>> LET'S GET STARTED.
GOT MORNING, GOOD AFTERNOON,
GOOD EVENING EVERYONE EMPLOY
WELCOME TO THE NIH DNA REPAIR
INTEREST GROUP VIDEO
CONFERENCES.
TODAY WE'RE REALLY GLAD TO HAVE
DR. GUO-MIN LI WHO IS A
DISTINGUISHED INVESTIGATOR AND
DIRECTOR FOR THE INSTITUTE OF
CANCER RESEARCH FOR CHINESE
INSTITUTES FOR MEDICAL RESEARCH.
DR. LI RECEIVED HIS Ph.D. FROM
THE UNIVERSITY AND WORKING WITH
[INDISCERNIBLE] IN A POST DOC
FELLOWSHIP, LATER HE STARTED HIS
CAREER IN KENTUCKY UNIVERSITY
AND THEN MOVED TO UNIVERSITY OF
SOUTH CALIFORNIA AND THEN UT
SOUTHWEST MEDICAL CENTER.
HE SERVED AS DISTINGUISHED CHAIR
FOR CANCER RESEARCH AND DIRECTOR
OF TRANSLATIONAL RESEARCH IN
2023 DR. LI MOVED BACK TO CHINA
AND CONTINUE CANCER RESEARCH
INSTITUTE STUDIES.
DR. LI STUDIES THE MECHANISM OF
DNA MISTACH REPAIR AND ITS ROLE
IN CANCER AND HAS MADE SIMILAR
CONTRIBUTION TOWARDS THE
UNDERSTANDING OF THE MECHANISM
BY WHICH THE MISMATCH REPAIR
SYSTEM MAINTAINS GENOME
STABILITY AND CANCER AVOIDANCE.
SO, WITHOUT FURTHER ADO, WE WILL
HAVE DR. LI GIVE THE
PRESENTATION EMPLOY FOR THE
AUDIENCES IF YOU HAVE ANY
QUESTION, PLEASE LEAVE IT IN THE
CHAT BOX AND WE WILL GO THROUGH
AFTER THE SEMINAR.
THANK YOU.
DR. LI?
>> THANK YOU SO MUCH CHUN FOR
THE CANDID INTRODUCTION AND I
WOULD ALSO LIKE TO THANK KAREN
AND [INDISCERNIBLE] FOR PATIENT.
I HAVE BEEN WORKING ON DNA
MISMATCH REPAIR FOR THE PAST
30-SOMETHING YEARS.
I STARTED WITH [INDISCERNIBLE]
OUT OF DUKE UNIVERSITY AT
[INDISCERNIBLE] SO I WILL BE
TODAY, MAINLY FOCUS ON 1 OF THE
KEY MISMATCH REPAIR FACTOR MLH1
NEXT, SLIDE, PLEASE.
SO AS YOU ALL KNOW, THE DNA
MISMATCH REPAIR IS A WAYS DNA
REPLICATION TO INSURE
REPLICATION FREE DAMAGING AND
ALTHOUGH WE KNOW, REPLICATION
PROCESS IS VERY UPDPRAID, EVEN
FROM THE LOCATION AND
OCCASIONALLY MAKE A MISTAKE BY
DOING MISSING CORPORATION WHICH
GENERATE BASE-BASE MISMATCH,
THAT SHOW HERE IN THE MISMATCH
OR INTRODUCE INSERTION DELETION
MISPAIR WITHIN THE SIMPLE
REPETITIVE DNA SEQUENCE, SO THIS
MISPAIRS HAS TO BE CORRECTED TO
AVOID MUTATIONS.
AND UNLIKE OTHER DNA DAMAGES
WHICH ARE VERY OBVIOUS, BUT
MISMATCH REPAIR I MEAN, ACTUALLY
THEIR NORMAL, MOST SPACES ARE
NORMAL DNA COMPOSITION, SO IN
ORDER TO REMOVE [INDISCERNIBLE]
THE MISMATCH REPAIR SYSTEM HAS
TO KNOW WHICH 1 IS 1 AND THIS IS
ACTUALLY PROVIDED BY A SIGNAL IN
THE LIGAND BREAK AND THIS
CONTAINS A FRAG
M-TEBURKEULOSEISOT MEDIAN STRAND
YOU ALSO HAVE THE 3 PAIR FOR THE
END THOSE ENDS OR BREAKS PROVIDE
OPPORTUNITIES FOR MISMATCH
REPAIR TO TARGET THE REPAIR ON
THE NEW [INDISCERNIBLE].
SO THAT THE MISPAIRED ARE BASED
OR THE INSERTION ON THE
[INDISCERNIBLE] DRIVE OR BOTH
MEDIUM STRAND WILL BE REMOVED TO
INSURE REPLICATION FOR DAMAGING.
NEXT SLIDE.
SO MISMATCH IN HUMAN CELL HAVE
BEEN RECONSITUTED INVITRO USING
ALL PURIFIED PROTEINS BY SEVERAL
LABORATORIES INCLUDING MYSELF
AND [INDISCERNIBLE] AND WE KNOW
THAT [INDISCERNIBLE] PROTEIN IS
THE 1 THAT RECOGNIZE THE
MISMATCH AND THEN LATER ON IT
RECRUITS DUE TO L-PROTEIN TO THE
DNA PROTEIN COMPLEX, AND I'M
SURE [INDISCERNIBLE] LABORATORY
MUTE ANT L-ALPHA IN HUMAN CELLS
HAS THE ENDONUCLEUS ACTIVITY,
AND IT CAN [INDISCERNIBLE] DNA
ON A NEW STRAIN, MODIFIED
PLACES, HOWEVER, WE RECENTLY
SHOWED THAT ONLY IN THE STRAND
BREAK THAT CLOSE THE 2 MISMATCH,
[INDISCERNIBLE] FOR THE MISMATCH
WILL BE SERVED AS A PLACE FOR
AXON 1 RECRUITMENT AND THEN THE
AXON 1 OR STARTING AT THIS
STRAND BREAK AND THEN FROM
[INDISCERNIBLE] TO 3
[INDISCERNIBLE] TO REMOVE THE
MISMATCH, AND THEN ONCE YOU
REACH TO THE MISMATCH, WHERE THE
MUTE ANT HAS A PLACE AND THE
MUTANT SA SLIDING AWAY FROM THE
MISMATCH, SO HERE ARE THE WAY TO
ASK THE NUCLEUS 1 TO REMOVE THE
MISPAIRED BASE, SO THEN THEY
REPLICATE THE POLYMERASE SUCH AS
[INDISCERNIBLE] AND CONJUNCTION
WITH OUR RP A CANNED WHAT AND
THE NEED TO FIND THE
[INDISCERNIBLE] LIAISON GAS 1.
SO THIS COMPLOATS THE REPAIR OF
MISMATCH.
NEXT SLIDE, PLEASE.
SO TODAY, I WILL TRY TO
EMPHASIZE ON HOW MUTANT L-ALPHA
OR 1 OF THE K-MISMATCH REPAIR
COMPONENT WHEN IT CONSISTS OF
MH1, AND PMS2, AND HOW THIS MH01
PLAY A CRITICAL ROLE IN MISMATCH
REPAIR AS WELL AS IN NONMISMATCH
REPAIR FUNCTION, NEXT SLIDE IT
WAS THE FIRST MISMATCH REPAIR
IDENTIFIED WHICH CAN ACTUALLY,
AND THE IDENTIFICATION OF MUTANT
L-ALPHA AND THE MUTANT BEATA AND
THE OTHER HOMOLOGUES.
NEXT SLIDE.
SO THIS IS THE WAYS
IDENTIFICATION OF
[INDISCERNIBLE] IS THE STABILITY
IN COLORECTAL CANCER BY
DR. [INDISCERNIBLE] LABORATORY
AS WELL AS 3 OTHER LABORATORY
INCLUDING [INDISCERNIBLE] AT
MAYO CLINIC.
SO WHAT THEY SHOWED IN THEIR
STUDY IS THAT THEY FIND TUMOR
CELLS DISPLAY SIMPLE REPETITIVE
SEQUENCE IN COMPARISON WITH
NORMAL CELLS AND AS YOU CAN SEE
IN THIS PICTURE THAT TUMOR CELL
EITHER THEY HAVE EXPANSION OF
SIMPLE REPETITIVE DNA SEQUENCE
OR THEY HAVE A CONFRACTION,
CONTRACTING THE SIMPLE
REPETITIVE DNA SEQUENCE.
SO AT THAT TIME, THEY ONLY
KNOW--THEY DON'T KNOW EXACTLY
WHAT'S HAPPENING, THEY CONCLUDE
THIS IS DUE TO REPLICATION ARROW
SO YOU CALL IT THE RER+ OR THE
RER- TUMOR PLEASE.
