>> GOOD AFTERNOON, EVERYBODY.
WELCOME TO TODAY'S
CLINICOPATHOLOGIC GRAND ROUNDS.
BEFORE I ANNOUNCE OUR SPEAKERS,
LET ME JUST ADDRESS A COUPLE OF
HOUSEKEEPING ISSUES. FOR THOSE
WHO REQUIRE CME THE CME CODE FOR
TODAY'S 57907, YOU KNOW THE
NUMBER TO TEXT AND IT IS ON THE
SLIDE THERE SO TEXT THAT CODE
YOU SHOULD BE ABLE TO RECEIVE
CME. WE REQUEST THAT YOU PLEASE
PROVIDE US FEEDBACK AT THE END
OF THE TALK. YOU CAN DO THAT BY
SCANNING THE QR CODE THAT WILL
APPEAR ON THE CME SLIDE AT THE
END. FOR THOSE WHO APPLIED FOR
CME YOU WILL GET AN EMAIL LINK
TO TAKE YOU BACK TO YOUR
FEEDBACK FORM. WE APPRECIATE
HEARING FROM YOU. AFTER THE
PRESENTATION, IF YOU HAVE ANY
QUESTIONS PLEASE PROCEED FOR
THOSE IN THE LIVE AUDIENCE
PROCEED TO THE MIKES IN THE
AISLES AND FOR THOSE WHO ARE
VIEWING ONLINE YOU CAN SEND US
QUESTIONS THROUGH THE LIVE
FEEDBACK BUTTON AND WE WILL TRY
TO ADDRESS THEM AS TIME PERMITS.
SO WITH THAT, I WILL INTRODUCE
OUR ESTEEMED SPEAKERS TODAY. WE
HAVE DR. MAXWELL LAWS, A
NEUROSURGERY RESIDENT AND
POST-DOC IN THE FUNCTIONAL
NEUROSURGERY SECTION OF NINDS.
WE ALSO HAVE DR. NADIA BIASSOU,
WHO IS SENIOR CLINICIAN IN THE
RADIOLOGY IMAGING SCIENCES HERE
AT THE CLINICAL CENTER. WE HAVE
DR. PJ CIMINO, HEAD OF THE
NEUROPATHOLOGY UNIT SURGICAL
NEUROLOGY BRANCH OF NINDS AND
DR. DESMOND BROWN, ACI AND HEAD
OF NEUROSURGICAL ONCOLOGY UNIT
IN THE SURGICAL NEUROLOGY BRANCH
NINDS. THEY WILL PRESENT A CASE
OF ANNA PLASTIC AND ASTRO
CYTOMA. WITH THAT I WILL HAND IT
TO DR. LAWS. ANAPLASTIC
>> THANK YOU VERY MUCH,
EVERYBODY. SO WE HAVE NO
DISCLDISCLOSURES. OUR PATIENT
INITIALLY PRESENTED TO OUR
SERVICE IN SEPTEMBER OF 20 --
2011, HE WAS A 30-YEAR-OLD AT
THE TIME, THIS WAS FALL
FOLLOWING A PARTIAL RESECTION OF
A WHO GRADE 3 ANAPLASTIC ASTRO
CYTOMA PERFORMED AT GEORGETOWN
HOSPITAL. HE HAD RECEIVE HAD
STANDARD OF CARE RADIATION
THERAPY AND CURRENT
TILAZOLAMIDE. HE INITIALLY
INVOLVED FIELD RADIATION AND
FOLLOWING THAT HAD TEN CYCLES OF
TEMZOLAMIDE STOPPED IN SEPTEMBER
OF 2012 DUE TO FATIGUE AND LOW
NEUTROPHIL COUNT. THE PATIENT
UNDERWENT LATER PATHOLOGIC
TESTING THROUGH THE NIH AND IT
SHOWED HE HAD A TP 53 MUTATION
AND IDH MUTATION AS WELL AS CDK
4 COPY NUMBER GAIN. THE PATIENT
WAS FOLLOWED BY THE NIH
NEUROONCOLOGY BRANCH FOR THE
NEXT TEN YEARS. AND HE HAD
STABLE SCANS AND HE WAS DOING
WELL CLINICALLY. HE HAD AN
INTERVAL SCAN PERFORMED IN MAY
OF 2023 WHICH SHOWED A NEW
11-MILLIMETER ENHANCING NO DUAL
WITHIN THE PREVIOUS RESECTION
BED WHICH WAS CONCERNING FOR
RECURRENCE OF HIS DISEASE. IN
JUNE OF THAT YEAR, THE PATIENT
UNDERWENT A REDO RIGHT FAN TOLL
CRANEIOTMY FOR RESECTION OF
TUMOR WITH DR. BROWN. THIS
PATHOLOGY WAS CONSISTENT WITH
HIGH GRADE GLIOMA AND PATIENT
WAS SEEN IN FOLLOW-UP CLINIC A
MONTH LATER WITH A REPEAT MRI.
AT THAT TIME, WE FELT THAT IT
WAS APPROPRIATE TO MANAGE
CONSERVATIVELY WITH GETTING
REPEAT IMAGING IN OCTOBER AND
THEN CONSIDERING RESTARTING THE
PATIENT ON TEMOZOLOMIDE IN
OCTOBER. SO WE HAD OBTAINED THAT
SCAN AND IT SHOWED A LARGE
ENHANCING MASS WITHIN THE PRIOR
RESECTION CAVITY. SO THE PATIENT
UNDERWENT SURGERY AGAIN. ON
OCTOBER 12TH. AND THE PATIENT
HAD A HOSPITALIZATION SOMEWHERE
AROUND 12 DAYS AND WAS
DISCHARGED TO REHAB. SO THE
PATIENT THEN CONTACTED OUR
CLINIC AND OR THE PATIENT
HEALTHCARE PROVIDERS CONTACTED
OUR CLINIC AND INFORMED THE
PATIENT DEVELOPED NEW SOME LENS
AND VOMITING. SO THE PATIENT WAS
TRANSFERRED TO THE WASHINGTON
HOSPITAL CENTER AT THE
WASHINGTON HOSPITAL CENTER THE
PATIENT WAS NOTED TO HAVE
HYDROCEPHALUS, CONCERNING FOR
EITHER TUMOR ASSOCIATED
HYDROCEPHALUS AND SO WE
TRANSFERRED THE PATIENT BACK TO
THE NIH FOR CONSIDERATION OF A
CENTRIC LAR PERITONEAL SHUNT. SO
WE PLACED A SHUNT ON NOVEMBER 20
WITHOUT ANY ISSUE AND THE
PATIENT WAS ADMITTED TO ICU.
