>>WELCOME TO NIH WEDNESDAY
AFTERNOON LECTURE SERIES
IT'S REALLY A GREAT PLEASURE TO
INTRODUCE DR. ANDREA CALIFANO TO
YOU.
I'VE KNOWN ANDREA BECAUSE OF A
JOINT INTEREST IN BOTH GENOMICS
AND EARLY ON HIS INTEREST IN D
CELL LYMPHOMAS, NOW INTERESTED
IN ALL CANCER, AND HE'S GOING TO
BE DISCUSSING SYSTEMS BIOLOGY
NETWORK-BASED METHODS FOR
IDENTIFYING THERAPEUTIC TARGETS.
I THINK HIS FOCUS IS TO USE
THESE GENOMIC CYST TIM'S BIOLOGY
WAYS TO ACTUALLY DO NEW CLINICAL
TRIALS WHICH HE'LL TELL US
ABOUT.
OUR SPEAKER IS EXTREMELY
WELL-KNOWN WITHIN THE CANCER
COMMUNITY AND MORE GENERALLY IN
SYSTEMS BIOLOGY COMMUNITY.
HE IS THE CLYDE AND HELEN WU
PROFESSOR OF CHEMICAL SYSTEMS
BIOLOGY, CHAIR OF THE DEPARTMENT
OF SYSTEMS BIOLOGY AND DIRECTOR
OF THE J.P.SULZBERGER COLUMBIA
GENOME CENTER AT THE COLUMBIA
UNIVERSITY MEDICAL CENTER.
ORIGINALLY A PHYSICIST BY
TRAINING AND HE'S TAKEN IN MY
VIEW QUITE INNOVATIVE APPROACHES
TO IDENTIFYING LIKELY FACTORS
INVOLVED IN CANCER PROGRESSION,
CANCER PHENOTYPES IN GENERAL,
DRUG RESPONSE AND RESISTANCE,
AND MORE RECENTLY I THINK AS
HE'LL TELL YOU ABOUT TODAY,
USING SINGLE CELL DATA FROM
CANCER PATIENTS TO PREDICT
PATHWAY ACTIVATION.
THIS INVOLVES A LOT OF MATH,
WHICH ANDREA IS VERY GOOD AT,
AND IT INVOLVES INTEGRATING INTO
NETWORK MODELS TO MAKE A TOTAL
MODEL OF THE BEHAVIOR OF AT
LEAST PARTS OF THE CANCER CELL.
HE'S SHOWN I THINK HIS MAJOR
CONTRIBUTION IN MY VIEW TO
CANCER BIOLOGY LITERATURE AS THE
IDEA OF THE MASTER REGULATOR.
WHICH IS BASICALLY THESE ARE
NODES WITH -- REGULATORY NODES
WITHIN CANCER CELLS THAT HE CAN
IDENTIFY COMPUTATIONALLY THAT
INTEGRATE A LOT OF SIGNALS AND
THEN HAVE OUTPUT TO MANY
DOWNSTREAM TARGETS AND THAT
THESE ARE CALLED MASTER
REGULATORS THAT IF YOU INTERFERE
WITH THESE, THEN THEY HAVE SUCH
A BROAD EFFECT ON THE CANCER
CELL THAT TYPICALLY THAT INDUCES
CELL DEATH OR A MAJOR CHANGE IN
PHENOTYPE.
AND THIS THEN LEADS OBVIOUSLY TO
A NEW SET OF THERAPEUTIC TARGETS
IN CANCER THAT CAN BE ADDRESSED
BY A NUMBER OF MEANS,
PHARMACOLOGICALLY.
MANY OF THESE ARE TRANSCRIPTION
FACTORS SO THEY MAY REQUIRE
THINGS LIKE PROTACS TO BE DONE
TO BE TARGETED.
SO JUST A LITTLE BIT MORE
DETAIL, HIS UNDERGRADUATE DEGREE
AND PH.D. WAS IN PHYSICS AT THE
UNIVERSITY OF FLORENCE IN ITALY,
HIS PH.D. WAS ON THE STUDY OF
CHAOTIC BEHAVIOR IN HIGH
DIMENSIONAL SYSTEMS, HAVING READ
THE BOOK "CHAOS," THIS DOES NOT
MEAN THAT HIS DISSERTATION WAS
CHAOTIC, BUT IT WAS A TYPE OF
BROAD ANALYSIS OF COMPLEX
BEHAVIOR OF DYNAMICAL SYSTEMS
THAT I THINK IS A GOOD WAY TO
THINK ABOUT HOW BIOLOGICAL
SYSTEMS MIGHT ACTUALLY WORK.
HE THEN MOVED ON TO POSTDOCTORAL
TRAINING IN FLORENCE, MIT AND
FINALLY SPENT QUITE SOME YEARS
AT THE IBM WATSON RESEARCH
CENTER, IN THE 1980s, BECOMING
A STAFF MEMBER THERE, AND THEN
MOVED ON TO COLUMBIA BACK INTO
FULL ACADEMIA, SHALL WE SAY,
ALTHOUGH I THINK HE WAS DOING
ACADEMICS AT THE WATSON RESEARCH
CENTER.
HE MOVED IN 2003 AND EVENTUALLY
BECAME PROFESSOR OF BIOMEDICAL
INFORMATICS, AND NOW AS I
MENTIONED, DIRECTOR OF THE
SALZBERGER COLUMBIA GENOME
CENTER.
HE'S A VERY ENERGETIC GUY IF YOU
GET TO KNOW HIM, AND HE'S
FOUNDED SEVERAL COMPANIES SUCH
AS FIRST GENETIC TRUST AND
THERASIS.
HE'S GOT MANY HONORS, VERY
RIGHTFULLY IN MY OPINION HAS
BEEN AWARDED THE NCI OUTSTANDING
INVESTIGATOR AWARD, HE RECEIVED
THE RUTH SIEGEL AWARD IN
PANCREATIC CANCER RESEARCH,
MEMBER OF THE IBM ACADEMY OF
TECHNOLOGY, AND NOTABLY OF THE
NATIONAL ACADEMY OF MEDICINE.
THE TIE TITLE OF THE TALK IS
ELUCIDATION AND PHARMACOLOGICAL
TARGETING OF NON-ONCOGENE
DEPENDENCIES AT THE SINGLE-CELL
LEVEL.
ANDREA, IT'S A PLEASURE TO HAVE
YOU COME TO THE NIH, AND I'M
ALWAYS EXCITED TO LEARN FROM
YOUR LECTURES, SO WE'RE REALLY
LOOKING FORWARD TO YOUR
PRESENTATION.
THANKS A LOT.
>> THANK YOU FOR THAT WONDERFUL
INTRODUCTION.
I ALSO WANT TO THANK YOU AND
MICHAEL GOTTESMAN FOR INVITING
ME TO GIVE THIS LECTURE.
I THINK THE INVITATION CAME IN
MARCH 2020, AND THEN WE KNOW
WHAT HAPPENED, SO I REALLY AM
GLAD WE'RE FINALLY ABLE TO DO IT
EVEN IF NOT IN PERSON.
THE REASON I HAVE THESE LOGOS
HERE IS BECAUSE MY ENTIRE FIELD
WOULD NOT HAVE DEVELOPED CANCER
SYSTEMS BIOLOGY WITHOUT
TREMENDOUS EFFORTS BY THE NIH
AND NCC AND HOW THE NCI CAN
REALLY FOSTER OF KIND OF STUDIES
THAT I'LL BE DISCUSSING TODAY.
YOU'VE SEEN MY CONFLICT OF
INTEREST ALREADY.
THIS IS THE LATEST COMPANY I
STARTED IN 1916 THAT LICENSED
SOME OF THE ALGORITHMS I'LL BE
DISCUSSING TODAY.
SO IT ALL STARTED FROM WHAT WE
HAVE COME TO CALL THE CANCER
PARADOX.
AND THIS WAS PUBLISHED IN NATURE
REVIEWS, CANCER, AND BASICALLY
THE PARADOX STATES SOMETHING
QUITE OBVIOUS THAT A LOT OF US
HAVE KNOWN FOR AGES, BUT THE
DIFFERENCE HERE THAT WE ACTUALLY
TRY TO FIND POTENTIAL
COMPUTATIONAL SOLUTION TO
ADDRESS THIS PROBLEM.
SO THE PARADOX IS THE FACT THAT
IF YOU LOOK AT THE
TRANSCRIPTIONAL STATE OF CANCER
CELLS, TUMORS BREAK DOWN INTO
SUBTYPES THAT ARE SO TIGHTLY
REGULATED THAT THEY'RE NO MORE
DIFFERENT THAN EQUIVALENT NUMBER
OF SAMPLES TAKEN FROM NORMAL
TISSUES OF THE SAME NUMBER OF
INDIVIDUALS.
SO THE REVIEWERS FROM THAT
PERSPECTIVE DIDN'T BELIEVE THAT
STATEMENT, SO THEY ASKED US TO
GENERATE -- I'M JUST SHOWING YOU
FOUR TUMOR TYPES, AND YOU CAN
SEE OF COURSE BLUE IS TUMOR, RED
IS NORMAL TISSUE, AND YOU CAN
SEE BASICALLY THE DISTRIBUTION
LOOKS VERY SIMILAR.
SO TRANSCRIPTIONALLY, THE CELLS
ARE VERY, VERY SIMILAR.
AND YET AT THE GENETIC LEVEL,
THE MUTATIONS ARE LITERALLY ALL
OVER THE PLACE, SO -- IN TRIPLE
NEGATIVE BREAST CANCER, THERE'S
REALLY NO TWO PATIENTS THAT HAVE
THE SAME SET OF MUTATIONS.
SO WE ARGUED FOR A WHILE ON HOW
WE COULD POTENTIALLY FIND A
SOLUTION TO THIS PROBLEM THAT
WOULD EXPLAIN EX-WHY THAT EXPLAIN WHY THA T IS AND WE
WENT BACK TO -- ONLY ONE
POSSIBLE WAY IN WHICH THIS COULD
BE HAPPENING IS IF ALL THE
MUTATIONS OCCUR UPSTREAM OVER A
PIECE OF MACHINERY THAT IN
CANCER WE CALL THE TUMOR
CHECKPOINT IN FELON CANCER
TISSUE WE CALL THE REGULATORY
CHECKPOINT MODULE, WHERE A VERY
SMALL NUMBER OF PROTEINS WE
DUBBED MASTER REGULATORS, ARE
ACTUALLY INTEGRATING THE
ABERRANT SIGNALS THAT COME FROM
THE MUTATIONS BUT ALSO FROM BOTH
THE DISTAL AND TUMOR ENVIRONMENT
RELATED SIGNALS.
THEN THESE PROTEINS ESSENTIALLY
ARE INCREDIBLY TIGHTLY
OVERREGULATED AND THEREFORE
THEIR STABILITY IS DETERMINED BY
THEIR INTERACTIONS, AND TOGETHER
THEY REGULATE THE
TRANSCRIPTIONAL STATE OF THE
CANCER CELL.
SO THIS PARADIGM WHICH ARE
CALLED AN ONCOTECH TOUR
BASICALLY STATES WHETHER IT'S
REALLY OCCURRING IN CANCER IS
NOT THAT MUCH THE MUTATIONS BUT
RATHER -- THIS IS NOT JUST IN
CANCER BUT ALSO IN COMMON -- BUT
RATHER THE ARCHITECTURE, THE
SIGNALING AND TRANSCRIPTIONAL
ARCHITECTURE THAT IS RESPONSIBLE
FOR THEIR INTEGRATION AND THEIR
TRANSLATION INTO A
TRANSCRIPTIONAL STATE OF THE
CELL.
