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>> GOOD AFTERNOON, EVERYONE.
WELCOME TO THE WEDNESDAY AFTERNOON LECTURE
WE ANTICIPATE SOME SORT OF A
SNOW STORM BUT I'M GLAD YOU ALL
ARE HERE AND THOSE WATCHING ON
THE WEB PROBABLY IN GREAT
NUMBERS BECAUSE WE HAVE A
SPECIAL SPEAKER TODAY,
DR. CHRISTINE SEIDMAN KNOWN TO
ALL OF US WHO KNOW KNOW HER WELL
AS FRIEND AS KRICKET.
IN MEDICINES AND W. SMITH
GENETICS AT HARVARD, ALSO THE
GENETICS DIRECTOR AT BRIGHAM AND
INVESTIGATOR AT HOWARD HUGHES
MEDICAL INSTITUTE.
SHE DID HER UNDERGRADUATE WORK
AT HARVARD.
GOT HER MD AT GEORGE WASHINGTON
UNIVERSITY.
DID RESIDENCY AT JOHNS HOPKINS
WHERE SHE WAS INFLUENCED
PARTICULARLY BY ONE VICTOR
McCUESICK IN TERMS OF HOW TO
BRING SCIENCE AND MEDICINE
TOGETHER IN INTERESTING WAYS.
AND BOY HAS SHE LIVED UP TO THAT
EXHORTATION BY DR. McCUEBIK.
SUBSEQUENT TO THAT SHE DID
RESEARCH FELLOWSHIP IN MEDICINE
IN THE CARDIAC UNIT IN HARVARD
AND SHE'S BEEN CONTINUING SINCE
THAT TIME IN BOSTON AS I
MENTIONED NOW AS PROFESSOR IN
BRIGHAM AND WOMEN'S HOSPITAL IN
HARVARD MEDICAL SCHOOL.
SHE, MORE THAN ANYONE ELSE, HAS
REALLY DEFINED THE
CARDIOVASCULAR GENETICS FIELD AS
IT RELATES TO CARDIAC FAILURE,
BUT ALSO TO CARDIAC
DEVELOPMENTAL ABNORMALITIES AND
TAKEN A FIELD LARGELY
DESCRIPTIVE AND CAUSED IT TO
BECOME MOLECULAR WITH MANY
CONSEQUENCES BOTH FOR DIAGNOSIS
AND INTERVENTIONS TO ASSIST
PEOPLE WHO'S FAMILIES ARE
AFFLICTED BY THE DISORDERS.
IT'S A GREAT PLEASURE TO HAVE
THE OPPORTUNITY TO HAVE HER HERE
FOR THIS WEDNESDAY AFTERNOON
LECTURE.
PLEASE WELCOME KRICKET SEIDMAN.
[APPLAUSE]
>> DR. COLLINS, THANK YOU SO
VERY, VERY MUCH AND TO ALL OF
YOU WHO STOOD WITH ME AGAINST
THIS NASTY STORM THAT IS COMING
FROM THE SOUTH AND HOPEFULLY NOT
GOING ALL THE WAY TO BOSTON, I
REALLY APPRECIATE YOU COMING
THIS AFTERNOON.
IT'S AN INCREDIBLE HONOR TO BE
HERE.
SOME OF YOU MAY KNOW THAT I
SPENT A VERY IMPORTANT YEAR IN
MY LIFE AFTER LEAVING JOHNS
HOPKINS IN NICHD BUILDING 6 WHEN
IT WAS A LITTLE BITTY SMALL
PLACE AND IT'S ALWAYS FUN TO SEE
HOW MUCH THIS HAS GROWN AND
PARTICULARLY FUN TO SEE HOW MANY
INCREDIBLY TALENTED, BRIGHT,
SMART, AND OFTEN FEMALE
SCIENTISTS ARE ARE HERE TODAY.
THANK YOU FOR HAVING ME COME
BACK.
WHAT I'D LIKE TO DO IN THE NEXT
45 MINUTES OR SO IS TO TELL YOU
A LITTLE BIT ABOUT HOW WE'RE
REALLY USING GENETICS TO
UNDERSTAND CAUSES OF HEART
DISEASES THAT OCCUR IN RARE
FAMILIAR FORMS AND ALSO IN
POPULATIONS, AND EQUALLY
IMPORTANT, HOW WE ARE SEEING THE
COMPLEXITY OF USING THIS NEW
TECHNOLOGY TO TRY AND UNDERSTAND
HOW SIMILAR PHENOTYPES THAT
ARISE IN A SINGLE INDIVI
MIGHT BE INTERPRETED USING
GENOME SEQUENCING.
FINALLY N THE LAST LITTLE BIT, I
HOPE TO BE ABLE TO TELL YOU A
LITTLE BIT ABOUT HOW THERE ARE
NEW STRATEGIES.
I'M GOING TO TELL YOU ABOUT ONLY
ONE BUT MANY ARE ARE STILL UNDER
DEVELOPMENT, TO TRY AND USE THAT
GENETIC INFORMATION TO ACTUALLY
USE AND CREATE GENETICS THEY
WERE P PI.
A STUDY UNIT F UH YOU WILL, IS
GOING TO BE AS DR. COLLINS SAID,
ON CARDIO MYOPATHY OF SOMETHING
THAT I'VE BEEN INTERESTED IN FOR
A VERY LONG TIME.
I WILL REMIND ALL OF YOU THAT
THIS IS BY FAR AND AWAY THE MOST
BEAUTIFUL ORGAN IN YOUR BODY.
NOT ONLY DOES IT FUNCTION 24/7
BUT FOR THOSE OF YOU WITH A
STETHOSCOPE YOU KNOW IT SING
TOSS YOU.
IF EVER YOU'RE UNSURE OF WHETHER
YOU WANT TO CONTINUE IN YOUR
PATH OF MEDICINE OR NOT, JUST GO
LISTEN TO A HEART WITH A GREAT
MURMUR AND I'M SURE YOU'LL COME
ALONG TO THE CARD AWAYSIDE OF
THINGS.
UNFORTUNATELY FACED WITH THE
MANY INSULTS WE THROW TO OUR
HEARTS AND ENTIRE BODIES, THIS
ORGAN OFTEN UNDERGO A LOT OF
REMODELLING, BUT UNLIKE MANY
OTHER DIFFERENT TISSUES, IT
REALLY TAKES ONE OF TWO DISTINCT
COURSES.
EITHER RESULTS IN CARDIAC
HYPERTROPHIES WITH THE WALLS
-- AND CAVITY SIZE IS
DIMINISHED.
THIS CAN BE A GENETIC CONDITION
OR SPONTANEOUSLY OCCURRING
CONDITION OR THE WALLS BECOME
THIN AND THE INTRAVASCULAR
VOLUME IS INCREASED.
FOR YEARS WE'VE KNOWN THAT LOT
OF SECONDARY CAUSES OF THIS
MIGHT BE DUE TO HYPER TENSION,
VALVE HEART DISEASE OR
MYOCARDIAL ISCHEMIA BUT IN MANY
INDIVIDUALS WE'VE KNOWN THAT
THESE ARISE SPONTANEOUSLY AND
OFTEN IN YOUNG PEOPLE WHICH
REPRESENTS FINGERPRINTS OF
GENETIC DISEASES.
OVER THE PAST DECADES THERE HAVE
BEEN THE DISCOVERY OF MANY GENES
THAT GIVE ARISE TO EACH OF THESE
AND WE CALL THESE INTRINSIC
CARDIO MYOPATHIES.
THOSE CAUSING REMODELLING I'VE
SHOWN IN THE UPPER LEFT.
THE MOST IMPORTANT OF THESE ARE
MUTATIONS IN THE SASHGMINUTE
GENE.
SIMILARLY, WE HAV IDENTIFIED
MANY OTHER GENES THAT CAN GIVE
RISE TO PRIMARY VENTRICULAR
DILATION.
ONE HAS BECOME QUITE A STAND OUT
AND THAT'S [INDISC].
I'D LIKE TO TELL YOU A LITTLE
BIT MORE ABOUT THESE TWO
DIFFERENT CLASSES OF PROTEINS
AND ARE REMODELLING STRUCTURES.
THE WAY IN WHICH WE KNOW THAT
THESE ARE DEFINITIVE CARDIO
MYOPATHY GENES REALLY IS BASED
ON LOTS OF DISORDERS IN WHICH
INDIVIDUALS WHO ARE KNOWN TO
HAVE A GENETIC RELATIONSHIP,
THEIR SISTERS, BROTHERS, AUNTS,
UNCLES, HAVE BEEN IDENTIFY BY
ROUTINE IMAGING STUDIES
SOMETIMES ASSOCIATED WITH
UNFORTUNATELY SUDDEN CARDIAC
DEATH EVENTS WHICH WE CAN SEE
CARDIO MORPHOLOGY AND THIS ENA
ABLES VERY LARGE PED DRI
DEBRISES.
THIS LED TO GENETIC LINKAGE
ANALYSES.
REASON WE KNOW THIS IS SO
DEFINITIVELY TRUE IS BECAUSE THE
STATISTICAL POWER OF A VERY
LARGE FAMILY IS QUITE REMARKABLE
AND A LOT OF YOU CAN PREDICT HOW
RANDOMLY SUCH A COINHERITANCE
WOULD OCCUR ALL IN THE 40 PEOPLE
INFECTED ON THIS SCREEN AS
COMPARED TO THOSE WHO ARE NOT.