THIS PATIENT HAD PERSISTENT
GNASHIA AND NEUROLOGIC DECLINE.
WE OBTAINED INTERVAL IMAGING
GROWING A GROWING SUB DO YOU
RECALL FLUID CONNECTION FOR
TUMOR ASSOCIATED CYST. THE
PATIENT DISTIER YOUR RATED
NEUROLOGICALLY SO WE PERFORM --
DETERIORATED NEUROLOGICALLY SO
WE PERFORMED HOLE ASPIRATION AND
AND CSF TO DIALED THE SHUNT FROM
5 TO 7 TO SLOW THE CSF. THE
PATIENT CONTINUED WITH
NEUROLOGIC DECLINE AND SO WE
PLACE ADN'T RIGHT FRONTAL EVD TO
ADDRESS PATIENT ALTERED MENTAL
STATUS AND HYDROCEPHALUS. THE
CFS FROM THE ASPIRATION GREW
CANDIDA PARAPSILOSIS SO WE WERE
CONCERNED THE PATIENT HAS FUNGAL
MENINGITIS AND STARTED ON FLEW
CON SOLVE. WE HAD SAMPLES FROM
THE EVD, THOSE CONTINUED TO GROW
CANDIDA AND FUNGAL CULTURES. SO
WE ALSO CONCERNED THE TUMOR
RECURRED AND CONFOUNDD BY THIS
FUNGAL INFECTION AND WE DID
PERFORM EXTERNAL BEAM READIATION
IN EARLY DECEMBER TO TRY TO GET
TUMOR CONTROL. THE PATIENT
CONTINUED TO HAVE CHALLENGES
WITH THIS INFECTION SO WE
EXPLANTED THE SHUNT AND REPLACED
THE EVD TO TRY TO EXCHANGING THE
HARDWARE AND PLACED A RIGHT SUB
DO YOU RECALL DRAIN TO TRY TO
DRAIN THE TUMOR ASSOCIATED CYST.
CSF CONTINUED TO BE -- HAVE
EVIDENCE OF INFECTION. AND THEN
WE -- THE PARENTED WAS HAVING
FURTHER ISSUES WITH THESE THE
SHUNT AND THE EVD, BASICALLY WE
HAD TO CONTINUOUSLY TRY TO
EXCHANGING THE FLUID AND FLUSH
THE DRAINS. THE PATIENT BECAME
MORE PROGRESSIVELY SOME LENT AND
WE CHANGED CODE STATUS TO DNR
DNI. EVENTUALLY TRANSITIONED THE
PATIENT TO HOSPICE CARE DECEMBER
14 AND HE EXPIRED DECEMBER 29. I
WILL TRANSITION TO DR. BIASSOU
TO TALK ABOUT THE RADIOLOGY FOR
THE PATIENT.
>> THANK YOU. AS MENTIONED, THE
PATIENT ORIGINALLY WAS TREATED
AT GEORGETOWN UNIVERSITY
HOSPITAL SO THE INITIAL SET OF
IMAGING HERE IS FROM GEORGETOWN.
AND I JUST WANTED TO POINT TO
YOU THE APPEARANCE OF THE MASS
PRIOR TO THEIR SURGICAL
RESECTION. SO THIS WAS -- IF YOU
LOOK ON THE FIRST IMAGE ON THE
LEFT THIS IS A NON-CONTRAST T 1
POST CONTRAST T 2. THERE IS A
FLAIR WEIGHTED SEQUENCE HERE.
AND DWI WHICH BASICALLY ONE USES
TO TRY TO UNDERSTAND WHETHER
THERE'S DIFFUSION RESTRICTION.
WHAT WE SEE HERE IS A
NON-ENHANCING OR FAINTLY
ENHANCING MASS THAT HAS PERT
NACHOS MATERIAL THAT DOES
RESTRICT. THE PATIENT UNDERWENT
SURGICAL RESECTION. THIS IS THE
PRE-CONTRAST T 1 POST CONTRAST T
1 P PRE-OPERATIVE, THIS IS THEIR
POST SURGICAL T 1 POST CONTRAST
IMAGE. WHAT WE SEE IS A SURGICAL
BED HERE. AND WE SEE A LITTLE
ENHANCEMENT SO THE QUESTION
ARISES IS TO WHETHER YOU GOT
EVERYTHING OR IF THERE'S SOME
RESIDUAL TUMOR THAT'S LEFT. AND
WAY WHICH WE CAN ANSWER THAT
QUESTION IS TO LOOK AT THE
PRE-CONTRAST T 1 POST-OPERATIVE
STUDY. SO HERE THERE IS NO
CONTRAST. WE DO SEE WHAT LOOKS
LIKE HYPERSIGNAL. THIS IS
CONSISTENT WITH HEMORRHAGE
NON-ENHANCEMENT AND THIS IS
CONFIRMED ON THE SUSCEPTIBILITY
WEIGHTED IMAGE SEEN BELOW. AS
DR. LAWS MENTIONED THE PATIENT
WAS FOLLOWED WITH MULTIPLE
SUBSEQUENT MRI STUDIES,
BASICALLY FOUND TO BE DOING
QUITE WELL AND NO RECURRENCE
UNTIL 2023 ON ROUTINE FOLLOW-UP
SCAN THERE WAS A ENHANCING NO
DUAL HERE WE SEE THE RESIDUAL
SURGICAL CAVITY AND EVIDENCE OF
PRIOR SURGERY BUT WE DO SEE
ENHANCING NO DUAL HERE. SO THE
QUESTION IS WHAT IS THAT, IS
THAT RECURRENCE? IT LOOKS LIKE
IT IS. WE CONFIRMED THAT WITH A
PERFUSION SCAN. THIS IS
PERFUSION SCANS ARE DONE
DIFFERENTLY, WE LEAK AT THE
CURVE OF UPTAKE OF ENHANCEMENT.
HERE WE DO T AS A MAP AND THIS
IS A MAP OF THE PRE-CONTRAST OF
THE POST CONTRAST PERFUSION
MINUS PRE-CONTRAST PERFUSION
SCAN. SO IT REALLY SHOWS YOU
VERY CLEARLY THE HYPERPERFUSION
OF THIS ENHANCING NO DUAL. ONE
OF THE THINGS YOU MAY READ ABOUT
IN THE A LITERATURE IS
HYPERSENSE TVTY IS CONFUSED WITH
RESTRICTION, THAT'S WHAT WE
EXPECTED TO SEE ON THE WHI
SEQUENCE WHERE THE HIGHLIGHTER
IS POINT BUD WE DON'T SEE THAT.