SO WE HAVE PUBLISHED EXTENSIVELY
ON THESE POINTS AND BASICALLY
USED THE ALGORITHMS TO MAKE
PREDICTION AND GOING TO THE LAB
AND MAKING EXPERIMENTAL
VALIDATION ASSAYS THAT REALLY
DEMONSTRATED THIS PARADIGM IS
ACTUALLY -- HOLDS TOGETHER
PRETTY MUCH EVERY TUMOR TYPE
THAT WE'VE LOOKED AT AT THE
MOLECULAR LEVEL AND SORT OF
MECHANISTIC, AND I THINK I'LL
TRY TO EXPLAIN WHY WE CALL THIS
KIND OF A QUASI MECHANISTIC
MODEL.
THE MORE IMPORTANT THING IS THAT
THESE STUDIES HAVE NOW YIELDED
QUITE A LARGE NUMBER OF CLINICAL
TRIALS AND CLINICAL STUDIES,
CLINICAL END POINTS THAT ARE
CURRENTLY RUNNING, SOME OF THEM
ARE COMPLETED AND I'LL SHOW YOU
THE RESULTS, AND OTHERS WILL
START -- THEY'RE NOT PRESENTED
HERE BUT THEY'LL START IN THE
NEXT FEW MONTHS.
SO ONE OF THE NICE THINGS ABOUT
HAVING A MODEL, HAVING THIS
ONCOTECH TOUR IS WE CAN NOW
START DEVELOPING ALGORITHMS
WHERE WE'RE FIRST GOING TO
RECONSTRUCT THIS ARCHITECTURE
AND WE KNOW IT'S DIFFERENT IN A
WAY THAT IS COMPLETELY
CONTEXT-DEPENDENT SO THEONICS
TEXTURE OF A B CELL IS
COMPLETELY DIFFERENT FROM
THEONICS TECH TOUR OF GLIOMA
CELLS, SO WE USE THIS
METHODOLOGY AND ALL OF THEM HAVE
BEEN EXTENSIVELY VALIDATED
EXPERIMENTALLY, ALL OF THEM HAVE
A 70 TO 80 PERCIVAL DAITION RATE
IN THE EXPERIMENTAL LAB AND THE
FIRST ONE IS AN ALGORITHM THAT
ALLOWS YOU TO TURN A LARGE
NUMBER OF GENE EXPRESSION
PROFILES INTO A FAIRLY ACCURATE
MAP, ABOUT 70 PERCIVAL DAITION
RATE OF THE UNDERLYING
TRANSCRIPTIONAL REGULATION
MECHANICS.
THEN THIS VIPER USES
INTERACTOMES TO MEASURE QUITE
PRECISELY THE TRANSCRIPTIONAL
ACTIVITY OF PROTEINS.
IN THE UPPER PART OF THAT HOUR
GLASS BOTTLE TO IDENTIFY QUITE
EFFECTIVELY ALL THE PROTEINS
THAT HAVE A POTENTIAL MODULATORY
ROLE WHERE BASICALLY THEIR
ACTIVITY OR ABUNDANCE CAN
MODULATE THE ABILITY OF ONE OF
THESE MASTER REGULATORS TO
REGULATE ITS TARGETS AND THEN
FINALLY, FRAPPE WAS THE FIRST IN
DEVELOPING IN COLLABORATION WITH
BARRY TO ACTUALLY PREDICT
PROTEIN-PROTEIN INTERACTION
USING STRUCTURAL BIOLOGY DATA.
LET ME SPEND A COUPLE MINUTES
ON -- BECAUSE IT'S THE WORKFORCE
OF THE LAB AND A LOT OF THE
THINGS I'LL WILL SHOW YOU ARE
BASED ON IT, SO IF I DON'T
EXPLAIN THESE IN A WAY THAT IS
UNDERSTOOD, YOU MAY MISS THE
REST OF THE TALK.
THE WAY IT WORKS IS AKIN TO A
HIGHLY MULTIPLEX GENE REPORTED
ASSAY.
IF YOU USE A -- PROMOTER AND
THEN MEASURE LUMINESCENCE, IF
YOU HAVE LUMINESCENCE AND
RESPONSE TO MAKE -- IF YOU SEE
NONE -- THAT WORKS WELL IF MIC
IS THE ONLY THING THAT MOVES IN
THE CELL, OBVIOUSLY IN RESPONSE
TO OTHER TRANSCRIPTIONAL
REGULATORS, IT WILL NOT WORK
WELL IF EVERYTHING MOVES.
SO WHAT WE TRIED TO DO IS TO
GENERALIZE THIS CONCEPT OF A
GENE REPORTER ASSAY SO THAT WE
NOW CAN USE ABOUT 100 TARGETS OF
EVERY REGULATORY PROTEINS BOTH
POSITIVELY REGULATED AND
REPRESSED AS THE REPORTER ASSAY.
SO FROM THE EXPRESSION OF THESE
TARGETS, ACTUALLY FROM THE
DIFFERENTIAL EXPRESSION OF THESE
TARGETS IN TWO DIFFERENT STATES
OF THE CELL, WE CAN MEASURE THE
DIFFERENTIAL ACTIVITY OF ANY
REGULATORY PROTEINS IN THOSE TWO
COMPLEX.
SO YOU COULD USE TWO ISOGENIC
CONTEXT, ONE WITH -- DRUG
SENSITIVE VERSUS DRUG RESISTANT
AND WE'VE DONE ALL THE STUDIES
AND PUBLISHED THEM AND IN EVERY
SINGLE CASE, WE'VE BEEN ABLE TO
IDENTIFY THE MASTER REGULATORS
AND VALIDATE THEM
EXPERIMENTALLY.
BUT ALSO AS YOU'LL SEE, EVEN IN
VERY EARLY PAPERS, WE SHOW THAT
WE CAN ACTUALLY REPROGRAM THE
STATE OF THE CELL BY TARGETING
THE MASTER REGULATORS BECAUSE
THESE ARE ESSENTIALLY THE
PROTEINS THAT MECHANISTICALLY, I
SHOULD SAY QUASI MECHANISTICALLY
BECAUSE ONLY 70% OF THEIR
TARGETS ARE CORRECT, BUT
MECHANISTICALLY REGULATE THE
STATE OF THE CELL.
SO THERE ARE THREE MAIN HIGH
POTASSIUM THESE THAT
HYPOTHESES THAT STEM FROM THIS
ARCHITECTURE, MAJOR HYPOTHESES
THAT WE HAVE POSTULATED.
THE FIRST ONE IS THAT MASTER
REGULATORS FORM THESE TUMOR
CHECKPOINTS, IN CANCER, WE CALL
THEM REGULATORY CHECKPOINTS WHEN
IT'S NOT IN CANCER, RESPONSIBLE
FOR -- THEY CAN ESSENTIALLY
ADAPT TO ANY PERTURBATION AND
MAINTAIN THE STATE LARGELY
INDEPENDENT OF THESE
PERTURBATIONS.
THESE ARE PAPERS WHERE THAT HAS
BEEN SHOWN, THERE'S MORE -- I'M
JUST SHOWING SOME EXAMPLES.
THE SECOND ONE IS THAT THE
MASTER REGULATORS IN THESE TUMOR
CHECKPOINT MODULES CANNIZE THE
EFFECT OF MOST FUNCTIONAL
MUTATIONAL EVENTS.
THESE PAPERS, PROBABLY BRINGS UP
MEMORIES TO LOU.
THIRD IS MASTER REGULATORS IN
TUMOR CHECKPOINT MODULE
REPRESENT A NEW CLASS OF
PHARMACOLOGICALLY ACTIONABLE
NON-ONCOGENE DEPENDENCIES.
THE PROTEINS THAT ARE -- AND
ALMOST NEVER HARBOR MUTATION
THAT TURN THEM INTO
ESSENTIALLY -- ACTIVE PROTEINS
BECAUSE OF THE MUTATION.
SO LET ME SHOW YOU A CASE STUDY
IN PANCREATIC CANCER.
THIS HAS BEEN LED BY PASS QAL
LAISE IN MY LAB.
TWO OF THE HYPOTHESES ARE
ESSENTIALLY DISCUSSED IN MY DATA
HERE.
PANCREATIC CANCER HAS BEEN
CHALLENGING TO CLASSIFY IN
STRUCTURE BECAUSE IF YOU LOOK AT
THREE DIFFERENT STUDIES DONE
FROM GENE EXPRESSION PROFILES,
THE CLASSES THAT ARE GENERAL
LATED HAVE NO OVERLAP BETWEEN
THE THREE OF THEM, SO YOU CAN
SEE HERE THAT ACCORDING TO FOUR
DIFFERENT STUDIES IN TUMOR AND
STROMA -- OR IN THE TUMOR CELLS,
THERE WERE ZERO OVERLAPPING
GENES THAT WILL DETERMINE THE
IDENTITY OF THE SUBTYPES
IDENTIFIED BY EACH STUDY.
AND WE THOUGHT THAT MAYBE THIS
WAS BECAUSE, ONE, THERE IS
TREMENDOUS STROMAL INFILTRATION,
IN FACT, SOMETIMES THE TUMOR
CELLS ARE ACTUALLY A MINORITY,
OR EVEN A SMALL MINORITY OF ALL
THE CELLS IN THE TUMOR, AND
NUMBER TWO, BECAUSE WITHIN THE
TUMOR ITSELF, THERE MAY BE
MULTIPLE COMPARTMENTS OF CELLS
THAT HAVE STRIKINGLY DISTINCT
MOLECULAR IDENTITIES.
SO TO DO THIS, WE RESORTED TO
STUDY PANCREATIC CANCER AT
SINGLE CELL LEVEL.
I WANT TO SHOW YOU WHAT THE
CHALLENGE IS WHEN YOU GO TO
SINGLE CELLS.
THIS IS DAY TAI DATA PUBLISHED RECENTLY
IN CELL CARCINOMA, IN
COOPERATION WITH CHUCK DRAKE,
PH.D. STUDENTS IN OUR LAB, WHERE
BASICALLY WE TOOK SINGLE CELLS
FROM THESE TUMORS, -- AND THEN
PERFORMED ANALYSES USING EITHER
A CYTEK WHICH USES A PANEL OF 19
MYELOID AND 19 LYMPHOID
ANTIBODY, I'M ONLY SHOWING THE
LYMPHOID HERE, YOU CAN READ THE
PAPER FOR THE OTHER ONES, THEY
BASICALLY TELL THE SAME STORY,
AND THOSE IDENTIFIED FOUR
POPULATIONS THAT YOU WOULD
EXPECT TO FIND IN THESE CELLS.
HOWEVER, IF YOU TAKE THE SAME
CELLS AND SEND THEM TO CHROMIUM
AND YOU LOOK TO THE GENES THAT
ENCODE FOR THESE PROTEINS, YOU
SEE ABSOLUTELY NOTHING.
ESSENTIALLY THERE'S COMPLETE
LOSS OF THE MRNAs THAT ENCODE
FOR THOSE PROTEINS AND,
THEREFORE, YOU CANNOT --
ANYTHING.