AS YOU CAN IMAGINE THAT
STATISTIC SHOWS THAT THAT WOULD
BE EXTRAORDINARILY UNLIKELY TO
OCCUR BY CHANCE.
THINK ABOUT WHERE WE ARE TODAY.
TODAY, INSTEAD OF USING THAT
ROVING CIRCLE THAT TAKES SEVERAL
POSTDOCTORAL YEARS TO THIN, WE
CAN GO DIRECTLY TO ACTUALLY
SEQUENCING NOT ONLY XOEMS BUT
INCREASINGLY GENOMES WITH ONE
BOX THAT CAN SIT ON YOUR TABLE
TOP.
WE KNOW THE THIS HIGH CAPACITY
NOT ONLY GIVES US THE ACTUAL
GATC SEQUENCE BUT IT ALLOWS US
TO REALLY ROBUSTLY INTERROGATE
WHETHER THE NUMBER OF EACH OF
THOSE SPACE PAIRS ARE
APPROPRIATELY TWO COPIES, ONE
FOR EACH OF THE AUTOSOMES OR IF
THERE ARE A COPY NUMBER OF
INVARIANTS IN GENES A NEW SOURCE
OF HIDDEN PATHOLOGIES.
IF WE HAVE AN INDIVIDUAL WHO'S
XOEM OR GENOME WE CHOOSE TO
SEQUENCE FROM A PEDIGREE SUCH AS
THIS AND FIND A VERY RARE
MUTATION, WE MIGHT HAVE A FAIR
DEGREE OF CONFIDENCE THAT THAT
MIGHT CAUSE THE DISEASE.
IT BECOMES ALL THE MORE POWERFUL
IF WE LOOK AT ANOTHER DISTANTLY
RELATED INDIVIDUAL WHICH SHARED
ONLY ONE 32nd OF HIS GENOME WITH
HIS COUSIN OVER HERE AND THAT
MAKES IT HIGHLY UNLIKELY THAT
THESE SINGLE RARE VARIANCES
IDENTIFIED IN BOTH OF THESE
INDIVIDUALS WOULD HAVE ARISEN BY
CHANCE.
SORT OF THE XOEM E -- WHAT
HAPPENS WHEN WE DON'T HAVE NO
FAMILY HISTORY OF DISEASE AND A
SINGLE PATIENT WHO COMES IN?
CAN WE MAKE THOSE SAME
PREDICTIONS THAT VARIANT WE IE E
DENT FIE IN A GENE, EVEN ONE
KNOWN TO CAUSE DMZ THE CONTEXT
OF FAMILIES, DO WE KNOW THAT
ACCOUNTS FOR THE DISEASE IN THIS
INDIVIDUAL?
I WANT TO BRIEFLY REMIND YOU
THAT IN ADDITION TO HAVING
VARIANCE WE INHERIT, WE ALSO
HAV THE OPPORTUNITY TO ACQUIRE
OUR OWN PERSONAL VARIANCE.
IT'S ESTIMATED THAT THERE WAS
APPROXIMATELY ONE RARE DE NOVO
VARIANT IN EACH AND EVERYONE OF
US THAT ABOUT ONE IN 50 OF US
WILL CARRY A COPY NUMBER OF
VARIANTS AS WELL AND IF WE LOOK
NOT JUST AT XOEM SEQUENCES BUT
INDEED ACROSS THE GENOME, WE SEE
THERE ARE UH NEW VARIANCE ARISEN
IN US.
THE SIGNIFICANCE BECOMES
DIFFICULT TO INTERPRET WITHOUT
FAMILY MEMBERS AND WITHOUT
KNOWING A WHOLE LOT MORE ABOUT
THE GENE.
HOW DO WE TAKE OUR ABILITY TO
KNOW GENETIC CAUSES OF DISEASE
AND BRING IT TO POPULATIONS AND
PATIENTS IN A CLINIC?
HOW CAN WE DISCERN THE
SIGNIFICANCE DIFFERENCE IN EACH
AND EVERY PATIENT BEFORE US?
THAT'S GOING TO REQUIRE A VERY,
VERY BIG DATABASE SO I HOPE THAT
NCBI WILL GET MORE FUNDING,
DR. COLLINS.
I THINK THAT CERTAINLY THAT'S
GOING TO REQUIRE A SUBSTANTIAL
HARMONIZATION OF PHENOTYPE SO
THAT WE KNOW WHEN WE SAY THAT
THIS VARIANCE IS FOUND IN THIS
COLLECTION OF PHENOTYPES THAT
WE'RE ALL TALKING ABOUT EXACTLY
THE SAME THING, AND REMEMBER
THAT A LOT OF OUR PHENOTYPES ARE
PRETTY NON-SPECIFIC.
A CHEST X-RAY PROBABLY SEEMS
VERY SIMILAR LIS TICK BUT IT'S
NOT THAT VERY DISSIMILAR FROM
DOING AN ECHO OR MRI OF THE
HEART.
IT'S NOT EXACTLY HIS-SELF LOGIC
ANALYSIS.
SIMILARLY WE'LL COLLECT ALL OF
THE DEFINITIVE MUTATION AND
COMPARE THEM TO THE VARIANCE IN
THE SAME GENE.
ADD IN WHAT HAPPENS WHEN YOU
HAVE DIFFERENT LIFESTYLES,
EXPOSURES, ENVIRONMENTS, AND
ULTIMATELY COME OUT WITH SOME
KIND OF INTEGRATED ANALYSES.
FIRST PART OF THIS TALK I'D LIKE
TO TELL YOU SOME OF THE
STRATEGIES WE'RE TRYING TO USE
TO TRY AND OVERCOME THESE VERY
BIG OBSTACLES AND IN A SECOND
PART HOPEFULLY CONVINCE YOU IT'S
WORTHWHILE BECAUSE I CAN LEARN
SOME INTERESTING BIOLOGY.
SO WHEN WE THINK ABOUT THE
CONTRACTILE APPARATUS, MANY OF
YOU MAY BE REMEMBERING ON THE
SLIDE TO THIS IN WHICH YOU KNOW
THAT THE MOTOR PROTEINS IN ALL
MUSCLE CELLS ARE COMPOSED OF A
HEAVY CHAIN -- COOKING, EXCUSE
THIN FILAMENT AND SLIDE ACROSSH
E KNOWN FOR A LONG TIME THAT.
MUTATIONS IN THE EIGHT MAJOR
COMPONENTS OF THE MOLECULAR
MOTORS OF CARDIAC CELLS CAN GIVE
RISE PRIMARILY TO HYPER TROPHIC
CARDIO MYOPATHY.
THESE ARE FROM FAMILIAR
STUDIES -- AND WE HAVE ROBUST
STATISTICS TO SAY THESE ARE
DEFINITIVE DISEASE GENES.
WE ALSO KNOW THAT CARDIAC
HYPERTROPHY THAT REMODELLING I
SHOWED YOU ON THE FIRST SLIDE IS
FOUND IN GENERAL POPULATION.
4% OF THE GENERAL POPULATION HAS
UNEXPLAINED CARDIAC HIGH PERCH
FI AND LARGELY ATTRIBUTED TO
DIABETES, INCREASE IN BLOOD
PRESSURE AND LOTS OF OH COMMON
CARDIOVASCULAR DISORDER.
WE THOUGHT IT'D BE INTERESTING
TO KNOW IF SASHG MERE -- THIS
STUDY A LOLY COMMUNITY BEING
BURIED IN SNOW AND AS ALL OF YOU
KNOW IT'S HAD ABOUT 50 YEARS OF
LONGITUDINAL STUDY OF OVER
THOUSANDS OF SUBJECTS WHO HAVE
KINDLY PARTICIPATED IN SHOWING
US LOTS OF IMPORTANT PIECES OF
INFORMATION OVER MORE THAN
EIGHT, NOW, DIFFERENT SERIAL
EXAMINATIONS.
WE'VE LOOKED AT THE JACKSON
HEART STUDY, AT LEAST NOW
ONE-AND-A-HALF ROBUST PROFILES
IN WHICH WE HAVE CARDIAC IMAGES
AND LOTS OF ANCILLARY PHENOTYPES
AS WELL.
WHEN YOU TAKE THESE THREE
THOUSAND INDIVIDUALS AND YOU SAY
LET'S JUST SEQUENCE ALL OF THOSE
EIGHT MAJOR SASHG MERE GENES
THAT CAUSE MICRO TROPICAL
MYOPATHY, WE HAD EXPECTED WE
WOULD FIND SOME VARIANCE AND
THAT THAT WOULD CORRELATE WITH
THEIR DEGREE OF HIGH PERCH FI.
WE FOUND 11% OF THE GENERAL
POPULATION HAS A CHANGE IN THE
SEQUENCE THAT ALTERS THE PROTEIN
OF THESE EIGHT SASHG MERE GENES
AMONGST THESE TWO POPULATIONS.
MOREOVER, BECAUSCE THE
DISCOVER ARE RI OF THESE GENES
AS CAUSING HYPOTROPHIC CARDIO
MYOPATHY, A LOT OF CRITERIA HAS
BEEN DEVELOPED TO ACTUALLY
DISCERN WHICH ARE PATHOGENIC
ONES AND WHICH ARE COMMON
VARIANTS.