THIS IS SOMETHING I MENTIONED TO
FOLLOWS WHEN THEY ARE ROTATING
THROUGH GEORGETOWN AND GW THAT
WITH CERTAIN TUMORS YOU DON'T
ALWAYS SEE DIFFUSION
RESTRICTION. YOU SEE WITH SOME
TUMORS, LYMPHOMA, TIGHTLY PACKED
CELLS, THOSE USUALLY PRESENT
WITH DIFFUSION RESTRICTION BUT
NOT NECESSARILY GLIOMA. SO IF
YOU DON'T SEE DIFFUSION
RESTRICTED FOCUS ASSOCIATED WITH
A NODULE PLEASE DON'T THINK YOU
CAN THROW THAT AWAY, IT IS
SOMETHING THAT NEEDS TO BE
FOLLOWED UP. THE SECOND THING
THAT WE HAVE TO ALSO THINK ABOUT
IF YOU DON'T SEE ENHANCEMENT
ELEVATED PERFUSION WHICH WE
DON'T BUT IF WE DO YOU HAVE TO
SEE IF THAT'S SOMETHING THAT
MIGHT BE ASKING THE ELEVATED
PERFUSION. SO THAT'S WHY WE LOOK
AT THE SWI WITH SUSCEPTIBILITY
IMAGE SO SEE IF THERE IS
HEMORRHAGE ASSOCIATED THAT MIGHT
BE MASKING YOUR ABILITY TO SEE
THE ELEVATED PERFUSION. SO WE
DON'T SEE THAT, IF YOU SEE
ELEVATED PERFUSION YOU DON'T SEE
A REASON, THIS IS A HIGHLY PER
FUSED LESION LIKELY HIGH GRADE
GLIOMA. IF YOU LOOK CLOSE, I
DON'T KNOW FOR THOSE OF YOU IN
THE AUDIENCE IF YOU LOOK CLOSE
YOU CAN SEE THAT THIS PATIENT
HAS HAD PRIOR RADIATION THERAPY
BECAUSE WE DO SEE POST RADIATION
ANGIOPATHY, SO HEMORRHAGE WITH
THE BRAIN PARENCHYMA AND
COINCIDES WITH THE HISTORY OF
BRAIN RADIATION THERAPY. AS DR.
LAWS SAID THIS LESION WAS
RESECTED ON FOLLOW-UP SCANS IN
OCTOBER OF 2023. THERE IS
ANOTHER MASS THAT'S MUCH BIGGER,
THERE IS A LARGE AMOUNT OF
SURROUNDING EDEMA ASSOCIATED
WITH THAT. IF OWE LOOK AT THE
PERFUSION SCAN WE SEE IT IS
HIGHLY PER FUSED SO A HIGH --
PERFUSED SO A HIGH GRADE GLIOMA
UNTIL PROVEN OTHERWISE BY
RADIATION CRITERIA. THE PATIENT
UNDERWENT SURGICAL RESECTION YES
WANT TO MAKE SURE WE HAVE GOTTEN
ALL THE TUMOR, THE POST CONTRAST
T # SHOWS ENHANCEMENT ALONG THE
PERIPHERY OF THE SURGICAL CAVITY
AND YOU ALWAYS CONCERN THAT
MAYBE THERE IS A LITTLE BIT OF
RESIDUAL TUMOR THERE. WE LOOK AT
PRE-CONTRAST STUDY AND SEE THAT
IS REALLY ASSOCIATED WITH
POST-OPERATIVE HEMORRHAGE WHICH
IS CONFIRMED ON THE SWI
SEQUENCE. THE IMMEDIATE POSTOP
HEAD CT THAT WAS DONE SHOWS POST
SURGICAL CHANGES, RESIDUAL
HEMORRHAGE BUT I WANT TO POINT
TO YOU IMMEDIATELY POSTOP WE
DON'T HAVE EVIDENCE OF
HYDROCEPHALUS. THE LATERAL
VENTRICLES ARE NORMAL SIZE.
THERE IS A LITTLE POST-OPERATIVE
PNEUMOSYPHILOUS. BUT ONE MONTH
LATER THE PATIENT COMES BACK
NON-CONTRAST HEAD CT IS
PERFORMED. THERE'S LARGE AMOUNT
OF HYDROCEPHALUS, THIS IS THE
SURGICAL CAVITY AND WE SEE A
SMALL SUB DURAL FLUID
COLLECTING. THE PATIENT HAD VP
SHUNT PLACED IN THE VENTRICULAR
SYSTEM. YOU STILL SEE THE
SUBDURAL FLUID COLLECTION, THERE
IS A SECOND VP SHUNT PUT IN
PLACE TO TRY TO REDUCE THAT AND
THERE IS A SMALL RESIDUAL
IMPROVEMENT, I WANTED TO SHOW
THERE IS LIKELY EPIDURAL AND
SUBDURAL FLUID COLLECTION AND
THE LEFT TEMPORAL HORN APPEARED
ENTRAPPED. AFTER MULTIPLE SCANS
THAT HAPPENED BETWEEN THE LAST
SCAN I SHOWED AND THE ONE HERE,
ON THE LEFT WHERE ALL THE DRAINS
HAVE NOW BEEN ALL THE VP SHUNTS
HAVE BEEN REMOVED PROBABLY
BECAUSE THERE'S BEEN EVIDENCE
FROM CSF THERE IS INFECTION.
THEY REMOVED ALL OF THAT. WE CAN
SEE THE SUBDURAL FLUID
COLLECTION INCREASED THE
VENTRICULAR SIZE ENLARGED
DESPITE MANY WEEKS OF HAVING THE
VP SHUNT IN PLACE TO HELP DRAIN
THAT. THEY THEN PUT IN A RIGHT
FRONTAL SURGICAL DRAIN AND WE DO
SEE AN IMPROVEMENT OF THE
EPIDURAL AND SUBDURAL FLUID
COLLECTION AS WELL AS SOME
IMPROVEMENT IN THE SIZE OF THE
VENTRICLE, IT STILL REMAINS
ENLARGED BUT AFTER ABOUT A FEW
WEEKS, THIS IS ON DECEMBER 14,
YOU CAN SEE THAT THOSE FLUID
COLLECTIONS ARE STARTING TO
REACCUMULATE. THEN THE PATIENT
IS SENT TO HOSPICE. I WANT TO
INTRODUCE DR. CIMINO WHO WILL
TALK ABOUT PATHOLOGY.