HOWEVER, WHEN YOU NOW RUN VIPER
ON THESE DATA, NOT ONLY YOU CAN
CONSTITUTE BASICALLY THAT WAS
FOUND BY ANTIBODIES, THE
STANDARD VERSUS THE MEAN BECOMES
DRAMATICALLY BETTER WITHIN THE
SUBTYPES, SO CD4 IN THIS CASE,
THE ANTIBODY LEVEL, CD14 FOR
MONOCYTE MACROPHAGES, AGAIN,
VERY NOISY, YOU CAN'T SEE IT IN
THE ANTIBODIES -- FOR THE
CD4 CELLS WHICH IS VERY NOISY
AGAIN.
AND THERE ARE ALSO THINGS LIKE
CD19 WHICH WE DIDN'T DETECT
WELL, WHICH MEANS WE HAVE TO
REFINE -- PROPOSAL WE HAVE FOR
THE NEXT FEW YEARS ADDRESS THAT
QUESTION EXPERIMENTALLY BY
COMBINING -- WITH SINGLE CELL
SILENCING DATA.
SO BASICALLY WE CAN NOW USE
VIPER TO TURN SINGLE CELL DATA
ALMOST LIKE IF IT WERE BULK
TISSUE, SO WE USE DATA FROM A
PRIOR PUBLICATION FROM TUVESON
WHERE WE FOCUS ON THE
FIBROBLASTS, MULTIPLE DIFFERENT
TYPES THAT ARE
IMMUNOSUPPRESSIVE, YOU CAN SEE
HOW SMALL IT IS COMPARED TO ALL
THE CELLS THAT WE IDENTIFIED, WE
VALIDATED THESE WERE ACTUALLY
TUMOR CELLS AND THEN PERFORMED
OUR ANALYSES PERFORMING VIPER
AND CLUSTER, BUT THE CLUSTERING
WAS DONE NOT BY GENE EXPRESSION
PROFILING BUT RATHER BY PROTEIN
ACTIVITY.
WHY IS THAT IMPORTANT?
BECAUSE IF YOU DID THE CLASS --
EXPRESSION YOU WOULD NOT SEE
THESE THREE POPULATIONS, IN
FACT, FOUR OF THEM, SO PROTEIN
ACTIVITY GENERATED FOUR
SUBTYPES, POPULATION OF SINGLE
CELLS WITH A VERY STRONG SCORE
THAT IS DRAMATICALLY IMPROVING
GENE EXPRESSION.
YOU CAN SEE WE DID ADDITIONAL
ANALYSES, SO THESE ARE THE
DIFFERENT COLORS, BUT YOU CAN
SEE THAT WITHIN THIS CLUSTER OF
GREEN CELLS, THERE WERE ACTUALLY
THREE PEAKS.
SO WE NOW DO THIS PEAK ANALYSIS
BECAUSE ESSENTIALLY FROM THE --
DISTRIBUTION -- DENSITY OF THE
CELL CORRESPONDS TO A STABLE
STATE, AND THE STABLE STATE ARE
VERY IMPORTANT BECAUSE THE CELLS
ARE NOT UNDER PEAKS.
THEY'RE ESSENTIALLY MOVING VERY
RAPIDLY FROM ONE STABLE -- TO
ANOTHER BUT THEY EVENTUALLY WILL
GET THERE OR THEY HAVE TO
START -- THE ONES IN STABLE
STATE TEND TO PERSIST FOR A LONG
PERIOD OF TIME SO THOSE ARE THE
ONES YOU WANT TO TARGET
PHARMACOLOGICALLY.
DIFFERENTIATE FOR -- STATE AND
THIS IS A VERY INTERESTING --
PROGENITOR STATE.
WHEN YOU LOOK AT THE -- YOU CAN
SEE YOU HAVE LINEAGE, APS AND
MOS CONVERGING ON THE DIFFERENT
STATE AND THIS IS CONSISTENT
WITH THE VIEW BY -- OF THIS
POPULATION WITH THE APS BEING
THE MOST STEM LIKE AND LIN
AND -- WE LOOK AT THE PRESENCE
OF THESE POPULATION WITHIN
DIFFERENT PATIENTS AND YOU CAN
SEE HERE THAT ESSENTIALLY ALL
THE PATIENT PRESENTED AT LEAST
THREE OF THE FOUR AND IN SOME
CASES ALL FOUR OF THEM, THE APS
ARE MORE RARE, CONSISTENT WITH
THE FACT THAT PROBABLY MORE
STEM-LIKE POPULATIONS OR IN SOME
PATIENTS MAYBE WE HAVEN'T SEEN
THIS BECAUSE WE DIDN'T SEQUENCE
ENOUGH CELLS.
BUT THEN MORE IMPORTANTLY, IF
YOU LOOK AT THE BIOLOGY OF THE
CELLS, YOU CAN SEE THAT THERE
ARE REALLY MARKERS THAT ARE
HIGHLY DISTINCT.
THESE ARE MARKERS THAT YOU CAN
ACTUALLY USE TO -- THE CELLS AND
DIVIDE THE SUBPOPULATION THAT
DISTINGUISH THESE FOUR DIFFERENT
SUBPOPULATIONS, AND JUST AS A
SUMMARY HERE, YOU CAN SEE THAT
THIS POPULATION HERE ARE VERY,
VERY SMALL NUMBER OF CELLS IS
ACTUALLY EXTREMELY DISTINCT AND
THESE ARE -- THREE OF THESE
MARKERS ARE CLASSICAL MARKERS OF
ADM, YOU CAN SEE HERE
REPRESENTED -- AND THESE CELLS
THE ONE CUT WHICH YOU CAN SEE
HERE BASICALLY ACTIVATED ONLY
THE CELLS AND THE SOX 9.
SO THIS ANALYSIS REALLY ALLOWS
YOU TO DISCOVER BIOLOGY THAT
THEN CAN BE VALIDATED THAT WE
MAY HAVE COMPLETELY MISSED IF
YOU WERE TO USE GENE EXPRESSION
PROFILE.
WE THEN WENT TO TWO -- OF SINGLE
CELLS.
CSY.
YOU CAN SEE SINGLE CELLS FROM
THIS DATASET COMPLETELY
VALIDATED THE STRUCTURE IN THE
FOUR CLUSTERS WITH THE
DIFFERENTIATED PROGENITOR CELLS
FORMING THE MAJORITY OF THE
CELLS AND THEN LINEAGE --
REPRESENTING THE REST.
AND THIS WAS VERY HELPFUL
BECAUSE IN EVERY SINGLE STUDY
WE'VE DONE, WHEN WE DO THESE
ANALYSES AT THE PROTEIN LEVEL AS
HE OH POSE TODAY GENE EXPRESSION
LEVEL WE FIND TREMENDOUS
CONSISTENCY IN DIFFERENT
DATABASES.
SO THESE WERE 35 PATIENTS,
21,000 SINGLE CELLS.
SO THEN WE WENT TO THE BULK
COHORT AND ASK IF WE CAN
RECAPITULATE SOME OF THESE --
SUBTYPES SO IF YOU ACTUALLY JUST
DO STRATIFICATION BY BULK
PROFILE BASED PROTEIN ACTIVITY,
YOU SEE THAT BOTH ICGC WHICH IS
THE GREEN AND TCGAs 25
ACCORDING TO THESE PROTEINS HERE
AND AGAIN THESE WERE INCREDIBLY
ENRICHED IN THE MASTER
REGULATORS OF THE LINEAGE AND
MORPHOGENIC SUBTYPE.
HOWEVER, THE -- SIMPLY COULD NOT
BE IDENTIFIED IN ANY OF THE
DATASETS.
THIS WAS NOT A DATASET GENERATED
BY US, BY LASER MICRO RESECTION
IN CONNECTION -- 200 PANCREATIC
CANCER PATIENTS, BASICALLY YOU
CAN SEE THE PROTEINS YOU FIND IN
THE DATASETS ARE
DIFFERENTIATING -- ARE
INCREDIBLY OVERLAPPING IN TERMS
OF THEIR MASTER REGULATOR.
SO AGAIN, ALL THE OTHER SINGLE
CELL ACTIVITY MACRO FILE NOW
INSTEAD OF FINDING GENE
EXPRESSION PANELS THAT HAVE NO
OVERLAP, MASTER -- PANELS THAT
ARE VIRTUALLY IDENTICAL ACROSS
THESE TWO DIFFERENT TUMORS.
SO WE WENT TO CELL LINES BECAUSE
YOU NEED CELL LINES TO DO SOME
OF THE EXPERIMENTS SO WE
IDENTIFIED THAT IN A COLLECTION
OF EIGHT DIFFERENT CELL LINES,
WE COULD FIND SINGLE CELL
REPRESENTATIVE LINEAGE, DPS, APS
AND MORPHOGENIC SUBTYPES AND
THIS GIVES YOU THE IDENTITY OF
THE LINES, FOR INSTANCE -- WAS
PREDOMINANTLY LINEAGE AND DPS,
YOU CAN SEE ESSENTIALLY ALONG
THIS AXIS, WHILE KP4, VERY
AGGRESSIVE CELL LINE, WAS MOSTLY
MORPHOGENIC -- STORY, THIS
LINEAGE IN -- THIS IS
MORPHOGENIC IN DPS.
THEN WE DID LINEAGE BARCODE --
TRACING ASSAYS BECAUSE WE WANTED
TO KNOW WHETHER THE PLOT I
SHOWED YOU BEFORE BASED ON
ANTEROPEE WAS HOLDING --
DIFFERENTIATION SO WHETHER THE
PLASTICITY WAS GOING TWO WAYS OR
ONLY ONE WAY.
SO THIS IS IN KP4 CELL, ALSO
FOR -- TWO CELLS THAT ARE THEIR
LINEAGE, BASICALLY -- THESE ARE
BARCODES THAT YOU SEE AT TIME 1,
48 HOURS AFTER THE START, AND
ONLY IN THIS POPULATION, AND NOW
YOU CAN SEE THAT TIME .2, WHICH
IS ABOUT 25 DAYS, YOU SEE THAT
THESE BARCODE ARE REPROGRAMMED
TO THE DPS CELLS.
SO OBVIOUSLY WE CAN SEE THERE IS
TREMENDOUS -- BUT WHEN YOU
ACTUALLY LOOK AT THE OPPOSITE,
THESE BARCODES THAT WERE IN THE
DPS CELLS THEN FOUND IN THE
MORPHOGENIC CELLS, YOU ALSO SEE
THAT.
SO THIS SHOWS THESE CELLS ARE
REALLY SPON TAPE SPONTANEOUSLY
REPROGRAMMING -- DIFFERENT
KINETICS.
SO WE THEN MOVE TO SELECTING THE
THREE MOST MORPHOGENIC AND THE
THREE MOST LINEAGE CELLS --
COLLECTION OF PANCREATIC CANCER
CELLS, YOU CAN SEE THE THREE
THAT ARE MORPHOGENIC AND THE
THREE THAT ARE MORE
LINEAGE-LIKE, AND THEN WE
PERFORMED CRISPR I-BASED
SILENCING WITH THE LIBRARY OF
ALL THE TRANSCRIPTIONAL
REGULATORS AND COTRANSCRIPTION
FACTORS AND NEGATIVE CONTROL, OF
COURSE, AND THEN WE USED THIS
LIBRARY TO LOOK AT TWO THINGS.
THE FIRST ONE WE ASKED, ARE
THERE MASTER REGULATORS THAT --
THERE IS ACTUALLY
SUBTYPE-SPECIFIC.
SO WE GENERATED TO COMPARE THE
LINEAGE VERSUS MORPHOGENIC CELLS
AND WE ASKED WHETHER THEY WERE
ENRICHED -- KNOCKOUT AND BY
CRISPRI AND YOU CAN SEE STRIKING
ENRICHMENT, 10 TO THE MINUS 6 IN
TERMS OF THE P VALUE OF THE
ENRICHMENT OF THE PREDICTED
MASTER REGULATORS, IN ONE
LINEAGE VERSUS THE OTHER.