SO WE HAVE WAYS THIS, I MEAN
PUBLICATIONS, AND THE WAY WE
BELIEVE THAT PROBABLY RELATED TO
MISMATCH REPAIR DEFACT, SO WE
GOT A CELL ON THE
[INDISCERNIBLE] MINOR CELLS FROM
THE VOGELSTEIN AS WELL AS HIS
POST DOC RAMON PARSONS AND WE
GOT THE HRER+ H6, AND WE DIDN'T
KNOW WHAT IT WAS AND AT THE TIME
WE WERE TOLD THAT IT WAS A CELL
GENERATED FROM [INDISCERNIBLE]
LABORATORY AND LATER ON WE KNOW
THIS ACTUALLY THE
[INDISCERNIBLE] 6, SO IT WAS
SHORT FOR H6, SO WE MEASURED A
MISMEASURED REPAIR ACTIVITY IN
BASE H6 AND SO WHICH STANDS FOR
S WO480, AS CAN YOU SEE IN THIS
MISMATCH REPAIR ASSAY, THE H6
CELL LINE CANNOT GENERATE TOO
SMALL FRAGMENTS, THE REPAIR PULL
BACK OR SO CAN DO VERY WELL, AND
AT LEAST 50% REPAIR ACTIVITY,
YOU CAN SEE IN SO, SO THIS SHOWS
FOR THE FIRST TIME THAT HUMAN
CELL TUMOR CELL DISPLAY,
REPLICATION DEFECTIVE IN THE
MISMATCH REPAIR.
NEXT SLIDES PLEASE.
SO WE THEN KNOW THAT THIS MUST
BE MISSING SOMETHING, RIGHT?
CRITICAL MISMATCH REPAIR
COMPONENT.
SO I STARTED TO TRY TO IDENTIFY
OR PIEWNIFY ACTIVITY TO RESTORE
MISMATCH REPAIR TO H6 CELL LINES
AND AS YOU CAN SEE, IN THE LEFT,
THIS IS PUTIFICATION PROFILE.
THERE WAS A VERY SHARP PEEK WITH
DO THE EXTRACT AND THIS WE PUT
ALL THE DATA TOGETHER AND THEN
PUBLISHED THIS WORK IN CELL, AND
THAT WAS A LONG TIME AGO, SO 30
YEARS AGO AND NEXT SLIDE,
PLEASE.
SO I TARTED WORKING ON THE
PURIFY THIS AND I PURE PIE THIS
ACTIVITY THROUGH CHROMATOGRAPHYS
AND AS YOU CAN SEE IN THE RIGHT
GEL AND THOSE ARE 2 BANDS, I
REALLY CANNOT SEPARATE THIS FOR
THAT.
AND THEN WE HAD NO IDEA WHAT
THIS COMPONENT IS, BECAUSE
COMPARED WITH THE
[INDISCERNIBLE] OR MISMATCH
REPAIR COMPONENT THE E.COLI ARE
FULL OF SINGLE PEPTIDE, SO 1
[INDISCERNIBLE] DILL O DALTON
PROTEIN AND THE MUTANT IS ALSO
SINGLE POLYPEPTIDE, SO IN ORDER
TO KNOW WHAT THIS 2 BANDS ARE,
SO WE PERFORMED A SEQUENCE AND
ASSAYS ON DEGRADATION AND THAT
YOU CAN SEE IN THIS TABLE AND WE
FOUND AND WE FOUND THE PEPTIDES
FROM THESE 2.
THEY ARE HEREWITH HOMOLOGY WAYS
OR HUMAN AT THAT TIME,
IDENTIFIED AND ALSO HAS SOME
YEAST YEENS AND THEN THE BIGGER
1, ALTHOUGH THEY WERE--WE FOUND
THERE WERE SEVERAL, SEVERAL OF
THIS GENES BY AT LEAST WE KNOW
THIS PROBABLY JUST COME FROM A
SINGLE, MAYBE A SINGLE
POLYPEPTIDE IN PMS2, SO BOTH OF
THIS PMS2 AND THE MLH1 HAD THE
SAME PHENOTYPE, SHOWING THE SAME
PHENOTYPE, WE BELIEVE THIS
PROBABLY MUTANT L HYDRODIMER SO
WE PUBLISHED THIS WORK AS SHOWN
THIS IS FOR A HUMAN MUTANT, AND
IT IS A HYDRODIMER AND THE NEXT
SLIDES PLEASE EMPLOY SO AS I
TOLD YOU ABOUT THIS MUTANT EXTRA
HELP TO DEFINE THE HUMAN
MISMATCH REPAIR POSSIBLY, THAT'S
AS WELL AS IDENTIFICATION OF
HUMAN MUTANTS, THE REASON I SAY
THAT, NEXT SLIDE, PLEASE EMPLOY
I ACTUALLY PURE PIED A
HYDRODIMER 6 MONTHS AT
DR. [INDISCERNIBLE] LABORATORY
AND THIS HYDRODIMERRIZED SHOWN
ON THE RIGHT, THIS 160 AND 105
KILODALTON RATINGS AND WE, I
PURIFIED THIS USING AFFINITY
COLUMN, OLIGO CAN CONTAIN A
SINGLE MISMATCH AND I COUPLE
THEM TOGETHER AND THEY ARE
COUPLED TOGETHER, WITH THE DNA
MOLECULES INTO [INDISCERNIBLE]
AND WE ARE PAST THE
[INDISCERNIBLE] YOU COULD
EXTRACT THE SMOOTHEST COLUMN,
AND IT DID 1 PART, IT DOESN'T
MATTER, I MEAN HOW--IT'S THE
WHOLE CELL EXTRACT, THE
[INDISCERNIBLE] JUST PURIFIED
THE FRACTIONS AND THE CANNED
WHAT THEY CAN GET IN THESE 2
BANS AND I WAS SO EXCITED BY THE
DATA I COULD SHARE THAT 1 IS NO
WAY--I MEAN IF YOU THINK OF THIS
HAS A MISMATCH, WE GOTTA SAY, 1
BAND, [INDISCERNIBLE] SO I'M
STUDYING WORKING ON THIS FOR
ANOTHER 6 MONTH, I REALLY CANNOT
SEVER THIS 2 BAND, SO AT THIS
TIME, WE CANNOT REALLY DEFINE
WHAT IT IS.
SO WE JUST PUT A PUTATIVE
PROTEIN IN THE FREEZER AND
WORKING ON SOMETHING ELSE.
AND FROM THIS STUDY, I REALLY, I
MEAN, LEARNED HOW TO DO SCIENCE
FROM DR. [INDISCERNIBLE].
SO IF YOU ARE NOT A
[INDISCERNIBLE] ON SOMETHING,
DON'T PUBLISH, TRY TO FIGURE IT
OUT BEFORE YOU PUBLISH THIS
STUFF.
SO BECAUSE OF THE IDENTIFICATION
OF MUTANT L ALPHA WHICH IS THE
HETERODIMER AND THEN THIS
IMMEDIATELY MUST KNOW THAT THE
MAYBE THE 1 I PURIFIED 4 YEARS
AGO, IT IS NOT THE HYDRODIMER,
AND THAT INDEED, WE FOUND FROM
BOTH THE [INDISCERNIBLE]
ANTIBODIES ALREADY, AVAILABLE,
SO WE TO INVEST IN
[INDISCERNIBLE] AND SHOWING THAT
[INDISCERNIBLE] BAND 105 WAS
[INDISCERNIBLE] AND THEN BY
SEQUENCING ANALYSIS, WE FOUND
THAT [INDISCERNIBLE] AND AT THIS
TIME WE DON'T KNOW WHAT IS MHS6,
THAT WE NAME OUR 160 KILODALTON
PROTEIN WHICH IS A MUTANT S
ALPHA, WE NAMED IN THIS SCIENCE
PAPER.
NEXT SLIDE, PLEASE.
AND IN ADDITION TO THAT, I
SHOWED HERE, AND ALL THE MUTANT
S CELLS ARE HYDRODIMERS, THIS
ALL STARTED A WAYS
IDENTIFICATION OF MUTANT L
ALPHA, WHETHER I SHOWED YOU
ALREADY, THE [INDISCERNIBLE] FOR
THE DIMER AND THERE ARE ALMOST,
ALL ASSOCIATED WITH THE CANCER
OR OTHER DISEASES.
THAT'S NOT--I'M NOT GOING TO
TALK ABOUT MSH3, WHICH IS REALLY
INVOLVED IN NEURON DEGENERATIVE
DISEASES.
NEXT SLIDE, PLEASE.
SO WHILE I'M GOING TO FOCUS ON
MUTANT L-ALPHA ACTIVITY AGAIN,
MUTANT L-ALPHA CONTAINS A LOT OF
SIMILAR BIOCHEMICAL ACTIVITIES
INCLUDING ATP ASE ACTIVITY WHICH
WAS INITIALLY SHOWED BY DR. YANG
IN E.COLI AND LATER
DR. [INDISCERNIBLE] LABORATORY
SHOWING HUMAN L-PROTEIN AS ATP
ASE ACTIVITY AND I MENTIONED
EARLIER DR. [INDISCERNIBLE]
ENDONUCLEUS ACTIVITY OF MUSEUM
TABT L-ALPHA WHICH IS CRITICAL
TO PROVIDE THE INITIAL STRAND
BREAK TO RECRUIT ALE 1 TO THE
SIDE TO REMOVE THE MISMATCH.
AND THEN TODAY I'M GOING TO SHOW
YOU 1 OPPORTUNITY WE IDENTIFIED
AFTER THIS MUTANT L-ALPHTHIS
ACTIVITY LIKELY REGULATING THE
AXON NUCLEUS 1, ACTIVITY AND I
WILL SHOW YOU HOW THIS NUCLEUS
ACTIVITY INVOLVED IN THE
MUTANT-LOR MISMATCH REPAIR
DEFICIENCY, MEDIATED CANCER
THERAPY AND ALSO THE--INVOLVED
IN THE NEURON DEGENERATIVE
DISEASE INCLUDING HUNTING TON
DISEASE.