>> THANK YOU. SO I'M GOING TO GO
OVER THIS PATIENT'S PATHOLOGY IN
A TIME LINE FORMAT BECAUSE IT
GETS COMPLICATED AS WE GO OVER
TIME SO I WANT TO START WITH THE
INITIAL EXCISION THAT HAPPENED
IN GEORGETOWN IN 2011.
UNFORTUNATELY DON'T HAVE ANY
PATHOLOGY SLIDES TO REVIEW SO
I'M GOING TO DISCUSS THE
PATHOLOGY WHAT WE ARE SEEING. ON
THE ACTUAL HISTOPATHOLOGY WE SAW
DIFFUSE ASTROSITIC GLIOMA,
ELEVATED MITOTIC ACTIVITY AND NO
MICROVASCULAR PROLIFERATION OR
NECROSIS. I WILL COME BACK HOW
WE GRADE THE TUMORS AND WHY THE
FEATURES ARE IMPORTANT. ON
IMMUNOHISTOCHEMISTRY IT HAD A
GLIAL PHENOTYPE SO POSITIVE FOR
GLIAL ASITIC PROTEIN OR GFAPP.
POSITIVE FOR R 312H MUTATION ONE
OF THE MAIN FEATURES NOW THAT WE
USE TO CATEGORIZE DIFFUSE
GLIOMAS IS THE PRESENCE OF IDH
MUTATION. ATRP WAS RETAINED P53
PROLIFERATED AND ELEVATED 8%
INDICATING MORE PROLIFERATIVE
THAN LOWER GRADE TUMOR. THEN
WHEN WE THINK ABOUT IDH MUTANT
DIFFUSE GLIOMAS, IT CAN BE
PRESENT IN AL GO DEN DROVE
GLIOMA AND HOW WE DISTINGUISH IS
ROOKING FOR PA P 19 Q CODE
DELETION BACK IN TIME IN SITU
HYBRIDIZATION WAS PERFORMED AND
IT WAS NEGATIVE FOR Q CODE
DELETION. THAT'S WNS WHO GRADE
3, THE FORMER TERM WAS ANN
PLASTIC CYTOMAPPED A LITTLE HOW
WE GRADE, IN 2008 THE IDH
MUTATION WAS INTRODUCED AS
CRITERIA FOR GRADING BUT
OFFICIALLY 2016 THERE WAS ADDED
TO THE WHO CRITERIA. SO IT WAS
THAT IF DIFFUSE ASTRO SIGH
MOMMA, NO OTHER FEATURES GRADE
2. WITH INCREASED MITOTIC
ACTIVITY THEN BUMPS UP TO GRADE
3 ANN PLASTIC ASTRO CYTOMA
MUTANT, IF IT HAD NECROSIS
NEITHER WHICH THE PATIENT HAD IT
MOVES TO THE GLIAL BLASTOMA IDH
MUTANT. THIS IS NO LONGER
ACCEPTED NOMENCLATURE, IN
SEPTEMBER 2021 IT WAS CHANGED SO
GLIOBLASTOMA IS ASSOCIATE WITH
THE WILD TYPE TUMORS SO NOW WE
HAVE A COUPLE OF WAYS TO GRADE.
HISTOLOGICAL OR MOLECULAR
CRITERIA. SO AGAIN ASTRO CYTOMA,
IDH MUTANT WHO GRADE 2 IF NO
FEATURES IF ELEVATED MITOTIC
ACTIVITY, YOU GET TO GRADE 3.
THEN EITHER MICROVASCULAR
PROLIFERATION OR NECROSIS ON
HISTOLOGY OR CDK AND HOMOZYGOUS
DELETION BASED UPON MOLECULAR
FINDINGS ELEVATE TO WHO GRADE 4
AND BECAUSE I WANT TO BRING THIS
UP BECAUSE THERE IS THE TIME
SPAN OF WHEN THIS WAS DIAGNOSED
AND RELEVANT NOMENCLATURE
CHANGES EVERYTHING I WANT TO
TALK MORE ABOUT THE RELEVANT
CHANGES IN THE 2021 CNS. SO HERE
I SHOW YOU THE COVERS FOR THE
WHO OVER TIME AND REALLY WAS
MORE HISTOLOGY AND IMMUNOCHEST
CHEMISTRY BASED UNTIL RECENTLY
WHEN WHEN WE STARTED USING
GENETICS AND EPIGENETIC
SIGNATURES WHICH I WILL TALK
ABOUT IN ORDER TO CLASSIFY THE
TUMORS. SO THEY MOVE FROM
RATHER THAN ROMAN NUMERALS FOR
GRADING. THEY TURNED TO IN CLASS
GRADING. SO INSTEAD OF DIFFUSE
ASTRO CYTOMA TO ANN PLASTIC SAY
TOE MA TO GLIAL BLASTOMA, IT IS
ASTRO CYTOMA GRADES 2, 3 AND 4
SO ANN PLASTIC IS REMOVED. WE
HAVE COMBINED HISTOLOGIC AND
MANY LOOK ALREADY GRADING FOR
TUMORS AND THERE WAS FIRST DONE
WITH IDH MUTANT TUMORS. FOR
WHAT WILL BE RELEVANT FOR THE
PATIENT RESORNS, WE HAVE COME UP
WITH THESE -- RECURRENCE WE HAVE
COME UP WITH NOT HE WILL
CLASSIFIED THE THE PATIENT TUMOR
DOES NOT FIT INTO A NICE WHO
CATEGORY. SO THE INITIAL TUMOR
WE CAN CONSIDER IDH OR ASTRO
CYTOMA IDH MUTANT WHO GRADE 3
AND MAKES MORE SENSE IF WE THINK
ABOUT THE LONG LIVE SURVIVAL FOR
THIS IT IS THE IKH MUTATION
DRIVING PROLONGED SURVIVAL
RATHER THAN ANY TYPE OF GRADING
SO SECOND EXCISION PERFORMED
HERE, BY DR. BROWN, WE SEE
HEMOTOXIN HED STAIN SLIDE YOU
HAVE TO LOOK AT ENOUGH TO TRUST
ME THIS IS MINIMAL
HYPERCELLULARITY AND WE SEE A
LOT OF REACTIVE CHANGES RATHER
THAN ACTIVE TUMOR SO I JUST
HIGHLIGHTED HERE SOME HYPERPLAST
TICK BLOOD VESSELS AS WELL AS
RARE FRACTION SEEN AS