THEN WE ACTUALLY ASKED THERE ARE
SOME COMMON DEPENDENCIES THAT
CAN BE FOUND BY, FOR INSTANCE,
COMPARING BOTH MORPHOGENIC CELLS
TO THEIR NORMAL PANCREATIC
TISSUE COUNTERPART IN G TECH,
AND SO THIS IS A SIGNATURE OR
ESSENTIALLY A CANCER PROGRESSION
AND AGAIN THERE WAS REALLY A
STRIKING ENRICHMENT OF THE TOP
MASTER REGULATORS IN GENES THAT
ARE ESSENTIAL IN ALL THE SEQ
CELL LINES AS OPPOSED TO JUST
ONE SUBTYPE VERSUS THE OTHER.
SO THIS BASICALLY SUPPORTS THE
HYPOTHESIS OF THE NEW CLASS OF
NON-ONCOGENE DEPENDENCIES.
THEY'RE VERY RICH, I'LL GIVE YOU
A PAPER ON THAT, WHERE THIS HAS
BEEN SHOWN.
THE MORE INTERESTING THING IS TO
ACTUALLY SHOW THAT THESE GENES,
ACTUALLY THESE PROTEINS
MECHANISTICALLY CONTROL THE
STATE OF THE CELL.
TO DO THAT, WE ECTOPICALLY
EXPRESS THE MASTER REGULATORS OF
THE LINEAGE SUBTYPE IN KP4 CELLS
WHICH ARE STRONGLY MORPHOGENIC.
SO THIS IS THE SIGNATURE, THIS
IS THE ACTIVITY OF THE PROTEINS
IN THE KP4 CELLS, ON THE
SIGNATURE -- MORPHOGENIC VERSUS
LINEAGE, AND THIS IS THE
SIGNATURE OF THE LINEAGE
SUBTYPES.
SO YOU CAN SEE HERE THAT --
VIRTUALLY EVERY MASTER
REGULATORS SIGNIFICANT
ACTIVATION OF THE PROTEINS THAT
WERE INHIBITED, DOWNACTIVATED IN
THE MORPHOGENIC SUBTYPES BUT
THEY HAD A MIXED RESULT OF THE
PROTEINS THAT WERE ACTIVATED,
BUT ONE OF THEM ACTUALLY INDUCE
COMPLETE REPROGRAMMING SO THESE
CELLS AFTER INDUCTION LOOKED
EXACTLY LIKE LINEAGE CELL,
MORPHOGENIC CELL.
WE ACTUALLY HAD THE RNA SEQ
AFTER EACH ONE OF THESE ECTOPIC
EXPRESSIONS, YOU CAN ACTUALLY
RECONSTRUCT THE FULL
ARCHITECTURE OF THE MASTER REG
LATER MODULES AND YOU NOW
UNDERSTAND MECHANISTICALLY
WHY -- IS RESPONSIBLE FOR
DRIVING THAT TRANSITION
BECAUSE -- ACTUALLY REPRESSES
ALL THE MASTER REGULATORS OR A
LARGE NUMBER OF THE MASTER
REGULATORS OF THE MORPHOGENIC
LINEAGE AND ACTIVATE A LARGE
NUMBER OF MASTER REGULATORS OF
THE LINEAGE, AND THE ONES THAT
IT ALSO REGULATES INDIRECTLY
REGULATES THE ONES THAT ARE
MISSING.
SO TAKEN TOGETHER, THIS INDUCES
REALLY DRAMATIC REPROGRAMMING.
WE THEN ASK, IS THIS ACTUALLY
DUE TO THE -- WE DIDN'T SEE THE
REPROGRAMMING BECAUSE THE EFFECT
OCCURRED ONLY IN A SUBSET OF THE
CELLS, SOMETIMES WHEN YOU
REPROGRAM YOU HAVE LOW
EFFICIENCY SO WE REPEATED THESE
AT THE SINGLE CELL LEVEL AND WE
OVEREXPRESSED GENES WITH AN MY
OF 1 SO THAT SOME CELLS RECEIVED
MORE THAN ONE CDNA, SO YOU CAN
SEE HERE THAT ESSENTIALLY A LOT
OF DIFFERENT COMBINATIONS AND
SINGLE GENES ACTUALLY
STATISTICALLY SIGNIFICANTLY
REPROGRAM THE CELL BUT WITH
DIFFERENT EFFICACY AND
EFFICIENCY.
SO OL2 IS HERE, F1A DRAMATICALLY
INCREASES THE EFFICACY OF THE
REPROGRAMMING, AND THIS IS JUST
A SUBSET OF THE CELLS THAT WE
CALL REPROGRAMMED TO THE LINEAR
SUBTYPE.
SO THIS, I HOPE, DEMONSTRATES
THAT THESE PROTEINS ARE
RESPONSIBLE REALLY FOR HOMO
STATICALLY MAINTAINING THE STATE
OF THE CELLS AND BY GENETICALLY
INTERFERING WITH THEM, YOU CAN
REPROGRAM A CELL AS YOU WANT.
SO THIS WAS PUBLISHED IN CELL
ABOUT EIGHT MONTHS AGO, WHERE WE
ESSENTIALLY ASKED AND
GENERALIZED THESE TWO TUMORS,
CAN WE USE THE ONCO -- IGNORING
ANYTHING WE KNOW ABOUT SUBTYPES,
ANYTHING WE KNOW ABOUT TUMOR
TYPE, ET CETERA, OTHER THAN IN
USING OBVIOUSLY THE RIGHT
REGULATORY NETWORKS FROM A
TISSUE THAT IS MAPPED.
-- NOW AN INDEPENDENT
INVESTIGATOR AT BARCELONA,
AND -- FROM MY LAB, MARIANO IS
THE CHIEF SCIENTIFIC OFFICER, SO
WHAT YOU SEE HERE IS THAT
INSTEAD OF WHAT DOING WHAT WE
DID IN A CELL PAPER, GOING FROM
GENE EXPRESSION TO THE MASTER
REG A LITTLE TORE USING
REGULATOR -- BASICALLY IDENTIFY
ALL THE POSSIBLE MUTATION THAT
CAN AFFECT THE ACTIVITY OF THE
MASTER REGULATORS, WHAT WE DID
HERE, WE PUT THE MASTER
REGULATORS IN THE CROSSHATCH OF
BOTH -- GENETICS IN THE
DOWNSTREAM TRANSCRIPTIONAL
REGULATION.
THIS ALLOWED US TO IDENTIFY, TO
STRATIFY THE ENTIRETY OF TCJ1
SAMPLE AT A TIME, IN 112 TUMOR
SUBTYPES, AND THE FIRST TRACKING
THINGS WAS THAT WHEN YOU LOOK AT
EACH ONE OF THE TUMOR TYPE, ALL
OF THEM EXCEPT -- IDENTIFY
SUBTYPES THAT HAD STATISTICALLY
SIGNIFICANT DIFFERENTIAL
OUTCOME.
THIS HAS BEEN VERY
CHALLENGING -- IN PROSTATE
CANCER I WOULD CHALLENGE YOU TO
FIND ANY -- THAT IDENTIFIES -- J
SUBTYPES THAT HAVE DIFFERENT
OUTCOME.
THIS IS AN EXAMPLE FOR KIDNEY
CANCER, WHERE WE FOUND FIVE
SUBTYPES AND YOU LOOKED AT
S3 VERSUS 5, REALLY STRIKING
DITCH RENGS
DIFFERENTIAL OUTCOME.
THE OTHER -- BY PROTEIN
ACTIVITY, THESE ARE THE BLUE
CLUSTERS, THE BLUE -- PLOT, THIS
IS A SILHOUETTE SCORE THAT TELLS
YOU HOW TIGHT THESE CLUSTERS
ARE.
THE RED LINE, THE DOTTED LINE IS
BASICALLY THE LINE FOR
STATISTICAL SIGNIFICANCE.
ANYTHING ABOVE IS SIGNIFICANT,
THE RED PLOT, WHAT YOU GET
FROM -- EXPRESSION ANALYSIS.
SO THE CLUSTERING OF PCJ BY GENE
EXPRESSION REALLY DOES NOT
PRODUCE CLUSTERS THAT ARE VERY
TIGHT.
SO THIS IS KIND OF THE ONE SLIDE
IN TERMS OF SHOWING THAT THE
GENETIC IS UPSTREAM OF THE
MASTER REGULATORS, BECAUSE
BASICALLY WE ASKED THE QUESTION
AND WE ASK THIS QUESTION ONE
SAMPLE AT A TIME, HOW MANY
GENETIC ALTERATION THAT WE CAN
IDENTIFY IN THAT SAMPLE BY GI
STIC 2 AND CHASM CAN BE PLACED
UPSTREAM OF THE SAMPLE.
EACH COLOR LINE IS ONE OF THE
112 SUBTYPES.
OVARIAN CANCER WHERE THERE'S AN
ENORMOUS NUMBER OF PASSENGER
MUTATION AND STRUCTURAL EVENTS,
THE SATURATION WAS DOUBLY FAST,
BUT YOU GET TO EXPLAIN ABOUT
100% OF THE MUTATION WITH
SOMEWHERE AROUND 15 TO 25 MASTER
REGULATORS.
SO THIS BASICALLY IS A
SATURATION CURVE, TELLS YOU HOW
MUCH OF THE GENETICS IS UPSTREAM
OF HOW MANY OF THE MASTER
REGULATORS.
THIS SIDE IS VERY IMPORTANT
BECAUSE IT TELLS US THAT WHEN WE
WANT TO REALLY KILL A TUMOR AND
WE WANT TO DO THIS BY TARGETING
THE MASTER REGULATORS THAT ARE
BARELY ACTIVATED AS A RESULT OF
THE MUTATIONS IN THAT SAMPLES,
WE REALLY HAVE TO FOCUS ON
AVERAGE OF THE TOP 25 MOST
ACTIVATED AND THE 25 MOST
INACTIVATED.
IMPORTANTLY, THESE ANALYSES
IDENTIFY ONLY 407 MASTER
VALIDATORS AND WE EVALUATED 141
THAT ARE ACTUALLY SUBTYPE
ESSENTIAL.
35%.
A PRETTY STRIKING ENRICHMENT.
SO NOW I WANT TO TELL YOU HOW WE
TARGET THESE THINGS IN THE
CLINIC BEFORE SHOWING YOU HOW WE
DO IT AT THE SINGLE CELL LEVEL.
SO THIS IS A LARGE COLLABORATION
THAT'S WITHIN GOING
THAT'S BEEN GOING ON FOR SEVEN
YEARS, AND NOW -- PURSUING IT.
SO A THIS IS BASED ON TWO
ALGORITHMS.
THE FIRST ONE IS NEW YORK AND
CALIFORNIA DEPARTMENT OF HEALTH
APPROVED AND CLIA COMPLIANT.
THE FIRST ONE IS A REALLY VERY
SIMPLE MINDED ADAPTATION AND
EXTENSION OF THE ONCOGENE
ADDICTION POSTULATE THAT IS
ACTUALLY TUMOR MUTATION
AGNOSTIC.
HOW IS IT AGNOSTIC?
BECAUSE ALL WE KNEW IS RANKING
ALL THE PROTEINS IN A TUMOR FROM
THE MOST INACTIVATED TO THE MOST
BARELY ACTIVATED AND THEN WE
WALKED DOWN THIS LIST UNTIL WE
HIT ONE FOR WHICH A HIGH
AFFINITY PHARMACOLOGICAL
INHIBITOR ALREADY EXISTS.