SO WE APPLIED THOSE TO APPROVE
CLASSIFICATION AND WE COULD
RESTRICT THOSE EIGHT RARE
VARIANCE DOWN TO ABOUT 8.6 WHO
WOULD BE DEEMED PATHOGENIC.
THAT MAY NOT SEEM LIKE A VERY
BIG NUMBER TO YOU BUT THAT'S
ABOUT THREE TIMES MORE PREVALENT
THAN ALL GIVEN REPORTS THE
DISEASE INCIDENCE IS IN A
GENERAL POPULATION WHICH WOULD
MAKE YOU CONCLUDE ONE OF TWO
THINGS.
ONE, [INDISCERNIBLE] VARIANT, OR
TWO, WE'RE MISSING A LOT OF THE
DIAGNOSTIC CRITERIA IN THE
SO WHEN WE LOOKED IN DETAIL AT
THE PHENOTYPES OF THESE VARIANT
CARRIERS.
WE FOUND ONLY ABOUT ONE IN TEN
OF THEM HAD ANY CRITERIA, EVEN
THE [INDISCERNIBLE] OF HAVING
PHENO TIPIC MANIFESTATION.
IT'S NOT AS IF THESE PEOPLE ARE
LIKELY TO DEVELOP IT OVERTIME.
IT'S REALLY IMPLIES THAT WE'RE
NOT CALLING ALL OF THE VARIANCE
IN A CORRECT MANNER.
IT DOESN'T MEAN HOWEVER THAT
THESE VARIANCE ARE COMPLETELY
SILENT BECAUSE AGAIN,
CAPITALIZING ON LONGITUDINAL
PHENOTYPES WHEN WE LOOKED AT
ADVERSE MYOCARDIAL EVENTS THAT
OCCURRED IN CARRIERS IN
COMPARISON TO THOSE IN FRAMING
HAM WHO DID NOT CARRY THE
VARIANT WE SAW ALMOST AN
INCREASE IN EVENTS.
THIS IS AFTER ACCOUNTING THE
USUAL RISK FACTORS.
SO TAKEN TOGETHER WHAT AYE TRIED
TO TELL YOU IS THAT WE'VE KNOWN
FROM DECADES NOW OF RESEARCH
RAL NICK PATHOGENIC NETRANT
MUTATIONS AND THIS OCCURS IN
GENES WE CAN BEGIN TO STUDY IN
GENERAL POPULATION.
WE FIND LOTS OF RARE VASHTS BUT
THE PENETRANTS AND
[INDISCERNIBLE] APPEARS TO BE
SIGNIFICANTLY LOWER.
IT'S NOT ABSENT, IT'S JUST
LOWER.
ONE OF THE BIG CHALLENGES THEN
BEGINS TO SAY WHAT ARE THOSE
MODIFIERS THAT ARE INFLUENCING
THE EXPRESSIVITY; GENETIC?
BACKGROUND?
EXPOSURE?
CAN WE USE THAT INFORMATION TO
TAKE AND MODIFY THE IMPACT OF
THESE PATHOGENIC MUTATIONS?
THAT'S A LONG WAY INTO THE
FUTURE RIGHT NOW, BUT I THINK
IT'S SOMETHING THAT'S GOING BE
IMPORTANT FOR US TO BEGIN TO
GRAPPLE WITH.
I'M GOING SHIFT GEARS NOW AND
TELL YOU ABOUT OTHER FORM OF
CARDIAC REMODELLING.
I WANT TO INTRODUCE WHAT IS
BECOMING THE MOST POPULAR GENE
IN MY PARTICULAR LABORATORY,
TITAN.
TITAN IS NOT TIE-TAN, IT IS THE
LARGEST GENE AND LARGEST PROTEIN
IN THE HUMAN BODY, BUT IT'S A
TITIN AND IT'S HUGE IN TERMS OF
SIZE.
IT ACTUALLY IS APPROXIMATELY A
HUNDRED THOUSAND BASE PAIRS LONG
FOR THE PROTEIN ENCODING
SEQUENCES.
THE MOLECULE IS ALMOST ONE MICRO
METER AND SPANS ONE HALF OF THE
ENTIRE [INDISCERNIBLE].
THIS IS TITIN HAS BEEN LONG
KNOWN TO BE CRITICAL -- IT'S
BELIEVED TO PARTICIPATE IN
DEVELOPING AND SERVING AS A
SCAFFOLD FOR THE DEVELOPMENT OF
THE SASHG MERE DURING
EMBRYOGENESIS.
IT CLEARLY IS IMPORTANT NOT
PARTICULARLY IN FOURTH
GENERATION BUT IN FOURTH
TRANSMISSION AND BECAUSE IT'S
BEEN RECOGNIZED THAT THIS PURPLE
DOMAIN HERE DESIGNATED M, THE M
BAND, HAS KINASE ACTIVITY UH BUT
IT MAY ALSO BE INVOLVED IN
SIGNALING.
AT THE FIVE CRIME END OF THE
MOLECULE, THE Z BAND IS
INTERACTING REGION ANOTHER AREA
KNOWN TO BE CRITICAL FOR
ACTUALLY TRANSDUCING SIGNALS.
SEVERAL YEARS AGO WE BEGAN TO
ASK WHETHER WE COULD FIND CAUSES
OF ID OWE E PATHIC DIELTED
REMODELLING BY LOOKING AT SOME
SMALL SMALLIES AND INCREASINGLY
BECAUSE THERE WERE NOT LARGE
LARGE KINDRE FOR ANALYSES TO
ICE LATED PATIENTS.
WE FOUND RARE FAMILIES THAT WE
COULD IDENTIFY FEW MUTATIONS IN
THE GIANT PROTEIN TITIN.
WE THEN ASKED SINCE WE'VE NOEP
THAT THIS IS A SINGLE GENE THAT
IS VERY BIG BUT MAYBE WE COULD
ALSO BEGIN TO GLEAN STRATEGIES
TO INTERPRET VARIANTS IN SINGLE
INDIVIDUALS BY DOING A DEEP DIVE
OF SEQUENCING THIS GIANT
MOLECULE IN LARGE COHORTS OF
PATIENTS.
AS YOU CAN IMAGINE, WE TOOK THE
SAME STRATEGY I TOLD YOU ABOUT
BEFORE WITH A LITTLE EXCEPTION.
FIRST OF ALL, WE TOOK ABOUT FIVE
HUNDRED INDIVIDUALS WITH DIELTED
CARDIO MYOPATHY WHO HAS BEEN
EXCLUSIVELY SEEN IN TIME.
WE'VE HAD EXPENSIVE MRI WHICH
HAS HELPED TO INFORM SOME OF THE
FEE KNOW TIP I CAN
MANIFESTATIONS.
WE BEGAN SEQUENCE BOTH FRAMING
HAM AND JACKSON OVER THREE
THOUSAND INDIVIDUALS AND BECAUSE
WE KNOW THAT THERE'S A VERY RARE
FORM OF PEDIATRIC ONSET WE HAD A
CHANCE TO LOOK AT A SMALL SUBSET
OF PATIENTS.
WE DID IS DO ROBUST SEQUENCING
USING AN ALUMINUM PLATFORM AND
USE THE STANDARD PIPELINE FOR
ANALYSES.
WHEN WE USE THIS AND THEN BEGAN
A VARIANT CALLING, WE HAD THE
UNFORTUNATE CIRCUMSTANCE OF
RECOGNIZING THAT THE NUMBER OF
VARIANTS IN THIS GENE ALONE ARE
ABSOLUTELY ASTRONOMICAL.
THERE IS ONE SIS SENT VARIANT
THAT IS RARE, NOT IDENTIFIED IN
THE VAST MAJORITY OF DATABASES
OR OCCURRING IN LESS THAN 1% OF
THE POPULATION IN EACH AND
EVERYONE OF US.
WITHOUT A FAMILY HISTORY,
WITHOUT THE CAPACITY TO DEEPLY
EXAMINE THE CONSEQUENCES OF EACH
ONE OF THOSE, THERE ARE GOING BE
A LOT OF VARIANTS THAT ARE OF
UNKNOWN SIGNIFICANCE IN THIS
GENE FOR A VERY LONG TIME AHEAD.
WE'VE RESTRICTED OUR ANALYSES TO
THOSE VARIANCE WHO APPEAR TO
HAVE A MORE PROFOUND CAPACITY TO
CHANGE WHAT TITIN DOES.
THOSE ARE THE ONES THAT ACTUALLY
PREMATURELY TERMINATE THE
ENCODED PROTEIN EITHER BECAUSE
OF A PREMATURE SEQUENCE, A
NONSENSE, OR AN ESSENTIAL SPLICE
SITE MUTATION THAT RESULTS IN
FRAME SHIFT AND ALTERNATIVE
SPLICING.
WHEN WE EMPLOYED ONLY THE
ANALYSES ON THOSE TRUNCATING
MUTATIONS, WE FOUND VERY
INTERESTING RESULTS AS SHOWN
HERE.
