SO WELCOME, I AM VICKI AND IT'S
A PLEASURE TO HAVE YOU WITH US
TODAY.
WE ARE EXCITED TO HAVA A
WONDIRBFUL SET OF SPEAKERS
TALKING ABOUT XIST AND THE
XISTING DATA ON WHY AUTOIMMUNE
DISEASES MAY BE MORE COMMON IN
WOMEN.
COUPLE OF HOUSEKEEPING THINGS
JUST TO GET US STARTED, THIS
EVENT IS BEING RECORDED AND WILL
BE VIDEOCAST, IT WILL BE
AVAILABLE IN THE VIDEOCAST
ARCHIVE, IF YOU HAVE TECHNICAL
CHALLENGES, PLEASE USE THE CHAT
FUNCTION TO CHAT WITH I.T.
SUPPORT.
IF YOU HAVE ANY TECHNICAL
DIFFICULTYYS, AND THEN ALSO BE
THOUGHTFUL AND CONSIDERATE IN
YOUR CHAT IF YOU ARE TALKING TO
FELLOW PARTICIPANTS.
A COUPLE HOUSEKEEPING ISSUES
REGARDING Q&A, WE ARE GOING TO
HAVE QUESTIONS FOR THE PANELISTS
AT THE END OF ALL 3 TALKS, IF
YOU CAN SUBMIT KEQUESTIONS
THROUGH THE Q&A FUNCTION, WHICH
IS RIGHT NEXT TO THE CHAT
FUNCTION ON YOUR ZOOM, IF YOU
THERE ARE QUESTIONS YOU WOULD
REALLY LIKE TO HAVE ASK ASKED,
WE WILL MODERATE THOSE THROUGH
THE Q&A FUNCTION.
SO I WANTED TO GET STARTED, JUST
QUICKLY INTRODUCING OUR 2
MODERATORS FOR TODAY, DR. STACEY
FERGUSON IS THE CHIEF OF THE
AUTOIMMUNE AND PRIMARY IMMUNO
DEFICIENCY SECTION AT THE
NATIONAL INSTITUTE OF ALLERGY
AND INFECIOUS DISEASES AND
DR. MARIE MANCINI IS PROGRAM
DIRECTOR OF THE SYSTEMIC
AUTOIMMUNE DISEASE BIOLOGY
PROGRAM AT THE NATIONAL
INSTITUTE OF ARTHRITIS AND MUSK
LO SKELETAL AND SKIN CONDITIONS.
IT'S A PLEASURE TO HAVE THEM
WITH US TODAY.
I WANT TO INTRODUCE OUR 3
SPEAKERS, DR. [INDISCERNIBLE] IS
IN THE DEPARTMENT OF BIOMEDICAL
SCIENCES AT THE UNIVERSITY OF
SCHOOL OF VETERINARY SCHOOL
MEDICINE, SHE IS A MEMBER OF THE
EPIGENETICS INSTITUTE, INSTITUTE
FOR IMMUNOLOGY AND THE INSTITUTE
FOR REGENERATIVE MEDICINE AT
PENN.
Y RECEIVED HER MASTER IN
[INDISCERNIBLE] AND HER Ph.D.
FROM CORNELL, SHE WAS DID HER
POST GRAD AT HARVARD WHERE SHE
DID INACTIVATION OF THE
X-CHROMOSOME.
DR. ANG UERA HAS STUDIED IMMUNE
RESPONSES IN RESPONSE TO
INFECTION AND MORE RECENTLY IN
AUTOIMMUNITY AND IT'S A GREAT
PLEASURE TO HAVE HER GIVE THE
OPENING LECTURE OF THIS
SYMPOSIUM TODAY EMPLOY OUR
SPECCED SPEAKER WILL BE
DR. DIANEA DOU, SHE IS A N IAMS
FELLOW, SHE AND HER B. S. DEGREE
WITH HONORS FROM CALTECH AND
DOUBLE MAJORED IN BOTH BIOLOGY
IN BIOLOGY AND BUSINESS
MANAGEMENT.
SHE THEN WAS INTRODUCED TO RNA
BIOLOGY AS BOTH AN UNDERGRADUATE
RESEARCH AND SUBSEQUENT LOW IN
THE INTRAMURAL AND RESEARCH
TRAINING BACCALAUREATE TRAINING
AT NIH IN DR. FAUCI'S LAB,
DR. DOU WENT ON TO COMPLETE A
Ph.D. IN MOLECULAR BIOLOGY
INVESTIGATING HEMATOPOIETIC STEM
CELLS AND CURRENTLY STUDYING THE
STEM CELLS OF NONLONG CODING
RNAs AND ACCESSIBILITY IN
AUTOIMMUNE DISEASE, RECENTLY SHE
DISCOVERED A NOVEL ROLE EXISTS
IN THE RIBONUCLEAR PROTEIN AT
THE DRIVER PROTEIN USHED LYING
THE DOMINANCE OF AUTOIMMUNE
DISEASE, AND THEN OUR THIRD
SPEAKER TODAY WOULD BE
DR. BRENDAN ANTIOCHO, HE IS A
PHYSICIAN SCIENTIST AT JOHNS
HOPKINS SCHOOL OF MEDICINE.
HE HAS PRIMARY RESEARCH INTEREST
IN THE INNATE IMMUNE SYSTEM IN
THE PATHOGENESIS OF RHEUMATIC
DISEASE, HE RECEIVED HIS
BACHELOR'S DEGREE FROM DARTMOUTH
UNIVERSITY AND HIS MEDICAL
SCHOOL FROM DARTMOUTH MEDICAL
SCHOOL.
HE THEN COMPLETED RESIDENCY IN
INTERNAL MEDICINE AT OREGON
HEALTH SCIENCE UNIVERSITY
FOLLOWED BY RHEUMATOLOGY
FELLOWSHIP AT JOHNS HOPKINS
SCHOOL OF MEDICINE WHERE HE IS
NOW AN ASSISTANT PROFESSOR OF
RHEUMATOLOGY.
HE STUDIES THE IDENTITY OF
SPECIFIC INDOAJ NOWS NUCLEIC
ACIDS AND SINCE THE LIGAND
INTERACTIONS RESPONSIBLE FOR
INTERFORON RESPONSES IN LUPUS
AND SHOGREN'S SYNDROME.
IT IS A GREAT PLEASURE TO HAVE
ALL 3 OF OUR SPEAKERS WITH US
TODAY AND WITH THAT I WILL HAND
THE MICROPHONE TO DR. ANG UERA,
TO OPEN US UP WITH HER TALK ON
X-CHROMESOME ACTIVATION AND
IMMUNE SYSTEMS.
DOCTOR ANG UERA IN CAN YOU HEAR
ME?
CAN YOU HEAR ME NOW IN.
>> YES NTHANK YOU SO MUCH VICKI
AND THANKS TO EVERYONE FOR THIS
OPPORTUNITY EVERYONE AT THE
OFFICE FOR RESEARCH ON WOMEN'S
HEALTH FOR THIS CHANCE TO SHARE
SOME EXCITING PUBLISHED AND
UNPUB LIBRARY FOUNDATIONED DATA
WITH YOU TODAY.
SO IN MY LAB AT PEIN, N, WE'RE
VERY INTERESTED IN X-CHROMOSOME
ACTIVATION AND SEX DIFFERENCES
AND HOW THE GENETICS AND THE
EPIGENETICS CONTRIBUTE TO FEMALE
BIAS WITH IMMUNE RESPONSES AND
ALSO AUTOIMMUNE DECS, SO JUST A
LITTLE BIT OF BACKGROUND ABOUT
LUPUS OR SYSTEMIC LUPUS
ERYTHEMAITOSEIS, THIS A SYSTEMIC
RHEUMATIC AUTOIMMUNE DEC THAT
AFFECTS A VARIETY OF ORGANS ANDS
ISS, LUPUS IS CHARACTERIZED BY A
VARIETY OF PHENOTYPES WHICH I
WILL HIGHLIGHT--
>> CAN I JUST YUSM IN--
>> WE'RE NOT SEEING YOUR SLIDES,
CAN YOU SHARE?
>> UH, OH, OKAY, SORRY!
OKAY.
FOR SOME REASON, I CAN'T SEE ...
YOU CAN SEE YOUR SLIDES NOW, SO
SHARE SCREEN BUT SHARE THE 1
WITH JUST ALL YOUR SLIDES.
NWITH THE PRESENTER?
>> WITHOUT PRESENTER.
>> NOW YOU CAN SEE THE
PRESENTER, SO SELECT PRESENTER.
>> DO YOU SEE MY SLIDES NOW.
>> PERFECT.
>> YES, WE DO NPERFECT.
PERFECT.
GREAT.
THANK YOU SO MUCH, THANK YOU.
OKAY, SO LUPUS, SO LUPUS DISEASE
IS CHARACTERIZED BY A VARIETY OF
DIFFERENT PHENOTYPES, SO LOOKING
IN LUPUS PATIENTS AT THE SERUM,
SLE PATIENTS HAVE ELEVATED
LEVELS OF AUTOANTIBODIES
SPECIFICALLY, ANTIDOUBLE
STRANDED DNA, OR DSDNA,
AUTOASPECT BODIES AND ALSO
ANTISMITH, RMP COMPLEX
ANTIBODIES, LUPUS DISEASE IS
ALSO KNOWN TO AFFAIRS TEAM
LEADERRER THE B-CELL COMPOSITION
WHERE SLE PATIENTS HAVE
INCREASED NUMBERS OF ACTIVATED
B-CELLS AND THERE'S ALSO
INCREASED NUMBERS OF AGE
ASSOCIATED B-CELLS, LUPUS
PATIENTS ALSO HAVE HIGH NUMBERS
OF PLASMA CELLS.
SLE PATIENTS ARE ALSO SOMETIMES
DEVELOP KIDNEY PATHOLOGY, SO
DPLU MARRIAL NEPHRITIS, WHERE IT
WILL INFILTRATE INTO THE KIDNEYS
RESULTING IN TISSUE BREAK DOWN,
SO WE KNOW THAT B-CELLS ARE
IMPORTANT OFFICIATORS AND
EFFECTORS OF NORMAL IMMUNE
RESPONSE, AND ALSO KNOWN TO BE
IMPORTANT PLAYERS, SO AUTOBODY
PRODUCTION AND DETECTION IS KEY
TO MAKING A DIAGNOSIS OF LUPUS
DISEED AND OFTEN TIMES,
AUTOANTIBODIES CAN BECOME
ELEVATED IN THE PATIENT EVEN
BEFORE THERE'S EVIDENCE OF
CLINICAL DISEASE.
NOW B-CELLS ALSO SURVEYS CRETE
AUTOANTIBODIES BUT THEY PLAY
IMPORTANT ROLES IN ANTIGEN
PRESENTATION AND ALSO IN
CYTOKINE PRESENTATION.
SO RHEUMATOLOGISTS HAVE BEEN
MAKING THE OBSERVATION FOR
DECADES THAT IF YOU LOOK AT
AUTOIMMUNE DISEASES, THE
MAIORITY OF PATIENTS ARE
BIOLOGICALLY FEMALE AND IN THIS
GRAF, IT SHOWS 25 AUTOIMMUNE
DISORDERS, WHERE GREATER THAN
50% OF THE PATIENTS ARE FEMALE
AND THESE 25 DIFFERENT
AUTOIMMUNE DISEASES ARE SHOWN
ALONG THE X-AXIS HERE AND THE
PERCENTAGE OF PATIENTS THAT ARE
EITHER MALES OR FEMALES IS SHOWN
IN THE XAXIS AND IT WAS THOUGHT
THAT SEX HORMONES ALONE WERE
RESPONSIBLE FOR THE SEX BIAS,
BUT NOW, WE KNOW THAT THE SEX
SCROAM STUDIES OF MULTIPLE
ENDOCRINE, IN PARTICULAR THE
GENETICS AND EPIGENETICS ALSO
PLAY AN IMPORTANT ROLE.
AND SO WHAT A STRIKING
OBSERVATION THAT REALLY CAUGHT
OUR ATTENTION IS THAT THERE'S AN
INCREASED RISK FOR DEVELOPING
SOME OF THESE SEX BIAS
AUTOIMMUNE DECS, THAT CORRELATES
WITH THE NUMBER OF X-CHROMOSOMES
THAT AN INDIVIDUAL HAS, SO
INDIVIDUALS WITH WOOR MORE
X-CHROMOSOMES, ARE AT INCREASED
RISK TO DEVELOP LUPUS, IN
ADDITION TO 4 OTHER AUTOIMMUNE
DISEASES THAT ARE PREDOMINANTLY
FEMALE BIASED EMPLOY SO 1 COULD
ASK, WHAT'S SO SPECIAL ABOUT THE
X-CHROMOSOME.
WELL, ON THE X-CHROMOSOME IN
HUMAN ANDS IN MICE, THERE'S A
IMPORTANT DENSITY OF IMMUNE
RELATED YEENS THAT ARE
PRESCRIBINGENT ON THIS
CHROMOSOME, AND SO, TODAY I JUST
BRIEFLY WANT TO TELL YOU ABOUT 1
OF THESE EXCELLENT YEENS CALLED
TLR 7, WE WILL HEAR MORE ABOUT
THIS FROM BRENDA IN THE LAST
PRESENTATION, AND SO, TOLL-LIKE
RECEPTOR 7 OR TLR-7, THE DOSAGE
OF THIS YEEN, OR THE EXPRESSION
LEVEL IS CRITICAL BECAUSE IF
THERE'S TOO MUCH OF IT CAN
RESULT IN THE DEVELOPMENT OF
LUPUS LIKE DISEASE IN A MOUSE
MODEL AND SO THE EXAMPLE OF THIS
MOUSE MODEL THAT I WANT TO SHARE
WITH YOU IS THIS BXSB MICE WHERE
A HUNDRED% OF THE MALE ANIMALS
DEVELOP LUPUS LIKE DISEASE AND
THESE ANIMALS HAVE ELEVATED
PRODUCTION OF AUTOANTIBODIES,
GLUE MARRIAL PATHOLOGY SIMILAR
TO LUPUS PATIENTS AND ALSO VERY
HIGH NUMBERS OF ACTIVATED
B-CELLS.
AND SO IT TURNS OUT THAT IN THIS
MOUSE STRAIN, THERE WAS A
TRANSLOCATION EVENT WHERE A
REGION OF THE X-CHROMOSOME
CONTAINING TLR 7 WAS
TRANSLOCATED ON TO THE Y.
SO IN THESE MALE MICE THEY HAVE
DOUBLE THE EXPRESSION OF TLR 7
THAT RESULTS IN THESE
PATHOLOGIES.
AND SO THEN 1 COULD ASK HOW IS
THE X-CHROMOSOME REGULATED?
SO IN THEORY IN MAMMALS HAVE
THIS REALLY SPECIAL AND EXCITING
WAY TO REGULATE EXPRESSION FROM
THE X-CHROMOSOME CALLED
X-CHROMOSOME AND ACTIVATION, SO
IF YOU LOOK AT THE NUMBERS OF
SEX CHROMOSOMES, BETWEEN THE
SEXES THEY'RE DEFINITE SO
FEMALES HAVE 2 Xs AND WOULD
EXPRESS TWICE AS MANY COMPARED
TO MALES SO WHAT THE MAMMALS
HAVE DONE IS DEVELOP THIS
MECHANISM WHERE 1 X CHOSEN AT
RANDOM FOR TRANSCRIPTIONAL
SILENCING AND THIS PROCESS
HAPPENS EARLY DURING EMBRYONIC
DEVELOP AM WHERE THE FUTURE
INACTIVE X-CHROMOSOME REGULATES
THIS NONCODING RNA CALLED XIST,
AND XIST WILL REMAIN TETHERED
FROM THE CHROMOSOME WHERE IT'S
EXPRESSED AND RESULT IN
TRANSCRIPTIONAL SILENCING, THIS
INACTIVE X-CHROMOSOME ACQUIRES A
NUMBER OF REPRESSIVE EPIGENETIC
MODIFICATIONS THAT AID IN THE
SILENCING AND SOME OF THESE
MARKS CAN BE VISUALIZED AT THE
SIGNIFY TO LOGICAL LEVEL, SO
THIS IS XIST R NA INSITU
HYBRIDIZATION IN FISH, AND THIS
IS XIST CHROMOSOME WITH THE RNA
ENCODING IT AND THIS IS THE SAME
NUCLEUS GOING FROM LEFT TO RIGHT
AND YOU CAN SEE THERE A H3
KTRI-METHYLATION AND ALSO H2A
UCIBOL WIDE JUMPSIN VENTILATION,
SO ALL THESE WORK TO SILENCE THE
X, HOWEVER, THERE ARE SOME GENES
THAT ESCAPE SILENCING FROM THE
INACTIVE X-CHROMOSOME INCLUDING
RNA XIST ITSELF, AND THESE ARE
ALL IMPORTANT TO WORK TOGETHER
IN ORDER TO MAINTAIN DOSAGE
COMPENSATION, SO IT WAS
TRADITIONALLY THOUGHT THAT ONCE
YOU FORM AN INACTIVE
X-CHROMOSOME DURING DEVELOPMENT,
IT'S MAINTAINED
TRANSCRIPTIONALLY SILENT IN THE
SAME WAY, NO MATTER WHAT TYPE OF
CELL YOU'RE LOOKING AT WHETHER
IT'S A MUSCLE CELL OR NEURON,
HOWEVER, WORK FROM OUR LAB HAS
FOUND THAT THE IMMUNE CELLS DO
ACTIVATION VERY DIFFERENTLY AND
SO IN B-CELLS RESTING NAIVE
B-CELLS FROM BOTH THE MOUSE AND
THE HUMAN LACK SIGNIFY TO
LOGICAL MODIFICATION FROM THESE
MODIFICATIONS ENCLOUDING XIST
RNA, APPROXIMATE WHEN YOU
STIMULATE THESE INVITRO, AND
THESE COME BACK TO THE
X-CHROMESOME AND WE ALSO FOUND
THIS PROCESS IS INDEPENDENT OF
TRANSCRIPTION OF THE XIST GENE,
SO THE LOCALIZATION OF XIST,
HAPPENS JUST GENETICALLY
DISTINCTION FROM ITS
TRANSCRIPTION EMPLOY AND O IN
OUR LAB, WE'VE BEEN
INVESTIGATING THE EPIGENETIC
MECHANISMS OF X-INACTIVATION IN
TYPES DIFFERENT TYPES OF IMMUNE
CELLS IN THE HEALTHY STATE BUT
WE'VE ALSO BEEN INVESTIGATING
HOW THESE MECHANISMS ARE
PERTURBED IN AUTOIMMUNE DISEASE
SUCH AS LUPUS, SO WE'VE BEEN
EXAMINING THE EPIGENETIC
FEATURES OF THE INACTIVE
X-CHROMOSOME IN BOTH ADULT AND
PEDEIATE RICK LUPUS PATIENT
LYMPHOCYTES AND WE MADE THE
OBSERVATION THAT IN B-CELLS
EXIST RNA, IS MISLOCALLIZED IN
ACTIVATED B-CELLS IN LUPUS
PATIENTS COMPARED TO ADULT,
HEALTHY INDIVIDUALS AND THIS IS
TRUE BOTH FOR ARK DULT PATIENTS
AND ALSO FOR PEDIATRIC PATIENTS.
AND WE'VE ALSO MADE THE
OBSERVATION THAT CIRCULATING
NAIVE B-CELLS AND ALSO IN VIVO
ACTIVATED B-CELLS FROM LUPUS
PATIENTS HAVE A BARREN
EXPRESSION OF GENES SUGGESTING
THAT THE MISLOCALLIZATION OF
XIST RNA MIGHT BE AFFECTING GENE
SILENCING AND ALSO GENE
EXPRESSION FROM THE INACTIVE
X-CHROMOSOME.
AND SO WORK WITH MOUSE MODELS OF
SPONTANEOUS LUPUS LIKE DISEASE,
THE NTBW F1, WE'VE SEEN THESE
FEMALE ANIMALS WITH LATE STAGES
OF DISEASE ALSO EXHIBIT
MISLOCALLIZATION OF XIST RNA
FROM THE INACTIVE X-CHROMOSOME,
AND SO TAKEN TOGETHER, WE'VE
PROPOSED THIS HYPOTHESIS THAT
PERHAPS IMPAIRED DYNAMIC
X-INACTIVATION MAINTENANCE IN
B-CELLS MIGHT CONTRIBUTE TO THE
SUSCEPTIBILITY FOR INDIVIDUALS
WITH 2 X-CHROMOSOMES TO DEVELOP
AUTOIMMUNE IMMUNE DISEASES.