NEXT SLIDE.
SO THIS ACTUALLY SHOW YOU, I
TRIED TO SHOW YOU HOW I
IDENTIFIED THIS MUTEITANT -
-MUTANT L-ALPHA
ACTIVITY AND DURING THE
CONSTITUTION OF MISMATCH REPAIR
REACTION AND I SHOWED IT HERE WE
USE ALL PURIFIED PROTEIN AND
THAT THE RIGHT 1 SHOWS THE
MISMATCH REPAIR ASSAY USING ALL
PURIFIED PROTEIN EMPLOY AS CAN
YOU SEE IN 9-4 WHEN WE TAKING
OUTTA A MUTANT L-ALPHA IN THIS
REACTION, AND YOU CAN SEE, THAT
DEALING ACTUALLY, THE TOTAL
AMOUNT IS NARNLGINAL IN THIS
REACTION COMPARE WITH OTHERS,
EITHER THE UNREPAIRED POP BAND
OR IN THE EVEN THE REPAIR, THE
[INDISCERNIBLE] DIED USING THE
SMALLEST FRAGMENT SO WE WANT TO
FIND OUT WHAT IS HAPPENING IN
THIS REACTION.
NEXT, SLIDE, PLEASE.
SO WE MEASURED IN THE LEFT, CAN
YOU SEE, ACTUALLY, IN THE
[INDISCERNIBLE], YOU CAN SEE
AGAIN, THE DNA MOLECULES, AMOUNT
OF MOLECULES, MUCH, MUCH WORSE
COMPARED WITH OTHER ACTIONS
BECAUSE THIS REACTION IS NO
MUTANT L-ALPHA AND WE FINDALLY
FIND OUT THIS IS PROBABLY DUE TO
THE REASON THAT THE EXCISION
STUDY FROM THE STRAND BREAK AND
THE AXLE NUCLEUS WORKED FROM TOP
TO THE BOTTOM AND THEN FOR THIS
IS [INDISCERNIBLE] ASSAYS, YOU
CAN SEE, ACTUALLY REMOVED ON THE
SIDE FOR THE PROBE AND THAT'S
WHY WE COULD BT DETECT THE
MORNING CREWS ANYMORE, SO IT
SHOWS THE AMOUNT OF DNA SO IN
THE RIGHT, WHAT I WANT TO SHOW
YOU IS THAT IN THE PRESENCE OF
MUTANT L-ALPHA IN THE LEFT,
THE--YOU CAN SEE [INDISCERNIBLE]
IS IMMEDIATELY TERMINATED WHEN
THE EXO1 REMOVES THE MISMATCH.
AND ON THE RIGHT WHEN YOU DON'T
HAVE A MUTANT L-PROTEIN, OR IN
ABSENCE OF MLH1, THE EXCISION IS
VERY EXTENSIVE AND THIS MUTANT
L, THIS MUTANT L TO STOP THE
NUCLEUS ACTIVITY, SO WE GENERATE
LARGE AMOUNT OF SINGLE STRANDED
DNA, WHICH COULD BE A PROBLEM AS
I'M GOING TO SHOW YOU LATER,
NEXT SLIDE, PLEASE.
JUST ACTIVITY IN THE WORLD IN
CANCER IMMUNOTHERAPY.
AND THAT WE KNOW IN 1999-2015,
IT HAS BEEN SHARED BY SEVERAL
LABORATORY THAT MISMATCH REPAIR
DEFICIENCY BENEFITS
IMMUNOTHERAPY AND THIS HAS BEEN
REPUTED IN ANIMAL MODEL AS WELL.
NEXT SLIDE.
AND THEN THE MECHANISM AT THAT
TIME WAS--ESSENTIALLY CONDITION
CLUEDED THIS SLIDE, SO IF YOU
DON'T HAVE A MISMATCH REPAIR
ACTIVITY, YOU ARE GOING TO BE
MUTATE A LOT OF MUTATIONS, THE
FRAMING SHIFT OF MUTATIONS WHICH
CAN CAUSE EITHER SHORTER
PEPTIDE, BECAUSE YOU MAY
GENERATE A STOP CODON OR YOU
CODING FOR SOMETHING THAT IS NOT
ALSO NEEDED, THERE IS NOT A NEED
THIS PROTEIN, SO THIS PROTEIN
WILL BE DEGRADED WHICH WILL
GENERATE MORE PEPTIDE, SO THAT'S
GOING TO USE ITS NEOANTIGEN, SO
THE IDEA IS THAT THE CANCER
CELL, OR MISMATCH REPAIR
DEFICIENT TUMOR CELL WILL NOT
HAVE A NEOANTIGEN WHICH WILL NOT
BE RECOGNIZED BY OUR T-CELL SO
THAT THIS MUTATED TUMOR CELL
WILL BE TREATED AS A FOREIGN, A
FOREIGN AND THEN PROBABLY GOING
TO BE KILLED BY T-CELL.
NEXT SLIDE.
INTERESTINGLY ALTHOUGH THIS
EXPERIMENTATION IS REALLY NICE,
I MEAN IT CAN EXPLAIN AT LEAST
50% OF CASES THAT PATIENTS
RESPONDED TO IMMUNOTHERAPY, BUT
OTHER 50% OF TUMORS, MISMATCH
REPAIR DEFICIENT TUMORS, THEY DO
HAVE NEOANTIGEN, THEY DO NOT
HAVE RESPONDED TO PD1, PDL
ANTIBODY IMMUNOTHERAPY AND
SUGGESTING SOMETHING ELSE MUST
ENROLL IN THIS, SO WE ARE REALLY
INTERESTED IN THIS AND WE TRY TO
FIND THE ISO FOR THIS.
NEXT SLIDE.
SO THEN GOING INTO THE
LITERATURE AND FIND THAT GENOME
INSTABILITY IS ASSOCIATE WIDE
THE DNA PRODUCTION AND THE CGAS
EVALUATION, SO THIS IS A REVIEW
BY MY FORMER COLLEAGUE JIM CHEN
AND HEITATED HERE THAT IF YOU
HAVE A DNA BREAKS OR CHROMOSOME
BREAKS, YOU ARE ANYTHING TO
GENERATE [INDISCERNIBLE] DNA AND
IT WILL BE RECOGNIZED BY CGAS,
CGAS WILL BE ABLE TO GENERATE
CCAMP--HELPING IMMUNOTHERAPY AS
HONE BY CHEN IN ANIMAL MODEL, IF
HE SHOWS THAT CGAS IS INVOLVED
IN PD1, PDL1, PLOK TREATMENT, I
MEAN TRY TO KILL CANCER, CANCER
CELLS.
SO WE BELIEVE THAT A MISMATCH
REPAIR DEFICIENCY CAN INDUCE
GENOMIC INSTABILITY THAT TRIGGER
CYTOSOLIC DNA [INDISCERNIBLE]
AND ACTIVATE CGAS POSSIBLY
IMMUNOTHERAPY, THIS IS WORK BY
[INDISCERNIBLE] AND THIS WORK WE
HAVE PUBLISHED A COUPLE YEARS
AGO, NEXT SLIDE.
CELLS SHOWING THE DNA BUT ONCE
YOU PUT MHL1 BACK AND REDUCED A
LOT, DOESN'T MATTER, YOU
JUSTIFIED CELL ITSELF OR TREAT
IT WITH RADIATION WHICH CAN CAN
TRIGGER A LOT OF DNA PRODUCTION
BUT WHEN YOU PUT IT THE MH1
BACK, THE AMOUNT IS DRAMATICALLY
ARE I DUCED NEXT SLIDE PLEASE SO
THIS IS THE CELL LINE, I MEAN
YOU ESTABLISH IT ON 50 OR 60
YEARS AGO, WHAT ABOUT JUST KNOCK
OUT A CELL MHL1 FROM NORMAL OR
NOT A NORMAL MISMATCH REPAIR
PROFICIENT CELLS AND WE ACTUALLY
DID THAT, AS CAN YOU SEE HERE
WHEN YIEW KNOCK OUT MHR1 WHERE
MIDDLE THEN, YOU CAN SEE THE
DEFICIENCY CELLS SHOWING
CYTOSOLIC POINTED BY THE
[INDISCERNIBLE] AND YOU PUT THE
GENES BACK ON THE CELL LINES,
YOU RESTORED THE WILD-TYPE
PHENOMENON.
AND IT'S--WHEN YOU TREAT IT,
TRADE A CELL WITH IONIZING
RADIATION, WITH THE DNA
PRODUCTION, I SHOW IT IN MH6
CELLS SO THIS EVALUATING THE
NEXT SLIDE, PLEASE SO NOW WE
WANT TO SAY WEATHER OR NOT,
DETISHT CELL CAN IMENERATE A
CCAMP AND IN UNTREATED
AND--INCREASED CCAMP PRODUCTION,
MHL1 DEFICIENT CELLS WHICH YOU
CAN SEE UNDER THE LIGHT THERE
FOR QUANTIFICATION, AND SO
SUGGESTING THAT THIS IS
ACTIVATED AND PRODUCED THE CCAMP
NEXT SLIDES.