FOR THOSE PATIENTS WITH N SAGE
REALLY PROFOUND HEART FAILURE --
THESE ARE INDIVIDUALS WHO HAVE
BEEN REFERRED FOR
TRANSPLANTATION OR WHO ARE
UNDERGOING BAG TREATMENTS WE
FOUND SIGNIFICANT ENRICHMENT IN
THE TITIN TRUNCATION MUTATIONS
FOUND NAH N THAT POPULATION AS
COMPARED TO HEALTHY FOLLOW
VOLUNTEERS AND FRAMING HAM IN
JACKSON.
WE'RE NOT YET PROGRESSED TO
HEART FAILURE, THEY WERE ALSO
SIGNIFICANTLY ENRICHED IN
COMPARISON TO THIS OTHER
POPULATION.
BUT AS YOU'LL SEE, THERE WERE
NOT ZERO PATHOGENIC SOUNDING OR
LOOKING MUTATIONS IN THE GENERAL
POPULATION.
THEY WERE CERTAINLY PRESENT ARE,
ALBEIT SOMEWHERE BETWEEN 1-3%.
THAT MAY NOT SEEM LIKE A VERY
HIGH E PERCENTAGE, BUT IF IT WAS
YOUR HEART, I THINK SOME OF U.S.
MIGHT BE WORRIED ABOUT IT.
WE BEGAN TO ASK WHETHER THERE
WERE OTHER STRATEGIES THAT WE
COULD USE TO KEEP OUT WHICH ARE
THE MEANINGFUL VARIANTS AND
WHICH ARE NOT.
WE TRIED TO ASK WHETHER THERE
WAS A DISTRIBUTION OF THESE
VARIANTS ALONG THE PROTEIN THAT
MIGHT BE INFORMA
SHOWN HERE ON THE TOP ARE THE
AMBULATORY SEVERE PATIENTS IN
THE JACKSON FRAMING HEART
DISTRIBUTED ALONG THIS REGION.
IT'S NOT A VERY IMPRETIVE
CLUSTERING.
SO THE NEXT THING THAT WE ASKED
WAS WHETHER THERE WERE
DIFFERENCES IN THE LOCATION OF
THESE VARIANCE NOT ALONG THE
LONGITUDINAL DISTRIBUTION OF
THIS LARGE MOLECULE BUT THE
DEGREE WITH WHICH THE EXXON IN
EACH OF THOSE ARE EXPRESSED.
LIKE MANY GREAT PROTEINS WITH
CAPACITIES TO PERFORM DIFFERENT
PROCESSES, TITIN UNDERGOES LOTS
OF DIFFERENT ISOFORM SWITCHES --
NOT ISOE FORMS, EXCUSE ME --
EXXON USES.
SO WE RESTRICTED OUR ANALYSIS TO
ASK WHICH EXXONS ARE HIGHLY
EXPRESSED AS COMPARED TO THOSE
LOWLY EXPRESSED.
THE HIGHER THE GRADE PEAK, THE
HIGHER OF THE LEVEL OF
EXPRESSION.
WHAT YOU'LL SEE IS THAT ALONG
THIS REGION HERE, THERE'S
CONSIDERABLE DROPOUT BECAUSE
THESE EXXONS HAS MUCH, MUCH LESS
EXPRESSION WITHIN THE HUMAN
HEART.
YOU'LL ALSO NOTE THAT A LOT OF
THE VARIANTS THAT ARE NOT FOUND
IN OUR DILATED IN -- SO IF WE
ELIMINATE THOSE FROM ONLY THE
LOWLY EXPRESSED EXXON YOU GET A
FAR MORE CLUSTERING KIND OF
APPROACH WITH SUBSTANTIAL
ENRICHMENT ALONG THE A BAND, AND
THAT IS SIGNIFICANTLY ENRICHED
AMONG THOSE WITH DCM COMPARED TO
GENERAL POPULATION WHERE THOSE
THAT ARE ENRICHED IN THE GENERAL
POPULATION TURNED OUT TO BE IN
THE LOWLY EXPRESSED EXXON.
SO THIS IS IDIOPATH DIELTED
HEART IN WHICH THERE IS A GLOBAL
VENTRICULAR DECREASE IN FUNCTION
BUT THERE'S NO REGIONAL WALL
ABNORMALITITY, NOTHING
PARTICULARLY REMARKABLE ABOUT
THE INTENSITY OF THE IMAGE
QUALITY AROUND HERE AND THE HIS
TOE PATHOLOGY IS WHAT I LIKE TO
CALL QUITE VANILLA.
IT DOESN'T TELL YOU EITHER THE
IDEOLOGY OR WHETHER IT'S GOING
TO PROMOTE THE ARRHYTHMIAS IN A
PATIENT FROM WHOM IT'S OBTAINED
OR NOT.
WE ASKED WHETHER KNOWING THIS
MUTATION WOULD ACTUALLY INFORM
PHENOTYPE OUTCOMES BASED ON
DETAILED INFORMATION WE HAD
ABOUT THEM.
FROM OUR PATIENT WITH MRI WE
LOOKED AT THOSE WHO HAD THE
M SHOWN HERE VERSUS THOSE
WHO DID NOT.
YOU SEE A SLIGHT DECREASE IN
LEFT VENTRICULAR EJECTION
FRACTION.
PERHAPS MORE IMPRESSIVE TO ME
WAS THE FACT THAT THEIR WALL
THICKNESS WAS DECREAS
COMPARISON TO OTHER FORMS OF
DIELTED CARDIO MYOPATHY AND
SADLY THE NUMBER OF ARRHYTHMIAS
THEY HAD WERE SUBSTANTIALLY
INCREASED.
THIS IS PROVIDED TO US FOR THE
FIRST TIME THAT WE'RE BEGINNING
TO TEASE OUT A PHENOTYPE THAT
MAY BE GENOTYPE CHECK SLEKTIVE
IN THE DIELTED POPULATION.
DILATED POPULATION.
MOREOVER, WE HAD THE SENSE FROM
THE DISTRIBUTION OF MUTATIONS
AND WHONS THAT ARE HIGHLY
EXPRESSED IN THAT CLUSTERING
THAT THOSE WERE MORE LIKE TO
OCCUR IN INDIVIDUALS WHO HAS HAD
THE SWER -- NOT SURPRISING THAT
DISTRIBUTION ALWAYS IMPLIES THEY
WOULD HAVE THE POOREST EJECTION
FRACTION.
THE FURTHER THE TRUNCATION
OCCURRED TOWARDS THE THREE PRONG
END OF THE MOLECULES T WORST THE
VENTRICULAR FUNK.
THAT WAS TRUE IF YOU HAD DIELTED
MYOPATHY OR IF YOU WERE A
FRAMING HAM HEART SUBJECT AND
SIMPLY CARRIED ONE OF THESE
MUTATIONS SO YOU HAD A DECREASE
IN YOUR FRACTION IN COMPARISON
TO THOSE WHO DID NOT AND IT WAS
WORST THE MORE THREE PRONG TO
THE GENE OR TO THE MORE CAR
BOTTLE END OF THE MOLECULE THAT
BECAME.
WE'VE DONE THOSE SAME KIND OF
ANALYSES IN CHILDREN WHO HAVE
PEDIATRIC ONSET OF CARDIO
MYOPATHY BEFORE THE AGE OF 16.
THIS IS A DEVASTATED DIS ORDER
THAT LEADS TO RAPID DEATH IF
THEY HAVEN'T DONE THEIR
TRANSPLANTATION.
WE'VE NOW LOOKED AT ABOUT 84
GENES THAT HAVE BEEN IDENTIFIED
FROM FAMILY STUDIES IN PATIENT
COHORT AND SUFFICE IT TO SAY
THAT A FEW MUTATIONS THAT HAVE
BEEN IDENTIFIED, NOT VERY MANY.
THERE'S A LOT MORE TO DO IN THIS
COHORT OF PATIENT.
MOST IMPORTANTLY, WE SEE VERY
FEW MUTATIONS WITHIN TITIN,
SIGNIFICANTLY LESS THAN WE DO
WITHIN THE ADULT POPULATION.
TAKEN TOGETHER, I THINK WHAT
THIS INFORMATION HAS GOWN TEASE
OUT IN TERMS OF INFORMATION
THAT'S BEEN RELEVANT TO
INTERPRETING TITIN VARIANTS IS
THAT MOST OF THESE APPEAR TO
TRUNCATE THE PEPTIDE T
PATHOGENIC ONES, THAT THEY OCCUR
IN HIGHLY EXPRESSED EXXONS
WITHIN THE LEFT VENN TRI CALL.
THEY ARE MORE REGIONALLY LOCATED
THAN RANDOMLY DISTRIBUTED ACROSS
THE MOLECULE, AND THOSE THAT ARE
CLUSTERED THE FOR AB END OF THE
MOLECULE ARE CLEARLY ASSOCIATED
WITH MORE SEVERE DISEASE, POORER
LEFT VENN TRICK CALL EJECTION
ACTION AND THEY ARE UNLIKELY IF
YOU FIND THEM IN A PEDIATRIC
ONSET DIELTED CARDIO MY MY OP
THINMYOPOTHY, -- THE FIRST THING TO
DO IS UNDERSTAND MECHANISM.
WHEN WE THINK ABOUT A
POLYPEPTIDE THAT'S TRUNCATED WE
OFTEN ASSUME IT HAD A DIMINISHED
CAPACITY TO PARTICIPATE IN ITS
USUAL ROLES.