SO WHEN THEY RETURN TO THE
ACTIVE X TO MAINTAIN
TRANSCRIPTIONAL SILENCING FOR
THESE IMPORTANT IMMUNE
REGULATORY GENES ON THE INACTIVE
X-CHROMESOME, HOWEVER, IN THE
CONTEXT OF LUPUS LIKE DISEASE
AND FEMALE IMMUNITY DISEASES,
THE TETHERS OF THE X-CHROMESOME
IS COMPROMISED AND THERE'S ALSO
ENRICHEDMENT OF THE
MODIFICATIONS ALONG THE
CHROMESOME, SO THE CHROMATIN OF
THE INACTIVE X-CHANGES AND
BECOMES PERMISSIVE FOR CERTAIN
GENES TO BECOME REACTIVATED FROM
THIS CHROMESOME RESULT NOTHING
THE OBSERVATION THAT THERE'S
INCREASED EXPRESSION OF SOME
IMMUNE ACTIVATED GENES.
SO ALL THESE OBSERVATIONS TODAY
HAVE BEEN CORRELATIVE SO WE
WANTED TO LOOK AT CAUSALITY AND
MECHANISM, SO IN ORDER TO DO
THIS WE WANTED TO TURN TO A
MOUSE MODEL WHERE WE COULD
IMPAIR OR DISRUPT X-INACTIVATION
MAINTENANCE AND SO WHAT I WILL
TELL YOU ABOUT TODAY IS THE WORK
OF A VERY TALENTED MD-Ph.D.
STUDENT CLAUDIA LEVEL WHO HAS
BEEN WORKING ON THIS IN MY LAB
AND THIS IS ALL UNPUSH PUBLISHED
AND WE'RE EXCITED TO SHARE THIS
WITH THE COMMUNITY BECAUSE WE
THINK IT'S AN EXCITING NEW
MECHANISM, THAT WILL COMPLEMENT
THE OTHER MECHANISMS THAT WE'RE
GOING TO HEAR ABOUT FROM DIANEA
AND BRENDAN IN THE LATER PART OF
THE SEMINAR.
SO X-INACTIVATION TAKES PLACE IN
THE MOUSE, DURING THE
PREIMPLANTATION STAGE EMBRYO
WHERE 1 X IS RANDOMLY CHOSEN FOR
TRANSCRIPTIONAL SILENCING AND SO
IN ORDER TO MAKE A B-CELL
SPECIFIC EXIST DELETION, WE
MATED XIST 2-LOCK ANIMALS THAT
EXPRESS THE N1 CREE RECOMBIN
ACE, SO THIS,A LOWS FOR
EXPRESSION, SPECIFICALLY DURING
B-CELL DEVELOPMENT STARTING AT
THE PRO B-CELL STAGE, SO WE
GENERATED THROUGH VARIOUS
INTERPROCESSES, HOMOWRIGGOUS
EXIST ANIMALS WHERE EXIST IS
DELETED ON BOTH X-CHROMOSOMES,
AND NEXT WE VERIFIED THAT EXIST
WAS DELETED AND HOMOZYGOUS KNOWN
ANIMALS USING QPC R AND WE ALSO
PERFORM SEQUENTIAL EXIST RNA
FISH FOLLOWED BY IMMUNO
FLUORESCENCE TO LOOK AT 1
HETEROGENEOUS ORY CHROMEATIC
MARK ENRICHED ON THE INACTIVE
X-CHROMOSOME, H2 KA19 UBIQUITIN,
YOU SEE THIS RNA XIST CLOUD THAT
FOCUSED WITH THE H2 K119.
AND THEN HOMOZYGOUS EXIST
DELETED MICE DO NOTEE IT IN
EXIST CLOUD AND ALSO SEE THERE'S
NO FOCUS FOR FHKAUBIQUITIN, THE
NOWLET DELETION OF EXIST HAS
BEEN DONE BY OTHER BROWPS AND
THE DELETION CAN RESULT IN A
RANGE OF DIFFERENT PHENOTYPES
THAT DEPEND ON WHERE AND WHEN
THE DELETION HAS TAKEN PLACE.
SO FOR EXAMPLE, IF YOU DELETE
EXIST, USING A GENERAL DELETER,
FEMALE ANIMALS, EMBRYOS DO NOT
SURVIVE, IT'S EMBRYONIC LETHAL.
AND WHEN XIST IS DELETED IT CAN
IMPACT GENE EXPRESSION
SPECIFICALLY FROM THE INACTIVE
X-CHROMESOME RESULT NOTHING
PARTIAL REACTIVATION OF THIS
CHROMESOME.
HOWEVER, THE NUMBER OF X-LINKED
YEENS AND ALSO THE IDENTITY OF
THESE X-LINKED YEEN SYSTEM CELL
TYPE SPECIFIC DEPENDING ON WHERE
YOUR DELETING HAPPENS, SO
THERE'S A PHENOTYPE THAT RESULTS
FROM THIS DELETION, SO FOR
EXAMPLE, IF YOU DELETE EXIST IN
THE SKIN WITH A KERATIN ARE 14
CREE, THE ANIMALS ARE FINE AND
THERE'S NO NOTABLE PHENOTYPE, IF
YOU DELETE IT IN THE GUT,
THERE'S NO STEADY STATE, HOWEVER
IF YOU INDUCE POLARIZED UP FORM
EGG IN THESE ANIMALS YOU SEE
THERE'S MORE SEVERE DISEASE WHEN
XIST IS DELETED AND THEN FINALLY
DELETION IN HEMATOPOIETIC STEM
CELLS USING A BABCREE, RESULTS
IN FEMALE SPECIFIC BLOOD CANCER
WHERE THERE'S A HUGE EXPANSION
OF THE MYELOID COMPARTMENT AND
THE FEMALE MICE DEVELOP A
MYELOID DRIVEN CANCER
PHENOTYPES.
OKAY, SO WHAT DID WE FIND?
SO WHEN WE DELETE EXIST IN THE
B-CELL COMPART AM, THE FEMALE
HOMOZYGOUS ANIMALS EXHIBIT
EQUIVALENT VIABILITY, SAME LIFE
SPAN AS WILD-TYPE LITTER MATES,
WE SEE THERE'S NORMAL B-CELL
DEVELOPMENT IN THE BONE MARROW,
AND SO, NEXT WE LOOKED IN THE
SPLEEN AND WE SEE THAT FEMALE
EXISTS AND THE FEMALE 6 MONTHS
OF AGE, THE TOTAL NUMBER IN THE
SPLEEN IS NOT AFFECTED, HOWEVER
IF YOU LOOK CAREFULLY ACROSS
B-CELL SUBSETS WE SUE --SEE THEE
ARE SOME ELEVATIONS WITH THE
ACTIVATED GL7 POSITIVE, B-CELLS
WHERE XIST KNOCKOUT MICE ARE
SLIGHTLY HIGHER, SIGNIFICANT,
MORE OF THESE ACTIVATED B-CELLS,
THEY ALSO HAVE MORE SHORT LIVED
PLASMA CELLS AND ALSO MORE CLASS
SWITCH B-CELLS.
NOW, BECAUSE THE EXIST KNOCKOUT
MICE HAD MORE ACTIVATED B-CELLS,
WE MONITORED THE FEMALE MICE,
FOR AUTOANTIBODIES SPECIFIC TO
DOUBLE STRANDED DNA, SO DSDNA,
AND AS A POSITIVE CONTROL WE USE
SERUM 42 FEMALE FDBW MICE WHICH
SPONTANEOUSLY DEVELOP LUPUS LIKE
DISEASE AND THE POSITIVE ARE
SHOWN HERE IN YELLOW AND WE SEE
FOR A RANGE OF AGES OF FEMALE
MICE BETWEEN 6 WEEKS OF AGE AND
12 MONTHS OF AGE, THE MAIORITY
OF XIST HOMOZYGOUS NULL MICE DO
NOT DEVELOP ELEVATED LEVELS OF D
SDNA.
HOWEVER, WHEN WE LOOK AT ANIMALS
BOTH WILD-TYPE AND XIST XOK OUT
MICE THAT WERE 12 MONTHS OF AGE
OR OLDER, WE SEE THERE WERE SOME
MICE THAT DEVELOPED VERY HIGH
LEVELS OF ANTIDSDNA IN THEIR
SERUM, SO WE FOCUSED OUR
ANALYSIS ON AGE MATCHED
WILD-TYPE AND ALSO EXIST
HOMOZYGOUS NULL DELETED FEMALE
MICE AND DISTINGUISHED BETWEEN
XIST KNOCK OUT ANIMALS THAT HAD
EITHER LOW LEVELS OF ANTIDSDNA
AUTOANTIBODIES OR ANIMALS THAT
HAVE HIGH LEVELS OF DSD NA
AUTOANTIBODIES.
HAVE SIGNIFICANTLY MORE NUMBERS
OF DSDNA SECRETING CELLS
COMPARED TO WILD-TYPE CELLS ALSO
EXIST KNOCK OUT LOW MICE, SO
THESE ARE LA-SPOT EXPERIMENTS
WHERE WE QUANTIFIED THE NUMBER
OF POSITIVE CELLS IN THE PLEEN O
SIGHTS THAT WERE RESPONSIBLE FOR
PRODUCING THE DSD NA
AUTOANTIBODIES.
SO NEXT WE WANTED TO ASK WHAT
OTHER AUTOANTIBODIES WERE
PRESENT IN THE SERUM OF THE
KNOCK OUT HIGH AND LOW MICE AND
WE USED AN AUTOANTIBODY
MICROARRAY TO PROFILE ACROSS 120
DIFFERENT AUTOANTIIENS AND WE
FOUND THERE ARE 16 DIFFERENT
AUTOANTIGENS THAT ARE ELEVATED
IN THE SERUM FROM EXIST KNOCKOUT
MICE, HIGH KNOCKOUT MICE,
COMPARED TO LOW KNOCKOUT MICE
AND AS EXPECTED WE SEE THAT
ANTIDNA IS ELEVATED ALONG WITH
ANTISMITH COMPLEX, R& P
PROTEINS WHICH ARE ALSO KNOWN TO
BE ELEVATED IN SLE PATIENTS, WE
ALSO OBSERVED THERE'S OTHER
AUTOANTIGEN THAT ARE STRONGLY
ASSOCIATE WIDE OTHER FEMALE BIAS
AUTOIMMUNE DECEASES SUCH AS MDA
5 AND ALSO ANTIRO.
SO NEXT WE EXAMINED KIDNEYS FROM
XIST KNOCK OUT LOW AND HIGH MICE
TO LOOK FOR EVIDENT OF
PATHOLOGY.
SO WE SUBMITTED THE KIDNEY
SECTIONS FOR PLIENDED
PATHOLOGICAL ASSESSMENT BY THE
PENN VETS COMPARATIVE PATHOLOGY
CORE.
AND OUR PATHOLOGY HALEY FOUND
THAT XIST KNOCK OUT HIGH MICE
HAD HIGHER PATHOLOGY SCORES
COMPARED TO THE XIST KNOCK OUT
LOW MICE AND THIS IS AN EXAMPLE
OF A KIDNEY SECTION RIGHT HERE
SO THIS IS THAT STAINING AT 20 X
RESOLUTION, AND WE ARE SHOWING 3
GLOMULAR MARRIAL EYE I, AND WE
HAVE 3 EPITHELIUM AND ALSO 2
GLOMULAR MERUE LIE THAT ARE
HYPER TROPHIC, THERE'S ALSO MY O
STIRTIAL INFLAMMATION, AND ALSO
EVIDENCE OF TUBEULAR EPITHELIAL
DEGENERATION AND ALSO LOSS WITH
INTERTUBEULAR CAST SHOWN HERE.
SO NEXT WE WANTED TO SHOW WELL
WELL ARE THERE CHANGES TO THE
B-CELL SUBSET IN THE KNOCKOUT
MICE, SO WE ISOLATED SPLEANS AND
BONE MARROW TO DO FLOW CYTOMETRY
AND SO TODAY BECAUSE OF TIME,
I'M JUST GOING TO SHOW YOU THE
RESULTS FROM SPLEN AND TISSUE,
SO WHAT WE FOUND WAS THAT EXIST
KNOCK OUT HIGH MICE, SHOWN HERE
IN THE PURPLE BARS HAVE MORE
SPLEEN X CD11 C AGE ASSOCIATED
B-CELLS BOTH IN FREQUENCY AND
ALSO IN TOTAL NUMBERS.
XIST KNOCK OUT HIGH MICE HAVE
MORE GH7 POSITIVE ACTIVATED
B-CELLS BOG IN FREQUENCY AND
ALSO TOTAL CELL NUMBER.
EXIST KNOCK OUT HIGH MICE ALSO
HAVE SHORT LIVED PLASMA CELLS,
THEY ALSO HAVE HIGHER NUMBERS OF
LONG LIVED PLASMA CELLS, AND
ALSO HIGHER NUMBERS OF CLASS
SWITCHED B-CELLS.
SO, NEXT WE WANTED TO ASK--SO
TAKEN TOGETHER, THE XIST
HOMOZYGOUS B-CELL MICE HAVE MORE
ACTIVATED B-CELLS, SIMILAR TO
THE WILL YOU PLEASEUS PATIENT
ANDS WE WANTED TO ASK HOW IS
GENE EXPRESSION AFFAIRS TEAM
LEADERRERRED IN THESE B-CELL
SUBSETS SO BECAUSE OF TIME I
WILL JUST SHOW YOU THE RNA
SEQUENCING RESULTS FROM 1 SUBSET
THAT EXPANDS AND THE XIST KNOCK
OUT HIGH MICE AND THESE ARE THE
CD11 AND B-ASSOCIATED C-CELLS
AND LOOKING SPECIFICALLY AT THE
X-CHROMESOME WE FOUND THERE ARE
10 X-LINKED GENES THAT ARE
ABERRANTLY UPREGULATED IN THE
MICE, SO 1 THAT CAUGHT OUR
INTEREST IS TASL, IT'S THE
ADAPTER PROTEIN THAT REGULATING
ACTIVATION OF IRF 5 DOWN STREAM
OF TLR8, 7 AND 9 SIGNALING WHICH
RESULTS IN THE PRODUCTION OF
TYPE 1 INTERFERONS CROSS LINKING
SO IT'S KNOWN THAT STIMULATION
OF SIGNALING PATHWAY IS INVOLVED
IN THE PATHOGENESIS OF SLE, O
LOOKING AT AUTOSOMAL GENES WE
OBSERVE XIST KNOCK OUT HIGH MICE
HAVE KNOCKED OUT DIFFERENT
EXPRESSIONS OF AUTOSOMAL GENES
SO THEN TO SUMMARIZE WHAT WE
FOUND IS THAT A B-CELL SPECIFIC
XIST DELETION IN BALB C-MICE CAN
RESULT IN SPONTANEOUS LUPUS LIKE
DEC IN SOME ANIMALS SO ANIMALS
OVER 12 MONTHS OF AGE HAVE
ELEVATED AUTOANTIBODY
PRODUCTION, INCREED GLOMULAR
MARRIAL PATHOLOGY AND ALSO
INCREASED NUMBERS OF ACTIVATED
B-CELLS AND MEMORY ACTIVATED
B-CELLS.
SO NEXT WE WANT TO ASK, WELL
DELETION EXIST IMPACT FEMALE
MICE AND INDUCE MODEL OF LUPUS
LIKE DISEASE, SO BECAUSE THE
DISEASE PEN TRAPTS FOR THE XIST
KNOCKOUT MICE IS LOW YET SIMILAR
TO LUPUS PATIENTS WE WANT TO
LOOK AT MECHANISMS OF DISEASE
DEVELOPMENT SO WE TURN TO THIS
MODEL OF USING PRIVATEAIN, SO
PRIOF THEAIN INJECTIONS IS
ELEVATED LEVELS OF ANTIDSDNA
AUTOANTIBODIES AND ALSO
INCREASED PRODUCTION OF
ACTIVATED B-CELLS AND GLUE
MERULAR NEPHRITIS, THIS SHARES
SOME FEATURES BUT NOT ALL
FEATURES WITH LUPUS PATIENTS SO
WE WANT TO ASK THE QUESTION, HOW
DID THIS DELETION AFFECT THIS
PROCESS, SO FOR THESE
EXPERIMENTS, WE INJECTED VARIOUS
COHORTS OF FEMALE MICE, BOTH
WILD-TYPE AND EXIST KNOCK OUT,
WITH 1 TIME INJECTION OF PRIOF
THEAIN AND THEN WE MONITOR THE
ANIMALS FOR DEVELOPMENT OVER
TIME BY COLLECTING BLOOD AND
URINE, AND AT 3 DIFFERENT TIME
POINTS WE HARVESTED THE KIDNEYS
AND THE BLOOD FROM THE ANIMALS
TO EXAMINE CHANGES WITH THE
B-CELL SUBSETS.
AND SO, FIRST WE LOOKED AT
SERUM, FOR AUTOANTIBODY
PRODUCTION AND AS EXPECTED WE
SEE THAT PRISTANE TREATED
WILD-TYPE ANIMALS PRODUCED MORE
ANTIDSDNA AUTOEABT BODIES OVER 7
MONTHS OF TIME WHEREAS
NONTREATED WILD-TYPE ANIMALS
LACK DETECTION OF ANTIDSDNA.
AND WHAT WE OBSERVED WAS THAT
EXIST KNOCKOUT MICE THAT WERE
TREATED WITH PRISTANE PRODUCED
HIGHER LEVELS OF DSDNA AUTOEABT
BODIES COMPARED TO THE WILD-TYPE
PRISTANE TREATED MICE.
SO NEXT WE EXAMINED THE LEVELS
OF B-CELL SUBSETS OVER TIME AND
WE FOUND THAT PRISTANE TREATED
XIST KNOCKOUT MICE DEVELOPED
MORE AGE ASSOCIATED B-CELLS AT 9
MONTHS OF AGE SHOWN HERE IN THE
BINK BARS AND THEN LOOKING
SPECIFICALLY AT SHORT LIVED
PLASMA CELLS WE SEE THAT EXIST
KNOCKOUT MICE TREATED WITH
PRISTANE AND THESE LEVELS AT 7
MONTH ANDS 9 MONTHS POST
INJECTION.
PRISTANE TREATED XIST KNOCKOUT
MICE HAD HIGHER TOTAL NUMBERS OF
GL7 POSITIVE ACTIVATED B-CELLS
BOTH AT 7-MONTHS AND 9 MONTHS
POST INJECTION.
AND WE ALSO DETECTED INCREASED
NUMBERS OF CLASS SWITCH B-CELLS
FOR EXIST KNOCKOUT MICE THAT
WERE TREATED WITH PRISTANE.
SO TAKEN TOGETHER, PRISTANE
INDUCED DEC REVEALS THAT XIST
DELETION CAN IMPACT B-CELL
NUMBERS AND INCREASE B-CELLS
THAT ARE ACTIVATED AND ALSO THE
B-CELLS THAT ARE RESPONSIBLE FOR
PRODUCING ANTIBODIES.
SO NEXT WE ASKED WHAT X-LINKED
GENES ARE AFFAIRS TEAM
LEADERRERRED IN B-CELLS IN THE
CONTEXT OF PRISTANE INDUCED
DISEASE AND AND I WILL SHOW YOU
THE DATA FOR 2 B-CELL SUBSETS,
ABC CELLS OR ABCS AND ALSO GL7 +
ACTIVATED B-CELLS, SO WOOK
LOOKING AT THE CHROMESOME, WE
SEE THERE ARE 10 X-LINKED GENES
THAT ARE UPREGULATED AND KNOCK
OUT PRISTANE TREATED MICE AND
THIS INCLUDES TASL WHICH WAS
ALSO OVEREXPRESSED IN THE AGE
ASSOCIATED B-CELLS FROM EXIST
KNOCKOUT MICE WITH SPONTANEOUS
LUPUS LIKE DISEASE, AND NOW IN
GL7, POSITIVE ACTIVATED B-CELLS
LOOKING SPECIFICALLY AT THE
X-CHROMESOME, WE ALSO SEE, THAT
TASL IS UPREGULATED ALONG WITH
OTHER X-LINKED YEENS IN THESE
B-CELL SUBSETS.