WE SHOWED YOU, WE CAN CLEARLY
SHOW THAT THIS IS PHOSPHORYLATED
AND THEN IT IS ATTENUATE BUT
BECAUSE OF TIME, I WILL SKIP
THAT BECAUSE IT'S PUBLISHED
ALREADY, IF YOU ARE INTERESTED
YOU CAN LOOK AT THOSE 2 PAPERS.
SO NEXT QUESTION, WE WANT TO ASK
IS PRETTY TOUGH.
SO HOW DOES MHL1 DEFICIENCY CAN
INTRODUCE CYTOSOLIC DNA AND THEN
THIS IS HAS COME TO THE 1 I
SHOWED YOU EARLIER, RECONSITUTE
MISMATCH REPAIR, PAPER WE
PUBLISHED IN 2005.
WE KNOW THAT MLH1 CAN ISHT ACT
WITH EXOMUSEUM CLE ACE 1 AND CAN
NEGATIVELY REGULATE EXO1 MUSEUM
CLE ACE ACTIVITY, SO WIEW KNEW
THIS REGULATION IS CRITICAL FOR
WHETHER OR NOT THE CELL CAN
GENERATE THE CYTOSOLIC DNA.
NEXT SLIDE, PLEASE EMPLOY SO
THEN WE THINK IF THAT'S THE
CASE, IF WE ARE KNOCKING OUT
CERTAINLY--CERTAINLY XO1, SO YOU
MAY NOT BE ABLE TO STAY IN THE
DNA IN THE EMERNG DEFICIENT
CELLS AND THAT'S EXCITING WHAT A
WAY WE SAW AS YOU CAN SEE HERE,
IN THE KNOCK OUT ITS OR IMOWBL
DOCK OUT, MLH1 AND IMOWBL DOCK
OUT AS SHOWN ON THE RIGHT, YOU
REALLY CANNOT SEE THE CYTOSOLIC
DNA AND AGAIN, ARRESTING SHOWS
THAT YOU CAN SAY IMMEDIATELY
THEY ARE ACTIVATION OF CGAS
IMPOSSIBLY WHEN YOU HAVE MLH1
KNOCK OUT BUT A DOUBLE KNOCK
OUT, YOU KNOW PHOSPHORYLATED, SO
SUGGIESTING THAT ACTIVATION FOR
THE INTERNET, IN THE SECOND
PATHWAY, NO MAN CAN EXIST WHEN
YOU KNOCK OUT [INDISCERNIBLE] 1.
NEXT SLIDE, PLEASE.
SO FROM LITERATURE, WE ALSO KNOW
THAT MLH1 IS INVOLVED IN TOP O
STRAND BREAK REPAIR AS SHOWN IN
THIS PAPER, NEXT SLIDE, PLEASE.
SO WE BELIEVE THAT MLH1 IS
CONTROL ACTIVITY IN THE DOUBLE
STRANDED BREAK REPAIR WITH THE
FOUNDATION, AND MISMATCH REPAIR
AS I SHOW YOU EARLIER.
SO IN DOUBLE STRANDED REPAIR, IN
THE RESECTION, AND ACTUALLY 1 OF
THE ENZYME I HAD BEEN HOEING IN
THIS ENDORESECTION, AND WE
BELIEVE MLH1 OR MUTANT L-ALPHA
ALSO CONTROLS THIS PROCESS, IF
YOU DON'T HAVE MUTANT LR-5 SHOWN
IN THE RIGHT, THE EXCISION WILL
BE REALLY EXTENSIVE AND GENERATE
A LARGER QUANTITY OF SINGLE
STRANDED DNA WHICH ACCRUED, GAVE
THE CHROMOSOME, BIGGER PROBLEMS,
SO THEY WILL BE CHROMOSOME BREAK
OUTS.
SO NEXT SLIDE.
SO WE WANT TO SAY WHETHER OR NOT
IF THAT'S REALLY THE CASE
BECAUSE MLH1 AND EXO1 INTACT,
WITH EACH OTHER AND THE DOMAIN
HAS BEEN AMENDED BY SEVERAL
LABORATORY, PARTICULARLY
[INDISCERNIBLE] LABORATORY
SHOWING THEY ARE INTO
INTERACTIVE DOMAINS, MLH1, AND
EXO1, ARE SHARED BY THIS GREEN
AREA.
SO WE TRY TO MARK OUT THIS
INDUCTION DOMAIN AND WE TAKE
EITHER TAKING THE FF, WHICH IS
THE MIDDLE ACTION, OR TAKING OUT
THE WHOLE C-TERMINAL OR COMBINE
THIS 2 TOGETHER, SO I SHOWED
HERE, THE PURIFIED PROTEIN IN
THE BOTTOM LEFT AND THEN THERE
ARE 3 MUTANT, I SHOWED HERE AND
THEN ON THE RIGHT, BOTTOM RIGHT,
AND THIS IS ACTUALLY
SHOWING--TRIED TO SHOW
INTERACTION OF THIS 3 MUTANT
PROTEIN WITH MUTANT L-ALPHA AS
YOU CAN SEE IN THE FAR RIGHT,
THE DOWNL MUTANT, WHERE WE
KNOCKING OUT THE C-TERMINAL AND
ALSO THE FF [INDISCERNIBLE]
AA YOU DO REALLY SEE THE
INTERACTION BETWEEN MUTANT
L-ALPHA AND EXO1 AND THAT'S
EXACTLY WHAT WE NEED AND WE KNOW
WE ACTUALLY TASK THIS MUTANT,
ALL THIS MUTANT HAVE THE REALLY
ACTIVE NUCLEACE ACTIVITY, SO 1
PAST WITH THIS MUTANT
ENDORESECTION, CATALYZE THE
ENDORESECTION CAN BE CONTROLLED
BY THE ALPHA.
NEXT SLIDE, PLEASE.
SO THIS IS THE ENDOASSAY
NUCLEUS, THIS CREATED BY
DR. [INDISCERNIBLE] DIFFICULT TR
ME TO SAY LAST NIEM, I MEAN TRY
TO USE DUKE [INDISCERNIBLE] NAME
ALTHOUGH THEY ARE NOT EXACTLY
THE SAME NAME, SO AS YOU CAN
SEE, HERE, ON THE RIGHT, WE ARE
USING THE WILD-TYPE--EXO1, AND
THE MUTANT EXO1 TO DO THIS
ENDORESECTION IN THE PRESENCE OR
ABSENCE OF MUTANT L-ALPHA SO
WHEN YOU SAY, WILL YOU USE
WILD-TYPE AXON 1 IN THE ABSENCE
OF MUTANT LH1, CAN YOU ONLY SEE
THE PRODUCT OF VERY SHORT, WE
NAME THE PRODUCT 1 AND THEN WHEN
YOU INCREASE THE MUTANT-L
CONCENTRATION IN THIS REACTION,
CAN YOU SEE THE [INDISCERNIBLE]
COMING BACK, SO MUCH BIGGER SO
SUGGESTING IT CONTROLS THE
NUCLEUS ACTIVITY OF 1, HOWEVER,
IF YOU LOOK AT THE FAR RIGHT 1
WHICH WE SHOWED THIS 1 LITTLE IN
THE ACTION, CONTAIN THIS MUTANT
X-1 AND THE MUTANT L-ALPHA WHAT
DO YOU SEE, YOU REALLY CAN HAVE
THE SEE MUCH OF THIS WITH 2 IN
THIS REACTION, SO SUGGESTING
EXTENSIVE ECCISIONS IN THIS
MUTANT REACTION SO SUGGESTING,
GENERATING A LOT OF SINGLE
STRANDED DNA, NEXT SLIDE,
PLEASE.
SO WHEN WE LOOK AT THIS, WHETHER
OR NOT THAT--THEY'RE NOT SINGLE
STRAND DNA GENERATED AND AS YOU
CAN SEE HERE, IN MLH1 DEFICIENT
CELLS AND WE DO THIS BRDU
INCORPORATION AND YOU CAN SEE A
LOT OF BRDU INCORPORATED IN MLH1
CELLS COMPARED WITH WILD-TYPE
AND ALSO IN THE RPA BINDING, YOU
CAN SEE NICELY, MHL1 DEFICIENT
CELLS BUT NOT MANY IN WILD-TYPE
CELLS, SO THAT'S--I WAS THINKING
WHAT DO WE THINK IS HAPPENING IN
THERE IN VIVO IN THE SUCCESSES
SECTION PLEASE.
THIS WAS CHROMOSOME
ABNORMALITIES SO WE DID THIS
EXPERIMENT, CAN YOU SEE MLH1
DEFESHT CELLS AND ALSO IN HUMAN
CELLS WE SHOW EXACTLY THE SAME.