THAT MAY BE SO PROFOUND THAT
IT'S NOT INCORPORATED AND THAT
WE ARE HAVE LOSS OF ONE OF THE
FUNCTIONAL GENE COPIES OF THE
MOLECULE OR HAVE ONE
INSUFFICIENCY.
IF WE HAD -- BY THE MECHANISM BY
WHICH TITIN TRUNCATION CAUSED
DIELTED CARDIO MYOPOTHY, WE
WOULD NOT EXPECT A SELECTED
DISTRIBUTION OF DCM MUTATIONS
ARRIVING AT THE THREE-WRONG
PRONGED END OF THE MOLECULE.
RATHER THEY SHOULD BE ABLE TO
TRUNCATE ANYWHERE A ALONG THE
CHAIN.
I'VE TRIED TO CONVINCE YOU THAT
THOSE AT THE AMY KNOW TERMINUS
FAIR FOOR BE FAR LESS
DEVASTATING.
THAT RAISED THE CHANCE THAT THIS
IS -- BUT RATHER IT SUGGESTS
SOMETHING ELSE AND THAT HAS A
LOT TO DO WITH THE GIANT SIZE OF
THIS PEPTIDE.
THIS IS A VERY, VERY LARGE
TRANSCRIPT AND CHANCES ARE THE
FIVE-PRONGED SEQUENCES ARE ARE
TO BE TRANSL AND
ACTUALLY BEGIN TO BE
INCORPORATED INTO THE SAR KOE
MERE EVEN BEFORE THE
THREE-PRONGED SEQUENCES HAVE
BEEN GENERATE INTO PEPTIDES.
MOREOVER, WE KNOW THAT FROM VERY
RARE FAMILIES WHERE THERE ARE
TRUNCATION MUTATIONS AT THE VERY
END BAND T MOST CAR BOTTLE END
OF THE PROTEIN THAT IMMUNE
CHEMICAL STUDIES REVEAL THAT
THOSE PROTEINS ARE INCORPORATED
INTO THE SASHG MERE.
THIS IS A PROFILE OF THESE KINDS
OF PATIENTS.
ANTIBODIES THAT ARE SELECTED FOR
THE Z DISK REGION OF TITIN, THE
A BANDS REGION OF TITINS SHOW
ROBUST EXPRESSION SUGGESTING
THAT THESE PEPTIDES ARE
SUFFICIENTLY STABLE, THAT THEY
ACTUALLY ARE INCORPORATED.
I THINK THIS PROVIDES THE MOST
PROFOUND EVIDENCE THAT THESE ARE
LIKELY TO OPERATE BY A DOMINANT
NEGATIVE MECHANISM.
SO IF THEY'RE DOMINANT NEGATIVE,
WHAT DO THEY DO?
WE HAVEN'T HAD A A CHANCE TO DO
A WHOLE LOT OF WORK ON THIS YET.
I CAN TELL YOU THAT SOME OF THE
HYPOTHESES WE CAN PUT TO REST.
NAMELY THAT THEY MIGHT DISRUPT
FORMATION OF NORMAL SASHG
MEEZ STRUCTURE.
WE'VE DONE LOTS OF ELECTRON
MICROSCOPY AND AS BEST WE CAN
SEE STRUCTURE IS COMPLETELY
NORMAL DESPITE HAVING THESE
PATHOGENIC MUTATIONS.
WHAT WE DO SEE IS LOTS OF
INTERESTING CHANGES IN NUCLEAR
MOREOLOGY, SOMETHING THAT HINTS
THAT THERE MIGHT BE SIGNIFICANT
ABNORMALITIES.
WE KNOW THAT A LOT MORE WORK
WILL HAVE TO BE DONE TO TRY AND
BUILD ROBUST MODELS TO DO THIS,
I WANT TO OUTLINE FOR YOU HOW WE
COULD USE THIS INFORMATION, BUT
IN THIS SENSE, SORRY A
TRUNCATION OF TITIN IN IS ACTING
IN A DOMINANT NEGATIVE MODE OF
ACTION MIGHT PROVIDE A NEW WAY
FOR TREATING.
THE FIRST THING IS TO DEVELOP MODELS IN WH ICH WE CAN ROBUSTLY
LOOK FOR SUFFICIENCY VERSUS
DOMINANT NEGATIVE.
WE CAN DO ALL SORTS OF
TRANSCRIPTIONAL PROFILING THAT
IDENTIFY -- IN PARTICULAR
BECAUSE OF THE INBAND KINASE --
AND THEN IDENTIFY CANDIDATE
MOLECULES.
SEE HOW THEY PERTURB CARDIAC
BIOLOGY AND PHYSIOLOGY AND THEN
SEE IF ANY OF THESE THINGS MIGHT
PLAY OUT AS POTENTIALLY
BENEFICIAL TO OUR PATIENT.
WE'VE DONE A LOT OF THIS ALREADY
IN THE MUTATIONS THAT CAUSE
HYPOTROPHIC CARDIO MYOPATHY
BECAUSE WE'VE BEEN AWARE OF THE
BIOPHYSICAL PROCESSES FOR
SEVERAL YEARS NOW.
WE'VE MADE THE MICE AND DONE THE
STUDIES, AND I JUST WANT TO TELL
YOU LATE BUILT ABOUT WHAT THOSE
STUDIES HAVE INFORMED US.
I WANT TO REMIND YOU THESE ARE
IN THE CONTRACT HALL APPARATUS
THAT ARE GENERATING THESE
SACOMERE POWER AND FORCE BECAUSE
THEY OCCUR IN MYOSIN, ACTIN,
[INDISCERNIBLE] AND THEY RESULT
IN A VERY REMARKABLE HIS TOE
PATHOLOGY THAT IS QUITE
DIFFERENT FROM DILATED CARDIO
MYOPATHY.
THEY'RE ENLARGED, SEPARATED BY
MATERIAL -- THIS KIND OF THIS
MORPHOLOGY ULTIMATELY RESULTS IN
[INDISCERNIBLE] DYSFUNCTION AND
PRODUCES THE [INDISCERNIBLE] WE
SEE IN PATIENTS.
HOW COULD A MYOSIN MUTATION DO
THIS?
THE FIRST THING WE NEED TO DO IS
UNDERSTAND WHAT DOES THE SIGH
SIN MUTATION DO TO SACOMERE
PERFORMANCE?
W THE UNIVERSITY OF VERMONT
AND STANFORD, WE'VE BEEN ABLE TO
ENGINEER HUMAN MUTATIONS EITHER
INTO CELLS AND HAVE THEM PRO
BUSTLY EXPRESSED IN VITRO OR,
INDEED, FROM MOUSE MODELS IN
WHICH WE CAN ISOLATE THE MUTANTS
FROM THOSE ANIMAL HEARTS.
AND WHAT WE'VE SEEN IS THAT
QUITE SURPRISINGLY, MUTATIONS
THAT RESULT IN HYPOTROPHIC
CARDIO MYOPOTHY DOES NOT
DECREASE THE CAPACITY OF MYOSIN
TO PERFORM.
RATHER IT INCREASES IT.
SHOWN HERE WITH THE SLIDING
VELOCITY OF ACTIN AND FOURTH
GENERATION IN A WILD TYPE
COMPARED TO HYPERTROPHY
MUTATION.
ONE OF THE MORE PROFOUND USES OF
HYPOTROPHIC REMODELLING.
WE SEE TR'S ACTUALLY INCREASE IN
SLIDING VELOCITY ALONG ACTIN AND
APP A ACE AS WELL ANOTHER
MUTATION HAS SOMEWHAT DIFFERENT
DISTRIBUTIONS IN WHICH OF THOSE
BIOPHYSICAL PROPERTIES ARE
RETURNED, BUT BOTH OF THESE WILL
DO EXACTLY THE SAME THING.
THEY WILL INCREASE THE POWER OF
THE CONTRACTILE APPARATUS, NOT
DECREASE IT.
WELL IF THAT'S THE CASE F WE ARE
INCREASING POWER, HOW ARE WE
PRODUCING THOSE HISTONE CHANGES
THAT WE SEE?
WHAT WE THINK IS GOING ON RIGHT
NOW IS FROM THE INCREASE IN
POWER FROM THE SACOMERE GENE
MUTATION THAT WE ARE RESULTING
IN AN INCREASE OF SIGNALING THAT
IS TRANSMITTED TO THE
NON-MYOCYTE POPULATION,
NON-MYOCYTE BEING THE FIBER ONES
THAT ARE RESIDENT WITHIN ALL OF
OUR HEART CELLS AND THE
BIOPHYSICAL -- ARE ACTUALLY
TRANSDUCING SIGNALS IN THOSE
NON-MYOCYTE CELLS.
THIS IS BREASTING 'CUZ THESE
NON-MYOCYTE CELLS DON'T EVEN
CONTAIN THE MUTATION, BUT THE
ACTIVATION OF SIGNALING ON THEM
IS SO POWERFUL THAT WE SEE
INCREASES IN CALCIUM, HOMEOSTAS
SIS, INCREASED IN TGF BETA
SIGNALING THAT LEADS TO A
PROFOUND INCREASE IN THE
[INDISCERNIBLE] AND THAT
OVERTIME RESULTS IN DEPOSITION
OF THE EXTRA CELLAR MATRIX THAT
WE SEE IN HUMAN HEARTS.
