.

>> HEY.

>> YOU HAVE TO SLIDE AROUND

AND PUT YOUR SMILEY FACE IN

THE CAMERA.

REFLECTION.

>> LET'S PLEASE MUTE IN

BALTIMORE AND LET'S MOVE ON

TO UNIVERSITY OF NORTH

CAROLINA CHAPEL HILL, SINCE

WE WERE JUST TALKING ABOUT

DR. SAN GAR'S WORK.

>> YES, HI.

THIS IS BILL KOFFMAN.

IF YOU SEE, OUR ROOM IS

PRETTY MUCH EMPTY.

IT'S NOT THAT THEY WEREN'T

PEOPLE HERE THAT REALLY

ENJOYED THE TALK.

IT'S JUST THEY DON'T WANT TO

LISTEN TO MY QUESTION.

AND SO THEY'VE ALL LEFT.

I DO HAVE A QUESTION FOR

STEVE ABOUT THE BLAST OMA

WITH POLL KAPPA BEING AN

INDICATOR OF PROGNOSIS.

DO YOU KNOW IF THE DATA HAS

BEEN SUFFICIENTLY

WELL-EXAMINED TO SHOW THE

OVEREXPRESSION OF POLL KAPPA

IN THE HIGH-GRADE OR

PROGNOSIS TUMORS IS

UNRELATED TO THE

PROLIFERATIVE INDEX WITHIN

THE CELLS IN THE TUMOR OR

COULD IT JUST SIMPLY BE THAT

THE HIGHER THE KI 57 INDEX

THE HIGHER THE PROLIFERATIVE

INDEX, THE HIGHER THE LEGAL

OF EXPRESSIONS OF POLL KAPPA

AND WE'RE REALLY JUST

LOOKING AT PROLIFERATIVE

ACTIVITY OF THE CELLS AND

NOT SOME SPECIAL ATTRIBUTE

OF AN EXCESSIVE AMOUNT OF

POLL KAPPA PER S PHASE CELL?

>> PHIL, I THINK THAT'S A

GREAT QUESTION.

AT THIS POINT, I THINK IT IS

MERELY A MATTER OF

CONJECTURE AND SPECULATION.

I THINK THAT SIMPLY THE

OBSERVATION IS THAT THERE

DOES SEEM TO BE A HIGH

DEGREE OF CORRELATION

BETWEEN THE LONG TERM

PROGNOSIS OF INDIVIDUALS

DIAGNOSED WITH THE ABLIAL

BLASTOMAS VERSUS THE LEVELS

OF POLL KAPPA.

THERE IS NO INDICATION THAT

THE TIME THAT KAPPA IS --

OVEREXPRESS IS POTENTIALLY

CAUSATIVE, BUT THAT THE

POINT I THINK IT'S REALLY

VERY MUCH AN OPEN QUESTION.

AND WHETHER OR NOT GOING AND

BEING ABLE TO REDUCE POLL

KAPPA EXPRESSION IN SOME OF

THOSE CELLS NOW IS A THUJ

CHALLENGE DEALING WITH BLOOD

BLOOD-BRAIN BARRIER ISSUES.

BUT WHETHER OR NOT THAT

WOULD INCREASE A THERAPEUTIC

EFFICACY IS COMPLETELY AN

OPEN QUESTION.

I THINK THAT IT SIMPLY

APPEARS TO BE AN AREA OF

OPPORTUNITY TO TAKE A LOOK

AT AND THEN TO TRY AND

DESIGN EXPERIMENTS TO ANSWER

THE QUESTION THAT YOU ARE

ASKING.

>> THANKS VERY MUCH.

GREAT PRESENTATIONS.

>> OKAY.

THANK YOU.

LET'S PLEASE MUTE CHAPEL

HILL.

LET'S GO OFF TO UNIVERSITY

OF PITTSBURGH.

>> STEVE, HI.

ROB SUDDENLEY HERE AND STILL

A BUNCH OF US ENJOYING THE

QUESTION AND ANSWER PERIOD.

THANKS SO MUCH FOR THE

OVERVIEW AND PARTICULARLY

LIKE TO HEAR THE STORY ABOUT

KAPPA, SINCE WE HAVEN'T HAD

A CHANCE TO GO INTO THAT IN

DETAIL.

MY QUESTION REALLY IS FOR

THE BOTH OF YOU.