VACCINATION IN THE BACK. WE SEE
WHAT LOOKS LIKE REACTIVE ASTRO
CYTOSIS AND NOT A LOT OF TUMOR
CONTENT. WE DID NEXT GENERATION
SEQUENCING AS PARTS OF STANDARD
WORKUP, WE DID NOTE THAT THE IDH
1 MUTATION WAS LOST SO WILD TYPE
AND PROMOTER MUTATION AND TP 53
MUTATIONS AT VERY LOW FREQUENCY
SO HERE I'M SHOWING THE CHART,
THE VF FREQUENCY IS LOW
CONFIRMING THAT THE TUMOR BURDEN
IS LOW IN THIS PATIENT. THEN FOR
COMPLETENESS ONE OF THE THINGS
WE DO HERE AT THE NIH AND
BECOMING MORE PREVALENT IN
SURGICAL NEUROPATHOLOGY IS
LOOKING AT DNA METHYLATION BASED
CLASSIFICATION SO HERE I'M
SHOWING THE SEMINOLE PAPER BY
CAPPER AND IDLEBURG GROUP THAT
SHOWED WHAT YOU DO IS TAKE A
RANDOM MACHINE LEARNING RANDOM
CLASSIFIER, TAKE A WHOLE GENOME,
WHOLE GENOME DNA METHYLATION
ARRAY, FEED IT INTO THIS MACHINE
LEARNING PROGRAM AND YOU CAN
COME OUT WITH A CNS TUMOR BASED
CLASSIFICATION. HERE IS ONE OF
THE EXAMPLES THAT INCLUDES IDH
TUMOR MEDULLAR BLASTOMA, ET
CETERA, ET CETERA. THEN IN OUR
CLINICAL PRACTICE HOW WE
INCORPORATE DNA METHYLATION IN
SURGICAL NEUROPATHOLOGY, WE TAKE
DNA FROM TISSUE BLOCKS, PREPARE
THE DNA IN SUCH A WAY THAT IT IS
HYBRIDIZES TO THE DNA
METHYLATION ARRAYS AND IS READ
BY EYE SCAN. THEN PUT THROUGH A
BIOINFORMATIC PIPELINE AND WE
COME OUT WITH, WE USE CURRENTLY
A FEW DIFFERENT CLASSIFIERS, AND
THEN WE ALSO PUTS IT ON THESE
TWO DIMENSIONALITY REDUCTION
MAPS, YOU CAN USE U MAP OR TISNY
PLOTTING AND IT GIVES
APPROXIMATION WHERE OTHER TUMORS
WITH SIMILAR SIGNATURES MIGHT
LAND. FROM THIS WE ALSO GET COPY
NUMBER ALER RATIONS SO HERE I
SHOW YOU A -- ALTERATIONS. SO
HERE IS A COPY NUMBER PLOT ON
THE X AXIS WE HAVE CHROMOSOMES
LABELING FROM SMALL TO LARGEST
SO CHROMOSOME 1 ON THE BOTTOM
AND CHROMOSOME 22 OVER HERE. WE
CAN GET SPECIFIC GENE
METHYLATION FOR ESPECIALLY MGMT
PROMOTER METHYLATION STATUS,
NEUROONCOLOGIST USE FOR
TREATMENT DECISIONS. THEN WE
INTEGRATE THIS WITH THE CLINICAL
RADIOGRAPHIC HISTOLOGICAL AND
GENOMIC FINDI FINDINGS TO COME H
INTEGRATED DIAGNOSIS. THIS
SPECIMEN, METHYLATION CLASSIFIER
CAME BACK WITH NO MATCH. WE HAVE
THE U MAP DIMENSIONALITY
CLUSTER, THE PATIENT'S TUMOR
CLUSTERED WITHIN NORMAL OR
REACTIVE BRAIN WHICH IS
CONSISTENT WITH THE HISTOLOGICAL
APPEARANCE AND WE HAVE A FLAT
COPY NUMBER PLOT WHICH INDICATES
VERY LOW TUMOR BURDEN. THE FINAL
NEUROPATHOLOGICAL DIAGNOSIS FROM
THE INTEGRATED WORK UP IS
REACTIVE CHANGES WITH RESIDUAL
OR NON-ACTIVE GLIOMA. SO THAT
WAS THE GENE SPECIMEN AND THEN
-- OCTOBER. DR. BROWN WENT BACK
FOR ANOTHER EXCISION. AND HERE
WE CAN SAY HNE STAINS AGAIN THIS
IS HYPERCELLULAR OPPOSED TO THE
LAST ONE WITH TUMOR NUCLEI. WE
SEE ATYPICAL GLIAL CELLS, WE
HAVE A LOT OF WHAT LOOKS LIKE
MONSTROUS OR MULTI-NUCLEATED
GIANT CELLS SO IT LOOKS VERY
MUCH LIKE AGREE OWE MA AND HERE
I CIRCLED A BUNCH -- GLIOMA, I
CIRCLED MITOTIC, SO TO IS
MITOTICALLY ACTIVE AND WE SEE
NECROSIS IN SOME AREAS SO A LOT
OF HISTOLOGICAL HIGH GRADE
FEATURES. WE DID NGS SEQUENCING
ON THIS. AGAIN STILL IDH WILD
TYPE SO IT STILL HAS LOST IDH
MUTATION AND P53 MUTATION IS A
HIGHER BRAIN ALLELE FREQUENCY
WITH OUR HISTOLOGICAL FINDINGS
OF BEING HIGHER ACTIVE TUMOR
BURDEN. THE METHYLATION
CLASSIFIER IN THIS CASE STILL
GAVE BACK NO MATCH. THE U MAP
DID PUT IT INTO IDH MUTANT HIGH
GRADE ASTRO CYTOMARE GENERAL.
THE U MAP DIMENSIONALITY
REDUCTION NOT AS ROBUST AS THE
METHYLATION CLASSIFIERS AND
STOCHASTIC PLACING SO HARD TO
KNOW TOO MUCH WHAT THIS MEANS
OTHER THAN LOOKS SOME LEVEL
STILL RETAINS EPIGENETIC PROFILE
OF IDH GLIOMA, WE HAVE A LOT OF
ANEUPLOIDY WITH PROGRESSIVE
GLIOMA AND PDJF ONCOGENIC
AMPLIFICATION ON CHROMOSOME 3.