THEY'RE NOT ALL [INDISCERNIBLE]
IN THE SO WE CAN'T SO HOW THESE
ARE AWARDED TO 2, THE DNA BY THE
WAY WE BELIEVE, THAT'S PROBABLY
HAPPENING AND THAT'S WHERE I'M
LOOKING AT IT, BECAUSE I YOU CAN
IMAGINE HOW IT OCCURRED SO THE
MODEL, BASED ON THE DATA I SHOW
YOU, I CAN--I CALCULATE IT HERE
IS THAT, ON THE LEFT SIDE, WHERE
YOU HAVE THE MUTANT L-PROTEIN
HERE, SO YOU ARE ENDORESECTION,
IN DOUBLE STRANDED BREAK, IS
REALLY LIMITED ON THE RIGHT, AND
IF YOU HAVE LIMITED PROTEIN IN
THE CELL, OR BREAK FOR X-1
NUCLEUS ACTIVITY, SO THE
DECISION CAN BE REALLY
EXTENSIVE, GENERALLY A LOT OF
SINGLE STRAND DNA, AND YOU CAN
SOURCE THE RPA SO YOU WANT
SINGLE STRAND DNA CANNOT BE
PROTECTED SO THAT'S WHY YOU
CAUSE THE CHROMOSOME BREAK AND
YOU SET IT OF THE DNA WHICH CAN
INDUCE THE C-GAS DEACTIVATION
WHICH FACILITATED, AND IT'S THE
NATION FOR YOU FOR 50%.
FOR THE DEFICIENT 1S TO RESPOND
AND WE BELIEVE THAT CGAS IS
IMPOSSIBLE, PROBABLY
CONTRIBUTING TO THOSE SOME OF
THOSE BECAUSE MISMATCH REPAIR
DEFICIENCY CAN CAUSE ANY GENE
MUTATIONS, SO CGAS IS CAN THEN
PASS AWAY, ALL THE YEEPS CAN BE
MUTATED IF THAT HAPPENING, AND
THEN, THOSE TUMORS EVEN THEY ARE
DEFECTIVE IN MISMATCH REPAIR,
THEY'RE NOT GOING TO RESPOND FOR
THE IMMUNOTHERAPY, SO THAT'S WHY
I PUT THAT MARKER HERE, SO NEXT
SLIDE, PLEASE TO, EVALUATED THAT
SO WE, COME BACK, CONTACT
FOR--COME ON THERE OF THE WHO
INITIALLY SHOWED THE MISMATCH
REPAIR DEFICIENCY, HELP THE
IMMUNOTHERAPY.
SO WE HAVE TO HAVE PATIENTS
DEFECTIVE, WITH THE MH1 IS
TREATED WITH THERAPY AND THIS IS
THE PAPER, CAN YOU SEE, SO
WHENEVER THE PATIENT HAVE
REALIZED OR GO TO EXPRESSION OF
CGAS AS SHOWN IN THE LEFT OR
STING ON THE RIGHT, THEN THEY
RESPONDED TO THE PDL1 ANTIBODY
TREATMENT AS WELL, BUT IF THEY
DON'T HAVE THIS GOOD EXPRESSION,
THOSE PATIENT DO NOT RESPOND TO
IMMUNOTHERAPY AT ALL, SO THIS
CONFORMS OUR DATA
[INDISCERNIBLE] FOR BIOCHEMICAL
ASSAYS.
NEXT SLIDE, PLEASE, SO I HOPE I
HAVE SHOWED YOU THAT WE HAVE
ACTUALLY FOUND THE MLH1
DEFICIENCY, AND CGAS, AND THE
DEFICIENT OR NO EXPRESSION OF
CGAS THIS FACTOR CAN BE CAUSED
BY MISMATCH REPAIR DEFICIENCY
CONTRIBUTES TO THE 50% OR MAYBE
CERTAIN IN THIS MISMATCH REPAIR
ALLOW THEM TO EXIST WITH THE
IMMUNOTHERAPY AND WE ALSO SHOW
THAT ACTUALLY REALLY IMPORTANT
BUT CONTROLLING EXO1 ACTIVITY.
NEXT SLIDE, PLEASE.
SO NOWIME GOING TO SWITCH, AND
IT'S TALKING ABOUT THIS MLH1
MUTANT L-ALPHA CONTROL, THE EXO1
NUCLEACE ACTIVITY AND WE SHOW
THIS ACTUAL LOW, ATHE SAME
FUNCTION, SEEMS DOING AND
CONTRIBUTING TO EXO1 GENERATING.
SO NEXT SLIDE PLEASE.
SO I SHOWED OTHER IN THE
AUDIENCE, ALSO THE
[INDISCERNIBLE] AND THE
[INDISCERNIBLE].
SO AS YOU PROBABLY KNOW, YOU
HADITANT SYSTEM ACTUALLY BRAIN
DISORDER AND IT'S A GENE AND OUR
NORMAL INDIVIDUAL, YOU WOULD
HAVE REPEAT NUMBER AROUND THE
[INDISCERNIBLE] BUT ONCE THIS
CAKB SIN REPEATS, IT REACHES 36,
THEN YOU WILL HAVE ISSUES AND
THERE ARE PATIENTS, YOU KNOW
THAT, SO THIS EXPRESSION WILL
ALLOW THE WILD-TYPE HTT TO
MUTATE TO THE FORM OF HTT, AND
THE SO CALLED MHTT.
OKAY, THIS PROTEIN, THIS
MUTATING THERE, WITH THE
PROTEIN, HAS BEEN SHARED TO
INDUCE CGAS ACTIVATION, MEDIATED
APOPTOSIS BUT THE NECKANISM IS
NOT REALLY LOW, AND WE KNOW FROM
PREVIOUS TODAY, THAT MISMATCHED
REPAIR GENES, PARTICULARLY MSH3
AND MHSL3, ARE KEY GENETIC
MODIFIERS BUT NOW PEOPLE ARE
THINKING NOT JUST THE MODIFIER,
IT'S JUST THE DRIVERS OF HUNTING
TON'S DEC.
NEXT SLIDE, PLEASE.
SO WE WILL TRY TO--I MEAN, MLH3,
WHICH IS THE SUBUNIT OF THE
MUTANTS ALPHA THERE ARE MANY
LABORATORIES SHOWING THAT THE
MUTANT ALPHA, HOW MUTANTS ALPHA
CONTRIBUTED TO REPEAT THE
EXPANSION AND RELATED TO HUNTING
TON'S DEC AND ENCLOUDING
DR. [INDISCERNIBLE] SHE WAS IN A
MEETING, SHOWING HOW--I MEAN
MUTANT S-BETA IS BINDING AND
PROMOTE EXPANSIONS, THERE ARE
MANY OTHER LABORATORIES ALSO
WORK ON THAT, HOWEVER, HOW MLH1
CONTRIBUTED TO THIS IS NOT
REALLY KNOW, NOWHERE THE
MECHANISM OUT OF THERE.
SO WE HAVE A 2 POSTER FELLOWS
WHILE WE'RE INTERESTED IN THIS
PROJECT, AND TRIED LOOKING AT
HOW MLH1 DOING AND THEY BELIEVE
THAT MLH1 PROBABLY IPT GREATER
TACT WITH THE HUNTING TON
PROTEIN, NEXT SLIDE, PLEASE.
AND WITH THAT IN MIND, THEY
ACTUALLY TRY TO DETERMINE
WHETHER OR NOT MUTANT L-PROTEIN
IMPACT WITH HUNTING TON PROTEIN,
AND THEY DID WITH PURIFIED
PROTEIN, WITH IMMUNO
PRECIPITATION, WERE ALSO USING
THE CELL, I MEAN THEY JUST
[INDISCERNIBLE], THE MUTANT
L-PROTEIN AND DIFFERENT LENGTH
OF HUNTING TON DEC OR REPEATS
HUNTING TON PROTEIN IN CELL, IT
SHOWS REAL NICELY, SO THE
WILD-TYPE HUNTING TON PROTEIN
SHOWING IN THE WESTERN BLOT CAN
INTERACT WITH BOTH EXO1 AND
MUTANT L-ALPHA.
BUT NOT THE MUTATED HGT, SO BOTH
SHOWED THE SAME RESULT, AND
WE'RE ALSO USING THE PROXIMITY
MEDICATION ASSAY, SHOWN ON THE
RIGHT, YOU CAN SEE THAT THE
SHORTER 1, I MEAN, IF HTT ONLY
CONTAIN 23 CUE, I MEAN SAGE,
REPEAT, AND THEN THEY CAN BUT,
IF YOU INCREASE THIS REPEATS TO
73, AND THE INDUCTION
[INDISCERNIBLE] SO THIS DATA IS
REALLY CONSISTENT WITH OUR
WESTERN BLOT OF DATA, IMMUNO
PRECIPITATION DATA.
NEXT SLIDE, PLEASE.
NEXT WE WANTED TO KNOW WHERE
THIS HTT IMPACTS WITH EXO1, SO
WE CHOP DOWN THE X-WHY INTO
MODIFIED FRAGMENTS AND YOU CAN
SEE THEM AND THEY'RE REGULATING
HOW INTACT WITH HTT AND YOU CAN
SEE ON THE RIGHT AND THEN, WE
HAVE THIS NUMBER OF THE SAME AS
THE FLASH NUMBER ON THE LEFT,
AND FRACTION OR FRAGMENT 1-3,
1-4 AND ALSO SEVERAL AND YOU CAN
SEE SEVERAL ON THE LAB AND IT'S
A FRAGMENT IN BLUE, INTACT WITH
PROTEIN, VERY, VERY WELL, THAT'S
PROBABLY THE BEST, AND YOU HAVE
THAT AND THEN THE FULL
[INDISCERNIBLE].
SO WE BELIEVE THIS IS--THIS IS
THE REASON THE REASON HOW
HUNTING TON PROTEIN SOMEHOW ALSO
REGULATE THE EXO1 ACTIVITY, AND
INTERESTINGLY WE KNOW THIS
IS--THIS REGION, THIS FRAGMENT
IS SERVING IT IS IN THE NUCLEUS
DOMAIN, SO YOU CLUE CLE ACE
DOMAIN, WE SEE IT FOR MUTANT L
PROTEIN, SO NEXT SLIDE, PLEASE.