OKAY, SO TAKEN TOGETHER, I WILL
END WITH THE MODEL HERE, SO,
WHAT WE FOUND IS THAT XIST
DELETION CAN LEAD TO THE
UPREGULATION OF X-LINKED IMMUNE
GENES SPECIFICALLY TASL WHICH WE
THINK CONTRIBUTES TO OUR RESULT
IN PHENOTYPES THAT ARE OBSERVED
IN SLEPATES.
SO WHEN XIST IS DELETE FRIDAY
THE X-CHROMESOME IN B-CELLS
THERE'S A LOT OF FROM THE
X-CHROME COME IS ALSO LOTS OF
HETEROCHROMEATIC MARKS, WE SEE
THERE'S TASL, WE SEE INCREASED
FREQUENCY AND ACTIVATED NUMBER
OF B-CELL SUBSETS AND MEMORY
B-CELL SUBSETS.
WE SEE PRODUCTION OF VARIOUS
AUTOANTIBODIES AND ALSO ON A
NUMBER OF DIFFERENT GLUE MARRIAL
PATHOLOGIES AND IMPORTANTLY
THESE ARE ALL PHENOTYPES THAT
HAVE BEEN OBSERVE INDEED LUPUS
PATIENTS WHICH HAVE DISPURSED
PATTERNS OF RNA LOCALIZED AT THE
INACTIVE X-CHROMESOME SO WITH
THAT I WANT TO TAKE A MOMENT AND
THANK ALL THE PEOPLE IN MY LAB,
ESPECIALLY CLAUDIA WHO HAS
REALLY BEEN SPEARHEADING THIS
PROJECT WHICH HAS BEEN A VERY
TECHNICALLY CHALLENGING PROJECT
AND REALLY REQUIRES A LOT OF
PATIENTS AND DETERMINE NATION
BECAUSE THESE ANIMALS ARE QUITE
AGED WHEN THEY PRESENT WITH
PHENOTYPES.
SO IN PARTICULAR, DR. HALEY
AMERMAN, DOING ALL THE PATHOLOGY
BLINDED FOR THE KIDNEY SECTIONS,
UT SOUTHWESTERN FOR THE GENOMICS
AND MICRO,A RAY, AND FINALLY ALL
THE FUNDINGS SOURCES FOR THE
LAB.
SO WITH THEY WILL STOP SHARING.
>> THANK YOU DOCTOR, FOR ANY WHO
JOINED LATE, WE WILL TAKE
QUESTIONSA THE END AFTER ALL 3
SPEAKER VS SPOKEN, AT THIS TIME
I WOULD LIKE TO INTRODUCE THE
NEXT SPEAKER DR. DIANA DOU, FROM
HARVARD UNIVERSITY.
>> THANK YOU.
LET ME SEE IF I REMEMBER HOW
THORS TO
DO THIS.
ALL RIGHT, YES, THANK YOU FOR
THAT INTRODUCTION AND I AM A
POST DOC IN HOWARD CHANG'S LAB
AT STANFORD AND THANK YOU FOR
INVITING ME TO SPEAK ABOUT THE
ROLES THAT EXIST IN AUTOIMMUNITY
AND SHARE OUR STORY TODAY.
SO I ALWAYS LIKE TO START WITH
THIS CARTOON BECAUSE, WHAT DREW
ME TO HOWARD'S LAB WAS NONCODING
RNAs AND I LOOK THIS ANALOGY
OF LOOKING FOR THESE AT NIGHT,
AND TRADITIONALLY WE'VE ONLY
FOCUSED ON LOOKING FOR THESE DEC
IN THE WELL LIT AREAS OF THE
GENOME, SO PRIMARILY ENCODING
GENES BUT WHEN YOU CONSIDER THAT
IT'S LESS THAN 2% ENCODING WHAT
ALMOST IGNORING 99% OF THE
POSSIBLE PLACES WHERE YOU COULD
HAVE LOST YOUR KEYS OR WHERE
THREEZ IMPORTANT KEYS ARE, SO
THIS IS A GREAT ANIMATION FROM
HUMAN HEALTH AND DEC, SO I THINK
1 OF THE AREAS WHERE THIS
PRESENTS GREAT PROMISE IS THAT 1
OF THE BLACK BOXES OF MEDICINE
THAT WOULD BENEFIT FROM
SEARCHING FOR MORE KEYS ARE
AUTOIMMUNE DISEASES.
SO THIS IS LIKE A BASIC SLIDE,
SHOWING HOW THE IMMUNE SYSTEM IS
VERY COMPLEX, ALWAYS MAINTAIN
THIS BALANCE BETWEEN
SELF-TOLERANCE AND PROTECTION,
IF YOU HAVE TOO MUCH TOLERANCE,
YOU TEND TO NOT CLEAR OUT CELLS
AND YOU GET CANCER WHEREOT OTHER
END OF THE SPECTRUM, IF YOU HAVE
TOO MUCH ACTIVITY AND
PROTECTION, YOU DEVELOP
AUTOIMMUNITY.
AND THE PROBLEM WITH
AUTOIMMUNITY RIGHT MOW IS THAT
IT'S DIFFICULT TO DIAGNOSE AND
LIMITED THERAPIES BECAUSE, AT
LAST YEAR'S SCHEDULED FOUNDATION
MEETING 1 OF THE CLINICIAN
SCIENTISTS MENTIONED SOMETHING
THAT STUCK WITH ME WHICH IS THAT
THE FIELD'S AUTOIMMUNITY SEEMS
TO BE IN THE FIELD OF CANCER
WHERE AROUND 10 YEARS AGO WHERE
EVERYTHING WE'RE DOING
CLINICALLY IS MOSTLY SYMPTOM
BASES BECAUSE WE DON'T REALLY
UNDERSTAND THE CAUSE AND RISK
FACTORS FOR AUTOIMMUNITY
COMPLETELY, SO WHAT ARE THE RISK
FACTORS FOR AUTOIMMUNE DEC THAT
WE KNOW ABOUT.
WELL WE KNOW THERE'S A VERY
STRONG GENETIC COMPONENT BUT
THERE'S ALSO AN ENVIRONMENTAL
COMPONENT.
HOWEVER, THE SINGLE VARIOUS RISK
FACTOR FOR DEVELOPING
AUTOIMMUNITY IS SIMPLY
BIOLOGICAL FEMALES SO 80% OF THE
AUTOIMMUNE POPULATION IS FEMALE,
AND I THINK ALL OF US USE--ALSO
USE SOME OF THE SLIDE FROM THE
SAME REVIEW, IT'S A GREAT
REVIEW.
SO AGAIN HIGHLIGHTING THE RISK
FACTOR FOR DEVELOPING
AUTOIMMUNITY, FEMALES PRESENT A
MUCH HIGHER INCIDENCE AND RISK
FACTOR FOR DEVELOPING IT AND SO
THESE ARE JUST SOME OF THE
FEMALE BIASED IMMUNE DECS, I
WILL SEE IF I CAN GET MY LASER
POINTER UP HERE, YEAH, THE
FEMALE BIASED AUTOIMMUNE DECS
AND 1 OF THE MODELS WE USE IN
MICE, IS THE LUPUS, SO WE
CREATED THIS GREAT INTRODUCTION
WHICH SAVES ME A LOT OF TIME SO
SHE SHARED THAT AUTOIMMUNE
CORRELATES WITH THE DOUBLE
XX COMPLEMENT AND YOU KNOW, YES,
AT THE VERY BASAL LEVEL, THE
DRVESS BETWEEN BIOLOGICAL MALES
AND FEMALES IS THAT FEMALES HAVE
2 X-CHROMESOMES AND WHILE MALES
HAVE 1, AND WHAT'S INTERESTING
IS THAT IN DISEASES LIKE LUPUS,
THE RISK OF DEVELOPING
AUTOIMMUNITY, SUCH AS IN LUPUS
IN [INDISCERNIBLE], THERE'S 1
X-CHROMOSOME INCREASES
DRASTICALLY FOR A LEVEL THAT'S
SIMILAR TO WHAT WE SEE IN THE
FEMALES.
SO WHAT IS IT ABOUT HAVING 2
X-CHROMESOMES THAT IS ABLE TO
ELEVATE THIS RISK, SO
SIGNIFICANTLY IN BOTH MALES AND
FEMALES.
WELL, WE KNOW THAT THE
X-CONTAINS THE LARGEST NUMBER OF
IMMUNE DISEASE, AND KIND OF
BREEZE THROUGH IT AND SO THEY
SHOWED THEIR BEAUTIFUL TODAYS
SHOWING HOW X-GENE EXPRESSION
REALLY NEEDS TO BE TIGHTLY
CONTROLLED, AND THE WAY IT IS
CONTROLLED IS THROUGH THE RNA
CODING, XIST, AND IT SILENCES 1
OF THE 2 X-CHROMESOMES, SO SHE'S
SHOWN HER PATTERNS, IF THERE'S
INCORRECT PATTERNS IT CAN AFFECT
THE GENE EXPRESSION OFF THE
X-CHROMESOME, AND IF YOU GET
THESE YEENS YOU DEVELOP HIGH
AMOUNTS OF AUTOIMMUNITY AND SO 1
OF THE YEENS THAT YOU MENTIONED,
THAT'S AN X-GENE IS TLR 7,
HOWEVER, WHAT OUR WORK WAS
STUDYING IS XIST ITSELF, WHETHER
JUST HAVING XIST, AND THE
PARTICLE THAT IT FORMS IN THE
FUNCTION, IT'S NONCODING SO IT
NEEDS TO HAVE ALL THE PROTEINS
IN THIS COMPLEX TO ACTUALLY HAVE
AN ABILITY TO FUNCTION AND
SILENCE THE X-CHROMESOME.
SO MY FOCUS ON WHETHER THIS
EXISTS ON THE PARTICLE ITSELF,
IT IS A FACT THAT INCREASES THE
AUTOIMMUNE RISK.
SO THERE IS ACTUALLY PRESENT FOR
THE CONCEPT OF THE EXIST
RIBONUCLETIDE AFTER ALL MANY
RNPs ARE KNOWN AUTOANTIGENS
AND THE COMPLEXES DO ENGAGE
TLRs AND AUTOIMMUNE DISEASE,
FOR EXAMPLE, TLR7 OF ACTIVATION
OF DENDRITIC CELLS, AND WHAT IS
IN THE RNF, WELL THE LAB
DEVELOPED LIKE THIS DIRECT
PROTEOMICS TECHNIQUE CALLED TRIP
MS, USED TO PULL DOWN XIST
COMPLEX AND WE USE IT TO
IDENTIFY THE PROTEINS.
SO WHAT'S INTERESTING IS THAT WE
IDENTIFY 81 UNIQUE BINDING
PROTEINS IN THE XIST PULL DOWN
AND A ABOUT A THIRD OF THOSE ARE
KNOWN AUTOANTIGENS WITH KNOWN
ASSOCIATIONS WITH AUTOIMMUNE
DISEASEs, INCLUDING ANTILAW,
SSB, WHICH BRINGS US TO THE
HYPOTH THIS THAT THIS EXISTS AND
IT CAN BE SCAFFOLDING INTO THESE
LARGE AUTOPOLYMERRIC COMPLEXES
THAT'S ACTIVATING IMMUNE
RECEPTORS SO THAT IF YOU GET
PROLONGED ACTIVATION OF THIS
IMMUNE SYSTEM, PROLONGED
INFLAMMATION, YOU CAN DEVELOP
CHRONIC INFLAMMATION AND
DISEASE.
SO IN MY TALK, IT'S KIND OF
GOING TO BE SEPARATED INTO
PARTS, THE FIRST OF WHICH IS
WE'RE LOOKING INTO WHETHER
EXPRESSION OF THESE XIST RNPs
ELEVATES THE RISK OF AUTOIMMUNE
DEC AND FOR THIS WE USE A MOUSE
MODEL.
AND THENOT CLINICAL SIDE, WE ASK
LIKE ARE THE CONPOEMENTS OF THE
XIST RNP, SERUM AUTO, CAN WE
DETECT SERUM AUTOANTIBODIES SO
IS THE EXIST RNP AN ANTIGEN
COMPLEX, SO WE'RE FORTUNATE TO
WORK WITH ANTON WOODS IN ZURICH
AND SHOWING THAT IT BEHAVES
MODULARLY.
SO IF YOU TAKE OUT THE FIRST 900
BASE PAIRS THERE'S A REGION
CALLED REPEAT A, SO WITHOUT THE
REGION OF REPEAT A, YOU DON'T
GET SILENCING OF THE
S-CHROMESOME, SO IT'S REQUIRED
FOR YEEN SILENCING IS THAT WE
DID WITH ANTON WE CREATED THIS
INDUCIBLE TRANSGENE WITH THIS
REPEAT A MUTANT EXIST, AND WE
PUT IT UNDER PET OP PROMOTER SO
WE CAN CONTROL THE EXPRESSION
AND PUT IT INTO THE CHROMOSOME
LISTEN.
YOU WILL NOTICE THAT IN THIS
MODEL I PUT IT ON 1 CHROMESOME
BECAUSE XIST IS JUST ON THE
XCHROME COME SO TO MIMIC THAT WE
USED HETEROZYGOTES AND THIS,A
LOWS US TO STUDY THE FUNCTION OF
WHAT WAS REALLY A PHENOSPECIFIC
LINK RNA AND RNA AND RNP COMPLEX
IN A MALE BACKGROUND INDEPENDENT
OF HORMONAL CONTRIBUTIONS AND
DEVELOPMENT TO VERIFY WE WERE
SEEING INDUCIBLE EXPRESSION IN
THE SYSTEM.
WE USE FISH, WE ALSO USE QPC R,
THROUGH SYSTEMIC ORGANS, MANY
ORGANS IN THE MOUSE, SO YOU CAN
SEE IN THE FEMALE CONTROL THAT
WAS NOT TREATED WITH DOXYPSYCHE
LINE AND YOU SEE THESE BRIGHT
TYPES OF THE WILD-TYPE XIST IN
THE TRANSGENIC ANIMALS THAT WERE
NOT TREATED WITH PRISTANE, SO
YOU DON'T SEE IF IN THE NUCLEI,
HOWEVER, WHEN YOU ADD IT TO THE
TRANSGENIC MOUSE, NOW YOU'RE
ABLE TO SEE THESE CLEAR AND
BRIGHT IMAGES HERE.
SO WITH THE KEY TAKE AWAY, WE
ARE ABLE TO GET THE EXIST RNP
WITHOUT ANY SILENCING AND THIS,A
LOWS US TO STUDY WHETHER XIST
EXPRESSION ALONE IS SUFFICIENT
TO INCREASE AUTOIMMUNITY.
SO WE CHOSE THE VERY WELL
STUDIED INDUCED MODEL AND WE
SHARE MANY ATTRIBUTES OF PEASHTS
IN LUPUS INCLUDING A FEMALE SEX
BIAS, SO KIND OF THROWING BACK
TO THE TRAITS OF LUPUS IN
PATIENTS, THERE IS A VERY STRONG
FEMALE, INCIDENCE, IT'S
MULTIORGAN AS WAY OF THE RANGE
AND REFLECTED IN THE PRISTANE
MOUSE MODEL AS WELL, BOTH HUMANS
AND THIS MOUSE MODEL, IT'S
INTERFERON AND TLR7 DRIIVE, AND
TWEPS A WIDE VARIETY OF
AUTOANTIBODIES.
SO WITH THIS MODEL WE TESTED IN
BOTH AUTOIMMUNE RESISTANT AND
PERMISSIVE BACKGROUND.
SO WE INITIALLY CONDUCKED OUR
EXPERIMENTS IN THE BLACK 6 MICE
BECAUSE IN THE WILD-TYPE STUDIES
WE KNOW THAT THE MALES NEVER GET
ANY DISEASE, SO WE THOUGHT IT
WOULD BE A BIENAR READ OUT, AND
SO, LATER ON, IN THIS
PRESENTATION, I WILL EXPLAIN TO
YOU, YOU WILL SEE WHY IT'S VERY
IMPORTANT TO HAVE LIKE A
RHEUMATOLOGIST AS YOUR
COLLABORATORS.
SO THIS IS KIND OF BASIC SLIDES
JUST SHOWING YOU THE PATHWAYS
AND IMMUNE CELLS THAT ARE
INVOFFED IN AUTOIMMUNE SIGNALING
AND HOW IT CAN PROCEED 2 WAYS
TRADITIONALLY WITH THE B-CELLS
PRESENTING AND ACTIVATING THE
T-CELLS AND ALSO PRODUCED
ANTIBODIES, T-CELLS WILL YOU
KNOW ACTIVATE THE CD8 T-CELLS,
PRODUCE CYTOKINES BUT THIS
PATHWAY COULD ALSO BE MORE
LIKELY CELL OR INITIATED OR
DRIVEN AND PRESENTATION CAN
OCCUR FROM THE DRAINAGE CELLS
AND SO THIS SLIDE REALLY IS
EMPHASIZED THAT OUR INITIAL
STUDIES WERE CONDUCTED WITH THE
CD4 T-CELL, BECAUSE YOU CAN SEE
THE T-REGS ARE INVOF INDEED
SUPPRESSING THESE AUTOIMMUNE
PATHWAYS AND THE T-EFFECTOR
CELLS SEEM TO BE RIGHT IN THE
MITSD OF IT SO WHAT WE SAW WAS
THAT THE CD4 T-CELLS SEEM TO BE
THE HUB OF AUTOIMMUNE SIGNALING
SO OUR INITIAL SEQUENCING
EXPERIMENTS AND EVALLIA YOU
GUYSS WERE DONE ON THIS, I USED
TO GET THE QUESTION A LOT AND I
ON GO WHAT ABOUT B-CELLS AND
DENDRITIC CELLS, IT STARTED LIKE
THIS BECAUSE IT LOOKED LIKE THE
HUB WHERE MOST ACTIVITY WAS
HAPPENING SO LET'S GO BACK,
STARTING WITH THE RESISTANT
BACKGROUND, YOU NOTICE I SAID
HERE 52 WEEKS SO AFTER TREATMENT
WITH PRISTANE AND 1 TIME
INJECTION WITH PRISTANE, 52
WEEKS LATER I COLLECTED THE
SPLEEN AND ISOLATED OUT THE CD4
T-CELLS AND CONDUCTED BOTH
ATTACK SEQ ON THE CELLS FROM THE
SPLEEN 52 WEEKS AFTER TREATMENT,
AND SO HERE IS A POSITIVE
CONTROL, IT WAS THE FEMALE, THAT
WAS TREATED WITH PRISTANE AND
YOU CAN SEE THE ATTACK SEQ
SPLITS INTO 2 DISTINCT CLUSTERS
AND HERE ARE MALE CONTROL
GROUPS, SO THESE MALE CONTROL
GROUPS THIS IS THE WILD-TYPE
THAT WAS TREATED WITH PRISTANE
AND HERE'S A TRANSGENIC MOUSE
THAT WAS TREATED FOR STAIN BUT
NO DOXYPSYCHEY PSYCHE LINE, SO
THERE'S NO TRANSGENE IN THESE
HIGHER POPULATIONS AND SAME HERE
IS THE MALE, SO IN ALL THESE
MALE CONTROL MALE ANIMALS YOU
CAN SEE THE 2 DISTINCT CLUSTERS
HERE AND YOU CAN SEPARATE IT TO
NOW WHAT'S MORE OPEN IN THE
CLUSTER AND WHAT'S MORE OPEN IN
THE FEMALES, SO, WHERE DOES OUR
ACTUAL TEST SUBJECT FALL?