NOW FOR THE FINAL INTEGRATED
PATHOLOGICAL DIAGNOSIS RECURRENT
OR HIGH GRADE GLIOMA, IDH WILD
TYPE BECAUSE LOST THE MUTATION
AND NOT ELSEWHERE CLASSIFIED
BECAUSE THERE IS NO REAL
CRITERIA, IT HASN'T REALLY FIT
THE PROFILE OF ANY SPECIFIC WHO
CLASS AND THIS COMES FROM LOWER
GRADE GLIOMA. AND USUALLY ONE OF
THE QUESTIONS I GET, USUALLY IDH
MUTANT ASTRO CYTOMA WHEN THEY
PROGRESS TO HIGHER GRADE RETAIN
MUTATIONS, THIS DID NOT. I WANT
TO SHOW ONE SIMILAR TO THIS ONE
WHERE THE PATIENT HAD GRADE 2
ASTRO CYTOMA ID,H MUTANT, UPON
RECURRENCE LOST THE IDH MUTATION
AND GAINED PDFJRN AMPLIFICATION
LIKE THIS PATIENT DID. SO WE
HAVE RECURRING HIGH GRADE
GLIOMA. THEN WE GET TO THE
AUTOPSY POSTMORTEM FINDINGS.
THIS IS GOING TO SHOW A LITTLE
PATHOLOGY FOR THIS, SO THIS IS
WHERE THE FRONTAL LOBE RESECTION
CAVITY IS, YOU CAN SEE IT IS
MORE PALE ON THIS LOWER POWER
HERE. THAT PALE AREA IS FILLED
WITH A LOT OF MACROPHAGES ON HNE
THEN WE CAN CON EQUIPMENT WITH
CD 163 MARKER OF ACTIVATEDDED
MYELOID CELLS. N'T EXTENSIVE BUT
WE DID NOT SEE EVIDENCE OF
CANDIDA IN THESE SLIDES. SO
BASICALLY KIND OF LIKE THIS
CHRONIC INFARCT TYPE SITUATION
YOU GET WHEN THERE'S BEEN TISSUE
RESECTED. ADJACENT TO THAT IT
IS LIKE THE FIRST SET OF IMAGES
I SHOWED IT IS MINIMALLY
ATYPICAL, THERE IS SOME
SCATTERED ATYPICAL CELLS. DID
LOOK A LITTLE WHEN WE SAMPLE
THROUGHOUT THROUGHOUT THE BRAIN
DEEPER IN THE THALAMUS SAW
SCATTERED ATYPICAL CELLS IN A
SINGLE MITOSIS WHICH IS KNOWN
WHEN YOU GO AND LOOK AT AUTOPSY
SPECIMENS OF DIFFUSE GLIOOWE MAS
WE GET SINGLE CELLS THROUGHOUT
THE CNS NOT MAKE PICKED UP ON
RADIOGRAPHIC EXAMS SO I WANTED
TO SHOW THIS EXAMPLE. THE FINAL
NEUROPATHOLOGICAL DIAGNOSIS IS
PREDOMINANTLY REACTIVE CHANGES P
WITH RESIDUAL GLIOMA. I WASN'T
ABLE TO GET PICTURES BUT UPON
GROSS EXAMINATION WE CAESAR
BRAILLE DEMANDS NOTCHING
CONSISTENT WITH A TRANSTENTORIAL
LEARNIATION. ONE OF THE FACTORS
IN PATIENT PASSING. SO OVERALL,
WE HAVE TIME LINE OF IDH MUTANT
ASTRO CYTOMA WITH LONG SURVIVAL,
BECAME VERY RECURRENT HIGH GRADE
GLIOMA WITH UNUSUAL GENETIC AND
EPIGENETIC FEATURES BUT
OTHERWISE LOOKED LIKE WHEN IT
GOT THE RADIATION PRIOR TO HIS
PASSING THAT IT LOOKED LIKE THAT
WOULD HAVE CONTROLLED THE
GLIOMA. SO THAT IS IT FOR THE
PATHOLOGY. NOWLY TURN IT TO DR.
BROWN TO TALK ASSOCIATED
BIOLOGY.
>> THANKS PJ. THIS IS VERY
INTERESTING AND COMPLEX CASE.
I'M GOING TO SEGUE HERE INTO
SOME OF THE BIOLOGICAL
IMPLICATIONS AND TIE THAT INTO
SOME OF THE THINGS I WORK ON IN
THE LAB. BEFORE I DO, I KNOW
THIS IS A COMPLEX CASE. DOES
ANYBODY HAVE QUESTIONS ABOUT THE
CASE, THE IMAGING OR PATHOLOGY
THAT'S BEEN PREVIOUSLY
PRESENTED? SO ONE OF THE THINGS
I DO WANT TO POINT OUT IS THAT
YOU SAW THE TIME LINE ON THE
LAST SLIDE WHERE THE PATIENT
INITIALLY GOT DIAGNOSED WITH
ESSENTIAL WILL I LOW GRADE
GLIOMA IN 2011. THEN THEY HAD
RECURRENCE OF SOMETHING THAT HAD
LAST -- THAT IDH MUTATION AND SO
ON IN JUNE BUT BY DECEMBER THEY
HAD PROGRESSED SO MUCH THAT THEY
DIED. SO THAT'S KIND OF
IMPLICATIONS HERE IN TERMS OF
WHEN WE THINK ABOUT
GLIOBLASTOMA, THE HIGH GRADE
GLIOMAS WE TALK ABOUT. AND THE
FACT THAT NOT ONLY ARE THEY
UNIVERSALLY FATAL, BUT REALLY
AGGRESSIVE. AND THAT TIME LINE
IS NOT OUT OF THE ORDINARY.