NOW WE DO EXACTLY THE SAME
ENDORESECTION ASSAY AS I DID
EARLIER, AND THEN YOU CAN SEE,
EXO1 BY ITSELF CAN AJUST THE DNA
INTO MORE PIECES AND THEN, WHEN
YOU--WHEN YOU INCUBATE EXO1 WITH
THE MUTANT PROTEIN, AND YOU KNOW
NO LONGER SHOW THAT EXO 1 ITSELF
BUT IF YOU INCUBATE THE EXO1
WITH MUTANT L-ALPHA OR WILD-TYPE
HTT, THEN CAN YOU SEE THE
NUCLEACE ACTIVITIES IS REALLY
SUPPRESSED, NOW YOU CUT A LITTLE
BIT, NOT A--ALL THE WAY TO
THERE'S MORE PIECES, OKAY.
AND INTERESTINGLY, IF YOU
INCUBATE MUTANT L-ALPHA AND EXO1
AND THE MUTANT HTT, YOU SEND
MUTANT L-ALPHA 1 BY TOPPING IT.
SO NOW THE QUESTION IS, IF WE
THINK THIS EXO 1, AND MUTANT
THERE PROTEIN CANNOT CONTROL EXO
1'S ACTIVITY,--FIND THERE AND
HOW MUTANT L-ALPHA SHOULD BE
ABLE TO CONTROL THE NUCLEUS
ACTIVITY, SO WHY WE SEE THE
PROBLEM IN THE MUTANT CELLS, SO
NEXT SLIDE.
SO THEN WE SAY--I'M SORRY, THIS
IS SHOWING THE ENDORESECTION
ASSAY, WHERE WE WANT TO SAY, WE
ONLY HAVE 1, 1 DOUBLE STRANDED
BREAK AND WANT TO SEE HOW THE
RESECTION, ENDORESECTION GOES,
WE HAVE 3 DIFFERENT PLACES
CONTAINED THIS RESTRICTION SIZE
AND THEN WE WILL BE ABLE TO
MODULATING AMOUNT OF SINGLE
STRAND D NA GENERATED, AND AS
YOU CAN SEE IN THE BOTTOM THAT
YOU REALLY MUTANT CELLS THAT
HTT CELLS THAT DEGENERATE A LOT
OF SINGLE STRAND DNA HAD IS ON
THE RIGHT, THE RIGHT COLUMN, SO
SUGGESTING THAT WHEN HAVE YOU
THIS MUTATED HTT AND THEY'RE NOT
SINGLE STRANDED DNA GENERATED SO
WHAT HAPPENED TO THIS MUTANT L
PROTEIN, WORSE, MUTANT L GOING
INTO FUNCTION WITHOUT THE
PRESENCE OF MTT, SO NEXT SLIDE,
PLEASE.
SO WE HAD THE IMAGE OF THE HALF
LIFE OF THE MUTANT L-ALPHA
EMPLOY OUR HYPOTHESIS IS THAT
MAYBE THIS MUSEUM TABT PROTEIN
CAN DETROIT MUTANT L-PROTEIN,
THAT'S INDEED WHAT WE FIND, THE
HALF LIFE OF MLH1 IS MUCH
SHORTER IN THE MUTANT HTT CELLS
AND THAT'S SHOWED IN THE WESTERN
BLOT AND ALSO ON THE QUANTIFIED
AND THEN WE LOOK AT THE--THE--IN
THE ANIMAL MODEL, IN THE FAR
RIGHT, THIS IS--THIS IS WESTERN
BLOT SHOWING THE WILD-TYPE AND
ACTUALLY HUNTING TON MOUSE
MODEL, AND THEN WE MAJORED MLH1
EXPRESSION AND YOU CAN SEE IN
THE WILD-TYPE IT WAS MUCH HIGHER
COMPARED WITH THE HD-HUNTING TON
PATIENT ANIMAL MODEL.
SO THIS SHOWS THE SAME RESULT IN
CELL LINES SO WE BELIEVE THAT
INDEED THERE ARE MUTANT WITH THE
PROTEIN, PROBABLY, NOW THIS MLH1
DEGRADED THAT'S PROBABLY DO YOU
TO PROTEIN-PROTEIN INTERACTION
BECAUSE THE WILD-TYPE INTERACT
WITH THE PROTEIN BUT THE MUTANT
PROTEIN CANNOT WHICH NOW MAY BE
A NOW MLH1 TO BE DEGREATED IN
THE CELL.
NEXT UP, PLEASE.
OKAY, YEAH, WE SHOW THAT THE DNA
GENERATION AND IN THIS
HTT MUTANT CELLS EMPLOY NEXT
SLIDE, PLEASE AND THEN THE
ACTIVATION OF CGAS, POSSIBLY AS
WE SHOWED IN THE MLH1 DEFICIENT
TUMOR CELLS AND YOU CAN SEE
HERE, IT'S--IT'S REALLY
ACTIVATION OF WHEN YOU MARK IT
ON AGONIST 1, OF COURSE, YOU
ARE--YOU ARE SITTING
PHOSPHORYLATION HAS DISAPPEARED
AND THIS IS IN BOTH--ON
THE--YEAH, WE TRIED TO SHOW, I
MEAN THERE ACTIVATION OF CGAS
PATHWAY IN THE MUTANT CELLS AND
THIS ALSO CAUSE THE CELL DIES,
WHERE ON THE RIGHT WE SHOW THE
CASP A SE 3 AND 9, AND ACTUALLY
CLEAVED IN THE MUTANT CELLS BUT
WHEN YOU'RE KNOCKING OUT AGONIST
1, THIS IS NOW HAPPENING
SUGGESTING JUST ALIKE A MISMATCH
REPAIR DEFICIENT CANCER CELLS,
THAT WOULD REQUIRE EXO1 TO
ACTIVATE THE CYTOSOLIC DNA AND
ALSO CGAS [INDISCERNIBLE].
NEXT SLIDE, PLEASE.
SO HERE'S THE MODEL THAT I--WE
PUTTING TOGETHER AND ON THE LEFT
AND WITH THE NORMAL SITUATION,
AND AS I SHOWED EARLIER IS A
CONTROL OF THE EXO1'S TROLL WELL
BUT ON THE RIGHT, YOU CAN SEE
THE--THERE'S 2 PROBLEMS, 1 THE
HTT PROTEIN IS [INDISCERNIBLE]
TO THE MUTANT PROTEIN, THE
MUTANT PROTEIN DID NOT INTERACT
WITH EXO1 AND AT THE SAME TIME
MUTANT L PROTEINS DEGRADED SO
EXO1, SO MUTANT CONTROLS SO
THAT'S WHY CGAS IS IMPOSSIBLY
ACTIVATED.
IN NEURON CELLS IT CAUSE, I MEAN
OR ACTUALLY WITH APOP TOAZIS, SO
THAT'S WHY YOU CAUSING THE
PATHOLOGY TO THE PATIENT, NEXT
SLIDE, PLEASE.
WELL I'M ACTUALLY--IN THE NEXT I
WILL TRY TO GO FAST, SO IN THE
LAST MAYBE 2 MINUTES, I WILL TRY
TO TALK ABOUT ANOTHER ACTIVITY,
WE IDENTIFIED AND WE HAVE NOT
PUBLISHED YET WHICH ISY
SHORTENING THE MUTANT L,
FOUNDATION IN INFERTILITY.
NEXT SLIDE, PLEASE.
SO IT HAS BEEN SHARED ALMOST 30
YEARS AGO THAT MALE MICE
DEFECTIVE MLH1 OR PMS2 ARE
INFERTILE.
AND BECAUSE THESE ANIMALS CANNOT
HAVE A NORMAL MEIOSIS AND THEY
CANNOT HAVE A NORMAL SPERMAT O
GENESIS, AND THEY ALSO HAVE
SMALLER TESTIS, AND NOBODY KNOWS
WHAT REALLY--I MEAN, WHY THIS IS
CAUSED.
AND THEN AT A CERTAIN TIME, WE
NOTICE THAT THERE'S ANOTHER
ANIMAL THAT'S DEFECTIVE IN YAPP,
ARE ALSO SHOWING SOME MORE
TESTIS AND KIND THAT GENERATE
MATURE SPERM AND SO, IT'S ALSO
COMPLETELY INFERTILE.
SO NEXT SLIDE, PLEASE.
SO WE BELIEVE THAT THIS MAY BE
DOING SOMETHING, WELL IT MAY BE
DOING SOMETHING WITH YAPP, SO
FIRST ORDER IS THE YAPP, IT'S
ACTUALLY THE TRANSCRIPTION CO
ACTIVATOR AND IT'S A DOWN STREAM
EFFECT OF HIPPO PATHWAY WHICH
CONTROLS ORGAN GROWTH AND THE
QUEPMENT.
AND THIS IS WHAT A YAPP DOES.
SO YAPP NORMALLY CAN BE
PHOSPHORYLATED BY MASTER 1 OR 2,
AND THIS WILL BE DEGRADED IN
CYTOSOL, BUT OUR WORK GOES INTO
NUCLEUS AND THEN PLAY A CO
ACTIVELY, TRANSCRIBING CO ACTIVE
FUNCTION AND EXPRESS A NUMBER OF
YEENS INVOLVED IN ORGAN
DEVELOPMENT OR GROWTH EMPLOY SO
NEXT SLIDE, PLEASE.