SO ABOUT 20% OF METASTATIC
BREAST CANCER WE IDENTIFY AS
HDAC6 -- VERY HIGH INFINITY
INHIBITOR SO WE STARTED CLINICAL
TRIAL IN COLLABORATION WITH JOSE
SILVA AND -- NOW AT ST. JUDE AND
REALLY CAME FROM MY LAB, IN
BREAST CANCER.
WHAT YOU SEE HERE WAS REALLY
QUITE STRIKING.
BASICALLY EVERY SINGLE PATIENT
THAT RESPONDED ACTUALLY HAD
HIGHER -- ACTIVITY OF
HDAC6 COMPUTED BY VIPER ANALYSIS
COMPARED TO -- YOU CAN SEE
SIGNIFICANT RESPONSE, RESPONDERS
VERSUS NON-RESPONDERS AND THE --
CURVE WAS ESSENTIALLY 1.
NOT THAT THE STUDY IS GOING TO
SAVE PEOPLE'S LIVES NECESSARILY
YOU SEE RECURRENCE BECAUSE
YOU'RE TARGETING ONLY ONE
PROTEIN, BUT AT LEAST IT TELLS
YOU WE CAN PREDICT WITH VERY
HIGH ACCURACY WHO WILL RESPOND
CLINICALLY, NOT JUST IN A PDX
MODEL OR IN A CELL LINE.
SO THIS IS -- ARCHIVE AND IT'S
IN REVIEW.
SO THEN WE EXTENDED -- AND
BASICALLY NOW WE'VE DONE THIS
FOR EVERY COHORT NOT ONLY IN BCJ
BUT TARGET -- WE CAN THINK OF,
ESSENTIALLY WE RANK EVERY
PROTEIN THAT IS DRUGGABLE -- IT
IS THE MOST BARELY ACTIVATED
ONE.
SO FOR INSTANCE, WE FOUND THAT
EXPLORING ONE WHICH IS A TARGET
OF -- IS ACTUALLY REALLY BARELY
ACTIVATED IN -- TUMORS AND GERM
CELL TUMORS, IN FACT MORE SO
THAN IN ANY OTHER TUMOR COHORT.
SO YOU CAN SEE THE SENSITIVITY
IS 10 TO MINUS 8 MICROMOLAR
RANGE, THESE TUMORS WHEN
TRANSPLANTED RESPONDED
STRIKINGLY WELL, EE STENGSLY
COMPLETE RESPONSE TO THE DRUG
AND THEN SHOWING RELAPSE WHEN
YOU STOPPED ADMINISTERING THE
DRUG.
AND SO THESE ARE PATIENTS THAT
HAVE BEEN TREATED BASED ON THESE
FINDINGS, THIS IS A 70-YEAR-OLD
PATIENT THAT HAD A RECURRENT
LUNG TUMOR ESSENTIALLY FAILED TO
RESPOND TO PRETTY MUCH ANY
THERAPY THROWN AT IT,
NUMEROUS -- IN THE PERITONEAL
CAVITY AND REFUSED TO GET
ADDITIONAL CHEMO TOXIC THERAPY
BECAUSE OF SIDE EFFECTS.
HE WAS PUT ON -- AND ESSENTIALLY
HAS BEEN DOING SPECTACULARLY
WELL.
THERE'S BEEN NO DETECTABLE
LESION AFTER HIS LAST SURGERY,
AND HE'S BACK IN SCHOOL AND
PLAYING SOCCER.
THIS IS AN ULTRA-RARE TUMOR.
THERE'S ONLY 40 REPORTED CASES
OF THIS TUMOR, NOT PER YEAR IN, IN
TOTAL.
A 14-YEAR-OLD PATIENT PRESENTED
WITH THIS TUMOR, FAILED TO
RESPOND TO EVERY POSSIBLE
THERAPY, INCLUDING A THERAPY
MEANT FOR -- TUMORS THAT DID
ABSOLUTELY NOTHING FOR HIM, AND
ONCOTARGET FOUND THAT PDJ --
BETA WAS THE MOST -- ACTIVATED
PROTEIN AND -- ACTUALLY WELL
TOLERATED IN PATIENTS WAS THEN
SELECTED BY THE ONCOLOGIST TO
TREAT THE PATIENT AT MEMORIAL.
AND YOU CAN SEE REALLY THE
STRIKING RESPONSE AT THE LEVEL
OF THE IMAGING.
SO NOW LET ME TALK A LITTLE BIT
ABOUT THE MORE SOPHISTICATED
APPROACH.
SO I TOLD YOU THERE'S 25 MASTER
REGULATORS BARELY ACTIVATED, 25
THAT ARE INACTIVATED THAT
EXPLAIN THE GENETICS OF THE
SAMPLE.
WOULDN'T IT BE WONDERFUL IF WE
COULD FIND A DRUG THAT ACTUALLY
INVERT THEIR ACTIVITY, SO
BASICALLY CAN WE DO DRUG
PERTURBATION IN CELLS THAT
RECAPITULATE THE ACTIVITIES OF
THE MASTER REGULATORS WITH A
LARGE RANGE OF DRUGS AND THEN
FIND THE DRUG THAT IS ABLE TO
COMPLETELY SWITCH THE ACTIVITY.
NOW THIS SOUNDS INSANE BECAUSE
IT'S HARD ENOUGH TO FIND A DRUG
THAT TARGETS ONE PROTEIN, LET
ALONE 50, BUT THE REASON THIS
ACTUALLY WORKS AND WORKS QUITE
WELL, THIS IS A NATURE GENETICS
PAPER IN ENDOCRINE TUMORS OF THE
GUT, IS THAT AS I SHOWED YOU FOR
PANCREATIC CANCER, THESE MASTER
REGULATORS REALLY WORK AS A MOD AS A
MODULE, SO WHEN YOU HIT THE
RIGHT ENTRY POINT EVEN IN A WAY
YOU DON'T NECESSARILY
UNDERSTAND, THE ENTIRE MODULE
ACTUALLY SWITCHES OFF.
THAT'S WHAT CONTROLS THE STATE
OF THE CELL SO YOU EXPECT EERD
THE TUMOR TO EITHER REPROGRAM OR
TO DIE.
IT'S VERY IMPORTANT TO SELECT
MODELS THAT ARE ACTUALLY MATCHED
TO THE MASTER REGULATORS
OTHERWISE YOU GET COMPLETE JUNK
BUT WE'VE SHOWN THIS IN SEVERAL
PAPERS THAT WEECH WE'VE PUBLISHED.
SO THESE ARE GENOME-WIDE IN
COMPARISON TO CMAP, THEY'RE NOT
USING 1,000 -- BUT THEY'RE --
THEY'RE DYING CELL LINES, BASED
ON CONSERVATION OF MASTER
REGULATORS, AND THE PERTURBATION
WAS DONE AT THE CONCENTRATION OF
THE 48-HOUR IC20 CONCENTRATIONS,
BASICALLY THE HIGHEST SUBLETHAL
CONCENTRATION OF THE DRUG YOU
COULD ACHIEVE, SO THAT YOU DON'T
HAVE CONFOUNDING FACTOR,
CONFOUNDING EVENT FROM CELL
STRESS PATHWAYS OR CELL DEATH
PATHWAYS.
SO THIS REPERTOIRE CAN BE NOW
USED TO MAKE ONCOTREAT
PREDICTIONS IN VIRTUALLY ANY
PATIENT.
SO THIS IS THE N OF 1 STUDY,
AGAIN SEVEN YEARS IN THE WORKS,
WHERE WE ENROLLED ABOUT 130
PATIENTS ACROSS 18 DIFFERENT
MALIGNANCY, GENERATED 45 PDX
MODELS, 34 DIFFERENT DRUGS
PREDICTED BY EITHER ONCOTREAT OR
CONOCO TARGET.
SO THESE DRUGS WERE PREDICTED
EITHER BECAUSE THEY WERE
TARGETING A SINGLE MASTER
REGULATOR OR THEY WERE
TARGETING -- SIGNATURE.
SO I THINK THESE ARE PRETTY
STRIKING RESULTS.
YOU'VE SEEN THAT THESE TUMORS
WERE SELECTED SPECIFICALLY
ENROLLMENT CRITERIA TO HAVE
FAILED THREE TO SEVEN LINES OF
THERAPY, ESSENTIALLY CONSIDERED
TO BE UNTREATABLE.
YOU CAN SEE ALL OF THEM DOUBLE
IN VOLUME SIZE IN LESS THAN 30
DAYS.
HOWEVER, IN 16 ONCOTARGET DRUG
ARMS AND 18 ONCOTREAT DRUG ARMS,
EIGHT OF WHICH ARE PREDICTED BY
METHODOLOGIES.
YOU CAN SEE ONCOTARGET INDUCED A
RESPONSE IN ABOUT 60% OR MORE OF
THESE PDS MODELS, AND ONCOTREAT,
MORE 90%.
SO THIS WAS VERY ENCOURAGING,
BUT WE ALSO WANT TO KNOW, WHAT
ABOUT USING CONVENTIONAL THERAPY
AS OPPOSED TO LETHAL CONTROL.
SO WE DID A TEST WHERE WE
RANDOMLY -- COMPARED THEM TO
THE -- YOU CAN SEE HERE THERE'S
ABSOLUTELY NO RESPONSE.
AND THIS WAS DONE IN 13 NEGATIVE
CONTROL ARMS, SO THIS IN OUR
OPINION IS THE FIRST STUDY WHERE
THE RESULTS OF THE STUDY THAT
PREDICTS ACTIVITY AND
SENSITIVITY OF A DRUG IN VIVO,
DE NOVO, IS ACTUALLY VALIDATED
IN A WAY THAT IS STATISTICALLY
SIGNIFICANT.
AND YOU CAN SEE HERE PANCREATIC
CANCER, THESE ARE THE FIVE DRUGS
THAT WE PREDICTED AND YOU CAN
SEE THAT THREE OF THEM ACTUALLY
HAD PRETTY SIGNIFICANT EFFECT,
PARTIAL RESPONSE OR STABLE
DISEASE.
ONE OF THEM WAS BORDERLINE, ONLY
ONE FAILED TO RESPOND IN A
MOUSE.
THE MORE INTERESTING THING IS
THAT BY TAKING SAMPLES OF THREE
HOURS AFTER THE THIRD DOSE OF
THE DRUGS WHICH ARE BASICALLY
PHARMACODYNAMIC SAMPLES, WE
COULD VALIDATE THAT IN EVERY
SINGLE CASE OUT OF THE 18 DRUGS
WE TESTED WE DON'T TREAT, SO 15
OUT OF 18 TREATMENT EXACTLY
REVERSE -- IN VIVO THAT WE HAD
PREDICTED FROM CELL LINES.
SO THIS IS VERY IMPORTANT
BECAUSE IT VALIDATES THE FACT
THAT WHEN WE MATCH THESE CELL
LINES WITH THE ONCOMATCH, WE
ACTUALLY FIND MODELS THAT ARE
RELEVANT TO THE IN VIVO STUDY
AND WE DON'T CARE AT ALL WHETHER
THE DRUGS WILL KILL THE CELL
LINES BECAUSE THAT'S NOT
HAPPENING, BUT WE CARE ABOUT
UNDERSTANDING THE DRUG MECHANISM
OF ACTION BECAUSE THEN WE CAN
MATCH TO THE -- OF THE TUMOR IN
THE PATIENT.