WE CAN CHALLENGE THIS KIND OF
PATHWAY WITH LOTS OF FARM LOGIC
INTERVENTION, EITHER THOSE THAT
DECREASE MECHANICAL POWER OR
MORE DIRECTLY THOSE THAT INHIBIT
TGF BETA SIGNALING WITH
ANTIBODIES OR SOME OTHER
PHARMACOLOGIC AGENT.
WE'RE ABLE TO DIMINISH -- THAT'S
WELL AND GOOD BUT THAT'S A
SECONDARY CONSEQUENCE TO THE
ACTUAL OR PROBLEM OF THE HEART,
NAMELY THE MYOSIN MUTATION,
ITSELF.
SO ONE OF THE THINGS WE'VE ASKED
IS WHETHER WE COULD HARNESS
NEWER TECHNOLOGIES THAT REALLY
GRAPPLE WITH THE CLOSER
APPROXIMATE MALL PROBLEMS THE
ACTUAL GENE MUTATION, WHETHER WE
COULD IN FACT SILENT THE MYOSIN
MUTATIONS THAT IS ONE OF TWO
COPIES WITHIN THESE ANIMALS.
TO DO THAT, WE'VE EMPLOYED A
SYSTEM OFFING A KNOW ASSOCIATED
VIRUSES, IN PARTICULAR TYPE 9
WHICH IS KNOWN TO BE CARDIO
TROPIC.
WE HAVE MANIPULATED THAT SO THAT
WE HAVE IT GRIFFIN UNDER THE
EXPRESSION OF [INDISCERNIBLE] T
SO THAT THERE'S EXQUISITE -- --
WHAT WE CAN SEE IS GENERATE
LOVELY GREEN HEARTS THAT ARE NOT
RESULTING IN INCREASE OF EGFP
EXPRESSION IN OTHER TISSUES AND
I'LL TELL YOU THAT LIST LASTS
FOR AT LEAST FIVE MONTHS PERIOD
OF TIME AFTER A SINGLE INJECTION
OF THE AAD VIRUS ON DAY ONE OF
BIRTH.
IF NOW INSTEAD OF JUST HAVING
NON-SPECIFIC VECTOR SUCH AS
GREEN PROTEIN WE USE RNAI WE
THOUGHT THIS MIGHT PROVIDE US
CAPACITY TO DECREASE MUTANT
EXPRESSION.
IF WE'RE GOING TO DO THAT, WE
HAVE TO BE PRETTY CAREFUL
BECAUSE THIS IS A DOMINANT
MUTATION.
SINGLE NUKE YEAH TIDE CHANGE IN
ONE OF TWO ALLELES AND WOULD NOT
BE BENEFICIAL TO THE MOUSE AND
CERTAINLY NOT A HUMANS IF WE
KNOCKED OUT BOTH COPIES OF MICE
IN EXPRESSION.
WE KNOW THAT'S LETHAL PHENOTYPE
FROM KNOCKOUT MICE.
WE ASKED WHETHER WE COULD
SELECTIVELY KNOCK DOWN THE
MUTANT EXPRESSION WITH SHRNA
THAT IN PARTICULAR TAGGED THE
PARTICULAR MUTATION AND WHETHER
THAT WOULD BE SPECIFIC TO
DISCRIMINATE BETWEEN MUTANT AND
WILD TYPE.
WHICH N COMPARISON TO SCRAMBLE
HERE THAT OUR MUTANT SELECTIVE
SHRNA WAS WHOAFULLY INACCURATE.
THIS IS DIFFERENT FROM THE WILD
TYPE ALLELE BY SINGLE NEWICALLY
TIDE.
SO IF WE ADD ONE MORE VARIANT IN
THE SHRNA SO THAT WE ARE NOW TWO
NUCLEOTIDES DIFFERENT FROM WILD
TYPE AND ONE FROM THE DIFFERENT
ALLELE, WE RECHIEFED A
REMARKABLE DIFFERENT RESULT WITH
SUPPRESSION TO ABOUT 25% OF THE
PRIOR LEVEL OF MUTANT RNA
EXPRESSION.
IS THAT SUFFICIENT TO DO MUCH?
WELL, WE INJECTED THESE SHRNAs
INTO OUR HEARTS OF TO OUR MICE,
RATHER, AND EVALUATED THEM
APPROXIMATELY EIGHT WEEKS AFTER
THE INJECTION AND THE RESULT WAS
TRULY ASTONISHING.
RECALL NOW THAT WE'RE REDUCING
AT MOST BY 25% THE AMOUNT OF
MUTANT RNA EXPRESSION AND THIS
IS A SERIES OF CARDIAC
EVALUATION.
HERE'S THE WILD TYPE AND WHAT
YOU CAN SEE IS THE LEFT
VENTRICULAR WALL THICK SNESZ
ABOUT .7 MILLIMETERS.
IF WE REDUCE THE LEVEL OF
EXPRESSION OF THE MU ABOUT THE
ALLELE CONSIDERABLY, WE GET IT
BACK TO NORMAL L.
THAT'S A NICE NON-SPECIFIC
PHENOTYPE, THE IMNEW NIS TICK
ARE ARE FAR MORE IMPRESSIVE.
THIS IS ANIMAL TREATED ARE
[INDISCERNIBLE] I HOPE YOU CAN
SEE THAT THERE'S DISARRAY AND
ACCUMULATION OF FIBROSIS.
IN THE ANIMAL TREATED AFTER
EIGHT WEEKS WITH THE SH RSHG NA,
SEE NONE OF THAT.
SIMILARLY, THE KINDS OF INCREASE
EXPRESSION AND MARKERS OF
HYPERTROPHY ARE ENORMOUSLY
REDUCED WHEN WE HAVE MAXIMUM
VIRAL DELIVERY.
THE AMOUNT OF FIBROSIS IS
SIMILARLY REDUCED.
WE KNOW THAT WHEN YOU HAVE A A
HYPOTROPHIC REMODELLING, YOU
HAVE LOTS OF EKG CHANGES AND WE
ALSO SEE THOSE IN OUR MOUSE
PARTS.
SHOWN IS A WIDENING THAT OCCURS
QUITE COMMONLY IN THE HYPOTRO
IFIC MOUSE.
WE CAN NOW BEGIN TO KNOW SOME OF
THE MUTATIONS THAT ARE
DEFINITIVE CAUSES OF CARDIO
MYOPATHY.
WE CAN BEGIN TO SAY THAT THESE
ARE LIKELY TO BE ACTING THROUGH
A DOMINANT MECHANISM, AND THAT
WE HAVE A STRATEGY TO
POTENTIALLY SILENCE THEM IN AN
ALLELE-SPECIFIC WAY IN MICE.
COULD WE BRING THIS TO THE HUMAN
POPULATION?
I WISH IT WAS FRIDAY AFTERNOON
BECAUSE IN BOSTON THAT'S A TIME
WHEN THE LABS ALL GATHER
TOGETHER AND HAVE A BEER AND
ENJOY EACH OTHERS COMPANY AND IT
MAKES YOU DREAM A LITTLE BIT AND
WE CARDIOLOGIES ARE SOMETIMES
VERY BIG DREAMERS.
THINGS.OT AFRAID TO TACKLE
WHAT I'M SHOWING HERE IN THIS
SCHIZOPHRENIC ECHOCARDIOGRAM IS
AN IMAGE OF A HUMAN HEART IN
WHICH THE HEAD OF THE
INDIVIDUAL'S HERE, FEET OVER
HERE.
ECHO IS IS SHOWING THE BLOOD
GOING FROM THE LEFT ATRIUM TO
LEFT VENN TRI CALL AND THIS BIG
CYST AREA OVER HERE IS A HIGHLY
INTERVENTRICULAR ACCEPT TIM.
IT ACCOUNTS THE FOR ENORMOUS
AMOUNT OF SYMPTOMS AND CAN
INCREASE RISK OF SUDDEN DEATH IN
INDIVIDUAL WHO IS HAVE
PROFOUND -- WE KNOW THIS IS SO
PROFOUND THAT WE TAKE TWO HEROIC
MEASURES TO GET RID OF IT.
FIRST IS TO DO OPEN HEART SURGE
RAND TO REMOVE THAT HYPERTROPHY
SEPTUM.
LESS INFAY VA SIEVE WAY IS THE
PASS CATHETER INTO THE AROR TA
FROM THE LEG AND RETROGRADE TO
CAN LATE THE VESSELS THAT
SPECIFICALLY FEED THE
INTERVENTRICULAR SEPTUM, THE ONE
SHOWN RIGHT HERE CALLED THE
INTERVENTRICULAR ACCEPT TALL --
AND TO INJECT ALCOHOL IN IT SO
THAT WE'VE GIVEN THE INDIVIDUAL
A HEART ATTACK TO HAVE THAT HIGH
PERT THE FI SET AND DIE.
I WOULD SUGGEST TO YOU THAT I
DON'T THINK TOO FAR OFF TO THINK
ABOUT RATHER THAN KILLING THE
MYOCARDIUM, TO USE THESE TO
POTENTIALLY FUND MUTATION.
IF WE COULD ACHIEVE LONG-TERM
SILENTING OF EVEN 25-30%, WE
MIGHT SUBSTANTIALLY REDUCE THE
HYPERTROPHY IN THAT REGION AND
SPARE THE MYOCARDIUM FROM DEATH.