SO WE BELIEVE THAT MLH1 PROBABLY
CONTROLS TESTIS DEVELOPMENT BY
REGULATING PATHWAY, SO THIS
OTHER WORK TEND TO POST
[INDISCERNIBLE] AND THIS CATANOT
PUBLISHED YET.
NEXT SLIDE, I WILL GO REAL
QUICK.
OKAY, THE FIRST IMMEDIATE
INTERACTION BETWEEN MUTANT
L-PROTEIN AND THE YAPP, AS YOU
CAN SEE HERE THIS IS A POA ASSAY
AND YOU CAN SEE WHEN THEY HAVE
THE MLH1 ON WITH THE YAPP AND
YOU CAN SEE THEY ARE LINKED
TOGETHER.
AND THE IMMUNO PRECIPITATION
ALSO SHOWED ON THE RIGHT THAT
THOSE YAPP OR IRPT ACTION WITH
THE MLH1, OVER YAPP, I MEAN
MOUSE 1 IS THE KINASE FOR YAPP,
THOSE 2 PROTEINS TOGETHER BUT IF
YOU USE MLH1, IT'S A BASE TO
IMMUNO PRECIPITATION, AND YOU
OWN IT FOR YAB AND THAT'S' THE
1, SO THIS SHOWS THIS
INTERACTION BETWEEN MLH1 AND THE
YAPP.
NEXT SLIDE, PLEASE.
SO WE KNOW HOW THIS INTERACTION
WORKS AND WE HOE THIS
INTERACTION ACTUALLY INHIBITS
YAPP POSFORALATION FOR LATS1.
YOU CAN SEE IN THE LAB, IN THE
WILD-TYPE, THE YAPP IS VERY
LITTLE, WHEN YOU KNOCK OUT MLH1,
PHOSPHORYLATION YOU SEE A LOT OF
PHOSPHORYLATED YAPP, SO
SUGGESTING THE MLH1 YAPP CAN
KEEP YAPP UNPHOSPHORYLATED AND
MAYBE YAPP HAS THE CHANCE TO
BUILD A TRANSCRIPTION PHASE SO
THAT IT'S AFFECTUATED, AND THEN
THEY ARE TESTED TO HOW THIS
PROTEIN INTERACTOR WORKS IN THE
INTERACTION DOMAIN, AND WE FIND
OR IN THE RIGHT HERE, JUST HERE
AND THEN WE SHOWED ON THE
BOTTOM, MLH1 WITH YAPP IN THE
WW1 AND WW2 DOMAIN, MOST DOMAIN,
IT'S ACTUALLY THE SAME DOMAIN
THAT YAPP INTERACT WITH LATS 1,
SO IT'S THE KINASE AND WE THINK
ABOUT THAT AND THIS INTERACTION
BLOCKS YAPP, [INDISCERNIBLE]
BECAUSE IT'S COMPETING FOR
BONDING.
NEXT SLIDE, PLEASE.
SO WE ALSO KNOW THAT YAPP, WE
CHECK, YAPP HAS SO FAR AS
NO--NOT BEEN IDENTIFIED WITH THE
NUCLEAR TRANSLOCATION SEQUENCE,
SO THE INTERACTION PROBABLY HELP
YAB, LOCATED 2 NUCLEACE AND THAT
YOU CAN SEE HERE WHEN YOU ARE IN
THE WILD-TYPE YOU WILL SAY YAPP,
OR YAPP WAS RED, YAPP, LOCATED
IN THE NUCLEUS, VERY WELL, SO
YOU KNOCK OUT MLH1, AND MOST OF
YAPP IS OUTSIDE OF THE NUCLEUS,
RESILIENCE STORE MLH1, YOU GET
THE NUCLEUS BACK IN AND
SUGGIESTING NLH1 DOES HELP YAPP
TRANSLOCATE INTO A NUCLEUS.
ON THE RIGHT, IF YOU ARE
KNOCKING OUT THE TRANSLOCATION
SEQUENCE IN MLH1 AND YAPP CANNOT
GO THROUGH THE NUCLEACE, BUT NOT
HUNDRED PERCENT, ANY TO
NUCLEACE, THEY SHOW NOW, THAT
IT'S NEIGHBORED SO YOU CAN SEE
THE BOTTOM 1 WHEN YOU KNOCK
OUT--YOU DISRUPT THE NUCLEAR
SEQUENCE AND YOU SEE THAT YAPP
IS OUTSIDE SO SUGGESTING THAT
MLH1 HELP YAPP ANY INTO
NUCLEACE.
NEXT SLIDE, PLEASE.
SO NOW WHAT WE WANT TO KNOW
WHETHER OR NOT THAT'S REALLY
HELP YAPP CONTROLLED GENE IN THE
TOP LEFT, YOU CAN SEE, WHEN YOU
KNOCK OUT MLH1, YOU DO NOT
AFFECT THE YAPP EXPRESSION, AND
THEY ARE THE SAME, BUT IF YOU
LOOK AT A YEEN THAT CONTROLLED
IT BY YAPP, FOR TRANSCRIPTION,
THE MLH1 KNOCK OUT DRAMATICALLY
REDUCE THE EXPRESSION, SO THOSE
ARE THE 1S YOU KNOW IT'S
DEVELOPMENT SO THAT'S PROBABLY
THE REASON WHY MLH1 DEFICIENCY
CANNOT GENERATE MATURE SPERM OR
THEN DEVELOPMENT OF TESTIS.
AND NOW, THIS MLH1 DEFICIENT
CELLS TO MICE, GO INTO IN PHOTO,
NEXT SLIDE, PLEASE.
THIS IS THE SUMMARY SLIDE FOR
THIS 1.
AND IN TERMAL CIRCUMSTANCES, IT
SHOWED IN THE LEFT, YOU HAVE
MLH1 INTACT WITH YAPP, WHICH
COMPETE WITH THE 1 FOR BONDING
WITH YAPP, AND THEN PROTECT YAPP
NOT FROM PHOSPHORYLATION SO YAPP
CAN GO INTO NUCLEUS SO THEY
CHANGE SOME OF THE DEVELOPMENT
GENE CAN EXPRESS, SO NOW TESTIS
TO HAVE A NORMAL DEVELOPMENT BUT
IF YOU DON'T HAVE THE MLH1 AND
YOU ARE YAPP OR IT'S MOSTLY
DEGRADED AND THEN ONLY A FEW OF
THEM CAN GO TO A NUCLEACE, SO
THAT'S WHY YOU ARE TESTING, IT'S
NOT NORMAL--NEXT SLIDE, PLEASE.
SO THIS CONVERT MY TALK AND IN
THE FINAL THERE, I WOULD LIKE TO
THANK THE PEOPLE WHO DID THE
WORK, I WOULD MENTION SOME OF
THEM, AND THE HIGHLIGHTED IN THE
SLIDE DID THE MAIORITY OF THE
WORK AND ALSO LISTS ALL THE
COLLABORATORS, MANY, MANY
RECRUITING, MANY COLLABORATORS
AND SO MANY OTHERS.
THANK YOU SO MUCH FOR LISTENING.
YEAH.
>> GREAT.
THANK YOU DR. LI FOR THE
WONDERFUL TALK.
WE WILL TART THE Q&A SECTION.
LET'S SEE, THERE IS A--KAREN DO
YOU WANT TO START?
>> YES.
SO PHILLIP JORDAN'S ASKED GOING
BACK TO THE CGAS PART OF THE
TALK THAT BASED ON THE CGAS
EXPRESSION,--SORRY EXPRESSION,
FOR IMMUNOTHERAPY, ARE THERE
ATTEMPTS TO INCREASE THESE
PROTEINS EXPERIMENTALLY TO SHOW
IT CAN POSITIVELY INFLUENCE
THEIR RESPONSE?
>> WELL, YEAH, THE ANSWER IS
YES, WE HAVE LIKE H6 CELL, THE
HTT6 CELL LINE IT HAS WILD-TYPE
CGAS GENE BUT UNFORT MATILY THE
PROMOTER OF CGAS GENE IS
[INDISCERNIBLE] AND THE GENE IS
NOT EXPRESSED BUT WHEN WE PUT IN
THIS H6 CELL IN ANIMAL MODEL FOR
TRYING TO DO THE IMMUNOTHERAPY
IS A LOT OF WORK BUT APPROXIMATE
WE RESTORE THE EXPRESSION OF
CGAS AND THEN IMMEDIATELY THE
RESPONDED TO ANTIBODY AGAINST
THE PDL1, PD1, THERAPY COMING
BACK O THAT'S REALLY--THAT'S THE
ONLY DATA WE HAVE, BUT IN HUMAN,
IN HUMAN PATIENT AS I SHOWED YOU
IN 1 OF THE SLIDES, THOSE
PATIENTS EXPRESS CGAS OR STING
VERY WELL AND THEY RESPONDED TO
IMMUNO THERAPY ANDARY
IMMUNOTHERAPY VERY POSITIVE.
>> OKAY, I WILL GO WITH THE
SECOND QUESTION WHICH IS
ACTUALLY ALSO FROM PHILLIP
JORDAN.