WELL WE'RE VERY EXCITED TO SEE
THAT AT THE EPIGENETIC PROFILE
LEVEL IN THESE T-CELLS, THESE
MICE THAT HAD BOTH THE
CHALLENGES PRISTANE TO INDUCE
THE DISEASE AND ALSO INDUCTION
OF THE TRANSGENIC XIST SEEM TO
FALL MORE INTO THE FEMALE
PROFILE, SO EVEN THOUGH AFTER 52
WEEKS, WE DEPARTMENT SEE ANY
PHYSICAL DISEASE
CHARACTERISTICS, WE DO SEE THAT
EXPRESSION OF XIST IMPOSES A
FEMALE LIKE EPIGENETIC PROFILE
IN THESE MALE CELLS THAT WE
ASSESS, WHAT IS IN CLUSTER 2
THAT WE TERM THE FEMALE, LIKE
THE ACCESS IBLGHT CLUSTER, WELL,
IN CLUSTER 2 ITSELF, WE SEE A
LOT OF TLRs AND MYD88 CASCADES
WHICH ARE VERY IMPORTANT IN
LUPUS AND LUPUS PATHOGENESIS,
AND NOW SHOWING THE RNA SEQ DATA
AND IT'S QUITE SIMILAR TO THIS
IS, IT'S LESS, IT'S LESS SORT OF
DISTINCT BUT THE TREND IS VERY
SIMILAR.
SO THIS GAVE US INCENTIVE TO
CONTINUE TO A DIFFERENT MODEL,
AND SO, THE AUTOMODEL WE WENT
WITH WAS SJLJ, THIS WAS JIM
LAMBERT, SO IN BOTH THIS MODEL
BOTH MALES AND FEMALES WILL
DEVELOP THE DISEASE BUT THE
FEMALES WILL DEVELOP MORE SEVERE
AND MUCH HIGHER LETHALITY OF
THIS PRISTANE INDUCED LUPUS.
AND SO THESE ARE OUR COHORTS
AGAIN, SO WE HAVE--THIS TIME WE
USE THE WILD-TYPE FEMALE AND THE
WILE TYPE MALE AND AGAIN WE HAVE
THESE TEST GROUPS AND SO OUR
TEST GROUP IS AGAIN, THE MALE
TRANSGENIC MOUSE WITH BOTH
INDUCTION OF THE TRANSYEEN AND
THE CHALLENGE WITH PRISTANE AND
HERE JUST TO SHARE THERE ARE MOW
DIFFERENCES WE SHARED THIS
TREATMENT IN THE MODEL WILD-TYPE
AS WELL, OR JUST TO CONTRAST THE
DIFFERENCES WE DID THE SAME
TREATMENT IN THE WILD-TYPE MODEL
AS WELL, AND YOU CAN SEE HERE
THAT THE FEMALES START DYING
AROUND 16 WEEKS AND THIS
WILD-TYPE SURVIVAL CURVE, SO WE
CHOSE THE TERMINAL POINT OF 16
WEEKS POST INDUCTION, POST
INJECTION, TO AVOID LOSING ANY
MICE BECAUSE WE WANT TO SEQUENCE
THESE MICE WORKING WITH DISEASE
TISSUE, WOULD BE--WOULD NOT BE
IDEAL.
SO FINALLY IN THIS PERMISSIVE
MODEL, WE'RE ABLE TO SEE A VERY
CLEAR OF ALL THE AUTOANTIBODIES
IN THE MALES WITH A TRANSGENIC
EXPRESSION AND ALSO CHALLENGE
WITH PRISTANE AND JUST 16 WEEKS
POST INDUCTION, SO AGAIN, WE HAD
TO WAIT 52 WEEKS IN THE BLACK 6
AND WE REALLY WEREN'T SEEING ANY
SIGNIFICANCE, WE DIDN'T SEE
TRENDS BUT WE DID SEE
SIGNIFICANCE, AND AGAIN THIS WAS
DONE WITH OUR COLLABORATOR
PJ UTZ, AND HE WAS GREAT IN
CONSULTING THE PANELS FOR US ON
THE PHOSPHORYLATED MODELING SIZE
AND THE RHEUMATOLOGY SIDE AND
HE'S ALSO THE 1 WHO POINTED OUT
BLACK 6 IS AUTOIMMUNE RESISTANT
SO VERY IMPORTANT TO HAVE GREAT
COLLABORATORS EMPLOY SO ON THE
PATHOLOGY END, AGAIN WITH BLACK
6 AT 52 WEEKS WE WEREN'T REALLY
SEEING ANY PATHOLOGY WE COULD
SAY DISTINCTLY WAS RELATED TO
DISEASE BUT HERE AT JUST 16
WEEKS WE CAN SEE THAT IN THE
KIDNEY OF THESE PRISTANE TREATED
MICE, WE CAN SEE THICKENING IN
THE DEPOSITS IN THE KIDNEY AND
ALSO DAMAGE TO LIVERS OF THESE
EXPRESSING MICE, SO HERE'S A
WILD-TYPE FEMALE, WILD-TYPE MALE
WITH THE TREATMENT CONTROL AND
THIS IS THE TRANSGENIC WITH THE
FULL TREATMENT AS WELL.
AND SO THIS TOTAL PATHOLOGY
SCORE IS ACTUALLY COMPOSITE
SCORE FOR THE SUM TOTAL OF
DAMAGE FROM THE KIDNEY, THE
LIVER, THE SPLEEN, ADIPOST, LUNG
AND LIVE LYMPHNODES AND SO THESE
ARE ALL IMPACTED IN PATIENT
LUPUS, AS WELL.
SO FROM HERE, I ALSO HARVEST THE
SPLEEN SAMPLES FOR SEQUENCING
AND THIS TIME, YOU KNOW HOWARD
WAS LIKE, YES, NOW YOU CAN DO
SINGLE CELL AND THIS IS ALSO
WHERE I ENCOUNTERED THE
LIMITATIONS OF HHMI FUNDING
BECAUSE I DID ASK HOWARD IF WE
COULD SEQUENCE LIKE ALL 60 MICE
WE DID AND SHE SAID NO, THAT IS
WAY TOO EXPENSIVE, YOU WILL JUST
HAVE TO PICK, A FEW FROM EACH
SIGNIFICANT EACH REPRESENTATIVE
GROUP AND GO FROM THERE IFLET
MULTIOME SEQUENCING, SO WHAT WE
SAW HERE WAS THE PATHOLOGY SCORE
WAS NEVER LOWER THAN 10, SO 10
WAS THE SCORE, SO WE TURN THIS
DEC SEVERE AND INTERESTING
ENOUGH WITH THE WILD-TYPE MALES
WE KNEW WE WOULD SEE SOME
DISEASE IN THE WILD-TYPE MALES
AND WHAT'S INTERESTING IS THAT
THE WILD-TYPE SCORE REACHES 10,
SO WE TERM IT FEMALE LIKE OR LOW
LIKE SO LOW SLEEZE AND SORT OF
HIGH DISEASE AND I ALSO POINTED
OUT THAT IT'S INTERESTING THAT
WE HAVE THESE BIFURCATION OF
THESE 2 IN OUR TREATMENT GROUP.
SO TO ASSESS THIS AGAIN, HOWARD,
YOU DIDN'T LET ME SEQUENCE EVERY
SINGLE MOUSE BUT WE DID TAKE A
FEW MICE FROM EACH OF THESE
HIGHLIGHTED GROUP, SO THAT WE
COULD SEE THE DIFFERENCES AND
WHAT'S GOING ON THERE AND SO I'M
JUST SHOWING THIS SLIDE, THIS IS
IMPORTANT FOR KIND OF STAGING OR
THE DISEASE PROGRESS, SO ON THE
PLAQUE 6 WHERE WE SAW CLEAR
DISTINCT PATTERNS DISTINGUISHING
EXIST EXPRESSING AND AND
NONEXIST EXPRESSING MICE, I
DIDN'T REALLY SEE THAT HERE SO
EACH WHEN I GROUP THE MALE LOW
SLEEZE TOGETHER FOR A HIGHER
POWER FOR THE COMPARISON
PURPOSES, BETWEEN THE WILD-TYPE
TO THE COMPARISONS I DIDN'T SEE
VERY MANY DIFFERENCES BETWEEN
THE SEXES, AND THIS FINALLY MADE
SENSE TO ME BECAUSE OF COURSE,
IT SEEMS LIKE THE EPIGENETIC
MODELS HAS ALREADY HAPPENED SO
WE REALLY HAD TO GO INTO THE
YEEN EXPRESSION LEVEL, SO
STARTING AT THE GENE EXPRESSION
LEVELS WHERE WE TART TO SEE
THOSE DIFFERENCES AND WE SEE
HERE ON THE MAP, PRETTY CLEAN,
CELL TYPE CLUSTERS SO THIS IS
JUST TO SHOW THAT HERE, WITH THE
TGM WITH THE HIGH DEC, EXISTS
EXPRESSING MALES WITH THE HIGH
DISEASE AND THE WILD-TYPE
FEMALES ARE SORT OF CLUSTERING
TOGETHER, SO YOU'RE SEEING THE
OVERLAY OF THE RED AND THE
GREEN.
SO AGAIN AS I MENTIONED WITH THE
SINGLE CELL MULTIDATA SET, WE
DIDN'T SEE ANY CHANGES BETWEEN
THE MISTINE CHALLENGE MOUSE
GROUPS AND THIS MAKES SENSE IN
TERMS OF TIME BECAUSE THESE MICE
IN CONTRAST TO THE BLACK 6 ARE
DISPLAYING PHYSIOLOGICAL SENSE
OF DISEASE, AND THE CHROMATIN
REMODELING HAS ALREADY HAPPENED
AND STABILIZED BUT THE SEVERITY
IS STILL INCAPSULATED IN THE
TRANSCRIPTOME, SO, LOCATING
WITHIN THESE POPULATIONS, SORT
OF THE--SO WE HAVE THESE B-CELL
GROUPS AND B-CELL GROUPS SO
LOOKING BROADLY INTO THESE
B-CELL GROUPS WE SEE MARKERS
THAT ARE AGAIN ASSOCIATE WIDE
LIKE AGE RELATED B-CELLS OR
ATYPICAL B-CELLS AS WE TERM THEM
SO YOU SEE THE ELEVATION OF ZEB2
AND ASSOCIATE WIDE DOUBLE B
CELLS AND WE ALSO SEE THE
DECREASE IN THE T-CELL
POPULATION OF THESE IMMUNO
MODULARY SIGNAL SO NRCC1 IS
INVOLVED IN THE T-CELL INFLAMM
EGG, CD54 IS A IMMUNE REGULATOR
AND CD37 IS A COMPLEMENT
MEDIATED INTERACTIVE T-CELLS, SO
ZOOMING INTO 1 OF THE CLUSTERS
WHERE WE SEE PRIMARILY JUST
OVERLAW OF THE RED AND THE
GREEN, SO THE TRANSGENIC MALE,
WITH THE HIGH DISEASE AND THE
FEMALE, WILD-TYPE POSITIVE
CONTROLS, WE SEE THAT THIS KIND
OF CORRELATES WITH B-CELL
CLUSTER 3, SO, IN B-CELL CLUSTER
3, THERE ARE VERY CLEARLY
B-CELLS BECAUSE THERE'S HIGH
CD19 SIGNATURES BUT WE ALSO SAW
THIS MASSIVE INCREASE IN THESE
OTHER 2 TRANSCRIPTS AND CONCUR
EPTLY WE SAW THE DOWN REGULATION
OF CR2 WHICH INCORRODES CD21 AND
IN A DOWN REGULATION OF CDR5,
BOTH OF WHICH ARE HALLMARKS OF
ATYPICAL B-CELLS IN LUPUS
PATIENTS.
SO CD21 NEGATIVE AND CD27 DOUBLE
NEGATIVE POPULATIONS ARE TYPICAL
OF THE SIGNATURES OF B-CELLS,
AND IN A HALLMARK OF ATYPICAL
B-CELLS IN THE PATHOGENESIS OF
THIS, AND THIS IS EXCITING FOR
US TO SEE THAT THERE IS
SOMETHING WE CAN FIND REFLECT
INDEED THE PATIENT POPULATION AS
WELL.
SO THAT KIND OF CONCLUDES THE
PART OF OUR MOUSE MODEL PART
WHERE WE SHOWED THAT IF WE KIND
OF INDUCE, EXIST RMPs IN A
MALE BACKGROUND, ANIMAL
BACKGROUND WE'RE ABLE TO PROMOTE
FEMALE BIASED AUTOIMMUNITY IN
THESE MALES.
SO PHYSIOLOGICALLY WE'RE ABLE TO
DETECT INCREASED AUTOANTIBODY
PRODUCTION AS WELL AS MORE
SEVERE DISEASE ORGANS AND
AUTOIMMUNITY.
AND SO, THIS KIND OF BRINGS US
TO THE NEXT PART, LIKE WHAT IS
THE CLINICAL IMPACT OF THE XIST
ARM PIECE?
ARE AUTOIMMUNE PATIENTS ACTUALLY
REACTIVE TO COMPONENTS WITHIN
THE XIST RMP?
SO THIS LEADS US TO THE NEXT
PART OF THE TALK, SO THIS KIND
OF A BASIC SLIDE, YOU KNOW JUST
REMINDING US OF THE IMPORTANCE
OF AUTOANTIBODIES AND
AUTOANTIGENS IN THE CONTEXT OF
AUTOIMMUNE DEC AND DIAGNOSIS,
AND THEY ARE PRODUCED AGAIN, AND
HOW THE CELL ANTIGENS DURING
CELL DAMAGE OR INFLAMMATION, CAN
YOU GET THE RELEASE OF CELL
ANTIIENS AND THEY COULD BE
CLEAVED BO THIS UNRECOGNIZED
ANTIGENS WHICH THEN THE
SELF-RECOGNIZES AS FOREIGN,
INCORRECTLY, OR THEY COULD LOOK
VERY SIMILAR TO AUTOANTIGENS
THAT ARE ALREADY--THEY COULD
CROSS REACT WITH AUTOEABT BODIES
THAT EXIST BETWEEN PATHOGENS
MEAN THEY LOOK VERY SIMILAR TO
PATHOGENS.
AND THEY ACT AS CLINICAL
BIOMARKERS FOR AUTOIMMUNE
DISEASE AND IT'S AGAIN,
AUTOIMMUNE DECS CAN BE DIFFICULT
TO DIAGNOSE BECAUSE OF THE
HETEROGENEITY AND OVERLAPPING
SYMPTOMS VERY EARLY ON SO IT'S
IMPORTANT TO FIND AS MANY
CLINICAL BIOMARKER AS WE CAN TO
HELP DORPH SPECIFIC AUTOANTIIEN
PROFILES IMPROVE DIAGNOSIS AND
TREATMENT.
AND SO, ON THE CLINICAL
RELEVANCE SIDE, HERE IS WHERE WE
COLLABORATE WITH THE HUMAN
PROTEIN ATLAS PROJECT IN SWEDEN
WHO HELPED US DESIGN AN
ASSOCIATED PROTEIN ANTIGEN
ARRAY, AND PROTEINS WE IDENTIFY
THROUGH THE TRIPMIS, PULL DOWN
ASSAY I MENTIONED EARLIER.
YOU KNOW INITIALLY WE DORPHED 81
IN THE EXPERIMENTS AND WE GOT
SERUM SAMPLES FROM MULTIPLE LABS
AT JOHNS HOPKINS AND STANFORD,
WITH 3 DIFFERENT DISEASES WITH
SCLERODERMA AND OTHERS AND WE
EXAMINED THEM WITH THE SERUM WE
GOT PRACTICES THE STANFORD
DONATION CENTER AND WE SAW THERE
WERE 9 THAT WERE SHARED BETWEEN
ALL 3 DISEASE GROUPS AND THE
REST SEEM TO BE KIND OF DISTINCT
TO EACH DISEASE.
AND AT THE TIME OF THE
PUBLICATION OF THE MANUSCRIPT OF
THE 79 AUTOANTIGENS WE KIND OF
IDENTIFY IN THIS ARRAY, 28 OF
THEM WERE NOT DESCRIBED IN THE
LITERATURE AS ANTIGEN, SO THIS
OPENED THE PROFILES FROM
INCREASED SPECIFICITY AND KIAG
NUMBER OF THETICS.
AND SO, THIS IS KIND OF THE
OVERVIEW OF THE MODEL OF HOW WE
PROPOSE XIST LINK RNA CAN BE
FUNCTIONING SORT OF A TRIGGER
FOR INFERIOR BIASED
AUTOIMMUNITY.
SO IN THE PHOSPHORYLATED MODEL
IN THE FIRST PART OF THE TALK,
THE INCREASE EXPRESSION PROMOTES
AUTOIMMUNITY IN THE MOUSE MODEL.
WE SAW THIS BOTH EPIGENETICALLY
AND ON THE GENE EXPRESSION LEVEL
THERE WAS INCREASE
IMPEDIMENTSUNE ACTIVATION AND
DECREASED IMMUNE MODULATION.
WE ALSO SAW THIS INCREASED
AUTOANTIBODY PRODUCTION, SO CAN
YOU KIND OF SEE THE STAGING OF
DEC, THE TLB CELL, CHROMATIN
REMODELING WILL OCCUR FIRST
BEFORE YOU HAVE ANY CHANGES IN
THE EXPRESSION.
AND THEN USUALLY THERE'S ANOTHER
CHECK POINT, NEVER PROGRESS TO
THE DISEASE BECAUSE THERE ARE
CHECK POINTS THERE, HOWEVER, IN
OUR MODEL, AS WE SEE ALSO, MORE
SEVERE ORGAN DISEASE PENETRANTS
AND SEVERITY ASSOCIATED WITH THE
EXPRESSION IN OUR MALE ANIMALS
AS WELL.
AND ON A CLINICAL END, WE SHOWED
THAT AUTOIMMUNE DEC PATIENTS ARE
ACTUALLY SERO-REACTIVE TO THE
XIST RMP, AND HOPEFULLY ALL THIS
RESEARCH WILL HELP US IMPROVE
THERAPIES AND DIAGNOSTICS AS WE
CONTINUE EXPLORING THIS ANGLE OF
HOW XIST, THE PARTICLE WORKS AND
INFLUENCES THE IMMUNE SYSTEM OR
INTERACTING WITH THE IMMUNE
SYSTEM TO PROMOTE THIS
BACKGROUND OF INCREASED
AUTOIMMUNITY IN FEMALES, SO, AT
THIS POINT, I WOULD LIKE TO
ACKNOWLEDGE THE MANY PEOPLE
WITHOUT WHOM THIS WORK WOULD NOT
HAVE BEEN POSSIBLE, SO EVERYONE
IN THE CHANG LAB AND ESPECIALLY
HOWARD.
SO AS YOU CAN PROBABLY TELL FROM
THE SLIDES, THERE WAS A LOT OF
MOUSE WORK INVOLVED, A LOT OF
CROSSING, A LOT OF GENETIC
STUDIES AND SEQUENCING, EVEN
THOUGH HOWARD DIDN'T LET ME
SEQUENCE ALL 60 MICE, IT WAS
STILL VERY EXPENSIVE TO DO ALL
THE SEQUENCING AND HE WAS VERY
PATIENT.
THIS WAS A VERY LONG PROGEC,
AND, YOU KNOW HE WAS A GREAT
CHEERLEADER, AND VERY GOOD
MENTOR.
AND HE PAID FOR EVERYTHING, SO
THANK YOU HOWARD.
AND ON OUR VETERINARIAN SIDE,
KERRY WAS THE 1 WHO DID A LOT OF
THE NECROPSY END POINT WORK FOR
US, AND READ THE HISTOLOGY, AND
OUR CLINICAL COLLABORATORS WHO
CONTRIBUTED TO THE SERUM TO THE
STUDY, AGAIN, A PGA, THE PGA IS
ACTUALLY THE CO MENTOR FOR THE
K99 AND HE PROVIDED A LOT OF THE
RHEUMATOLOGY EXPERTISE, AND HE
LET ME GO TO HIS LAB AND HE WILL
PRESENT AND GAVE ME GREAT
FEEDBACK, OUR ADMIN WITHOUT WHOM
NOTHING WOULD REALLY GET DONE
AND ALSO SCILIFE LAB IN SWEDEN
WHO DESIGNED THE ARRAY AND
ALLOWED US TO HAVE STOCK UPS TO
THE HUMAN AT PLACENTAS PROTEINS
WHICH IS A LOT AND ALSO OF
COURSE, THE FUNDING, THE PROJECT
WAS LARGELY FUNDED BY THE
SCLERODERMA RESEARCH FOUNDATION
BUT I WAS ACTUALLY PERSONALLY
FUNDED BY THE NIH, NIAMS, SO I
WAS ON A TRAINING GRANT WITH
THEM INITIALLY WHEN I START MY
POST DOC AND I GOT THE K99
R00 TRANSITION FROM THE SAME
INSTITUTE SO THANK YOU VERY MUCH
NIH AND NIAMS AND I WOULD LIKE
TO CONCLUDE ON THIS SLIDE WHICH
IS A SUMMARY OF THE PAPER THAT
I--THE WORK I JUST PRESENTED WAS
THE CELL PAPER WE JUST PRESENTED
IN FEBRUARY, AND AGAIN WE WANT
TO HIGHLIGHT THAT THE END GOAL
IS TO IMPROVE THERAPIES AND
DIAGNOSTICS, SO IF YOU HAVE
SERUM AND WOULD LIKE TO BE PART
OF THESE STUDIES PLEASE GET IN
TOUCH WITH ME, IT WOULD BE VERY
COOL IF WE COULD ALSO FIND LIKE
SOME CLIENTELE SERUM AND CHANGES
TO THE XIST EXPRESSION OR
INCREASE DOSAGE OF XIMPEDIMENTS
PACTS AUTOIMMUNITY AS WELL, AND
KIND OF A SELF-PLUG HERE.