SO LET ME SHOW YOU HOW THIS CAN
BE MOVED IN SINGLE CELLS.
WE STUDIED THIS IN COLLABORATION
WITH JERRY -- NOW BMS AND
SEVERAL OTHER INVESTIGATORS BY
FOCUSING ON A POPULATION OF
TUMOR AND SHADING CELLS THAT CAN
BE ENRICHED FOR BY SORTING USING
CD49F AND -- THIS POPULATION,
SOME PEOPLE CALL IT ENRICH FOR
CANCER STEM CELLS, WE DON'T LIKE
THAT TERMINOLOGY, SO WE STARTED
ABOUT 75% -- AND TWO TRIPLE
NEGATIVE AND 25% WE BROADLY DID
EPCAM.
WE DID -- WE PREDICTED DRUGS FOR
THE DIFFERENT SUBTYPES WE
IDENTIFIED, TREAT IT IN VIVO AND
THEN AVEST SES ASSESS IF THE DRUG DID WHA T
IT WAS SUPPOSED TO DO.
YOU SEE HERE, IT'S REALLY
STRIKING BECAUSE FIRST OF ALL,
IF YOU DO GENE EXPRESSION BASED
CLUSTERING, YOU SEE EVERY
PORTION FORMS A DIFFERENT
CLUSTER.
SO PEOPLE CALL THIS A BATCH
EFFECT.
THIS IS NOT BATCH EFFECT.
THIS IS BIOLOGY.
THE BIOLOGY COMES FROM THE LARGE
COPY NUMBER ALTERATION THAT --
INDIVIDUAL PATIENT WHICH
CONTRIBUTES TO SOME DIFFERENCE
IN THE -- THE CELL BUFFERS THOSE
DIFFERENCES SO THE DIFFERENCE
THAT'S IMPORTANT IN ESTABLISHING
THE STATE OF THE CELL AS
TRANSFORMED CELLS ARE RETRAINED.
SO WHAT HAPPENS IS WHEN YOU GO
FROM GENE ECK PRETION TO VIPER
ACTIVITY, NOW ALL THE PATIENT
OVERLAP.
AND THIS REQUIRES NO
NORMALIZATION, NO BATCH EFFECT
CORRECTION.
AND WHAT YOU SEE NOW IS THE
DIFFERENCE BETWEEN THE TOP AND
THE BOTTOM, IS THE HORMONE
POSITIVE VERSUS THE TRIPLE
NEGATIVE.
SO BASICALLY YOU'RE RETAINING
IMPORTANT BIOLOGY AND YOU'RE
MISSING OUT THE THINGS THAT ARE
JUST OBTURATION THAT ARE NOT
FUNCTIONALLY RELEVANT.
AND ALSO YOU SEE THAT WHEN YOU
GO LEFT TO RIGHT, YOU SEE
STRIKING ENRICHMENT IN MARKERS
THAT HAVE BEEN PUBLISHED AS
ESTABLISHED MARKERS OF
STEMNESS -- IN FACT, BY
CYTOTRACE ANALYSIS, YOU CAN SEE
STRIKINGLY HIGH -- SO WHAT WE
DID, WE TOOK THE 10 MOST STEM
LIKE CELLS FROM EACH PATIENT AND
COMPARED THEM TO THE 10 MOST
DIFFERENTIATING CELLS FROM THE
SAME PATIENT AND WE DID THIS
INDEPENDENTLY ACROSS EACH
PATIENT, AND THEN WE INTEGRATED
THE RESULTS.
THE MASTER REGULATORS WERE
EXACTLY THE SAME ACROSS THE
PATIENTS SO THE INTEGRATION WAS
VERY, VERY SUCCESSFUL, THEN WE
USED ONCOTREAT PO TREE
DICTIONARY -- WHICH REALLY HAD A
STRONG CONSERVATION FOR THE
ACTIVITY OF THESE MASTER
REGULATORS.
AND WHAT WAS SURPRISING IS THAT
TWO DRUGS, THESE ARE ANTE --
TYPICALLY NEVER USED IN CANCER
BUT CAME UP REALLY -- IS THE
NUMBER ONE MOST STATISTICALLY
SIGNIFICANT.
THESE MICE TRANSPLANTED WITH --
THE TUMOR BIOPSY FOR A TRIPLE
NEGATIVE, VERY AGGRESSIVE
PATIENT, WITH -- THEY PROFILED
THEM AND SEQUENCED THEM, AND
WHAT YOU'RE SEEING HERE IS THAT
WE MEASURE THE ENRICHMENT
INCREASING DENSITY BY COMPARING
EACH TREATMENT TO THE --
CONTROL.
SO AS YOU EXPECT WHEN YOU TREAT
WITH PACLITAXEL WHICH IS KNOWN
TO TARGET DIFFERENTIATED
PROLIFERATIVE CELLS, YOU SEE
STRIKING DEPLETION OF THE CELLS
WHICH ARE IN BLUE, THE COOLER
TONES OF THE COLOR SPECTRUM, AND
DRAMATIC REPLENISHMENT OF THE
STEM-LIKE COMPARTMENT.
SO THESE TUMORS STOP GROWING
BECAUSE YOU'RE KILLING A LARGE
AMOUNT OF THE CELLS, BUT THEY
ACCUMULATE A LOT OF CELLS THAT
CAN RESTART AND REMOVE THE
TUMOR.
HOWEVER, WITH IVERMECTIN,
DRAMATICALLY BETTER WITH
ALBENDAZOLE -- MORE STEM LIKE
AND IN FACT INCREASE IN THE
CELLS IN THE POPULATION
DIFFERENTIATED AND PROLIFERATING
CELLS.
SO WE THOUGHT WE SHOULD USE
THESE TWO DRUGS IN COMBINATION,
MAYBE THEY SYNERGIZE.
INDEED WE DID THIS CLINICAL
STUDY, YOU CAN SEE THE ALBEN DA
SOL
ALBENDAZOLE DOES ABSOLUTELY
NOTHING ON ITS OWN.
WHEN YOU ACTUALLY TREATED WITH
COMBINATION, YOU GOT REGRESSION
AND THEN THE TUMOR WORKS
ESSENTIALLY TO A AN ESSENTIAL
STATE.
WE HAD TO STOP THIS CURVE HERE
BECAUSE THE TUMORS STARTED TO
GROW SO LARGE THAT SOME OF
THE -- STARTING TO HAVE TO BE
SACRIFICED SO WE COULDN'T
REPRESENT ALL THE MICE.
BUT IF YOU LOOK AT THE SPIDER
PLOTS, YOU NOW SEE THE MICE THAT
WERE ON THE COME BOW, FOUR OF
THE SIX HAD REALLY STRIKING
RESPONSE, WERE VERY DURABLE, AND
IN FACT THESE MICE WERE NOT --
BUT RATHER BECAUSE THEY STARTED
TO ULS RATE.
SO WE'RE NOW REPEATING THE STUDY
BY FIRST TREATING WITH PACK
PACLITAXEL, NOW FIRST -- WE TAKE
DOWN THE TUMOR VOLUME BEFORE IT
HAS A CHANCE TO DO TOO MUCH, AND
THEN ALBEN DA ALBENDAZOLE.
I WANT TO CONCLUDE IN THE NEXT
THREE MINUTES WITH SOMETHING WE
CONSIDER TO BE A REALLY EXCITING
STUDY BECAUSE INSTEAD OF
TARGETING ACTUAL MORE CELLS,
WE'RE NOW TARGETING HIGHLY
IMMUNOSUPPRESSIVE CELLS IN THE
TUMOR MICRO ENENVIRONMENT.
SO WE ISOLATED BOTH PRIF LAL PERIPHERAL
AND TUMOR -- IN FOUR DIFFERENT
TUMOR TYPES.
WE ASKED CAN WE SEPARATE THE
INFILTRATING VERSUS
NON-INFILTRATING T-REGS BY GENE
EXPRESSION PROFILE, NEITHER BY
TUMOR TYPE OR ACROSS ALL FOUR
TUMORS.
HOWEVER, NOW YOU FIND 15 MASTER
REGULATORS -- VERSUS THE
PERIPHERAL T REGS AND ADDITIONAL
TWO -- IN COMMON THAT ARE
ACTUALLY ALSO SPECIFICALLY
ACTIVATED IN THE INFILTRATING T
REGS BUT NOT IN THE NON-T REG
INFILTRATING T-CELLS WHICH --
YOU WANT TO MAINTAIN BECAUSE
OTHERWISE YOU LOSE THE --
RESPONSE IN THE TUMOR.
SO TO VALIDATE THIS, WE DID A
CHIMERIC ASSAY WHERE YOU DO
ESSENTIALLY A FOOL CRISPR
KNOCKOUT SCREEN THAT -- IN
HEMATOPOIETIC STEM CELLS AND USE
THEM TO REMOP LATE THE BONE
MARROW OF A MOUSE, THEN AFTER
ABOUT 10 WEEKS WHERE THE IMMUNE
SYSTEM IS COMPLETELY REBOOTED
AND NOW EVERY CELL AND ABOUT
35% -- KNOCK OUT ONE OF THESE
PROTEINS, YOU NOW IMPLANT --
VERY AGGRESSIVE COLON CANCER
MODEL, THEN YOU LOOK AT WHETHER
THERE WAS A DIFFERENCE IN
PERIPHERAL VERSUS TUMOR --
CELLS, SO WE DID THIS IN TWO
DIFFERENT COURSES, THE SECOND
ONE, FILTERED WITH A REPORTER
FROM THE FIRST -- AND THEN
REPLANTED AND YOU CAN SEE
STRIKING CORRELATION BETWEEN THE
RESULTS IN TERMS OF THE PROFILE
OF REPRESENTATION OF THE
DIFFERENT MASTER REGULATORS GENE
TARGETING.
THE BARCODES.
AND THEN YOU CAN SEE HERE THIS
AXIS IS EXTRAORDINARY -- THE
TOP -- STATISTICAL
SIGNIFICANT -- 10 TO THE 170 IS
SIGNIFICANT, CAN YOU SEE THEY
ARE SIGNIFICANT BOTH IN TERMS OF
PERIPHERAL T REGS WHICH WERE
SELECTED FROM THE SPLEEN AND
ALSO WITH RESPECT TO NON-T REG
CD4 T-CELLS FROM THE TUMOR
MICROENVIRONMENT.
SO WE REPEATED THE ASSAY BUT
THIS TIME WE ONLY SILENCED TRPS1 WHICH IS A GENE THAT
NOBODY KNOWS ANYTHING ABOUT IT
IN RELATIONSHIP TO T REGS AND
YOU CAN SEE THAT'S QUITE A
STRIKING EFFECT IN TERMS OF
TUMOR GROWTH IN THE FIVE MODELS
WITH THE NON-TARGETING CONTROLS,
AND SIX MODELS WITH TARGETING
PRPS1.
IN FACT IF YOU LOOK AT A SINGLE
MOUSE, THREE OF THE MICE --
WITHOUT ANY DRUG.
THIS IS A DIFFERENT MODEL
BECAUSE WE WANTED TO MAKE SURE
THIS WAS NOT SPECIFIC TO THAT
COLON CANCER MODEL, THIS IS A
HIGHLY IMMUNE RESISTANT AND
AGGRESSIVE SARCOMA.
SO WE THEN DID EXACTLY THE SAME
THING THAT WIDE DONE FOR SINGLE
CELL IN BREAST CANCER, WE
ASKED -- TO INVERT THE SIGNATURE
OF THE T REGS.