INHIBITORS ONLY SEEM TO BE

GIVING TWO-FOLD INCREASE

RESPONSE, SUGGESTING NOT

PROBABLY AS MUCH OF THE

SELECTIVITY AS YOU HAD HOPED.

CAN YOU COMMENT ON WHETHER

YOU THINK THAT THIS APPROACH

MAY SORT OF IMPROVE AS YOU

FIND BETTER INHIBITORS AND

AS YOU MOVE INTO ANIMAL

MODELS?

>> YEAH, THAT'STHAS

CERTAINLY THE HOPE ON OUR

PART.

IF YOU LOOK AT SIS PLAITIN,

THAT HAS ABOUT A THREEFOLD

-- IT'S CURRENTLY USED NOW.

IT'S GOT ABOUT A THREEFOLD

SELECTIVITY.

SO I THINK THAT TWO-FOLD

ISN'T TOO FAR AWAY FROM

THAT, BUT OF COURSE, THAT'S

NOT THAT GREAT.

BUT IN OUR CASE, WITH OUR

SCREENS, WE KNOW THAT WHAT

WE'VE COME UP WITH SO FAR IS

NOT SPECIFIC ENOUGH.

ONE OF THE DRAWBACKS OF

DOING THIS SCREEN THE WAY

WE'RE DOING IS IS THAT WE

DON'T HAVE A SPECIFIC TARGET

PER SE.

BUT THE STRENGTH OF IT IS

THAT WE HAVE THE WHOLE

SYSTEM PRESENT.

SO EVERY SCREEN HAS ITS

ADVANTAGES AND DISADVANTAGES.

BUT I'M CONVINCED THAT WE

CAN FIND A MORE SPECIFIC HIT

FOR UBQUINNATION, WHICH IS

WHY WE'RE GOING TO BRENT.

>> SO ROB, JUST TO ALSO

COMMENT.

I THINK NAEVERYTHING THAT WE

HAVE DONE TO THIS POINT IS

MINIMALLY TWO ORDERS, IF NOT

THREE ORDERS OF MAGNITUDE

OFF FROM WHERE ONE WOULD

WANT TO BE TO HAVE SOMETHING

THAT WOULD BE AS I A

SPECIFIC INHIBITOR.

YOU ALSO IDENTIFY THE OTHER

HUGE CHALLENGE IN WHETHER OR

NOT YOU ARE GOING TO BE

LOOKING AT INHIBITING

POLYMERASES OR PATHWAY OR

POSSIBLY IN THE BER PATHWAY.

THERE IS SUCH REDUNDANCY AND

YOU REALLY DON'T KNOW IF THE

IN THE CASE IF YOU SELECTIVELY

INHIBIT ONE OF THE ENZYMES

THAT IS POTENTIALLY

CONTRIBUTING TO MUTEOGENESIS,

YOU MAY THEN FORCE A PATHWAY

THAT COULD, IN FACT, BE

WORSE THAN THE ONE THAN YOU

ARE TRYING TO INHIBIT JOMT

ONE OF OUR CONCERNS,

ESPECIALLY IN TERMS OF SOME

OF THE WORK WITH POLL KAPPA

IS THAT IT IS

EXTRAORDINARILY

HIGH-FIDELITY IN REPLICATING

LESIONS AND SO IF YOU GO AND

INHIBIT THAT, NOW YOU ARE

GOING TO POTENTIALLY BRING

IN A TLS POLAR MEARS THAT IS

GOING TO HAVE A

SIGNIFICANTLY LOWER FIDELITY.

SO I THINK THAT THIS IS IN

EXTRAORDINAIRELY FORMATIVE

STAGES JUST TRYING TO LAY A

LITTLE BIT OF THE GROUNDWORK.

BUT I THINK THERE IS AN

EQUALLY CONVINCING ARGUMENT

COULD BE MADE THAT THE

IBINHIBITORS TO SOME OF

THESE POLYMERASES MAY HAVE

MORE DETRIMENTAL EFFECTS

THAN YOU WOULD ACTUALLY GET

BEFORE FROM.

I THINK THOSE TYPES OF

QUESTIONS AND THE

COMBINATIONS THAT ONE WOULD

USE FOR TREATMENTS IS JUST

TOO FAR DOWN THE ROAD TO

REALLY SPECULATE.