IF YOU'RE INTERESTED, I THINK I
CAN SAY PRETTY CONFIDENTLY THAT
THE DOU LAB WILL PROBABLY BE
OPENING IN FALL 2024, KIND OF
NARROW DOWN THE LOCATION RIGHT
NOW SO, YEAH, KEEP YOUR EYES
PEELED AND THANK YOU FOR
LISTENING.
>> THANK YOU SO MUCH DR. DOU.
WE WOULD LIKE TO INTRODUCE THE
NEXT SPEAKER DR. ANDIOS,.
>> THANK YOU, THANK YOU FOR THE
INVITATION TO BE A PART OF THIS
EVENT AND THANK TO YOU YOUR TEAM
FOR ORCHESTRATING 1 OF THE MORE
COMPLEX VOOM INTERACTIONS THAT I
HAVE BEEN A PART OF AND I JUST
WANT TO CONFIRM HERE THAT I CAN
SHARE MY SCREEN.
ALL RIGHT, OKAY, SO THANK YOU
AGAIN, I'M BRENDAN, I'M WITH THE
RHEUMATOLOGY DIVISION AT JOHNS
HOPKINS AND MY GOAL TODAY IS TO
REVIEW FOR YOU THE FINDINGS OF A
MANUSCRIPT THAT OUR GROUP
PUBLISHED LAST FALL.
THIS PAPER MADE 2 CONCLUSIONS
THAT I'M GOING TO HIGHLIGHT
WHICH ARE 1, WE IDENTIFIED XIST
AS AN ENDOGENOUS SOURCE OF RNA
LIGANDS THAT ACTIVATE TLR7 WHICH
FRAMES EXIST AS A DAMP AND TO
OUR KNOWLEDGE IS REALLY THE ONLY
EXAMPLE OF A FEMALE SCIENTIFIC
DAMP IDENTIFIED TO DATE.
THE OTHER MAIN FINDING TO REPORT
HERE IS THAT XIST UPREGULATION
OF CELLS IN PETRESSABLE MALE
PATIENTS WITH LUPUS IN
COMPARISON TO HEALTHY FEMALE
CONTROLS.
SO, TO INTRODUCE THIS TOPIC AND
REALLY EXPLAIN WHY WE CHOSE TO
FOCUS ON TLR 7, I AM GOING TO
FIRST REVIEW SOME OF THE DATA
BEHIND TLR 7 AS BEING A KEY
CONTRIBUTOR TO THE PATHOLOGY
OBSERVED IN SLE, SO THIS MAY BE
A REVIEW FOR SOME OF OUR
AUDIENCE BUT IT'S IMPORTANT TO
REVIEW THIS KIND OF FRAME WHY
THIS QUESTION WAS POSED IN THE
FIRST PLACE.
OKAY, SO, THESE CONCEPTS REALLY
DON'T NEED TO BE INTRODUCED NOW,
WE'RE HERE BECAUSE OF THE FACT
THAT WE KNOW THAT FEMALES,
EXPERIENCE, AUTOIMMUNITY
DISPROPORTIONATELY COMPARED TO
MALES, I WILL POINT OUT I HAVE
THE SAME DATA AS MY 2
PREDECESSORS, BUT SHOWING
DIFFERENT REVIEW ARTICLES WHICH
IS SHOWING THAT THE BULK OF IT
LIES WITH FEMALE ANDS THAT
INCLUDES ORGAN SPECIFIC
AUTOIMMUNITY, BUT ALSO SYSTEMIC
DISEASES THAT WE SEE IN
RHEUMATOLOGY, SUCH AS LUPUS AND
CLER DERMA AND OTHERS, AS HAS
BEEN MENTIONED IN LUPUS AND THE
BEST KNOWN RISK FACTOR FOR
HAVING LUPUS IS SIMPLY BEING
FEMALE WITH 9 TO 10 MORE TIMES,
AND THIS INTERESTING DOSE
RESPONSE RELATED TO THE
X-CHROMOSOME, AND MEN WITH
[INDISCERNIBLE], VERSUS WOMEN
WITH TURNER SIN TROAM, AND @ THE
SAME TIME AS HAS BEEN MENTIONED
THERE HAVE BEEN DIFFERENCES IN
THE HOST IMMUNE RESPONSES
BETWEEN FEMALES AND MALES, IT'S
SOMETHING THAT'S NOT EMILY THED
TO HUMANS, IT'S AVAILABLE IN
OTHER SPECIES BUT WHAT'S KNOWN
IN WOMEN IS THAT THERE'S
ENHANCED TYPE 1 INTERFERON
SIGNALING AS WELL AS ENHANCED
ANTIBODY RESPONSES, SO
DIFFERENCES IN THE INTERFERON
SYSTEM AND THE B-CELL
COMPARTMENT, WHICH GIVE FEMALES
AN ADVANTAGE IN SOME SCENARIOS,
AND SO THE WITH KNOWLEDGE THAT
THESE IMMUNE RESPONSES DIFFER
BASED ON SEX, THIS RAISES A
QUESTION WHETHER THERE ARE
SPECIFIC IMMUNE PATHWAYS THAT
COULD GIVE FEMALES AN ADVANTAGE
IN HOST DEFENSE BUT WHEN
DISREGULATED OR EXACERBATED,
COULD CONTRIBUTE TO AUTOIMMUNITY
AND THESE SEX BIASED RHEUMATIC
DISEASES THAT WE'RE ALL FAMILIAR
WITH IN RHEUMATOLOGY.
SO TO APPROACH THIS QUESTION IN
LUPUS, WE CHOSE TO FOCUS ON TLR
7.
SO TOLL-LIKE RECEPTOR 7 IS AN
INNATE PATTERN RECOGNITION
RECEPTOR THAT IS FOUND IN IMMUNE
CELLS, PARTICULARLY B-CELLS,
ANDA ANTIGEN PRESENTING CELLS OR
PDCs, TLR7 IS A SENSOR OF RNA
AND RNA DEGRADATION PRODUCTS
DETEBT INDEED THE ENDOSOME, AND
TLR 7LICATION IN B-CELLS
ACTIVATES PROLIFERATION IN
ANTIBODY RESPONSES, TLR
REGULATION IN DENDRITIC CELLS
TRIGGERRINGS TYPE 1 INTERFERON
PRODUCTION AND WE KNOW BOTH OF
THESE FETTURES DISREGULATED IN
PATIENTS WITH LUPUS, WELL KNOWN
THESE PATIENTS PRODUCE AN A
RANGE OF AUTOANTIBODIES.
WE HEARD ABOUT THE ATYPICAL
B-CELLS THAT RESPONSIBLE FOR
THAT, WHICH LIE DOWN STREAM OF
TLR SIGNALING AND THERE IS THIS
LONG RECOGNIZED INTERFERON
SIGNATURE PRESENT IN PATIENTS
WITH LUPUS AND PLASMA
SIGNIFYATOID DENDRITIC CELLS ARE
A CONTRIBUTOR TO THE SIGNALING,
THE ONLY CELL TYPE OF INTEREST
IN THAT INTERFERON SIGNATURE IS
INTERESTING IN THAT IT'S
UBIQUITOUS, IT'S PRESENT IN THE
BLOOD AND TARGET ORELIMINATEDIGE
AS WELL, SO IT'S KNOWN IS THAT
TLR 7 CAN CONTRIBUTE TO BOTH
THESE FEATURES OF THE DISEASE
AND THAT'S BEEN SHOWN BOTH IN
MOUSE MODELS AS WELL MORE
RECENTLY IN POLYMORPHISMS
DESCRIBED IN HUMANS WITH SLE, SO
JUST TO QUICKLY REVIEW THESE
CONCEPTS AGAIN, THERE ARE MANY
MOUSE MODELS OF SLE AND SEVERAL
OF THEM ARE TLR DEPENDENT, THE
YAA MODEL LONG STANDING USED
MODEL OF LUPUS RESULTS IN THE
DUPLICATION OF TLR7 ON A PORTION
OF THE X-CHROMESOME CONTAINING
TISSUING LR7 ON TO THE Y-GIVING
THIS THE TLR 7 LINKED PHENOTYPE,
THE PRISTANE MODEL IS ANOTHER
EXAMPLE OF A MOUSE MODEL THAT
GENERATES AUTOANTIBODIES, TYPE 1
INTERFERON AND END ORGAN DAMAGE
THAT MIMICS THE HUMAN DEC,
STUDIES IN TLR 7 KNOCK OUT MOUSE
HAVE SHOWN THAT IT TLR 7 SIGNAL
SUGGEST KEY FOR INDUCING
PATHOLOGY SEEN IN THAT MODEL.
AND MORE RECENTLY, KIND OF A
SIMPLER MODEL OF LUPUS HAS BEEN
DERIVED BY SIMPLY DIRECTLY
ACTIVATING TLR7 IN WILD-TYPE
MICE USING THESE ANALOGUES AND
THESE MICE ALSO DEVELOP
ANTIBODIES AND AGER AN DAMAGE
THAT MIMICS THE HUMAN DISEASE
EMPLOY SO SEVERAL EXAMPLES OF
MICE OF ENDOPATHOLOGY AND
AUTOANTIBODY PROFILES THAT
RESEMBLE THE HUMAN ILLNESS THAT
ARE ALL LINKED TO TLR SIGNAL 7
SIGNALING.
THE SAME TIME MORE RECENTLY
THERE'S BEEN EVIDENCE GAIN
INDEED HUMANS THAT GAIN OF
FUNCTION MUTATIONS IN TON LR 7
CAN BE FOUND IN PATES WITH ADULT
ONSET, SLE, SO THIS GROUP 2
YEARS AGO PUBLISHED THIS ON
ACTIVITIES AND PROJECTS INITIAL
CASE OF YOUNG PATIENT WITH
CLINICAL FEATURES OF LUPUS THAT
IDENTIFIED THAT MUTATION IN TLR
7 AS BEING RESPONSIBLE FOR THE
PHENOTYPE, THEY WENT TO TO
IDENTIFY 2 UNRELATED PHENOTYPES
IN PATIENTS WHO HAD EARLY SEVERE
SLE, NOW WHAT THESE PATIENTS HAD
IN COMMON IS THAT THEY LOWERED
THE THRESHOLD FOR T LR
ACTIVATION IN THE CELL SYSTEM,
THEY WEPT ON TO PUT 1 OF THESE
IN THE MOUSE AND GENERATED
ANOTHER MOUSE MODEL OF SLE, THE
FEATURED AUTOANTIBODIES IN ORGAN
DAMAGE THAT MIMIC THE HUMAN
DISEASE.
I WILL POINT OUT THAT MORE
RECENTLY JUST WITHIN THE LAST 2
MONTHS, A GROUP IN GERMANY HAS
DESCRIBED ARK DITIONAL MUTATIONS
NOT IN TLR 7 ITSELF, AND IN A
YEEN ENCODING TLR7 TURNOVER IN
THE ENDOSOME, SO THOSE MUTATIONS
WERE IDENTIFIED ALSO IN PATIENTS
WITH EARLY ONSET SEVERE SLE,
WITH WITH THESE MUTATIONS DO IS
TO PROLONG TLR 7 SIGNALING IN
THE ENDOSOME BY BLOCKING
DEGRADATION AND TURNOVER IN THE
SENSOR COMPARTMENT.
SO OBSERVATIONS MADE BOTH IN
HUMANS WITH THE DISEASE AND IN
MICE, GOING TO TLR 7, THE OTHER
POINT I WANTED TO MAKE ON THIS
SLIDE IS THAT IN THE PAST 10-20
YEARS THERE ARE BEEN MANY
POLYMORPHISMS IDENTIFIED IN
GENES IN THE INNATE IMMUNE
SYSTEM, WHICH CAN BE ASSOCIATED
WITH STERILE INFLAMMATORY
SYNDROMES, NOT ALL OF THOSE
MUTATIONS GENERATE CLINICAL
PHENOTYPES THAT PERFECTLY MIMIC
OUR RHEUMATIC DISEASES, SO FOR
EXAMPLE, GAIN OF FUNCTIONS IN
STING, A KEY COMPONENT OF THE
CELL INTRINSIC RESPONSE, IS
ASSOCIATE WIDE A CLINICAL
SYNDROME THAT HAS FEATURES OF
OUR DISEASES BUT DOES NOT LOOK
LIKE WHAT WE THINK OF AS
RHEUMATOLOGISTS AS IDIO TAGHTIC
SLE AND WHAT IS REALLY
COMPELLING ABOUT THESE MUTATION
SYSTEM THAT THEY'RE IDENTIFIED
IN PATIENTS, WHO HAVE FEATURED
THAT LOOK LIKE THE CLASSICALLY
THOUGHT OF IDIO PATHIC DISEASE.
SO WHAT WE SET OUT TO DO HERE
WAS RECOGNIZING THE FACT THAT
TLR 7 IS HEAVILY IMPLICATED IN
LUPUS PATHOGENESIS, AND
RECOGNIZING THIS FACT THAT LUPUS
IS SO HIGHLY FEMALE BIASED,
LET'S POSE THE QUESTION OF
WHETHER THERE ARE ENDOGENOUS RNA
LIGANDS THAT ARE FEMALE ENRICHED
OR FEMALE SPECIFIC THAT COULD BE
RESPONSIBLE FOR ACTIVATION OF
THE SENSOR IN THIS DISEASE.
AND SO THIS QUESTION IS KIND OF
POSED WITHIN A LARGER FRAMEWORK,
WITHIN THE LAST FEW YEARS,
INTEREST IN UNDERSTANDING
WHETHER THERE ARE SPECIFIC
ENDOGENOUS NUCLEIC ASITS THAT
DRIF THE IMMUNE PATHOLOGY THAT
DRIVE WHAT WE SEE IN SLE AND
OTHER INTERFERON LINKED DECS,
NUCLEIC ACIDS ARE POTENT LIGANDS
FOR ACTIVATING VARIOUS
COMPONENTS OF THE INTERFERON
SYSTEM, SO FOR EXAMPLE,
MITOCHONDRIA DNETWORKA IS A
SOURCE OF DOUBLE STRANDED DNA
THAT CAN ACTIVATE CGAS,
CYTOSTOOL DNAIC SENSOR, THIS HAS
BEEN OF INTEREST IN THE SLE
TODAY IN RECENT YEARS, THERE IS
A GAIN OF FUNCTION MUTATION,
DESCRIBED IN MDA 5 WHICH IS A
CYTOSOLIC SENSOR OF DOUBLE
STRANDED RNA, AN EFFORT TO
UNDERSTAND WHAT YOU THIS CLAYIC
ACIDS ARE RESPONSIBLE FOR
ACTIVATING THIS GAIN OF 5, THIS
MDA GAIN OF 5 FUNCTION MUTANT IN
PATIENTS WHO HAVE THIS IN THE
SETTING OF GENETICALLY DRIVEN
SLE, LED TO IDENTISK OF ALU,
CELLULAR DUPLEXS AS A SOURCE
THAT MDA 5 LIGAND.
SO THESE DIEWP PLEXS ARE
CAPABILITY OF ACTIVATING THIS
GAIN OF FUNCTION MUTANT BUT NOT
CAPABILITY OF ACTIVATING THE
WILD-TYPE AND THIS MUTATION WAS
IDENTIFIED IN PATIENT WHO IS HAD
SLE.
A THIRD REFERENCE THAT I'LL
POINT OUT IS THIS PAPER FROM
SOME YEARS AGO, THOUGHT TO
IDENTIFY SPECIFIC RNAs TO
INTERACT WITH RO 60 WHICH IS THE
HALLMARK ANTIIENS THAT ARE
FEATURED IN RHEUMATIC DECS, AND
HERE AGAIN, IT WAS IDENTIFIED AS
A SOURCE OF RNA FOUND IN
COMPLEXES WITH ROW 60, AND THE
MODEL HERE IS THAT
AUTOANTIBODIES CAN CONVEY
TAKEN--THEY ALU RNA TO ASPECT
GEN PRESENTING CELLS TO THE
IMMUNE COMPLEX.
SO EXAMPLES OF EFFORTS TO
IDENTIFY SPECIFIC NUCLEIC ACIDS
THAT CAN ACTIVATE THE INTERFERON
SYSTEM, BUT HERE WE WANT TO
APPLY THIS FILTER OF SEX BIAS,
AND THERE'S AN OPPORTUNITY TO DO
THAT, WITH TLR 7 BECAUSE IN
CONTRAST TO THESE OTHER SENSORS
I JUST MENTIONED, TLR 7 SHOWS
SOME SEQUENCE SPECIFICITY IN ITS
MODE OF ACTIVATION, SO, TLR 7 AS
2 BINDING SITES, 1 THAT IS BOUND
BY GUANA SIGN AND A SECOND SPIKE
IN THE SINGLE RNA FRAGMENTS AND
1 COMPONENT THAT'S BEEN IDENTIFY
INDEED GENERATING A POTENT
LIGAND TO ACTIVATE TLR7 AT THIS
SITE IS THE PRESENCE OF
UU SPECIFICITY AND THESE ARE IN
CONTRAST TO THE 2 OTHER SENSORS
I JUST MENTIONED, CGAS WHICH
RECK NICES THE SHAPE OF DOUBLE
STRAND THE DNA IN A INDEPENDENT
FASHION AND NDMA 5 THAT IS
DOUBLE STRANDED ON THE WAY TO
THE MOLECULE AND NOT THE
NUCLEOTIDE SEQUENCE, SO
UU RICHNESS IS A FACTOR WE CHOSE
TO FOCUS ON, WHEN LOOKING FOR
POTENTIAL ENDOGENOUS TLR 7
LIGANDS, THERE IS ANOTHER
EXAMPLE TO INDICATE THAT THE
DINUCLEOTIDES ALONE ARE NOT THE
SOLE DETERMINANT OF AN RNA'S
ABILITY TO ACTIVATE RNA7 BUT
THAT THE SURROUNDING MOTIFS MAY
INFLUENCE THAT AS WELL, SO THIS
A PUBLICATION PUBLISHED SEVERAL
YEARS AGO NOW FROM A GROUP THAT
WAS USING SiRNA TO KNOCK DOWN
GENE TARGETS IN THE ANTIGEN
PRESENTING CELLS AND FOUND
UNEXPECTEDLY THAT THESE COULD
TRIGGER INTERFERON PRODUCTION IN
PLASMA SIGNIFYATOID DENDRITIC
CELLS AND THEY MAPPED THEM OUT
AND IDENTIFIED SEQUENCES
INCLUDING THE 9 BASE SEQUENCE
WHICH THEY REFERRED TO AS
SiRNA 9.2 AND I WILL REFER TO
THIS AS WE MOVE AHEAD IN OUR DAT
AND THIS IS 2 FEATURES WE CHOSE
TO UTILIZE IN LOOKING FOR
POTENTIAL ENDOGENOUS TLR 7
LIGANDS.
SO THE STARTING POINT HERE WAS
GENE EXPRESSION DATA FROM THE
G-TEXT DATABASE, THIS IS DATA
DERIVE FRIDAY HEALTHY CONTROLS
AND WHAT WE DID WAS TO FIRST
LOOK FOR RNA MOLECULES THAT WERE
RICH IN THESE UU DISCIPLINARY
YOU DINUCLEOTIDE
REPEATS, WITH THE EXPRESSION
BIAS SO WE SEE ON THE PLATIS
THAT TRANSCRIPT ITS THE RIGHT
ARE FEMALE SPECIFIC AND EXIST,
HERE AN OUTLIER IN BEING
ENTIRELY FEMALE SPECIFIC AND
CONTAINING A LARGE NUMBER OF
THESE UU REPEATS.