SO WE STARTED WITH ABOUT 1500
DRUGS FDA-APPROVED, SELECTED THE
ONES THAT -- DID THOSE RESPONSE
CURVES FOR THOSE AND WE SELECTED
THE TOP 86, AND THESE WERE NOW
TO PERTURB INFILTRATING T REGS,
AND THEN WE GENERATED THE RNA
SEQ PROFILES AND -- WHAT YOU SEE
HERE THE THREE DRUGS, WE WERE
ESSENTIALLY ABLE TO INVERT
INFILTRATING VERSUS -- IN
VIRTUALLY EACH ONE OF THE 32
PATIENTS THAT WE HAD COLLECTED
EXCEPT FOR MAYBE ONE OR TWO, BUT
WHAT WAS EVEN MORE STRIKING IS
GENERAL GEM SITE BEAN IS THE
ONLY ONE YOU CAN REALLY VALIDATE
BECAUSE IT HAS A GRAD RULE
RESPONSE, -- IN A WAY THAT THEY
KEL T REGS BUT NOT THE
NON-INFILTRATING T REGS FOR THE
CELLS THAT ARE INVOLVED IN
IMMUNE RESPONSE.
SO THIS SHOWS THE RESPONSE OF
TUMORS TREATED IN VIVO WITH
12 MILLIGRAMS PER KILOGRAM WHICH
IS ABOUT 15 FOLD LOWER THAN WHAT
YOU WOULD NORMALLY USE FOR
CLINICAL STUDY, WHICH IS AROUND
150 MILLIGRAMS.
YOU CAN SEE THERE'S ABSOLUTELY
NO EFFECT IN A COMPROMISED --
STRONG EFFECT -- THAT BY ITSELF
HATION
THAT HAS ACTUALLY NO RESPONSE BY
ITSELF.
IF YOU GO TO 120 MG, YOU NOW SEE
THE NSG MODEL STARTING TO
RESPOND A LITTLE BIT, BUT YOU
STILL PRESERVE THE RESPONSE IN
THE IMMUNOCOMPETENT MODEL.
SO I'LL STOP HERE, JUST WANTED
TO SAY THAT FOR THE OTHER PEOPLE
THAT ARE ON THIS CALL THAT ARE
NOT INTERESTED IN CANCER, THIS
IS NOT THE WAY CANCER CELLS
WORK, THIS IS THE WAY ALL CELLS
WORK, AND SO WE HAVE USED THIS
MODEL TO ESSENTIALLY DO STUDIES
IN BIOLOGY, MANY DEGENERATIVE
DISEASES, CARDIOVASCULAR
DISEASE, JUST PUBLISHED WITH THE
RILEY GROUP, TYPE 2 DIABETES
JUST CAME OUT, AND THEN
SOMETHING THAT WE'RE VERY
EXCITED ABOUT IS THE FACT THAT
NOW THAT WE KNOW HOW TO DO THESE
ANALYSES, WE CAN LITERALLY TAKE
ANY CELL STATE THAT YOU START
FROM AND ANY CELL STATE YOU WANT
TO END IN AND DEVISE EITHER
GENETIC OR PHARMACOLOGICAL
RECIPE TO MAKE IT HAPPEN.
SO IF WE SAY THIS IS AN EXAMPLE
WHERE WE IDENTIFIED -- AS A
SINGLE TRANSCRIPTION FACTOR THAT
CAN BE PROGRAMMED -- HUMAN
LINEAGE PROGENITORS -- HSC STATE
THAT WAS JUST PUBLISHED.
SO JUST WANTED TO THANK A LOT OF
PEOPLE AND ALSO THE NIH FOR VERY
GENEROUS SUPPORT TO THIS
RESEARCH.
THIS IS NOT HAPPENING IN ONE
LAB, IT'S A HUMONGOUS NUMBER OF
COLLABORATORS THAT CONTRIBUTED
TO THESE AND FOUNDATIONS.
ALSO EVERYTHING WE DO IS MADE
AVAILABLE TO THE COMMUNITY,,
BOTH OUR -- THAT WE USE ARE ALL
AVAILABLE TO ADGENE, THE
SOFTWARE WE USE, OR THROUGH THE
WEBSITES IN OUR SHARED RESOURCES
OF THE DATA.
THANK YOU.
>> FANTASTIC.
THANK YOU VERY MUCH, ANDREA.
AS USUAL, I'VE GOT A LOT OF
QUESTIONS.
LET'S SEE IF I CAN STUMP YOU.
BUT I'LL JUST GET START AND THEN
PLEASE, ANYBODY IN THE AUDIENCE
WHO HAS A QUESTION, DO TYPE IN
YOUR QUESTIONS IN THE CHAT AND
THEN WE WILL -- THEN I'LL REPEAT
THEM.
WHERE TO START.
SO I'VE BEEN CONVINCED FOR A
LONG TIME THAT YOU CAN FIND
THESE MASTER REGULATORS, SO AS
YOU KNOW, WE TAKE KIND OF A MUCH
MORE TRADITIONAL, I WOULD SAY,
APPROACH TO FINDING AND
DEVELOPING NEW THERAPIES.
WHICH ALSO WORKS, I WOULD SAY.
SO --
>> IT WORKS REALLY WELL.
>> SO THE QUESTION IS, SO WHAT
WE'VE FOUND, GOING WITH MY FIRST
QUESTION, WHAT WE'VE FOUND IS OF
COURSE WHAT EVERYBODY HAS FOUND
IN CANCER, IS THAT SINGLE AGENT
TREATMENT OF ANYTHING, I DON'T
CARE WHAT, IS INSUFFICIENT.
OKAY?
IT'S VERY -- THERE ARE
EXCEPTIONS BCR-ABLE, BUT THEY'RE
RARE.
SO WE HAVE GONE TO DEVELOPING
FAIRLY COMPLEX MIXTURES OF
TARGETED AGENTS.
SO MY QUESTION IS,
COMPUTATIONALLY, HOW DOES THAT
WORK OUT FOR YOU?
CAN YOU -- SO YOU DO -- THERE'S
TWO DIFFERENT WAYS OF THINKING
ABOUT IT, YOU HAVE TWO PARALLEL
PATHWAYS THAT CONTROL DIRN
THINGS, YOU WANT TO TARGET EACH
OR YOU WANT TO TARGET ONE VERY
IMPORTANT PATHWAY TWICE, SO HOW
DOES YOUR MATHEMATICS SHOW YOU
THE WAY FORWARD?
WHICH IS THE BEST WAY TO COMBINE
DRUGS IN MULTI-TARGETED THERAPY?
>> SO LET ME FIRST ADDRESS THE
QUESTION ITSELF BECAUSE THERE'S
A LITTLE BIT OF PHILOSOPHY
BEHIND IT, NEEDS TO BE BROUGHT
TO BEAR.
THERE ARE REALLY THREE
FUNDAMENTAL REASONS WHY SINGLE
AGENT THERAPY FAILS.
ONE REASON IS THAT DRUGS, MOST
DRUGS HAVE ACTUALLY BEEN BUILT
TO TARGET INDIVIDUAL PROTEINS
THAT ARE ACTUALLY ONCOGENES.
OR THAT ARE INVOLVED IN AN
ONCOGENIC PATHWAY.
SO YOU'RE STARTING WITH DRUGS
THAT ARE ALMOST BUILT TO FAIL
BECAUSE WITHIN THE CELLS, WE
NEED THE MAP, YOU HAVE MORE
MUTATIONAL PATTERNS THAT CAN
INDUCE A TUMOR THAN ATOMS IN THE
UNIVERSE.
THERE'S ABOUT -- FROM THE
ONCOGENE WE ALREADY KNOW ABOUT
AND THERE'S 10 TO THE 80 ATOMS
IN THE UNIVERSE.
SO EVERY SINGLE CELL YOU HAVE IN
THE TUMOR WILL ACTUALLY ACCRUE
ADDITIONAL MUTATIONS THAT WILL
DEFEAT SINGLE CELL THERAPY
DIRECTED AT AN ONCOGENE.
THAT'S NUMBER ONE.
NUMBER TWO IS THAT YOU HAVE
WITHIN THE TUMOR A DIFFERENT
SUBPOPULATION THAT HAVE
COMPLETELY DIFFERENT DRUG
SENSITIVITY.
THIS IS ACTUALLY EPIGENETICALLY
DETERMINED.
AS I SHOWED YOU YOU CAN
LITERALLY REPROGRAM THE CELLS
WITHOUT HAVING TO CHANGE THEIR
GENETICS.
AND THE THIRD ONE IS THAT CELLS
ARE INCREDIBLY GOOD AT
PERFORMING WHAT'S CALLED
ADAPTIVE RESPONSE, SO THEY
REALLY REWIRE THEIR NETWORKS IN
RESPONSE TO THE DRUG, THIS
BEAUTIFUL STUDY THAT SHOWS THE
MAP KINASE PATHWAYS --
INHIBITORS, SO TAKEN TOGETHER,
THOSE THREE REALLY FORMULATE A
MANIFESTO FAILURE OF SINGLE
AGENTS, RIGHT?
SO WHAT WE HAVE OBSERVED IS THAT
WHEN YOU GO AND PERTURB THE
CELLS WITH DRUGS, YOU ACTUALLY
DON'T CHANGE THE POPULATION.
THEIR A STABLE POPULATION, YOU
ADD AT THE BEGINNING BEFORE
TREATMENT ARE ALMOST NEVER
CHANGED.
IN SOME CASES YOU DO, IN
PROSTATE CANCER, YOU GET A
COMPLETELY NEW POPULATION OF
NEUROENDOCRINE TUMOR CELLS WHEN
YOU TREAT THE OTHER WITH -- AND
YOU HAVE A -- MUTATION BUT IT'S
VERY RARE.
IN FACT I SHOWED YOU EVEN FOR
THE BREAST CANCER -- COMPLETED
FOR DRUGS THE STATES REMAIN THE
SAME.
SO WHAT WE DO IS INSTEAD OF
FOCUSING ON THE PATHWAY OR THE
ONCOPROTEINS, WE FOCUS ON THE
STATE OF THE CELL.
ESSENTIALLY ASK WHAT ARE THE
PROTEINS THAT MAINTAIN THE STATE
AND THOUSAND CAN WE HOW CAN WE TARGET THE M
PHARMACOLOGICALLY.
MANY OF THE PROTEINS YOU TRY TO
TARGET ARE IN FACT THE UPSTREAM
MODULATORS OF THE MASTER
REGULATORS IN THE CELL.
SO IF YOU REMEMBER FROM THAT
PAPER ON -- WITH RICARDO, WE DID
THIS ANALYSES IN A KIND OF POOR
MANUWAI AND WE SHOW THAT -- KB
WAS THE REPORTER FOR THE
ACTIVITY OF ALL THESE MUTATIONS
IN ITS SUBSTREAM PATHWAYS.
SO WHEN YOU FIND A DRUG THAT
ACTS ON -- TK YOU FIND A DRUG
THAT IS ACTUALLY DISABLING THE
MASTER REGULATORS THROUGH A
MECHANISM THAT IS COMPLEX
BECAUSE THERE'S A FEW STEPS TO
GET THERE.
NOW WE'RE DOING THIS IN
COLLABORATION WITH A LOT OF
PHARMACEUTICAL COMPANIES,
INSTEAD OF TARGETING THESE GENES
THAT ARE UPSTREAM ARE ACTUALLY
TARGETING DIRECTLY FROM -- TO --
TECHNOLOGY ARE TARGETING
DIRECTLY THE MASTER REGULATOR.