THE QUESTION IS, MLH1S OR 3,
INVOLVE IN CROSS OVER FORMATION
BETWEEN HOMOLOGOUS CHROMOSOME
DURING [INDISCERNIBLE] GENESIS
WHICH MEANS A SPERM O GENESIS
AND O-GENESIS, MLH1 OR 3
MUTATION RESULT IN ALMOST A
COMPLETE LOSS OF CROSS OVER
EVENTS WITH ALL SPERMAT O
SIGHTS, THIS LED TO ANUE
EMPLOYEDY AND INFER TITTY.
IS THE YAPP SHOWING ANY SIMILAR
LOCALIZATION, FOR EXAMPLE AT
CROSS OVER SITES OR DIM
SARDEFECTS WHERE GAP IS MUTATED.
NWOW, THAT'S A GOOD QUESTION,
BECAUSE I DO HAVE AN ASSAY, YET,
BUT I THINK--WELL I CANNOT BUT
GUESS BECAUSE YAPP IS DOING
SOMETHING, MANY NORMALLY IN
TRANSCRIPTION, BUT MLH1 I
BELIEVE, PROBABLY HELP, HELP
YAPP TO TRANSLOCATE FROM
CYTOSOLTO NUCLEACE BUT WITHOUT
THE MLH1, YAPP PROBABLY HAS A
VERY LIMITED AMOUNT OF YAPP OR
BE ABLE TO TRANSLOCATE IT INTO
NUCLEISOTOPE, SO THAT'S GOING TO
CAUSE THE--PARTICULARLY THE
TESTIS DEVELOPMENT ISSUES.
BUT I'M NOT SO SURE,--THERE
WAS--I WAS WITHOUT MLH1, MLH3, I
HAVE NO IDEA WHETHER OR NOT YAPP
INTERACTION WITH MLH1 ABSOLUTELY
OR PMS, 2, MLH3, I REALLY DON'T
KNOW WE HAVE NOT GOTTEN TO THAT
POINT YET BUT I WILL SAY THE
LOCALIZATION OF YAPP WITHOUT
MLH1, MOST OF THEM WILL BE IN
THE CYTOSOL, SO THEY'RE PROBABLY
IN THE CYTOSOL.
>> OKAY, NEXT QUESTION FROM
GEN-LI, VERY NICE TALK AND THERE
ARE 2 QUESTIONS.
HAVE YOU SHOWN THAT
[INDISCERNIBLE] ALPHA AND
HTTMODDULATE THE ACTIVITY TO
INCREASE THE SECTION, 2
QUESTIONS, WHICH PROTEIN IS
RESPONSIBLE FOR THE DECKER NA IN
RECOGNITION?
IS ANY [INDISCERNIBLE] PROTEIN
INVOLVED IN THIS PROCESS?
AND I WILL ASK YOU THE SECOND
QUESTION LATER.
>> SAY IT AGAIN, I DID NOT HEAR
CLEARLY.
>> WHICH PROTEIN IS RESPONSIBLE
FOR THE DNA END RECOGNITION?
DURING THE PROCESS OF WHERE
MUTANT L ALPHA IS BLOCKING HTT?
>> OH, MUTANT L-ALPHA, I THINK
IN THIS CASE MUTANT L-ALPHA
INTACT WITH HTTTHEY BOTH CAN
CONCUR NUCLEACE ACTIVITY, THEY
ONLY NEED 1 BUT IN CASE OF
MUTATING THE HTT, BECAUSE YOU
YOU NOTICE THE HTT ALREADY AND
NOW IT'S HTT AND THE
AGG NORMALLY, WILD-TYPE WITH THE
MLH1 WHICH WE BELIEVE WAS
STABILIZE MLH1.
BUT THE RESULT OF WILD-TYPE
PROTEIN, MLH1, PARTICULARLY I DO
KNOW IN BRAND CELLS MAYBE
FRAGILE TO DEGRADE IT--
>> [INDISCERNIBLE] PROTEIN
SYSTEM NOT INVOLVED?
[INDISCERNIBLE] PROTEINS.
>> [INDISCERNIBLE] PROTEINS WE
HAVE NOT TESTED BUT YES, IT
COULD INTERAT WITH THE L-PROTEIN
BUT THAT WILL REQUIRE A
MISMATCH, SO YOU HAVE TO HAVE A
DNA, SO THAT THOSE 2 PROTEIN
PROBABLY GOING TO IMPACT VERY
WELL WITH THE MISTACH REPAIR
REACTION IN THE ABSENCE OF
MISMATCH, I DO NOT KNOW HOW IT
PROTEIN WILL BE A CHANCE TO GET
IT TOGETHER.
>> OKAY, THE FOLLOW UP QUESTION
IS HTTISSUING INVOLVEMENT NEURON
CELLS SPECIFIC?
OR IS IT A GENERAL MECHANISM
THAT'S ACTIVE IN ALL CELL TYPES
>> YEAH, I AM NOT A
NEUROBIOLOGIST.
I CANNOT ANSWER THIS QUESTION,
BUT AT THE LEAST IN THE CELL, WE
ARE STUDYING ALTHOUGH WE ARE
TRYING TO USE THE ANIMAL MODEL,
I MEAN DERIVED FROM NEURON
CELLS, BUT IN OTHER CELLS, EVEN
WITHIN THE CELL, WE FIND, YOU
STILL HAVE HTT AND THOSE
REACTION, WHAT I TOLD YOU IS
ACTUALLY HAPPENING IN NONNEURON
CELL AS WELL.
>> MAY I ASK A QUESTION OF MY
OWN AT THIS POINT DO WE ARE
TIME?
>> YEAH.
>> SO, AS YOU KNOW MLH1 IS
REQUIRED FOR EXPANSION IN
HUNTING TON'S DEC, AND AUTOKNOWN
THAT EXPANSION CONTINUES IN
NEURONS THROUGHOUT THE LIFETIME
OF THAT NEURON SO OBVIOUSLY MLH1
REMAINS EVEN THOUGH MUTANTS
HUNTING TON'S IS THERE.
CAN YOU SPECULATE WHY THAT IS?
SO YOU MIGHT EXPECT THAT, WELL,
ARE YOU SUGGESTING THAT
INCREASED MUTANT HUNTING TON
WOULD EVENTUALLY LEAD TO
EXPANSION TOPPING IN THAT CELL
TYPE?
BECAUSE THAT DOESN'T SEEM TO BE
WHAT'S HAPPENING IN NYEAH, THE
EXPANSION OF HUNTING TON OR
EXPANSION GOING TO CAUSE--WELL,
I WILL SAY, MUTANT L-PROTEIN
WILL NOT BE PROTECTED WHEN YOU
HAVE 1 MUTATED HUNTING TON
PROTEIN BECAUSE WE KNOW THEY ARE
HTT AND MLH1 INTACT VERY WELL
ALTHOUGH WE DO NOT--ESTIMATION
THAT WE HAVEN'T TESTERRED
WHETHER OR NOT, IN BRAND CELLS,
THE MLH1 PROTEIN, INTACT WAYS
CLEARLY THROUGH MLH3, I BELIEVE
WILL BUT AT THE LEAST, I MEAN,
MAYBE SOME PART OF THE MLH1
PROTEIN IS STILL ENACT WITH
HTTBUT WHEN YOU HAVE A MUTATED
EDGE, THAT PART OF THE MUTANT
L-IS NOT IMPACTED WITH THE
DEGRADED THAT'S PROBABLY THE
REASON WE SEE IN OUR STUDY.
>> OKAY, SO SINCE TIME IS
RESTRAINED WE WILL GO TO THE
LAST QUESTION OF THE DAY SO THIS
IS FROM [INDISCERNIBLE].
DID YOU LOOK AT THE ROW OF POLO
LIKE CAIN ACE IN ACTIVATING MLH1
IN MALES IN STUDIES SUCH AS
CANNED MEN AND WOMEN ACTIVATES
MLH1 WITH THE INTERACTION WITH
EXO1, EXO1 IN THIS CASE MAY JUST
BE MEDIATE BE THE INTERACTION OF
CDC5 WITH MLH1?
>> WOW, THAT'S A QUESTION, I
REALLY, REALLY DON'T HAVE ANSWER
FOR.
SO WHAT THE ENZYME HE WAS
TALKING ABOUT, I DID NOT
HEAR--THE COMPONENT, THE ENZYME
>> THE COLO-LIKE PRK,.
>> NO, NO, I HAVE NO IDEA.
>> I DON'T HAVE A LOT OF
INFORMATION.
NO I CANNOT ANSWER THOSE
QUESTIONS.
>> OKAY.
OKAY, SO, I THINK THAT'S PRETTY
MUCH ALL THE QUESTIONS.
SO, AGAIN, PLEASE JOIN ME TO
THANK DR. LI AGAIN FOR THIS
WONDERFUL TALK AND PLEASE BE
AWARE THAT DR. LI IS IN CHINESE
TIME ZONE SO IT'S ALREADY
MIDNIGHT, SO, AGAIN, PLEASE
THANK YOU FOR THE WONDERFUL TALK
AND TO HAVE A GREAT GOOD REST.
>> OKAY, THANK YOU SO MUCH,
KAREN AND CHEN.
THANK YOU.
>> THANK YOU NTHANK YOU.
>> THANK YOU TAKE CARE.
>> BYE.