WE CONSIDER THE FACT THAT THE
LINK OF AN RNA WOULD INFLUENCE
ITS ABILITY TO CONTAIN THESE
UU-DINUCLEOTIDES AND THAT
PERHAPS SMALLER RNAs THAT
CONTAIN A HIGH DENSITY OF THOSE
REPEATS MIGHT BE MORE POTENT
THAN A REALLY LONG RNA WITH
JUDGE UOF THE A FEW SCATTERED
OVER A GREAT REGION AND SO WE
CALCULATED MAXIMUM UU RICHNESS
WHICH WAS THE MAXIMUM NUMBER OF
THESE MOTIFS PRESCRIBINGENT
WITHIN A 500 BASE REEJ OF ANY
RNA TRANSCRIPT AND AGAIN WHEN
COMPARED TO FEMALE VERSUS MALE
EXPRESSION, EXIST IS NOTABLE IN
BEING FEMALE SPECIFIC AND
CONTAINING A LOT OF THESE OR A
HIGH DEGREE OF UU RICHNESS.
WE THEN LOOK FOR TRANSCRIPTS
THAT CONTAIN THE 9 BASE SEQUENCE
THAT HAD BEEN PREVIOUSLY
IDENTIFIED AS A TLR7 AGONIST AND
WE IDENTIFY ROUGHLY 300 RNAs
IN THE DATA SET THAT CONTAIN THE
MOTIF AND ONLY 1 OF THEM WAS
FEMALE SPECIFIC AND THAT WAS
XIST, NOW I DON'T NEED TO REVIEW
WHAT XIST WE'VE BEEN TALKING
ABOUT IT FOR OVER AN HOUR BUT I
WILL POINT OUT YES THIS, IS THE
SAME XIST MOLECULE THAT'S BEEN
IMPLICATE INDEED AUTOIMMUNITY
FOR THE 2 REASONS WE JUST HEARD
ABOUT AND WHEN WE WERE IN THE
EARLY STAGES OF THIS WORK, WE
WERE SURPRISED TO SEE THIS
COMING UP IN THE SCREEN BECAUSE
THERE WERE ALREADY REASONS TO
IMPLICATE EXIST IN THE TWEPMENT
OF AUTOIMMUNITY.
SPECIFICALLY,
DR. [INDISCERNIBLE]'S WORK
SHOWING THAT BIOALLELIC
EXPRESSION COULD BE DRIIVE BY
LOSS OF SILENCING ENACTIVATE X
WHICH COULD CONTRIBUTE TO
UPREGULATION OF THE THE SENSOR
SO IN THIS MODEL WHERE TLR 7
COULD BE UPREGULATED IN PATIENTS
WITH AUTOIMMUNITY.
WE FOUND EXIST COMING TO THE POP
OF THIS LIST OF POTENTIAL
CANDIDATES OF BEING THE LIGAND
FOR THAT VERY SAME SENSOR.
AS WAS MENTIONED, BY DR. DOU,
DR. CHANG'S LAB HAD PUBLISHED A
DESCRIPTION OF PROTEIN
INTERACTORS WITH SEVERAL
AUTOANTIGENS ON THAT LIST, SO
THERE ARE REASON ALREADY TO
EXIST EXIST AS A CONTRIBUTOR TO
AUTOIMMUNITY BUT HERE FINDING IT
WHEN APPROACHING THIS QUESTION
FROM A DIFFERENT ANGLE.
SO NEXT WE CONSIDERED THE
OVERALL EXPRESSION LEVELS THAT
WOULD BE A SIGNIFICANT FACTOR IN
GIVING AN RNA MOLECULE
OPPORTUNITY TO KROWBT TO TLR 7
SIGNALING SO THESE ARE PLOTS
LOOKING JUST IN FEMALES IN THE
SAME PERIPHERAL BLOOD SATTA SET,
WE'RE LOOKING AT THE COMBINATION
LEVELS VERSUS THE PRESENCE OF
THESE UU REPEATS, AND FIND AGAIN
THAT WHEN ANALYZING THE DAILY
BASIS AT THIS WAY, EXIST
REMAINSA THE TOP OF THE LIST OF
CANDIDATE TLR 7 LIGANDS SO
HIGHLY EXPRESSED AND CONTAINS
THESE MOTIFS OF INTEREST.
SO THEN FINALLY WE DEVELOP WHAT
WE TERMED A TLR7 LIGATION SCORE
WHERE WE TAKE INTO ACCOUNT THESE
FEATURES, TOTAL FEMALE BIAS, THE
MAXIMUM UU RICHNESS AND
EXPRESSION LEVELS AND LOOK NOT
ONLY IN BLOOD BUT OF OTHER
TISSUES OF AND ALL TISSUES
PRESENT IN THE DATABASE AND YOU
CAN SEE HERE IN LOOKING ACROSS
ALL TISSUES REMAINS AN OUTLIER,
IT HAS THESE FEATURES THAT COULD
MAKE IT A FEMALE SPECIFIC TLR 7
LIGAND AND VERY UBIQUITOUSLY
EXPRESSED.
SO WE THEN WANTED TO FIGURE OUT
WHAT PORTIONS OF THE MOLECULE WE
COULD FOCUS ON, SO THIS IS A
PLOT OF THE UU RICHNESS ACROSS
THE MOLECULE, AND WHAT YOU CAN
SEE HERE IS THE DOTTED LINE THIS
IS THE AVERAGE UU RICHNESS OF
ALL TRANSCRIPTS PRESENT IN THE
DATABASE, SO KIND OF AN AVERAGE
UU RICHNESS, AND EXIST IS
REMARKABLE AND BASICALLY THE
ENTIRE MOLECULE LIES ABOVE THAT
LINE, THERE ARE 2 PARTS THAT
REALLY STAND OUT, THIS A-REPEAT
REAMINGION WHICH IS PARTICULARLY
DENSE WITH THE UU MOTIFS AND A
MORE DISTAL COMPONENT IN THE
VICINITY OF THE REPEAT.
HERE ON XIST 1 IS A FRAGMENT WE
CHOSE TO LOOK AT BASED ON PRIOR
IDENTIFICATION OF THIS ON A
PRIOR LIGAND, THAT WAS ON TLR 1,
AND WE TERM THIS XIST 1.1 AND WE
DO THESE INVITRO TO LOOK AT
THESE 2 PORTIONS OF THE MOLECULE
TO PROVE THESE AREAS OF EXIST
ARE CAPABLE OF ACTIVATING TLR 7.
SO WE STARTED WITH THIS
FRAGMENT, WE ISOLATED 20 BASE
SECTION SURROUNDING THIS 9 MER,
USE THE CONTROL POSITIVE 2 WHICH
HAD BEEN BREFIOUSLY PUBLISHED AS
WELL AS ANTISENSE AND POLY-A AS
WELL AS NEGATIVE CONTROLS.
PUTTING THESE IN HEALTHY DONOR
PLASMA SIGHT DENDRITIC CELLS, WE
SHOWED THIS XIST WAS ABLE TO
ACTIVATE INTERFERON EXPRESSION
BY USING PDCs AND WE USE THE
SAME FRAGMENTS IN A REPORTER
CELL LINE AND YOU CAN SEE IN
THIS DOSE RESPONSE EXPERIMENT,
IS THAT THE EXIST FREG
MENTORSHIP SKILL AND THIS
POSITIVE CONTROL BEHAVE
SIMILARLY.
THEN WE WANTED TO CONFIRM, A
DIRECT INTERACTION INVITRO WAS
RESPONSIBLE FOR THESE FINDINGS
IN CELLS SO WE USE FLUORESCENCE
WHICH WOULD BE BASED LIGAND
BINDING ASSAY THAT HOE THAT
EXIST IN THIS POSITIVE CONTROL
OLIGIO-INTERACT DIRECTLY WITH
THE FASHION IN SIMILAR AFFINITYS
SO INDEED THIS PART OF EXIST,
APPEARS CAPABLE OF BINDING TO
AND ACTIVATING TLR 7 IN PRIMARY
CELLS AND A REPORTER CELL
SYSTEM.
SO THEN WE LOOKED AT THE
A-REPEAT, WE ISOLATED A ROUGHLY
800 BASE FRAGMENT, FROM THAT
REGION OF EXIST, WHICH WAS THE
MOST UU DENSE PORTION OF THE
MOLECULE.
USED A CONTROL RNA AND AGAIN
WENT BACK TO OUR REPORTER CELLS
AND SHOWED THAT WE COULD
ACTIVATE TLR 7 IN THIS REPORTER
CELL SYSTEM, PUT THE A-REPEAT
INTO PRIMARY PDCs AND SHOWED
WE COULD GENERATE A PRIMARY
INTERFERON RESPONSE AND BY USING
HYDROXY CHLOR O QUINAND
INHIBITORS SHOWED THIS EFFECT
WAS INDEED DUE TO TLR 7
ACTIVATION.
SO THE A-REPEAT, IN ADDITION TO
THIS FRAGMENT ALSO APPEARS
CAPABLE OF ACTIVATING TLR 7 BOTH
IN THE REPORTER CELL SYSTEM AND
PRIMARY CELLS.
WE THEN WANTED TO KNOW WHAT WAS
THE RELATIVE CONTRIBUTION OF
EACH OF THESE COMPONENTS IN
TERMS OF POTENCY IN BEING A TLR7
LIGAND OR SOURCE OF TLR7.
SO WE EXAMINED THIS USING PICO
MOLAR EXAMINATIONS AND YOU COULD
SEE AS YOU WOULD EXPECT THAT THE
A-REPEAT IS FULL OF THESE
DINUCLEOTIDE MOTIFS IS A MORE
POTENT AGONIST, AND IN MORE
CAPABLE OF GENERATING AN
INTERFERON RESPONSE OF PDCs IN
A LOWER CONCENTRATION, SO THE
EVIDENCE THAT THE UU-ENRICHMENT
IS MORE IMPORTANT THAN THE 9-MER
SEQUENCE, SOAZ THESE WERE
SEQUENCES DONE INVITRO, WHERE WE
HAD ISOLATED PORTIONS OF THE
XIST MOLECULE ANDY WOO WANT TO
PUT IT IN CONTEXT AND ASK THE
QUESTION OF WHERE DOES XIST SIT
IN THE ENTIRE TRANSCRIPTOME OF A
FEMALE CELL IN TERMS OF THE
RELATIVE CONTRIBUTION TO TLR 7
ACTIVATION, SO TO DO THAT, WE
USED A FEMALE DERIVED
IMMORTALIZED CELL LINE WHICH IS
EXIST POSITIVE A431 CELL LINE
AND USE CRSPR EDITING TO DELETE
EXIST, WE FIRST GENERATED SOME
POPULATIONS OF CELLS THAT WERE
EITHER WILD-TYPE OR EXIST
NULLLIN VARYING DEGREES AND
ISOLATE FRIDAY THAT A CLONAL
POPULATION OF COMPLETE KNOCK OUT
CELLS, SO GENERATE--WE USE THESE
CELLS AS A SOURCE OF WHOLE
CELLULAR RNA WHICH WE THEN
ISOLATED AND FRAGMENTED INVITRO
DOWN TO THE LENGTH OF SEVERAL
DOZEN BASE PAIRS TO KIND OF
SIMULATE THE ENVIRONMENT OF THE
ENDOSOME WHERE TLR 7 WILL BE
EBCOUNTERRING RNA DEGRADATION
PRODUCTS AND WHAT YOU CAN SEE IN
THE TLR7 REPORTER CELL LINE HERE
IS THAT IN COMPARISON TO THE
WILD-TYPE AND THE CRSPER
CONTROL, THERE IS A DOSE EFFECT
BASED ON THE DEGREE TO WHICH
XIST HAS BEEN REMOVED FROM THE
TRANSCRIPTOME OF THESE CELLS,
AND THEN COMPARING RNA FROM THE
WILD-TYPE CELLS VERSUS COMPLETE
KNOCK OUT, WHAT WE SAW ON THE
REPORTER CELLS WAS GREATER THAN
50% REDUCTION IN THE ABILITY OF
THIS FRAGMENTED WHOLE CELLULAR
RNA TO ACTIVATE TLR 7 WHICH WAS
SURPRISING IN ITS MAGNITUDE,
REMOVING 1 RNA CAN CONTRIBUTE SO
MUCH TO THE ABILITY OF WHOLE
CELLULAR RNA TO ACTIVATE THE
SENSOR.
SO WE WANTED TO CONFIRM, THAT
THROUGH CRSPER EDITING WE HADN'T
KNOCKED DOWN ANY OTHER RNAs
WHICH MIGHT BE IMPORTANT SOURCES
OF TLR 7 LIGANDS SO WE USED RNA
SEQ TO LOOK AT CHANGES IN OUR
KNOCK OUT POOL VERSUS THE WILE
TYPE, WE IDENTIFIED TRANSCRIPTS
THAT WERE SIPPING 95 CANTILY
DOWN REGULATED AND THEN LOOKED
AT L UU CONTENT VERSUS
EXPRESSION LEVELS IF IN THE
WILD-TYPE CELLS TO TRY TO
DETERMINE WHETHER WE KNOCKED OUT
ANYTHING ELSE THAT MIGHT BE AN
IMPORTANT SOURCE OF THESE
UU FEATURES.
AND YOU CAN SEE IT IN FACT XIST
APPEARS TO BE THE MOST IMPORTANT
POTENTIAL SOURCE OF TLR7 LIGANDS
THAT WE'VE REMOVED BASED ON WHAT
WE'RE LOOKING AT HERE, BASED ON
THE UU-REPEATS, IT CONTAINS THE
NUMBER AS WELL, BUT YOU CAN SEE
IT'S EXPRESSED AT A MUCH LOWER
LEVEL THAN THE WILD-TYPE CELLS.
SO IN FACT IF LOOKED LIKE WE HAD
LOOKED LIKE WE REMOVED WHAT
SEEMED TO BE THE MOST IMPORTANT
SOURCE OF LIGAND SOUL CELLS, SO
HAVING SEEN THIS UNEXPECTED
RESULT, IT EXIST ITSELF LOOKS
LIKE IT CAN BE SUCH AN IMPORTANT
RESERVOIR FOR THESE TLR 7
ACTIVATING MOTIFS, WE THEN ASK
THE QUESTION OF WHAT EXIST
EXPRESSION LEVELS LOOK LIKE IN
LUPUS SUBJECTS, THE TLR7 DRIVEN
DEC EMPLOY SO TO DO THIS WE USED
AN RNAFLOW 7 ASSAY AND COMPARED
THESE FROM THE CENTER HERE AT
YAWNS HOPKINS VERSUS HEALTHY
CONTROL FEMALE DONORS, AND WHAT
YOU WILL SEE WHAT YOU CAN SEE IS
THAT IN PBMCs WE FOUND HIGHER
EXPRESSION LEVELS OF XIST IN THE
LUPUS COHORT COMPARED TO
CONTROLS.
AND THIS DIFFERENCE WAS MOST
EXAGGERATED IN THE B-CELL
COMPARTMENT, NOW, THOSE OF YOU
WHO ARE JUST LISTENING TO WHAT
THE DOCTOR TOLD US AT THE ONSET
HERE ABOUT WHAT HAPPENS WHEN YOU
DELETE XIST FROM A B-CELL, YOU
MAY BE SCRATCHING YOUR HEADS AND
SAYING HOW DOES THIS ALL ADD UP,
AND I'M NOT SURE I HAVE ALL THE
ANSWERS FOR YOU BUT HERE LOOKING
AT EXPRESSION LEVELS WE WERE NOT
O'CLOCKING AT XIST LOCALIZATION
EMPLOY I THINK IT WOULD BE
INTERESTING TO BE ABLE TO DO
THAT IT WILL BE NICE TO HAVE
THIS INITIAL ANALYSIS BUT IN
THINKING ABOUT THE XIST TO DO
LIST, I THINK THAT RECONCILING
EXPRESSION LEVELS VERSUS EXIST
LOCALIZATION IS SOMETHING THAT
COULD HELP US PUT THIS INTO
CONTEXT AND UNDERSTAND WHAT IT
MEANS.
IN T-CELLS AND MONOCYTES THERE
WAS A TREND TOWARDS HIGHER
EXPRESSION AS WELL BUT THAT
DIFFERENCE WAS MOST DRAMATIC
REDUCTIONTATTIC ON THE B-CELL
COMPARTMENT, USING THIS COHORT,
WE WERE USING THIS RELATION
BETWEEN EXPRESSION AND DISEASE
ACTIVITY AND THERE ARE 2
COMMONLY USED CLINICAL INDICES
FOR LUPUS ACTIVITY WE HAD ACCESS
TO AS A PART OF OUR COHORT WHICH
ARE PART OF THE THE PHYSICIAN'S
ASSESSMENT AND THE LEAD ON YOU
CAN SEE THERE'S A TREND TOWARDS
HIGHER XIST EXPRESSION AMONG
PBMCs AMONG THE EXPRESSIONS
WITH ACTIVE DEC CORRELATED POSE
TO THOSE WHO ARE CLINICALLY
QUESTIONNAIRES XET THERE'S A
POSITIVE CORRELATION BETWEEN
XIST EXPRESSION AND SLEDAI SEEN
IN THE COHORT.
SO THE FINDINGS HERE ARE THAT
EXIST LOOK TO BE UPREGULATED IN
THE PERIPHERAL BLOOD OF PATIENTS
AS COMPARED TO CONTROLS AND
AGAIN THERE'S A POSITIVE DISEASE
RELATIONSHIP ACTIVITY WITH THE
SINGLE POINT TIME ANALYSIS.
SO WE WANTED TO APPROACH THIS
USING ANOTHER DATA SET, AND ALSO
LOOK AT THE RELATIONSHIP BETWEEN
XIST EXPRESSION LEVELS AND
INTERFERON AND SO TO DO THAT, WE
USE THE AMP DATA SET WHICH IS A
MULTISITE STUDY WHICH COLLECTED
RENAL BIOPSY SAMPLES FROM
PATIENTS WITH LUPUS NEPHRITIS
AND PERFORMED SINGLE CELL
ANALYSIS ON IMMUNE CELLS
INFILTRATING THOSE RENAL
BIOPSIES AND 1 OF THE FINDINGS
OF THAT STUDY WAS THAT THERE IS
THIS POSITIVE RELATIONSHIP
BETWEEN INTERFERON EXPRESSION IN
THE TARGET ORGAN HERE BEING THE
KIDNEY AS WELL AS PERIPHERAL
BLOOD.
SO IN AN ANALYSIS OF THIS DATA
SET WE FOUND A STRONG POSITIVE
CORRELATION BETWEEN XIST AND
INTERFERON EXPRESSION AND AGAIN
IN THESE IMMUNE CELLS THAT IN
THIS ANALYSIS ARE IN THE TARGET
OREGON AFFECTED BY THE DISEASE
AS OPPOSE TO THE PERIPHERAL
BLOOD.
SO THAT'S AN INTRIGUING FINDING
AND 1 OF THE POTENTIAL
EXPLANATIONS WHICH WILL BE
PRETTY STRAIGHT FORWARD WILL BE
IF IT EXISTS ITSELF OR SIMPLY AN
INTERFERON INDUCED YEEN AND THAT
HADN'T BEEN PREVIOUSLY FORWARDED
BUT WE WANTED TO CONFIRM THIS
RELATIONSHIP WASN'T DUE TO THE
FACT THAT XIST ITSELF WAS
INTERFERON EXISTABLE AND WE TOOK
CELL LINES THAT EXPRESS EXIST,
PRIMARY PBMCs AND PRIMARY
KERATINOCYTES AND TREATED THEM
WITH INTERFERONS INVITRO AND
CONFIRMED WE'RE GENERATING
UPREGULATION OF INTERFERON
STIMULATED GENES IN THESE CELLS
AND SAW THAT THERE WAS NO CHANGE
IN EXIST LEVELS.
--XIST LEVELS, SOMETHING I WILL
NOTE THAT WASN'T INCLUDED HERE
THAT WE'VE DONE SENSE IS TO
STIMULATE THESE SAME XIST
EXPRESSING CELLS WITH OTHER
LIGANDS THAT WILL BE UPSTREAM OF
INTERFERONS SO FOR EXAMPLE,
USING POLYAIC OR ACTIVATING CELL
INTRINSIC ACTIVATING AND WE
HAVEN'T SEEN ANY RESPONSE IN
EXIST EITHER GOING UP OR DOWN,
IN RESPONSE TO THOSE STIMULI.