>> THAT MAKES SENSE FOR THE
FUTURE.
FOR SURE.
>> IN COMBINATION -- DRUGS THAT
TARGET DIFFERENT POPULATION AND
YOU COMBINE THEM TOGETHER, YOU
FIND THINGS THAT ARE
SYNERGISTIC.
>> SO FOR EXAMPLE, LET'S TAKE
SOMETHING I'M VERY FAMILIAR
WITH.
YOU HAVE A LOT OF THINGS THAT
DON'T WORK AS A SINGLE AGENT,
BUT IF YOU SIMPLY INHIBIT
BCL2 IN A LYMPHOMA, THEN THAT'S
THE BALL GAME.
THAT BASICALLY -- AND SO I DON'T
KNOW WHETHER YOU WOULD REFER TO
BCL2 AS A MASTER REGULATOR.
IT IS THIS MASTER THING IN THE
CELL.
SO HOW DOES THAT FIT IN YOUR
PHILOSOPHY?
>> THE PROBLEM WITH BCL2 AND
MASTER REGULATORS, A LONG TIME
AGO, I THINK IT WAS IN 2010, SO
IT DEFINITELY COMES UP AS MASTER
REGULATOR PROTEINS, BUT --
>> SORRY, IT'S NOT A MASTER
REGULATOR IN TERMS OF GENE
EXPRESSION.
>> IT'S NOT A DIRECT REGULATOR
BUT WHAT HAPPENS IS THAT
SUBSIGNALING PROTEINS ARE SO
CLOSE TO THE TRANSCRIPTIONAL
LAYER THAT ESSENTIALLY ACT AS --
TRANSCRIPTIONAL REGULATORS
BECAUSE THEY DIRECTLY
PHOSPHORYLATE OR -- A
TRANSCRIPTION FACTOR, SO YOU CAN
REALLY TRANSFER THEIR ACTIVITY
ALMOST AS IF IT WERE A
TRANSCRIPTION FACTOR.
SO THE ANALYSIS IS NOT AS GOOD
FOR THOSE PROTEINS AS IT IS FOR
TRANSCRIPTION FACTOR, 80% TO
ABOUT 65 PERCIVAL DAITION RATE 65% -- SO I T STILL DOES
REALLY WELL.
-- GLUCOCORTICOID
SENSITIVITY -- IS NOT A
TRANSCRIPTIONAL REGULATORS.
BUT IN GENERAL, WE WOULD SAY
THAT PROBABLY BCL2 IS
IMPLEMENTING AN EVEN MORE
TRANSCRIPTIONAL SPECIFIC CONTROL
MODULE THAT BCL2 IN SOME WAY
INHIBITS IN AN AMAZINGLY GOOD
WAY SO YOU ARE -- BY
ACTIVATING --
>> SO I HAVE ONE QUESTION FROM
THE AUDIENCE THAT GIBES WITH ONE
I WAS GOING TO ASK.
THE QUESTION WAS FROM -- YEN,
SORRY FOR BUTCHERING YOUR NAME,
AND IT SAYS MANY CANCER DRUG
COMPANION DIAGNOSTICS ARE
MUTATION-BASED.
HOW ARE THE MUTATION INFORMATION
USED IN COMBINATION WITH THE
MASTER REGULATORS?
I'M GOING TO BROADEN THIS OUT IN
GENERAL TO SAY IF YOU ARE GOING
FOR REGULATORY APPROVAL FOR A
DRUG THAT'S TARGETING MASTER
REGULATORS OR COMBINATION, WHAT
WOULD IT LOOK LIKE, WHAT WOULD
BE YOUR DIAGNOSTIC THAT WOULD
LEAD TO IDENTIFYING THE RESPONSE
OF PATIENTS?
AND HOW DOES THAT FIT WITH WHAT
HE OR SHE, I'M SORRY, ASKS ABOUT
MUTATIONS AS PART OF THE
DIAGNOSIS?
>> SO WE NOW HAVE, AS I TOLD
YOU, A CLIA CERTIFIED ASSAY THAT
CAN BE USED TO USE THE ACTIVITY
OF ANY PROTEIN AS A REPORTER FOR
THE ACTIVITY OF A DRUG.
SO WE HAVE ALREADY PUBLISHED
SEVERAL MANUSCRIPTS WHERE WE
ACTUALLY PRODUCED PREDICTORS OF
ACTIVITY OF DRUGS IN PATIENTS,
THIS IS NOT IN MY -- CELL LINES,
INCLUDING -- INCLUDING -- IN
AML -- AND GLIOMA, WE'RE NOW
ACTUALLY STARTING, JUST GOT
FUNDED TO START A STUDY IN
GLIOMA BASED ON THAT BIOMARKER,
AND SO -- LAB HAS PUBLISHED A
PAPER WHERE IT SHOWS THAT AMONG
ALL THE THINGS YOU CAN USE IN
THE -- LANDSCAPE TO PREDICT DRUG
ACTIVITY, GENETICS IS ABSOLUTELY
ON THE BOTTOM.
I FOUND IT VERY REFRESHING TO
SEE THAT THAT RESULTS -- SORT OF
PROPOSING FOR A LONG TIME CAME
FROM A LAB THAT IS AT THE BROAD
BUT BASICALLY REALITY.
THE -- MOST INFORMATIVE ONE IN
HIS ANALYSIS FOR GENE
EXPRESSION, WHAT WAS SHOWN IS
THAT THE PROTEIN ACTIVITY IS
DRAMATICALLY MORE EFFECTIVE THAN
GENE EXPRESSION IMPACT.
WHAT HAPPENS WITH GENE
EXPRESSION, WHEN YOU BUILD A
CLASSIFIER, THE MORE GENES YOU
USE, THE BETTER THE
CLASSIFICATION BECOMES.
THAT'S A CLASSICAL SIGN OF
OVERTRAINING, OVERFITTING.
WITH PROTEIN ACTIVITY, YOU
BASICALLY GET BETTER FOR THE
FIRST SIX OR SEVEN PROTEINS,
SOMETIMES THE FIRST FOUR, AND IT
ACTUALLY GETS WORSE OR DOESN'T
IMPROVE ANYMORE.
SO BASICALLY THIS TELLS YOU THAT
THAT'S THE MINIMUM NUMBER OF
MARKERS YOU CAN USE AND IT'S
VERY GOOD BECAUSE CLINICALLY,
USING ONLY FOUR PROTEINS IS A
GREAT WAY TO DO IT EVEN IN SITU
INSTEAD OF HAVING TO DO GENE
EXPRESSION AND LOOKING AT A
PANEL WITH -- SO THIS IS ALL
PUBLISHED, I CAN SEND YOU THE
REFERENCE.
>> NO, NO, NO, THAT'S FINE.
I'M GOING TO NOT LET YOU GET THE
LAST WORD ON THAT SUBJECT.
SO WHAT CLEARLY INDIVIDUAL GENE
MUTATIONS HAVE THEIR LIMITATIONS
IN -- BUT WHAT WE'VE BEEN
FOCUSED ON AS YOU PROBABLY KNOW,
AND IT'S RELATED, I THINK, TO
THE MASTER REGULATOR IDEA, IS JE
GENETIC SUBTYPES.
THESE ARE NOT INDIVIDUAL
MUTATIONS THESE ARE COLLECTIONS
OF MUTATIONS THAT SEEM TO
CO-OCCUR, AND I'LL BET YOU
DOLLARS TO DONUTS THEN THAT WHEN
WE DEFINE A GENETIC SUBTYPE,
BASICALLY WHAT WE'RE SEEING IN
AN INDIRECT WAY IS A DIFFERENT
CLASS OF MASTER REGULATORS THAT
GOES TO THAT SUBTYPE, BUT IN THE
PAPER WE JUST PUBLISHED IN
CANCER CELL, THESE THINGS HAVE A
100% PREDICTIVE POWER FOR
RESPONSE TO ABRUTE ANYBODY -- OR
BENEFIT FROM ABRUTANIB -- SO IT
MAY BE THAT WE HAVEN'T BEEN
SOPHISTICATED ENOUGH ABOUT
UNDERSTANDING HOW TO THINK ABOUT
THESE MUTATIONS AND MAYBE
THERE'S A NICE -- THERE'S A NICE
POINT IN THE MIDDLE THAT TAKES
INTO ACCOUNT BOTH OF THOSE BITS
OF INFORMATION.
>> I THINK THERE'S A RATIONALE
FOR THAT, LOU.
I THINK THERE'S TWO THINGS.
FIRST OF ALL, YOUR WORK WHICH IS
EXTREMELY SOPHISTICATED COULD
NOT HAVE HAPPENED WITHOUT VERY,
VERY PRECISE PATHWAYS, SIGNALING
PATHWAYS BUILT BY -- OVER --
YEARS.
>> ABSOLUTELY.
>> SO YOU'RE ACTUALLY BENEFITING
FROM SOMETHING THAT ALMOST
DOESN'T EXIST IN OTHER TUMOR
TYPES.
>> TRUE.
>> SO BASICALLY YOU'RE DOING
WHAT WE'RE DOING BUT YOU'RE
USING THE LITERATURE TO --
>> THE KNOWLEDGE BASE.
>> AND YOU -- BUT I FIND THIS
EXACTLY COALESCE INTO A PATHWAY,
ALL THE MUTATION IN THAT
PATHWAY, YOU CAN PREDICT.
THAT'S GREAT.
WHERE EVERY TUMOR HAS THE SAME
MAIN MUTATION IS -- AND THE
OTHER MUTATION ARE JUST
COMPLETELY COMPREHENSIBLE OR
MOSTLY COMPREHENSIBLE, YOU
REALLY DON'T KNOW WHAT TO DO,
YOU CANNOT USE THE APPROACH
YOU'VE DEVELOPED, LOU.
SO I THINK THIS IS WHERE THE
COMMON GROUND EXISTS BETWEEN
WHAT WE DO AND WHAT YOU HAVE
DONE.
TO SOME EXTENT, WE'VE TAKEN A
PAGE FROM YOUR BOOK AND THE WAY
WE HAVE STARTED TO DEVELOP THESE
THINGS PRECISELY --
>> BUT JUST FOR THE AUDIENCE,
AND WE'LL STOP IT THERE, WHEN
I'M SAYING A GENETIC SUBTYPE, I
THINK YOU SHOULD SORT OF THINK
OF, YOU KNOW, LUMINAL BREAST
CANCER VERSUS BASAL, THAT THAT'S
THE KIND OF BIG DISTINCTION, AND
THAT DOES TRACK WITH A BUNCH OF
DIFFERENT MASTER REGULATOR
DIFFERENCES BETWEEN THOSE TWO.
SO IT'S NOT JUST ONE PATHWAY,
IT'S A WHOLE CONSTELLATION OF --
IT'S SORT OF LIKE A SELIB ORIGIN
BASED TYPE OF CANCER THAT SEEMED
TO TRACK WITH THERAPEUTIC
RESPONSE, AND WHAT I'D LOVE TO
DO WITH YOU COLLABORATIVELY IS
TO SEE IF IT TRACKS WITH MASTER
REGULATORS.
SO WE'LL LEAVE IT AT THAT.
I HOPE THIS WAS -- I HOPE THE
LAST QUESTION AND ANSWER WAS
ENTERTAINING FOR EVERYBODY.
BUT THANK YOU, ANDREA.
I ALWAYS LEARN A LOT, AND THIS
WAS ANOTHER GREAT LECTURE.
THANK YOU SO MUCH.
>> THANK YOU, LOU.