THAT'S KIND OF WHAT WE SEE. SO
IT IS REALLY IMPORTANT THAT WE
STILL KEEP ENERGY ON THIS
DISEASE AND WE HAVEN'T MADE A
LOT OF PROGRESS HERE. I WILL
TALK ABOUT SOME OF THAT AS I GO
ON. SEGUEING TO THE BIOLOGY. SO
WE ARE GOING TO TALK ABOUT THIS
PATIENT HAD ULTIMATELY INFECTION
WITH CANDIDA PEREPSILOSIS. SO
THAT IS A FUNGUS THAT IS ON ALL
OUR SKINS. AND LIVES IN OUR GI
TRACT. TYPICALLY NOT SOMETHING
THAT CAUSES FATAL INFECTION
UNLESS YOU ARE
IMMUNOCOMPROMISED. SO AS I WILL
TALK ABOUT HERE, CANCER IN
GENERAL BUT GLIAL BLASTOMA IN
PARTICULAR ARE ADEPT AT LOWERING
YOUR ABILITY TO MOUNT AN IMMUNE
RESPONSE AGAINST TUMOR. SO THERE
IS TUMOR MEDIATED
IMMUNOSUPPRESSION THAT IS A
REALLY BIG FEATURE OF
GLIOBLASTOMAS, IN FACT AUSTIN
AND OTHERS HAVE WRITTEN ABOUT
SOMEBODY WITH HIGH GRADE OWE MA,
LOOKS LIKE -- GLIOMA, LOOKS LIKE
SOMEBODY WITH AIDS IN TERMS OF
T-CELL LYMPHOPENIA, AND THE
HALLMARK OF AIDS BEING
OPPORTUNISTIC INFECTIONS LIKE
THIS. SO THAT IS KIND OF ONE OF
THE BIG AREAS MY LAB STUDIES. SO
WE DO THIS BY TRYING TO
UNDERSTAND SIGNALING THROUGH
THIS ORGANELLE THAT ESSENTIALLY
ALL OF OUR CELLS HAVE, NUCLEATED
CELLS ANYWAY, HAVE A PRY PLAYERY
CILIUM. IT IS KIND OFs SEW
TEARIC SO I WILL SPEND A LITTLE
TIME TALKING ABOUT THIS, IT IS
DIDN'T THAN THE CILIA IN YOUR
TRACHEA THAT MOVE THINGS ALONG
OR OTHER MOW TILE CILIA LIKE IN
SPERM. THESE ARE NON-MOTILE
CILIA WITH 9 PLUS 0 ARRANGEMENT
YOU CAN SEE 9 MICROTUBULE DUB
DUBLET. WHAT WE KNOW NOW ABOUT
PRIMARY CILIA, THEY ARE
IMPORTANT ZIP CODES IN THE CELL
WHERE A LOT OF IMPORTANT
CANONICAL SIGNAL TRANSDUCTIONS
PATHWAYS THAT P HAPPEN. SO THE
PROTOTYPE IS SONIC HEDGEHOG
WHICH REQUIRES PRIMARY SILL YUM
TO WORK. SO -- CILIUM TO WORK.
WHEN THERE IS A PRIMARY CILIUM
YOU GET ACTIVATION PATHWAY AFTER
SMOOTHEN GOES UP AND IT COMES TO
THE NUCLEUS AND YOU GET
ACTIVATION OF HEDGEHOG
SIGNALING. IN MANY B A SENSE OF
CILIA THIS ACTIVATION OF
SMOOTHEN CAN OCCUR AND THE
GLIOMA PROTEINS DON'T GET
ACTIVATE AND YOU DON'T GET THAT
TO WORK. IN ADDITION TO
HEDGEHOG, THIS IS JUST ON THE
RIGHT, A SMATTERING OF ALL THE
DIFFERENT SIGNALING CASCADES
THAT ARE WELL WORKED OUT TO
REQUIRE PRIMARY CILIA TO CONTROL
THEM. IT IS DIFFERENT WAYS. IT
IS NOT THE CASE WHERE FOR
EXAMPLE, ALL OF THESE REQUIRE
CILIUM. WINT SIGNALING IF THERE
ARE DEVELOPMENTAL BIOLOGISTS IN
THE ROOM WINT SIGNALING IS VERY
IMPORTANT IN NEURAL TUBE
DEVELOPMENT, WINT SIGNALING IS
INHIBITED BY RENT PRESENCE OF
CILIUM. SO COMPLEX RELATIONSHIPS
BUT IN OUR PATIENT WE SAW
AMPLIFICATION OF THE PDGFRA
RECEPTOR TYPE ARE A. FOR EXAMPLE
THIS IS A VERY CILIA DEPENDENT
PATHWAY. SO LOTS OF IMPLICATIONS
HERE. SO AGAIN GLIOBLASTOMA, IT
IS A SUCKY DISEASE. LIKE I SAID
INVARIABLY FATAL. ONE THING I
WANT TO POINT OUT, AGAIN, GOING
WITH THE WHAT OCCURRED IN OUR
PATIENT. THIS IS STANDARD OF
CARE. AND SO I KNOW THAT DR.
LAWS MENTIONED THAT A LITTLE
BIT, BUT THESE PATIENTS COME IN,
THE FIRST ELEMENT AND TREATMENT
IS USUALLY WHAT WE SAY IS
MAXIMAL SAFE RESECTION. SO THE
MOST BRAIN YOU CAN SAFELY TAKE
OUT, THAT'S THE FIRST STEP.
WITHOUT HURTING THEM TOO MUCH.
THE NEXT STEP IS THAT THEY WILL
THEN GO ON TO THIS ONE MONTH
PERIOD OF DAILY RADIATION WITH
CHEMOTHERAPY FOLLOWED BY SIX
MONTHS OF CHEMOTHERAPY. TOGETHER
THAT IS THE TWO PROTOCOL. AFTER
THAT, WHAT YOU GET IS THIS
DIFFERENCE HERE IN LIFE
EXPECTANCY BETWEEN THE RED AND
THE BLUE. SO COUPLE OF MONTHS.
PEOPLE WHO DON'T GET THAT, DIE
RELATIVELY SOON WITHIN WEEKS.
YOU CAN EXTEND THAT TO A FEW
MONTHS WITHOUT REALLY HERCULEAN
TREATMENT. AND I WANT TO AGAIN
REITERATE HERE IS THIS IDEA THAT
THE IMMUNOSUPPRESSION LIMITS A
LOT OF TREATMENT INCLUDING THE
WHOLE NEW CLASS OF DRUGS FOR
CANCER, CALLED IMMUNOTHERAPIES
THAT HAVE REVOLUTION AS IORIZE
HOW WE THINK ABOUT OTHER --
REVOLUTIONIZED HOW WE SEE WE SEE
THE ADS ON TV, THOSE ARE CHECK
POINT INHIBITORS THAT BLOCK
CANCER ABILITY TO SUPPRESS THE
IMMUNE SYSTEM AND THOSE DON'T
WORK IN GLIOBLASTOMA. SO THAT A
BIG FUNCTION OF OUR LAB. THAT'S
MOSTLY WHAT I WILL TALK ABOUT
TODAY. SO WHAT DO WE KNOW SO
FAR ABOUT THE WAY THIS WORKS?