SO WE CONCLUDED FROM THIS, IS
THAT XIST ITSELF IS NOT
INFLUENCED BY INTERFERON, AND
THAT IN EXPLAINING THIS
RELATIONSHIP, POSITIVE
RELATIONSHIP BETWEEN INTERFERON
AND XIST EXPRESSION WE SAW IN
RENAL BIOPSIES, XIST IS MORE
LIKELY A DRIVER OF THAT THAN A
DOWN STREAM CONSEQUENCE OF THAT.
SO THAT BRINGS US TO OUR MODEL
WHICH AGAIN CENTERS ON THIS
FINDING THAT XIST IS AN
ENDOGENOUS SOURCE OF RNAs THAT
ARE CAPABLE OF ACTIVATING TLR7.
AS I MENTIONED, I THINK THIS IS,
YOU KNOW AS FAR AS WE KNOW THE
ONLY EXAMPLE OF A SEX SPECIFIC
DA AMP THAT'S BEEN IDENTIFIED TO
THIS IS SOMETHING THAT CAN
CERTAINLY CONTRIBUTE TO FEMALE
ADVANTAGE IN HOST DEFENSE
THROUGH SIGNALING THROUGH TLR 7
BUT IN LUPUS WHICH IS A DISEASE
CHARACTERIZED BY IMPAIRED
CLEARANCE OF APOPTOTIC DEBRIS AS
WELL AS AUTOANTIBODIES TARGETING
BINDING PROTEINS AND THE MODEL
IS THAT XIST WILL STIMULATE TLR
7 AND IMMUNE CELLS DOWN STREAM
OF THAT CRITICAL SENSOR, IN A
WAY THAT CAN CONTRIBUTE TO THE
PATHOGENIC IMMUNE RESPONSES THAT
ARE CHARACTERISTIC OF THIS
DISEASE EMPLOY --DISEASE.
I WILL POINT OUT SINCE I'M THIRD
WE'VE HEARD ABOUT 3 DIFFERENT
MECHANISMS OR FINDING INCLUDING
XIST ROLE IN GETTING
X-CHROMOSOME AND ACTIVATION, THE
INTERACTION WITH ANTIGEN
PROTEINS AND HERE THE IDENTITY
AS A TLR7 LIGAND, I THINK IT'S
EASY TO SEE HOW THESE FIT
TOGETHER AND CONTRIBUTE IN
MULTIPLE WAYS TO DWTMENT OF
AUTOIMMUNITY AND SLE AS WELL AS
OTHER FEMALE BIASED DISEASES.
THIS IS OUR GROUP, THIS PROJECT
WAS INITIALLY CONCEIVED OF BY
ERICA [INDISCERNIBLE], AND SHE
INVITED ME TO COLLABORATE ON
WHEN IT WAS IN ITS EARLY STAGES
AND JONATHAN CRAWFORD WORKED ON
THIS AS PART OF HIS Ph.D. ON
IMMUNOLOGY IN THE DIVISION,
WE'VE HAD A NUMBER OF
COLLABORATORS INCLUDING THE
LUPUS CENTER AND APPRECIATED
FUNDING FROM THE GREEN
FOUNDATION, NIDCR AND NIAMS.
SO THANK YOU AND I WILL LEAVE IT
THERE.
NOKAY, SO I WILL OPEN THE FLOOR
TO QUESTIONS AND I WANT TO
REMIND YOU IF YOU HAVE QUESTIONS
YOU WOULD LIKE ASKED DURING THIS
PORTION OF THE SESSION, FEEL
FREE TO PUT THEM INTO THE Q&A
AND WE WILL BE MONITORING THAT,
BUT FIRST, I AM GOING TO HAND
THE MICROPHONE OVER TO
DR. MANCINI TO GET US STARTED.
>> HI, THANK YOU ALL FOR THE
WONDERFUL TALKS SO WE HAVE A
LITTLE BIT OF A PLANTED QUESTION
WHICH I THINK WILL BE
INTERESTING AND GIVE YOU ALL AN
OPPORTUNITY TO TALK ABOUT THE
FUTURE, SO THE QUESTION IS HOW
DO YOU SEE XIST RESEARCH
EVOLVING OVER THE NEXT 3-5
YEARS?
SO CAN I ASK EACH OF THE
PANELISTS TO GIVE US THEIR POINT
OF VIEW ON THAT.
>> YOU WANT ME TO GO FIRST?
>> SURE.
>> OKAY.
YEAH, SO I LIKE THIS QUESTION
AND I SEE IT REALLY--WE'RE
STARTING TO HIT THE LOG PHASE OF
GROWTH FOR RESEARCH AND
INVOLVING XIST AND I THINK IT
WILL CONTINUE TO SKY ROCKET OVER
THE NEXT YEAR BECAUSE YOU CAN
THINK OF IT FROM THE PERSPECTIVE
OF AUTOIMMUNE DISEASE, SO TODAY
WE TALKED ABOUT SOME AUTOIMMUNE
DISEASES SO I HIGHLIGHTED LUPUS,
BUT THERE'S 25 AUTOIMMUNE DECS,
IS XIST GOING TO FUNCTION THE
SAME WAY IN ACROSS ALL 25 OF
THESE, ALL OF THESE ARE VERY,
VERY DISTINCT SO SO WE WILL SEE
A LOT OF DIVERSITY WITH THESE
MECHANISMS AND IT'S GOING TO
TAKE CREATIVE SCIENTISTS THIS
BRING TOGETHER, YOU KNOW REALLY
DIVERSE BACKGROUNDS AND
SCIENTIFIC TRAINING AND,A
PROACHING TO PROBLEM SOLVING
THAT ARE REALLY GOING TO BE KEY
TO FIGURING OUT HOW THE PUZZLE
PIECES ALL FIT ON A DISEASE BY
DISEASE BASIS.
>> THANKS.
DR. DOU, WOULD YOU LIKE TO ADD
ON?
>> YEAH, HIMARIE, SHE'S MY
PROGRAM OFFICER, SO YEAH, NO, I
AGREE, I THINK THIS IS A GREAT
CHANCE, THE COMMUNITY IS
GROWING, BUT I'M ALSO EXCITED
ABOUT THE IMPLICATIONS FOR THE
COMMUNITY UNITING BIOCHEMISTRY
AND EITHER HEALTH SCIENCES AND
BIOLOGY, BECAUSE [INDISCERNIBLE]
IS CORRECT IN SAYING THERE ARE
25 OTHER AUTOIMMUNE FEMALE
BIASED AUTOIMMUNE DECS BUT HOW
MANY LINK RNAs ARE ASSOCIATE
WIDE AUTOIMMUNE DISEASE THAT WE
DON'T KNOW THE FUNCTIONS OF YET
AS WE JUST SEE HIGH ASSOCIATIONS
IN THE DATA SETS SO THIS OPENS
THE AVENUE OF ALSO LIKE LOOKING
AT OTHER LINK RNAs AND HOW
THEY POTENTIALLY MIGHT HAVE A
FUNCTION IN AUTOIMMUNITY MAYBE
AS MOTEAR SCAFFOLDS, NOT JUST AS
BIOMARKERS WE DON'T KNOW THE
FUNCTION OF, BUT YEAH, I THINK
THIS IS A VERY EXCITING FIELD TO
BE IN AND I ALSO THINK THIS IS
THE PERFECT OPPORTUNITY TO GO
AND EVALUATE THE DATA SETS ON GO
BUT AND THERE ARE SO MANY
PATIENT STUDIES BUT WHAT HAS
BEEN DONE IN THE PAST IS THAT
THE X-CHROMESOME HAS BEEN
FILTERED OUT IN DISEASES THAT WE
KNOW HAVE A VERY STRONG SEX
COMPONENT WHETHER IT'S MALE OR
FEMALE, THERE'S SEX CHROMOSOMES
TEND TO BE FILTERED OUT SO WE
DON'T REALLY KNOW VERY MUCH
ABOUT THAT SO WE THINK THIS
PRESENTS A HUGE OPPORTUNITY TO
GO BACK AND COMPUTATIONALLY
EVALUATE THE DATA WE ALREADY
HAVE.
>> THANK YOU.
DR. ANTIOCHOS?
>> YEAH, SURE, I WILL ECHO THE
SENT I WANTS THAT YEAH, THERE
ARE ALL THESE FEMALE BIASED
DISEASES THAT WE DEAL WITH IN
RHEUMATOLOGY THAT SHARE CLINICAL
FEATURES, MAYBE ARE NOT
IDENTICAL IN THEIR PATHOGENESIS
BUT I THINK THERE'S AN
OPPORTUNITY TO TRY TO IDENTIFY
SUBSETS OF PATIENTS ACROSS THESE
DISEASES, WHERE EXIST IS PLAYING
AN IMPORTANT ROLE AND SO
ANALYZING THESE VARIOUS FACAS
NATIONAL LIBRARY OF MEDICINE NA,
X-INACTIVATION AND XIST PATTERNS
AUTOANTIBODIES XIST RNPs, XIST
EXPRESSION LEVELS, THESE THINGS
THAT BE APPLIED TO THE DISEASES,
LUPUS, ET CETERA TO IAT IDENTIY
PATIENTS THAT HAVE THE MECHANISM
RESPONSIBLE FOR DRIVING THE
DISEASE AND OF COURSE NEXT STEP
WOULD BE FIGURING OUT THERAPIES
THAT WILL BE MOST EFFECTIVE FOR
PATIENTS WITH DECS AND THAT'S
WHERE IT'S EXCITING TO SEE THAT
THERE ARE TLR 7 INHIBITORS THAT
ARE IN WILL EARLY STAGES OF
DEVELOPMENT SO THE QUESTION OF
WHETHER ANY OF THESE MARKERS WE
DISCUSSED TODAY COULD IDENTIFY
PATIENTS WHO WILL BE SUITABLE
FOR THAT OR OTHER THERAPIES THAT
INTERRUPT THE SIGNALING THAT MAY
BE RESPONSIBLE FOR DISEASE
PATHOLOGY.
I GUESS I WILL ALSO POINT OUT AT
THE SAME TIME THERE ARE THESE
EXCITING FIND XTION THERE'S A
LOT OF BASIC WORK LEFT TO
UNDERSTAND AND EXACTLY HOW ALL
OF THE VARIOUS THINGS IT CAN DO
BEYOND THE STAGES OF DEVELOPMENT
AND AT THE SAME TIME THERE'S
THIS OPPORTUNITY TO KIND OF MOVE
IN A MORE TRANSLATIONAL
DIRECTION, I THINK THAT THERE'S
STILL A NEED TO UNDERSTAND THE
NUT ANDS BOLTS OF WHAT XIST IS
DOG IN THESE QUEPMENTAL
DEPARTMENTS DURING THIS
DEVELOPMENTAL PHASE.
>> OKAY, I THINK GOING ON TO THE
NEXT QUESTION, I THINK IT MAY BE
A LITTLE REDUNDANT BECAUSE SOME
OF YOU ADDRESSED IT, PARTIALLY
IS WHAT HAPPENS FIRST WITH
ADVANCES OF SCIENCE, SO I GUESS
ANOTHER QUESTION I HAVE FOR YOU
IS WHAT ARE THE RESEARCH GAPS
THAT YOU THINK REALLY NEED TO BE
FILLED, YOU KNOW TO
ACTUALLY--EXCUSE ME, ACCELERATE
THE PROGRESS THAT YOU JUST
MENTIONED.
>> SO, I GUESS I WILL TART WITH
THAT, I THINK THERE'S A COUPLE
OF THINGS, FIRST, JUST AMOUNT OF
MONITOR THAT'S IS DEDICATED TO
RESEARCH THAT CONCERNS
AUTOIMMUNE DISEASE IS NOT THE
SAME AS CANCER FOR EXAMPLE, AND
THERE'S A LOT OF PATIENTS THAT
HAVE AUTOIMMUNE DECS.
I THINK IT'S ABOUT 5% OF THE
WORLD'S POPULATION HAS AN
AUTOIMMUNE DISORDER, AND SO, I
THINK--I THINK WE NEED A LOT
MORE FUNDING AND SORT OF
THIS--THE OTHER ASPECT OF THIS,
IS MY LAB DOES BASIC SCIENCE
RESEARCH INTO FUNDAMENTAL MECH
NIMS, AND I REALLY STRONGLY
BELIEVE THAT WE CAN LEARN SO
MUCH ABOUT UNDERSTANDING HOW THE
POLICIES TAKE PLACE IN THE
HEALTHY STATE, RIGHT AND THEN
HOW--USE THAT INFORMATION THEN
TO UNDERSTAND HOW THE PROCESSES
GO WRONG IN DIFFERENT DEC
CONTEXTS, RIGHT?
BUT HAVING THAT MECHANISTIC
UNDERSTANDING IS ABSOLUTELY
CRITICAL TO THIS, AND SO, I
THINK--I THINK IT'S REALLY
IMPORTANT TO HAVE A GOOD BALANCE
OF THE BASIC SCIENCE RESEARCH
ALONG WITH THE TRANSLATIONAL
COMPONENT FOR INVESTIGATING
THIS.
>> DRS. DOU AND ANTIOCHOS, DO
YOU HAVE ANYTHING TO ADD?
>> I KNOW MONTSERRAT IS PUT IT
REALLY ELOQUENTLY, AND GOING OFF
THE SECOND THING SHE SAID,
COLLABORATION AND COMMUNE KAIG
WILL BE KEY BECAUSE WE SEE
THINGS FROM A VERY BASIC SCIENCE
PERSPECTIVE BUT BRINGING IN
CLINICIANS IS IMPORTANT.
AND I THINK AS BASIC SCIENTISTS
WE HAVE A SPECIALTY BECAUSE IT'S
REALLY HARD TO EACH KEEP UP WITH
LIKE 1 SPECIALTY, GENOMICS OR IF
YOU LIKE T-CELL OR B-CELL OR
SOMETHING BUT I THINK THIS IS A
REALLY--NEED IS ALMOST THESE
HUBS THAT WE CAN BRING MULTIPLE
SPECIALTIES TOGETHER, MULTIPLE
SCIENTISTS TOGETHER, WE CAN
BRING BIOCHEMISTS IN, WE CAN
BRING PEOPLE IN WHO STUDY THE
STRUCTURE OF THE BIOPHYSICIST
PEOPLE PEOPLE WHO CAN REALLY
SOLVE THE STRUCTURE, FIGURE OUT
THE COMPONENTS OF EXIST AND IF
THERE ARE VARIATIONS IN
STRUCTURE THAT CAN IMPACT
FUNCTION AS WELL.
WE SEE THAT IN PATIENTS AS WELL
AND GENOMICS AND PEOPLE CAN
GENERATE THESE DATA SETS AND
ALSO LIKE PATTIALLY CONNECT THEM
AS WELL IN THE ORGAN AND
CLINICIANS ARE SO IMPORTANT
BECAUSE THEY CAN SEE THE ENTIRE
PATIENT, THE ENTIRE ORGANISM
WHERE SOMETIMES, YOU KNOW FOR
ME, I HAD THIS, I WAS LIKE,
OKAY, WE WILL LOOK AT THE WHOLE
SPLEEN BUT THEN WE WOULD LOVE TO
ASSESS THE KIDNEY AS WELL, WE
WOULD LOVE TO ASSESS LUNGS ALL
THESE ORGANS, SO I THINK THIS
LIKE WHOLE BODY ALMOST LIKE
WHOLE SCIENTIFIC COMMUNITY, NOT
JUST RESTRICTED TO BIOLOGY, BUT
EXPANDING OUT TO PHYSICS,
CHEMISTRY AND MATH, COMPUTER
SCIENCES, I THINK THAT WILL BE
REALLY KEY IN HELPING US
ADVANCE.
ANYTHING ELSE BEFORE I MOVE TO
THE NEXT QUESTION?
>> OKAY, I'M GOING TO PULL OUT A
QUESTION FROM THE CHAT.
WE HAVE LOTS AND LOTS OF
QUESTIONS IN THE CHAT AND THE Q,
ANDA AND WE'RE GOING TO TRY TO
PULL A FEW OF THEM SO TO GET YOU
STARTED, AND I GUESS WE WILL
STICK WITH THE SAME ORDER IN
TERMS OF RESPONSES SO WE TALKED
A LOT TODAY ABOUT LUPUS AND A
COUPLE FOLLOW UP QUESTIONS ON
THAT AND THEN A QUESTION ABOUT
THE MICE.
SO IN LUPUS, IS IT THOUGHT THAT
THERE'S A DEFECT IN OW THE
X-CHROMESOME IS BEING
ENACTIVATED INVOLVING XIST, IS
THERE A TRIGGER ASK WHAT MIGHT
WE KNOW ABOUT THE TRIGGER?
AND THEN YOU TALKED A LOT,
CERTAINLY DR. ANG UERA, TALKED
ABOUT THE B-CELLS WHAT'S
HAPPENING IN OTHER CELLS OF THE
BODY, SPECIFIC TO LUPUS AND
THIRD PART OF THAT QUESTION, DO
THE MICE GET ANYTHING ELSE?
WE KNOW IN OTHER AUTOIMMUNE DECS
THERE ARE A LOT OF CO OCCURRING
AUTOIMMUNE DISEASES, DO THE MICE
ONLY HAVE LUPUS, DO THEY GET ANY
OTHER AUTOIMMUNITY?
>> YEAH, GOOD QUESTIONS.
YEAH, SO THE FIRST QUESTION, SO
WHAT ARE OUR WORK HAS FOUND IS
THAT IN LUPUS PATIENTS THERE ARE
PROBLEMS WITH X-INACTIVATION IN
BOTH B-CELLS AND T-CELLS.
SO WHAT WE SEE IS THAT IF YOU
TAKE A PERIPHERAL BLOOD SAMPLES
FROM A PATIENT AND YOU TAKE--YOU
ISOLATE T-CELLS AND YOU ISOLATE
B-CELLS AND YOU INVITRO ACTIVATE
THESE CELLS, EXIST RNA DOES NOT
GO BACK TO THE INACTIVE
X-CHROMESOME AND FORM THIS
FLUFFY CLOUD, LIKE IT SHOULD,
AND INSTEAD WHAT WE SEE IS THAT
THERE'S PINPOINTS OF XIST
CLUSTERS THAT ARE ALL OVER THE
NUCLEUS, SO INSTEAD OF A FLUFFY
CLOUD, THERE'S PINPOINTS ALL
OVER THE PLACE AND THEN THE
SECOND PART OF THE QUESTION IS
ENACTIVATED X-CHROMESOME
DIFFERENT IN B-CELLS AND OTHER
PARTS OF THE BODY.
SO YES, IN IMMUNE SES IT TURNS
OUT THAT X-ACTIVATION IS
MAINTAIN INDEED A VERY DIFFERENT
WAY AND THERE'S A DIVERSITY OF
WAYS THAT THE INACTIVE X IS
MAINTAINED.
SO FOR EXAMPLE, IN B-CELLS AND
T-CELLS IN THE RESTING ESTATE,
THESE ARE CELLS THAT ARE
QUIESCENT, THEY DO NOT HAVE XIST
ERROR NA LOCALIZE AT THE ACTIVE
X, THEY DO FOCUS ON THE HAVE
MODISKS, CYTOLOGICALLY ENRICH.
WHEN THESE CELLS ARE INVITRO
ACTIVATED IN A DISH, XIST RNA
COMES BACK TO THE XCHROME SOM AS
DO THESE EPIGENETIC MODISHS TO
THE POINT WHERE WE CAN SEE IT
USING MICROSCOPY.
SO IF YOU LOOK AT A NUTRIFILL
FOR EXAMPLE, NUTRIFILLS EXPRESS
FROM FEMALES, MOUSE AND HUMAN
THEY EXPRESS RNETWORKA AND YOU
DO RNA FISH FOR EXIST, THEY ARE
NONAPOPTOTIC THE ON THE INACTIVE
CHROMESOME AND NUTRIFILLS, SAME
THING WITH PLASMA SIGNIFYATOID
DENDRITIC CELLS, SO THIS IS
EXPRESSED BUT NOT LOCALIZED ON
THE INACTIVE X-CHROMESOME, SO
WHY ARE THESE CELLS DOING IT THE
LIKE THIS.