BUT IT'S AN ENORMOUS

CHALLENGE WITH THE BACKUP

SYSTEMS.

>> LOOKS LIKE YOU'RE ON GOOD

TRACK, SO --

>> OKAY.

THANK YOU.

LET'S MUTE PITTSBURGH.

IS ANYONE THERE AT

UNIVERSITY OF KENTUCKY WITH

A QUESTION?POa

>> THERE IN LEXINGTON?

HOW ABOUT NIEHS?

>> HELLO?

CAN YOU HEAR ME?

>> YES.

OKAY.

DAVID.

NOW WE CAN HEAR YOU.

>> SORRY.

I MUST HAVE PRESSED THE MUTE

BUTTON TWICE IN SUCCESSION

REAL QUICKLY.

MY QUESTION IS FOR MAUREEN.

>> SURE.

>> YOU SHOWED REAL CLEARLY

THAT SOME ANALOGUES CAN

CAUSE ADDITIONAL CELL-KILLING

AND A.T.M. DEFICIENT

BACKGROUND.

I KNOW A.T.M. HAS MANY

TARGETS BUT OBVIOUSLY ONE IS

P 53.

HAVE YOU LOOKED AT WHAT

EFFECT P 53 MUTATION OR

DIFICIENCY MIGHT HAVE WITH

YOUR SOUTHLAND AND HOW THAT

MIGHT -- COMPOUND AND HOW

THAT MIGHT AFFECT

THERAPEUTIC USE OR

POTENTIAL?

>> YEAH, WE HAVEN'T LOOKED

AT THAT BECAUSE MY STRATEGY

HAS BEEN WE NEED OF IT A

MORE SPECIFIC INHIBITOR

BEFORE WE SPEND ANY MORE

TIME ON -- THESE EXPERIMENTS

CAN GO ON FOR QUITE A WHILE,

AND I KNOW THAT COR CUMIN

AND ANALOGUES ALSO HAVE A

NUMBER OF OFF-TARGET

EFFECTS.

SO RATHER THAN PURSUING

THOSE IN A P 53 BACKGROUND,

SOMETHING LIKE THAT, I

RATHER PREFER TO FIND

SOMETHING MORE SPECIFIC FOR

THE FRANC ONA PATHWAY FIRST.

I DON'T KNOW IF THAT ANSWERS

YOUR QUESTION.

>> THAT'S FINE.

>> OKAY.

THANK YOU.

PLEASE MUTE IN KENTUCKY.

LET'S TRY NIEHS.

>> HI, SCOTT.

THANK YOU BOTH FOR YOUR VERY

INTERESTING TALKS.

I HAVE A QUESTION FOR --

PROBABLY MORE A TECHNICAL

QUESTION.

YOU SHOWED A SLIDE THAT HAS

TWO HIGH-FREQUENT SCREENING

PLATES AND ONE HAS --

[INDISCERNABLE]

 THE HIGH-CONCENTRATION

PLATE HAD BOTH GRAY AND

WHITE SPOTS ON IT -- WHAT

WAS THE TECHNICAL ORIGIN OF

THOSE CHANGES?

>> YEAH, SO IN THE LIBRARY

OF PHARMACOLOGICALLY ACTIVE

COMPOUNDS, THAT THERE ARE

SOME NATURALLY FLUORESCENT

MOLECULES AND SO WHEN YOU GO

HIGHER AND HIGHER IN

CONCENTRATION, THOSE INTENSE

WHITE SPOTS ARE JUST PICKING

THE COMPOUND UP.

AND SO THEY PRESENT SOME

CHALLENGES IN THIS ASSAY

BECAUSE, IN FACT, IF YOU

HAVE YOU HAD AN INHIBITOR

WHERE YOU ARE LOOKING FOR

MAINTAINING A LOW LEVEL OF

FLUORESCENCE BASED ON

INHIBITING, IN OUR CASE THE

D.N.A. POLYMERASE, THAT

THOSE ARE GOING TO MASK

INHIX INHIBITION BECAUSE

THEY ARE NATURALLY

FLUORESCENT.

AT THIS POINT WE HAVE NOT

TRIED TO GO IN AND DO

INDIVIDUAL ASSAYS ON THOSE

COMPOUNDS, WHICH ARE NATURALLY

FLUORESCENT IN THE WAVE

LENGTHS THAT WE'RE MONITORING

TO SEE IF THEY HAVE ANY

INHIBTRY ACTIVITIES.