HOW MANY PEOPLE HAVE THAT
PARTICULAR MUTATION?
PRIVATE MUTATIONS ARE THE NAME
OF THE GAME IN LOTS OF IMPORTANT
DISEASES INCLUDING ALL THE
CARDIO MYOPATHY.
THEY'RE IN NOW MORE THAN A
THOUSAND DIFFERENT MUTATIONS
THAT HAVE BEEN IDENTIFIED TO
CAUSE HYPOTROPHIC CARDIO
MYOPATHY.
THERE WILL BE AN EQUAL NUMBER
THAT CAUSE DILATED CARDIO
MYOPATHY.
WHAT WE CAN DO IS CAPITALIZE ON
EXTENSIVE KNOWLEDGE THAT'S COME
BEGIN TO ASK WHETHER WE COULDND
INSTEAD OF TARGETING THE PRECISE
MUTATION, TO TARGET A NEARBY
VARIANT THAT IS IN LINKAGE
ASSOCIATION WITH THAT MUTATION.
IT'S ON THE NAME ALLELE, IT
ACTUALLY IS COINHERIT BD W THE
MUTATION.
TO SEE IF THAT WAS EVEN
FEASIBLE, WE WENT TO A FAVORITE
SOURCE OF US, THE NHLBI LOOKING
AT MYOSIN HEAVY CHAIN GENE AND
WE SEE HERE THAT THIS IS A
VARIANCE BUT THAT ONE TURNS OUT
TO BE IN AN INTRON WHICH WOULD
BE HEART TO SILENT FROM THE RNA
POINT OF VIEW BUT HERE'S A
CODING SYNONYMOUS VARIANT AND
HERE'S ANOTHER ONE OVER HERE.
SO EVERYWHERE IF -- IF WE WERE
TO TEST THAT STRATEGY, WE COULD
IMBRED MOUSE, BREED IT INTO
A DIFFERENT BACKGROUND AND NOW
TRY AND SILENT NOT THE MUTATION
BUT THE NEARBY POLYMORPHIC
[INDISCERNIBLE] WHEN WE DO SO
THE RESULTS ARE ESSENTIALLY THE
SAME.
WHAT I'VE TRIED TO TELL YOU IS
THAT WHAT WE THINK BY ACTUALLY
CONSIDERING THE USE OF NEARBY
SNPs THAT ARE SIGHTLY LINKED TO
MU TAN ATE A LEGALS THAT WE
COULD STILL A BATTERY OF
APPROACHES TO SILENCE MUTATION
IN A VERY SPECIFIC WAY THAT
WOULD LEAVE THE NON-MOW TATED
ALLELE TO MAKE THE PROTEIN AND
WHILE IT WOULD BE UNLIKELY THAT
WE COULD COMPLETELY REDUCE THE
MUTANT LEVEL TO ZERO THAT WE
MIGHT HAVE THE CAPACITY TO
REDUCE IT SIGNIFICANTLY TO
ATTENUATE THE REMODEL THAT
OCCURS.
WHERE ARE WE GOING WITH ALL
THIS?
WELL, I HOPE I'VE CONVINCED YOU
THAT THERE ARE LOTS OF
CARDIOVASCULAR DISEASE GENES.
THEY'VE BEEN POWERFUL IMPORTANT
IN TEACHING ABOUT SACOMERE
FUNCTION AND BIOLOGY.
I THINK THEY'RE INCREASINGLY
GOING TO BECOME VERY USEFUL IN
UNDERSTANDING PHENO TIP PICK
VARIANCE IN CLINICAL SETTING.
THERE'S A LOT MORE FOR US TO BE
ABLE DO THAT IN A VERY ROBUST
FASHION, BUT WITH THE INCREASE
USE OF GENETIC TESTING I'M
CERTAIN WE'LL GET THERE.
SHRNA IS ONLY THE BEGINNING OF
TRYING TO SILENCE MUTATION IN
SEMATIC TISSUES.
I THINK THERE'S ENORMOUS
CAPACITY TO USE GENETIC
ANTEDOTES FOR TREATING NOT ONLY
HEART DISEASE BUT A VARIETY OF
OTHER CONDITIONS, AND I WOULD
ENCOURAGE ALL OF YOU YOUNG
PEOPLE IN THE AUDIENCE TO THINK
ABOUT GETTING INVOLVED IN THIS
REALLY, REALLY EXCITING AREA OF
SCIENCE.
I HAVE TO ACKNOWLEDGE THIS
AMAZING TEAM I WORK WITH, IN
PARTICULAR ALEX THICK, DAN
HERMAN, PAULA KEY, BASIC
RESEARCHERS, AND ALSO ND Ph.D.
STUDENTS, CAR LYNN HULL ARK
WHOLE TEAM OF SEQUENCING
COLLEAGUE FROM THE BROAD, FROM
THE JACKSON AND FRAMING HAM
HEART COHORTS AND MY COLLEAGUE
FROM THE UK, STEWART COOK.
THANK YOU VERY MUCH.
[APPLAUSE]
>> THAT WAS TERRIFIC.
PLEASE DO USE THE MICROPHONES TO
POSE QUESTIONS SO PEOPLE
LISTENING ON THE WEB CAN HEAR AS
WELL.
GO AHEAD.
>> THANK YOU VERY MUCH.
WHEN THOSE [INDISCERNIBLE] [LOW
AUDIO].
>> I WILL.
>> THIS IS SA QUESTION ABOUT
[LOW AUDIO] LINK TO
[INDISCERNIBLE] --
>> TO THE QUESTION IS, CAN WE
REVERSE THE ESTABLISHED HIGH P
PERT FI USING SHRNA APPROACH
RIGHT NOW?
THOSE ARE EXPERIMENTS THAT WE
HAVE DONE ONLY IN A SHORT-TERM
SEQUENCE RIGHT NOW.
WE HAVE NOT DONE THEM IN A FULL
EIGHT-WEEK TO 12-WEEK PERIOD OF
TIME AND WE THINK THOSE ARE
IMPORTANT TO DO FOR A PROTRACTED
PERIOD OF TIME.
WHEN WE DO A ACCEPT L OBLATION,
IT TAKES MONTHS FOR THE
MYOCARDIAL TO REMODEL AND THAT'S
AFTER KILLING THE HEART MUSCLE.
WE WOULD ANTICIPATE IT COULD
TAKE MANY MONTHS TO REMODEL
HYPERTROPHY MOUSE HEART.
DO WE THINK IT'LL WORK, YEAH, I
THINK IT WILL.
WE WOULD EXPECT THERE TO BE
INCREASED AMOUNT OF WILD TYPE
ALLELE SHOULD ALLOW FOR THAT
INCREASE TO BE REACHED OVER
TIME.
SECOND REASON I THINK THAT IS
ANYBODY WHO'S TREATED A PATIENT
WITH HYPER TENSION AND HIGH P
PERT FI, IT TAKES TIME BUT OVER
TIME THAT LEFT VENTRICULAR WILL
RESOLVE.
WE JUST DON'T HAVE THE RESULTS
YET.
>> BASIC QUESTION ABOUT THE
TITIN PART OF YOUR TALK.
IN AN INDIVIDUAL WHO HAS HETERO
PSI GUS TRUNCATING MUTATION, ARE
THE TWO ALLELES TRANSLATED IN
50/50 PORTION OR DOES THE
TRUNCATED HAVE AN ADVANTAGE?
>> THAT'S A VERY GOOD QUESTION.
WE BEGUN TO LOOK AT THAT BY THE
ONE GOOD NEWS ABOUT TITIN IS
THERE'S SO MANY VARIANCE IN
THERE THAT YOU CAN LOOK TO SEE
FROM RNA SEEK ANALYSIS IF BOTH
IN FACT ARE EXPRESSED.
WE LOOKED AT THAT AND WE SEE NO
INCREASE NOR DECREASE IN
ALLELEIC-SPECIFIC SNPs AS WE
MARCH ALONG THE TITIN MOLECULE.
SO WE DON'T SEE AN ADVANTAGE NOR
A DISADVANTAGE.
>> [INDISCERNIBLE] HOW LONG
[INDISCERNIBLE] WHERE DOES THE
OUTCOME COME FROM CARDIAC
[INDISCERNIBLE]?
>> SO ALCOHOL ACCEPT TALL OWE
BLAGS IS JUST SO WE'RE CLEAR NOT
SOMETHING THAT I ROUTINELY DO
BUT MANY OF MY COLLEAGUES DO AND
IT IS USUALLY RECOMMENDED FOR
INDIVIDUALS WHO HAVE FIRST AND
FOREMOST A LOT OF SYMPTOMS THAT
HAVE NOT RESPONDED TO
PHARMACOLOGIC INTERVENTION, AND
FOR REASONS THAT THE PATIENT
OFTEN IS INVOLVED IN OR BECAUS
THEY'RE A A CLINICAL INDICA
THAT, PERSON IS NOT APPROPRIATE
FOR REFERRAL TO SURGERY TO OPEN
HEART SURGERY.
IT'S INCREASINGLY BEING USED
BECAUSE OF PATIENT PREFERENCE.