WE DON'T KNOW BUT THERE'S A
DIVERSITY OF FILTRATION THAT
EXIST FOR XIST AND INACTIVATION,
AND IT HIGHLIGHTS HOW MUCH WE
DON'T UNDERSTAND IN THIS PROCESS
IN IMMUNE CELLS WHICH MAKE ITS
REALLY EXCITING FOR SCIENTISTS
WHO ARE INTERESTED IN MECHANISM,
TO FIGURE OUT HOW DOES IT WORK,
WHY IS IT WORKING LIKE THIS?
THEN THE THIRD PART OF THE
QUESTION, REMIND ME AGAIN?
>> IT'S A QUESTION REALLY FOR
YOU AND MAYBE DR. DOU, AND THEN
WE'LL ASK DR. ANTIOCHOS, BECAUSE
HE'S MORE INTO HUMANS THAN
MOUSE.
NYEAH, GREAT QUESTION.
SO IN OUR XIST KNOCKOUT MICE BE,
WE LOOKEDDA THE AUTOANTIBODY
PROFILES AND WE WERE REALLY
SURPRISED BECAUSE WE DID NOT
EXIST TO SEE THAT THERE WAS
GOING TO BE ELEVATED
AUTOANTIBODIES DECIDES DSDNA AND
ANTISMITH.
SO THE MDA 51S WERE LIKE WHOA,
HOLD ON, WHAT'S HAPPENING?
SO BECAUSE OF THAT WE'RE NOW
LOOKING AT OTHER TISSUES IN THE
ANIMALS TO SEE IF THERE IS
EVIDENT OF PATHOLOGY THAT WOULD
BE SIMILAR TO OTHER AUTOIMMUNE
PATIENTS.
SO IN THE ORIGINAL LIKE COHORT,
BECAUSE AUTORANDOM RIGHT AND THE
PENETRANTS IS LIKE 15-18% OF
ANIMALS BEYOND THE AIMS OF 12
MONTHS WE ONLY LOOKEDDA THE
KIDNEYS IN THOSE ORIGINAL 3
MICE, AND SO WE JUST HAVE TO GET
LUCKY AND BE PATIENT AND THIS IS
WHY MY MD-Ph.D. STUDENT
CLAUDIA IS SO INEDIBLE -LY
AMAZING BECAUSE SHE SHOWING GRIT
AND PERSISTENCE TO FOLLOW THE
COHORT OF ANIMALS OVER TIME AND
MAKE SURE NONE OF THEM DIE ON
US, RIGHT, LIKE WE'RE GOING TO
BE CHECKING THEM EVERY COUPLE OF
WEEKS.
SO THAT WE CAN GET THEM AND THEN
DO THE EXTENSIVE PATHOLOGY TO
SEE WHAT ELSE MAY MIGHT BE GOOD
MODELS FOR.
>> YEAH, SO FOR US, WE ACTUALLY
TOOK THE MICE, I THINK IF YOU
RECALL, I SAID 16 WEEKS WE
WOULDN'T LOSE ANY FEMALES, WHAT
I DIDN'T SAY IS IF YOU LOOK AT
THE SAME CHART, THE MALES THAT
ARE WILD-TYPE DON'T DROP OFF
UNTIL AROUND 23 WEEKS SO FOR
THEM IT WAS KIND OF EARLY SO I
THINK WITH THE FEMALES WITH THE
SERUM, WE USE PJs SORT OF
LUPUS AUTOANTIGEN ARRAY BUT
THERE WERE SOME OVERLAPPED WITH
I THINK MYOCYTEIS AND MAYBE SOME
SCLERODERMA BUT AGAIN SOME OF
THE AUTOANTIBODIES TEND TO BE
SHARED ACROSS DISEASES AS WELL,
SO WE--I'M NOT A CLINICIAN, I'M
NOT A RHEUMATOLOGIST, WE HAVE
GREAT COLLABORATORS SO WE JUST
ASH ASSESSESSED FOR YOU KNOW
SIGNS OF LUPUS BUT I DON'T THINK
WE CAN RULE OUT THAT AND MAYBE
WE HAVE CHARACTERISTICS FROM
OTHER STEEZS AS WELL BECAUSE
IMEAN, SOME OF THESE SYMPTOMS IN
PATIENTS AS WELL, ARE HIGHLY
OVERLAPPING AND VERY ETICSER O
GENIUS AND AS FOR AS FEMALES
THERE WAS A RANGE OF SEVERITY OF
WHAT THEY WERE GETTING WITH THE
FEMALES AND WITH THE GROUPS IN
THE SAMPLE COHORT THAT WE WERE
TESTING ON AS WELL.
SO, I THINK IT'S A VERY
INTERESTING QUESTION AND
DEFINITELY WORTH EXPLORING.
>> OKAY, I WILL GIVE YOU A CHAND
TO ANSWER AS WELL BRENDAN.
>> YOU'RE MUTED.
>> SORRY WITH A QUESTION OF
WHETHER THERE ARE ANY SPECIFIC
FINDINGS, YOU KNOW WITHIN
SUBSETS OF HUMANS WHO SHOW
DIFFERING LEVELS OF EXISTING
EXPRESSION, WE HAVEN'T HAD
ACCESS TO MEASURE EXIST IN A
LARGE ENOUGH GROUP OF PATIENTS
TO REALLY KNOW IF THERE'S A
SPECIFIC PHENOTYPE, THAT'S ON
THE VERY LONG TO DO LIST, I
THINK OF THINGS THAT WOULD BE
USEFUL TO DO TO DETERMINE YOU
KNOW WHAT EXIST EXPRESSION
LEVELS LOOK LIKE IN LARGER
COHORTS OF PATIENTS,
LOCKITUDEINAL SAMPLES, IS THIS
SOMETHING THAT CHANGES OVERTIME
OR IS IT STATIC, IN THOSE VERY
VALUABLE PRECLINICAL COHORTS, IS
XIST SOMETHING THAT'S THERE
BEFORE HAND OR COMES AFTER
DISEASE DEVELOPED AND THERE'S A
LOT OF QUESTIONS THAT WOULD BE,
YOU KNOW INTERESTING TO RUN DOWN
WHICH ARE SIMILAR TO THE
QUESTIONS THAT HAVE BEEN PHRASED
WITH RESPECT TO THE INTERFERON
SIGNATURE ITSELF.
SO, MAYBE IF WE COME ACROSS SOME
OF THOSE FUNDS THAT DR. ANG
UERA WAS MENTIONING TO DO ALL
THIS WORK, THAT'S SOME OF THE
SORTS OF THINGS WE CAN COULD
ANSWER IN LARGER STUDIES.
>> AND I'M GOING TO PULL ANOTHER
QUESTION OUT FROM THE Q&A AS
WELL, AND IT'S REALLY SPEAKING
TO THIS DISCIPLINARY MORPHISM OF
RESPONSES.
AND SO, WE KNOW THAT WHILE THERE
ARE SOME MALE PATIENT WHO IS
HAVE AN ADDITIONAL X-CHROMOSOME
WHO GET LUPUS, WE KNOW THERE ARE
MALE PATIENTS WHO DON'T HAVE AN
ADDITIONAL MALE X-CHROMOSOME WHO
GET LUPUS WHAT MIGHT BE GOING ON
IN THE B-CELLS THERE?
ARE THERE OTHER X-LINKED IMMUNE
RELATED GENES BEING SEEN
UPREGULATED.
DO YOU THINK THOSE ARE
TRANSLOCATIONS OF GENERATED
THETIC MATERIALS, DO YOU HAVE
ANY THOUGHTS ABOUT HOW THAT
COULD BE STUDIED?
>> YEAH, SO WITH MALE PATIENTS
LIKE SAY ABOUT 15% OF LUPUS
PATIENTS ARE MALE IT'S REALLY
HARD TO RECRUIT THEM IN OUR
STUDIES, LIKE FEMALE PATIENTS,
ARE THE PREDOMINANT 1S, RIGHT
AND SO, WITH RESPECT TO LUPUS,
MALE AND FEMALES, HAVE PRETTY
SIMILAR DISEASE ACTIVITY, SO, WE
CAN'T REALLY SAY THAT LIKE, IT'S
DIFFERENT TO SAY FOR EXAMPLE,
SCLERODERMA WHERE THERE ARE MALE
PATIENTS PRESENT WITH MUCH MORE
DISEEEVER DISEASE AND THEN YOU
GD SAY, IS THERE SOMETHING
PROTECTIVE ON THE Y OR IN LUPUS
THAT IN THE CONTEXT OF
SCLERODERMA AGGRAVATES DISEASE
MORE, MAYBE?
PERHAPS WHAT THAT IS, WE'RE NOT
SURE, IS IT--HOW IS IT
MANIFESTING THIS, RIGHT IN IS
THROUGH PRODECLARATION OF
CYTOKINES?
IS IT THROUGH MEMBERS OF IMUBE
CELL POPULATIONS LIKE PROTECTED
1S, ADAPTIVE VERSUS INNATE, WE
REAL REALLY DON'T KNOW AND THEN
TO KIND OF GET BACK AT THE
QUESTION WITH LUPUS PATIENTS IN
MALES, REMEMBER THE SLIDE THAT
SOME--I THINK MAYBE ALL OF US
SHOWED THAT DOSAGE OF
X-CHROMESOMES IS LINKED TO
INCREASED RISK FOR DEVELOPING
THESE DISEASES SUCH AS LUPUS,
AND SO AN INDIVIDUAL MIGHT HAVE
TO COPIES OF AN X-CHROMESOME BUT
NEVER KNOW THAT, RIGHT?
SO AN INDIVIDUAL MIGHT BE X-Y
BUT MIGHT BE EXTINGUISHING
XY AND HAVE LUPUS LIKE DEC BUT
MAY NOT HAVE BEEN EVERY KERIO
TYPED TO PRESENT WITH THAT SO WE
HAVEN'T SEEN ANY--LIKE WE HAVE
RECRUITED ANY MALES IN OUR STUDY
WE WOULD WANT TO CHECK FOR
ABERRATIONS WITH X-INACTIVATION
MAINTENANCE BECAUSE IF THEY ARE,
AND XY THEY WOULD PREDICT THEY
HAVE MISLOCALLIZED EXIST, THEY
WOULD HAVE CHANGES IN EXCELLENT
GENE EXPRESSION, AND THEN AGAIN
IN THAT CONTEXT, IS THERE
SOMETHING PROTECTIVE ON THE
YRK-CHROMESOME IN THOSE
INDIVIDUALS, WE JUST DON'T KNOW,
WE DON'T KNOW YET.
>> GOING OFF LIKE WHY MALES,
SOME MALES MAY BE XY MAY
DEVELOP, I THINK THERE'S A
STRONG GENETIC CORRELATION
BECAUSE NOT ALL FEMALES WILL
DEVELOP AUTOIMMUNITY EITHER AND
WE CAN SEE AS I MENTIONED, WE
HAD AN AUTORESISTANT STREAM OF
BLACK 6 AND THEN WE HAD THE
AUTOIMMUNE PERMISSIVE AND WE
GOT VERY DIFFERENT IMMUNOLOGICAL
RESULTS AND THEY'RE THERE, SO
STRONGLY IN 6, THEY'RE NOT
PROGRESSING BUT IN THE DISEASES
BUT IN THE SDL, YOU SEE THAT THE
WILD-TYPE MALES ARE PROGRESSING
SO IT'S A CONTRIBUTION OF
FACTORS, I DON'T THINK-I DON'T
THINK THAT XIST IS THE ONLY 1
THAT IS INVOLVED, I THINK IT'S A
VERY STRONG COMPONENT AS WE'VE
SHOWN IN OUR TALKS TODAY BUT
ALSO GOING OFF THE SET, WAABOUT
THE OTHER LINK RNAs THAT ARE
OUT THERE THAT ARE HIGHLY
ASSOCIATED, MAYBE THEY HAVE SOME
KIND OF ROLE THERE BECAUSE I
THINK I SAW IN THE CHAT GROUP,
THERE WAS A QUESTION ABOUT
WHETHER WE SAW DIFFERENCES IN
THE REACTIVITY IN THE
MONOARRAYS, AND LIKE MONTSERRAT,
SAID, WE DIDN'T HAVE ENOUGH
MALES IN THE SERUM, SO I WAS
SAYING DO YOU HAVE ANY SERUM, SO
IF WE INCREASE THE SAMPLE SIZE,
I DO RECALL SEEING SOME OF THE
MALES HAVING GREATER THAN A BASE
LINE REACTIVITY, SEROREACTIVITY
TO THE SOME OF THE PROTEINS ON
THE XIST RNP, AND WHAT'S
INTERESTING ABOUT THE XIST RNP
ARE THESE PROTEINS ARE IN OVER
ABUNDANCE COMPARED TO XIST AND
THEY HAD BIOLOGICAL PROCESSES,
THEY ARE THERE FOR A REASON,
THEY'RE EXPRESSED FOR A REASON
AND PART OF IT IS TO BIND WITH
XIST, BUT NOT THE ONLY THING SO
THESE PROTEINS ALSO MIGHT BE IN
OTHER COMPLEXES OR HAVE OTHER
MECHANISMs AS WELL THAT
THEY'RE INVOLVED IN SEPARATE
FROM XIST SO IT COULD BE THAT
THIS SEROACTIVITY THAT WE'RE
IDENTIFYING COULD BE FROM, YOU
KNOW, THEY MIGHT BE IN ANOTHER
LINK RNA-RNP COMPLEX OR INVOLVED
IN A DIFFERENT WAY AS WELL.
SO I THINK GENETICS AND HORMONES
AND ALL OF THESE THINGS PLAY
VERY SIGNIFICANT ROLES.
NAND DR. ANT IOCH OS, I WILL LET
YOU HAVE THE LAST WORD AND THEN
I'M TOLD WE NEED TO WRAP IT UP.
>> OH, WELL, OKAY, I WILL ECHO
FACT THAT THEY'RE HETEROGENEOUS
ORY GENIUS AND IN LUPUS IN
PARTICULAR, THERE'S SUCH
VARIABILITY IN THE PATTERNS OF
INVOLVE AM, SEROLONLIC FEATURES,
ET CETERA AND SO, THE STUDY OF,
YOU KNOW MEN WITH THAT DISEASE,
YES, IMETTING SIGNIFICANT
NUMBERS ARE A PROBLEM, WE'VE
BEEN INTERESTED IN MEASURING
EXISTING LEVELS IN MEN AS WELL
BUT THERE JUST AREN'T MANY OF
THEM OUT THERE IN THESE COHORTS.
IN TERMS OF APPLYING FILTERS
THAT CAN MAKE IT EASIER TO
INTERPRET ALL THIS, JUST TO ECHO
WHAT DR. DOU JUST MENTIONED,
THERE ARE LOTS OF PATHWAYS THAT
CONTRIBUTE TO THESE DISEASES,
YOU KNOW IT'S UNLIKELY OF
COURSE, THAT EXIST IS THE MAIN
DRIVER IN MOST PATIENTS WITH
THESE CONDITIONS BUT AT THE SAME
TIME, YOU KNOW, THE FACT THAT
ANYONE VIEWING THIS TODAY WHO IS
TOTALLY UNFAMILIAR WITH XIST,
WOULD SAY WOW IT'S PRETTY
AMAZING THIS THIS 1 RNA CAN BE
IMPLICATED IN SO MANY AS
CONTRIBUTING TO THESE DISEASES,
SO I GUESS THAT I WILL EMPHASIZE
THE IMPORTANCE PERHAPS OF TRYING
TO,A CHIEF CORHOTTERS OF
PATIENTS WHO ARE REALLY--COHORTS
OF PATIENTS WHO ARE REALLY
GENIUS TO INTERROGATE A PATHWAY
LIKE THIS 1 IN A SIGNIFICANTLY
SIZED POPULATION WHERE PATIENTS
REALLY HAVE DISEASE PATHOLOGY
ATTRIBUTABLE TO 1 PATHWAY, TO
TRY TO COUNTERACT THAT
INCREDIBLE NOISE THAT THERE IS
IN RHEUMATIC DISEASES OF THIS
VARIABILITY AND THE CLINICAL
PATTERNS OF THE DISEASE.
>> YEAH, AND MAYBE, MAYBE I WILL
HAVE THE LAST WORD MYSELF, AND
SAY PERHAPS GOING UP STREAM OF
DISEASE AND PERHAPS PREE DEC
ALSO MIGHT BE A NICE PLACE TO
LOOK AND JUST SEE BECAUSE IF
IT'S A HUGE DRIVER, YOU WOULD
EXPECT TO SEE BEFORE THE OTHER
AUTOANTIBODY PERHAPS BECOME
MANIFEST, RIGHT?
>> OKAY, SO I WILL WRAP US UP
HERE, I WANT TO SAY A HUGE THANK
YOU TO OUR 3 SPEAKERS, THAT WAS
JUST A WONDERFUL SESSION, I REAL
LE ENJOYED ALL OF YOUR TALKS AND
A HUGE THANK YOU ALSO TO
DRS. MANCINI AND PERG-I SON FOR
MODERATING THE DISCUSSION AND
HELPING US WITH THE ORGANIZATION
OF THIS SYMPOSIUM.
IT'S NICE TO KIND OF LISTEN TO
THE SCIENCE SPEAK FOR ITSELF.
SO PLUGS FROM THE OFFICE OF
AUTOIMMUNE DISEASE RESEARCH, IF
YOU WOULD LIKE TO HEAR MORE
ABOUT OUR OFFICE AND THE WORK
THAT WE ARE DOING, PLEASE JOIN
OUR NEXT UPDATE SESSION, WE HAVE
THESE QUARTER LEE, THE NEXT 1
WILL BE ON FRIDAY MAY THIRD, IF
YOU SCAN THE QR CODE, IT WILL
TAKE YOU TO THE EVENT PAGE AND
YOU CAN SIGN UP FOR THE LINK.
IT'S VIRTUAL AS YULE NOTICE A
LOT OF OUR THINGS ARE ONLINE
ONLY, JUDGE UOF THE TO INCREASE
ACCESSIBILITY FOR OUR LISTENERS.
WE ALSO WANT TO GIVE A PLUG TO
THE EIGHTH ANNUAL VIVIAN
PENN SYMPOSIUM ON MAY 15th, IT
IS 1 OF THE NIH'S MECHANISMS BY
WHICH NIH SPONSORED WOMEN'S
HEALTH WEEK, WHICH FOR THOSE WHO
DON'T KNOW ALWAYS HAPPENS THE
WEEK AFTER MOTHER'S DAY SO MAY
15th, PUT IT IN YOUR CALENDAR,
WE HAVE A FABULOUS LINE UP OF
SPEAKERS RUNNING THE SPECTRUM OF
SCIENCE, CLINICAL, CARE, PATIENT
ADVOCACY, A REAL MIX, I WILL
SAY, REGISTRATION FOR THIS IS
LIMITED TO THE FIRST 1000 PEOPLE
SO YOU DON'T MISS A CHANCE, THE
QRR CODE WILL TAKE YOU TO THE
REGISTRATION SITE AND IT WILL BE
WEBCAST AS WELL AND THEN
FINALLY, IF YOU WOULD LIKE TO
STAY CONNECT WIDE US IN
PARTICULARLY HERE ABOUT FUNDING
OPPORTUNITIES ON THEN SIGN UP
OUR E-MAILS THIS QR CODE WILL
TAKE YOU TO THE E-MAIL SIGN UP
AND JUST TO HIGHLIGHT FOR YOU,
YOU NEED TO CLICK TO ASK TO SIGN
UP FOR SUBSCRIPTION TO THE
OFFICE OF AUTOIMMUNE DISEASE
RESEARCH, WE DO SEND OUT REGULAR
COMMUNICATIONS WHEN EVENT
REGISTRATIONS OPEN UP AND WHEN
WE HAVE NEW FUNDING
OPPORTUNITIES, OTHER EVENTS AND
LOTS AND LOTS TO HEAR ABOUT IN
THE WORLD OF AUTOIMMUNE DISEASE,
SO WITH THAT, I AM GOING TO WRAP
UP.
I WANT TO THANK YOU ALL FOR
COMING, WE HAD A FABULOUS TIME
AND I WANT TO THANK AGAIN, ALL
OF OUR SPEAKERS AND THE
MODERATORS FOR RUNNING A GREAT
SESSION.