BUT IT DOES PRESENT A

CHALLENGE WITHIN SOME OF

THESE LIBRARIES, WHERE YOU

HAVE THAT PROPERTY.

>> IS IT OKAY TO ASK A BRIEF

FOLLOWUP?

ABOUT WHAT FRACTION OF THE

LIBRARY -- AS BEING THE

NATURALLY FLUORESCENT IN THE

--

[INDISCERNABLE]

>> I DON'T HAVE A SPECIFIC

NUMBER.

I WOULD IMAGINE IT'S

SOMEWHERE IN THE RANGE OF

LESS THAN FIVE PERCENT,

THOUGH.

IN LOOKING AT THESE PLATES.

THAT WOULD BE A QUESTION I

WOULD JUST SHALL -- IF YOU'D

LIKE TO SEND AN EMAIL, I'D

BE MORE THAN I THINK FOLLOW

UP WITH THE GROUP AT NIECG

TO SEE IF THEY HAVE A NUMBER

LET'S SAY WHERE THE

FLUORESCENT ASSAY WOULD BE

BEING INHIBIT ORDER

UNINTERPRETABLE BASED ON THE

NATURAL FLUORESCENCE OF A

COMPOUND DOWN AROUND ONE

MICROMOLAR BECAUSE THAT'S

ABOUT THE SORT OF THE

MINIMUM THRESHOLD ONE TO TEN

MICROMOLAR WHERE ONE WOULD

TAKE A LOOK AT A SECOND

COMPOUND BUT I DON'T HAVE

THOSE NUMBERS AT MY DISPOSAL

AT THIS TIME.

>> THANK YOU.

>> OKAY.

THANK YOU.

LET'S GO TO ANN HARBOR,

MICHIGAN.

>> APPRECIATE THE NICE TALK.

LATE HOURKo WE ARE --

[INDISCERNABLE]

 APPRECIATE THE APPROACH OF

TAKING BASIC SCIENCE AND --

[INDISCERNABLE]

 THANK YOU VERY MUCH.

>> THANK YOU.

VERY GOOD.

IS THERE ANYONE THERE AT

BROOK HAVEN?

>> HI, KEN.

THIS IS PAUL WILSON.

ALWAYS GOOD TO HEAR WHAT'S

GOING ON IN OREGON BUT WE

DON'T HAVE ANY QUESTIONS FOR

THE SPEAKERS.

>> OKAY.

THANK YOU.

PLEASE MUTE.

I HAVE JUST ONE QUESTION.

IT'S VERY INTERESTING.

I WAS INTRIGUED BY THE WORK

WITH THE VARIANT CELLS.

WE SEE PATIENTS HERE WITH

THAT DISORDER AND SO

SOMETHING HE IS IS GIVING

YOU THE MUTATIONS AND THE

CANCER.

WOULD IT BE WORTHWHILE TO

THINK ABOUT INHIBITING POLL

KAPPA, WHICH MIGHT BE THE

MAIN -- OR IS THERE EVIDENCE

THAT THAT IS THE MAIN AREA

THAT'S GIVING YOU THE

MUTATION THAT'S LEAD TO THE

DMAERNS THESE PATIENTS?

WHAT ADVANTAGE OF PATIENTS

WITH SKIN CANCER IS THAT YOU

DON'T HAVE THAT BLOOD-BRAIN

BARRIER IF YOU ARE CONCERNED

ABOUT APPLYING THE COMPOUND.

>> MY UNDERSTANDING WAS THAT

A LIKE -- THE LIKELY

CANDIDATES ARE GOING TO BE

IOTA, THE POLL ZETA.

I HAVE NOT SEEN IN TERMS OF

THE MUTTOGENESIS ASSOCIATED

WITH THAT POLL KAPPA.

WE'D BE MORE THAN HAPPY TO

TALK MORE ABOUT IT.

>> OKAY, THANK YOU.

IS THERE ANYONE ELSE WHO HAS

QUESTIONS NOW?

IF NOT, I WANT TO THANK YOU

VERY MUCH FOR EXCELLENT

PRESENTATIONS.

[APPLAUSE]

AND HOPING THAT YOU FOLKS

WOULD GO TO THE WEBSITE SO

THAT PEOPLE CAN SEE ALL THE

RESOURCES.

THANK YOU VERY MUCH.