IT USUALLY TAKES A PERIOD OF SIX
WEEKS OR LONGER TO SEE
REMODELLING AND REGRESSION OF
THE INTERVENTRICULAR SEPTUM SO
THAT THE OUTFLOW TRACK BECOMES
MORE OPENED.
FROM THE LONG-TERM FOLLOW-UP WE
DO NOT SEE A GROWING BACK.
WHETHER THAT WILL HAPPEN IN FIVE
YEARS, I DON'T KNOW, BUT RIGHT
NOW WE HAVE NOT SEEN IT.
>> [INDISCERNIBLE] LIGATION [LOW
AUDIO] --
>> I'M SORRY.
OH, IT'S ONE-TIME, YES.
I WILL MENTION FOR PEOPLE WHO
ARE NOT INTERESTED IN THIS
CLINICALLY, THE ADVANTAGE OF
THIS AV THING IS IT'S ALSO
ONE-TIME USE AND YOU GET FIVE
MONTHS OF SILENCING THE GENE OF
YOUR CHOICE FOR ONE-TIME
INJECTION IN YOUR MOUSE.
THAT'S A GOOD EXPERIMENTAL
SYSTEM WHICH WE'RE USING FOR
LOTS OF OTHER INVESTIGATION.
>> THANKS.
>> JUST WONDERING ABOUT THE PART
OF THE TALK TITINS, THE
DIFFERENTIAL EXPRESSION OF THE
TITINS [LOW AUDIO] THOUGHT TO BE
PROTECTIVE AND HAVE YOU DEALT
WITH IT TO THE GENETIC SEQUENCES
MIGHT BE?
>> I'M SORRY.
>> WITHIN THE TITIN GENE, YOU
SHOWED DIFFERENT EXPRESSION
PATTERN INS AND DIFFERENT PARTS
MORE THAN OTHERS AND THAT
THERE'S CORRELATION WHICH --
[LOW AUDIO] COMPARED TO THE
EXPRESSION PATTERN.
JUST WONDER IS THAT EXPRESSION
PATTERN DIFFERENT HYPOTHESIZE
BEING PROTECTED?
>> THE REGIONS THAT WHERE THE
NUMBER L POPULATION HAS
ACCUMULATED SOME VARIANCE THAT
TRUNCATE THE MOLECULES ARE IN
PARTS POF, CAN BE INVOLVED IN
MAKING TITIN MOLECULES THAT ARE
VERY, VERY LOW-LEVEL EXPRESSION.
SO THEIR ALTERNATIVE EXXONS THAT
ARE SPLICED IN AND SPLICED OUT
BECAUSE THERE'S A LOT OF
DIFFERENT TITINS IN EACH AND ERE
HEART, BUT THE ABUNDANCE OF THEM
IS VERY, VERY LOW, AND INDEED
THE ONES THAT ARE INCORPORATED
INTO THE SACOMERE ALMOST BY
DEFINITION BECAUSE THEY STAND
FROM THE [INDISCERNIBLE] ARE
WHAT WE CALL FULL-LENGTH TITINS
REE FERED TO AND IT DOES NOT
INCLUDE THOSE LOWLY EXPRESSED
EXXONS.
THERE'S ALSO ANOTHER FOR THOSE,
CALLED THE KNOW VEX THREE
ISOFORM WHICH IS A VERY, VERY
SHORT FIVE THOUSAND BASE PAIR
TRANSCRIPT AND NO ONE ACTUALLY
KNOWS WHAT THAT DOES.
IF IT DOES ANYTHING.
>> WHOLE TITIN STORY GETS MORE
INTERESTING ALL THE TIME.
I GUESS YOU'RE REALLY CAUSING
MANY OF US TO DEPEND UPON NOT
JUST MEDIATED DECAY TO PREDICT
THE CONSEQUENCE OF ONSETS OR
FRAMESET MUTATION TO QUESTION
WHETHER WE SHOULD DEPEND ON THAT
IF THE GENE IS ANYTHING OTHER
THAN HEMOGLOBIN 'CUZ IT SOUNDS
AS IF MAYBE BECAUSE THE COUPLE
OF TRANSLATION, SPLICING THAT
YOU ALREADY PASSED THE POINT OF
NOT HAVING A PROTEIN BEFORE YOU
HIT THAT STOP CODE.
I ASSUME THOUGH THAT THERE WOULD
BE SOME RELATIONSHIP -- WE'RE
TALKING ABOUT THIS A MINUTE
AGO -- IN TERMS OF ABUNDANCE OF
THE MUTANT ALLELE DEPENDING ON
WHERE THE TRUNCATING MUTATION
WAS AND THAT IT WAS CLEARLY
EARLY ON THAT YOU WOULD GET
AMOUNTS OF TRANSCRIPT -- PART OF
THE QUESTION I WANTED TO ASK IS
WHAT IS THE PHENOTYPE OF A TRUE
APP LOW INSUFFICIENT TITIN
ALLELE AND IF IT'S NOT BAD DOES
IT SAY THAT WHAT YOU'RE DOING
FOR THESE HIGH PROTROPHIC CARDIO
MYOPATHIES MIGHT ALSO WORK FOR
TITIN MUTATION AND CAUSE DIELTED
CARDIO MYOPATHY OR TRYING TO
DIAL?
>> SO THE MOST DEFINITIVE
EVIDENCE, I THINK, OF HUMAN
PATIENTS -- NOT A MOUSE -- WITH
A TITIN INSUFFICIENT MUTATION
ANNUL HAS COME FROM SOME RARE
RE
RECESSIVE FAMILIES.
IT'S AT THAT THREE PRIME, IT'S
THOSE M BAN MUTATIONS.
THERE HAS BEEN NO REPORTS OF A
*** SIGH GUS OR HETERO PSI GUS
WITH OVER --
>> SO IT MIGHT BE [LOW AUDIO].
>> AT THE VERY, VERY AMY KNOW
TERMINAL SEQUENCE, THAT'S
CORRECT.
AND WHEN WE SEE WHERE WE FIND
THOSE CLUSTERS IN OUR RANDOM
PEOPLE, THEY'RE MORE LIKELY TO
BE AT THAT AMY KNOW TERM NANT.
SO THERE MAY BE NO ABNORMALITITY
OF THE HETERO PSI GUS LOSS OF
FUNCTION.
WHEN WE SHIFT TO MICE NOW AND
THERE IS ONE COMPLE MOUSE
MODEL THAT WAS A KNOCKOUT BUT IT
ALSO INTRODUCED A SIMULTANEOUS
MUTATION.
SUFFICE IT TO SAY THAT IT
APPEARS THAT THE *** SIGH GUS
IS LETHAL.
THE HETERO SIGH GUS IS FINE.
SO WE DO THINK THAT IF IT IS A
DOMINANT NEGATIVE MODE OF ACTION
SILENTING TITINS WOULD BE
BENEFICIAL.
>> WE HAVE PLENTY OF WAYS TO DO
THAT GIVEN THE TITIN SUCH A HUGE
NUMBER OF VARIANTS AND ALMOST
EVERYBODY'S GOING BE HETERO PSI
GUS --
>> RIGHT.
CORRECT.
WE WOULD LIKE TO HAVE THOSE THAT
ARE COMMON SO WE DIDN'T NEED TO
MAKE A THOUSAND OF THEM, JUST A
COUPLE HUNDRED.
>> ONE MORE QUESTION.
>> I READ AT THE BEGINNING WHAT
IS THE PERCENTAGE OF CARDIO
MYOPATHIES [INDISCERNIBLE] JE NE
SIX AND WHAT'S THE
[INDISCERNIBLE] LIFESTYLE OR
ENVIRONMENT FACTORS OR --
>> OR ALL OF THE ABOVE, RIGHT?
[LAUGHTER]
IF IF YOU TAKE INDIVIDUALS THAT
HAVE ONE FAMILY MEMBER WHO WAS
ALSO AFFECTED WITH A HYPER
TROPHIC PHENOTYPE, 75% OF THOSE
ARE GENETIC.
THAT'S HIGH.
IF YOU TAKE INDIVIDUALS WHO HAVE
NO FAMILY HISTORY OF CARDIAC
HYPERTROPHY AND THEY HAVE ALL OF
THE CLASSIC MANIFESTATIONS OF
HYPER TRO IF I CAN CARDIO
MYOPATHY BUT NO FAMILY HISTORY,
35% ARE GENETIC AT TOPS.
IF WE DO THE SAME EXPERIMENT
WITH DIELTED CARDIO MYOPATHY,
THE BEST EVEN IN A FAM MILL
Y'ALL CASE WE'RE DOING IS ABOUT
50% AND THAT'S INCLUDING TITIN
WHERE THAT'S 25%.
THAT'S THE BIGGEST PIECE OF THE
PIE RIGHT NOW AND EVERYTHING
ELSE IS A THOUSAND OTHER LITTLE
GENES.
SO THERE'S STILL MORE TO
DISCOVER THERE.
AND WHEN YOU TAKE AN ISOLATED
INDIVIDUAL SIT OWE PA THICK
CARDIO PAFK AND INCLUDE TITIN,
WE'RE AT ABOUT 15%.
>> WELL, DR. SEIDMAN WOULD BE
WILLING TO CONTINUE THIS
LIBRARY.ION AT A A RECEPTION IN
MEANWHILE, PLEASE JOIN ME IN
OUR SPEAKER AGAIN.