GLIOBLAHS STOMA, ONE OF THE
FIRST QUESTION YOU SHOULD ASK,
WE ARE NOT TALKING LUNG CANCER
OR BREAST CANCER, WHICH IS
ALREADY IN THE SYSTEMIC
CIRCULATION IN THE SYSTEM, WE
ARE TALKING ABOUT GLIOBLASTOMA,
WHICH ARE ISOLATED TO THE
CENTRAL NERVOUS SYSTEM SO HOW
DOES THAT CAUSE YOUR BONE MARROW
TO BE SUPPRESSED? BECAUSE BY
DEFINITION GLIOBLASTOMAS DON'T
SPREAD OUTSIDE THE CENTRAL
NERVOUS SYSTEM SO WHAT IS THAT
MECHANISM? SO US AND OTHERS
WORKED OUT THAT GLIOBLASTOMAS
LIKE MANY OTHER CANCERS RELEASE
VESICALES CALLED EXTRA CELLULAR
VESICLES. AND ONCE THOSE ARE
RELEASED THE EXTRA CELLULAR
VESICLES RELEASED FROM THE
TUMOR, BUT NOT TUMOR CELLS
THEMSELVES ARE ABLE TO ESCAPE
THE CENTRAL NERVOUS SYSTEM,
ENTER THE SYSTEMIC CIRCULATION.
WHEN THEY DO THAT, THEN THEY ARE
ABLE TO CAUSE A WHOLE HOST OF
IMMUNE RELATED EVENTS. ONE
EVENT IMPLICATED IN THE
GLIOBLASTOMA MEDIATED
IMMUNOSUPPRESSION IS THIS
CONVERSION OF YOUR NAIVE
MONOCYTES INTO THESE MYELOID
DERIVED SUPPRESSER CELLS. THOSE
ARE THE MDSCs. MDSCs IN
TURN ARE ABLE TO AFFECT YOUR
T-CELLS IN TWO WAYS. ONE IS THEY
BLOCK PROLIFERATION AND
EXPANSION OF T-CELLS NECESSARY
FOR YOUR T-CELLS TO TARGET NEW
ANTIGENS, THEY NEED TO REPRODUCE
AND GROW AND MDSCs BLOCK THAT
AND FOR THE, T CELLS AROUND THEY
NEED THE ACTIVATE TO ATTACK THE
ANTIGENS THEY SEE AND THE
MDSCs ALSO BLOCK THAT PROCESS.
SO NOW WHEN THE MDSCs ARE
PRESENT YOU HAVE A LOW T-CELL
COUNT AND T-CELLS PRESENT DON'T
WORK. SO YOU GET THIS DOUBLE
CLAMMWHAMMY. SO THIS IS NOT
NECESSARILY UNIQUE. TO
GLIOBLASTOMA. SO THIS IS FROM A
PRIOR STUDY THAT WAS PUBLISHED
LOOKING AT HEPATOCELLULAR
CARCINOMA, THAT ULTIMATELY YOU
CAN SEE MDSCs GET PRODUCED.
VERY SIMILAR TO THE PRODUCTION
WE TALKED ABOUT PREVIOUSLY. BUT
AND THEN YOU CAN SEE ULTIMATELY
YOUR T-CELLS GET EXHAUSTED. AND
AGAIN, YOU CAN SEE THAT THAT IS
CONSIDERED TO BE MEDIATED BY
THIS PD 1 PDL 1 CHECK POINT, THE
SO CALLED IMMUNE CHECK POINTS
WHICH ARE THE TARGET OF THESE
IMMUNE CHECK POINT INHIBITOR
DRUGS. SO WHAT CAUGHT MY
ATTENTION WHEN I SAW THIS, WAS
THAT IT WAS THOUGHT THIS
MOLECULE HERE CCRK, CELL CYCLE
RELATED KINASE, CDK 20, BECAUSE
IT IS THOUGHT BASED ON
SEQUENCING TO BE IN THE CDC
FAMILY BUT SOMEHOW SITS ATOP
THAT PATHWAY. THAT ULTIMATELY
LEADS TO CONTEXT OF IL 6 IN
HEPATOCELLULAR CARCINOMA, DRIVES
T-CELL PRODUCTION AND EXHAUSTS
YOUR T-CELLS. SO I THOUGHT THAT
WAS SERENDIPITOUS BECAUSE YEARS
EARLIER I CLONED THE FIRST CCRK
MOUSE. SO WHAT WE WERE LOOKING
AT AT THAT TIME WAS
UNDERSTANDING PRIMARY CILIA IN
CONTEXT OF NEURAL DEVELOPMENT.
THIS IS A WILD TYPE MOUSE AT
EMBRYONIC DAY 12.5 COMPARED TO
CCRK KNOCK OUT. THERE ARE
FEATURES LIKE EXEANCEPHALY. YOU
CAN SEE POLYSYNDACTYLY AND
SKELETAL DEFECTS. THE MORAL OF
THE STORY, THOSE DEFECTS LOOK
VERY SIMILAR TO WHAT YOU SEE IF
YOU KNOCK OUT SONIC HEDGEHOG IN
A MOUSE. BUT WE DO KNOW THAT AS
I TOLD YOU HEDGEHOG SIGNALING IS
THE SORT OF PROTOTYPE PATHWAY
FOR WHICH PRIMARY CILIA WERE
KNOWN TO BE IMPORTANT. SO WE
DECIDED TO SAY WHAT HAPPENS TO
PRIMARY CILIA IN OUR CCRK KNOCK
OUTS SO YOU CAN SEE SCANNING
ELECTRON MICROSCOPY HERE,
IMMUNOFLUORESCENCE LOOKING AT R
13B WHICH IS SPECIFIC TO THAT
MICROTUBULE STRUCTURE OF PRIMARY
CILIA AS WELL AS GAMMA TUBULIN,
WHICH STAINS THE BASAL BODY
WHICH THE PRIMARY CILIA EMANATE.
YOU CAN SEE MORPHOLOGICALLY HOW
THEY ARE DIFFERENT. YOU CAN SEE
THE ROUNDED STRUCTURES HERE AND
THIS IS TEM ON THE OTHER SIDES.
AND IMPORTANTLY WHAT WE WERE
ABLE TO CONFIRM, SO I SHOWEDDED
YOU EARLIER HOW WHEN THE PRIMARY
CILIA ARE PRESENT AND HEDGEHOG
SIGNALING THE WORKING, THAT YOU
GET ACTIVATION OF THESE SO
CALLED GLIOMA PROTEINS. ONE
THICK YOU CAN DO AS READ OUT HOW
WELL THIS IS WORKING IS ASK
WELL, WHAT ABOUT THIS ACTIVATION
OF GLI 1. YOU CAN SO E IN THE
CONTEXT OF THE CRRK KNOCK OUT
YOU CAN'T GET ACTIVATION OF GLI
1 IN THE WAY YOU DO WILD TYPE.
SO WHAT WE FOUND IN A VERY
SEPARATE CONTEXT WAS THAT CCRK
IS IMPORTANT FOR CILIA
MORPHOLOGY AND BECAUSE OF ITS
ROLE IN CILIA MORPHOLOGY, WHEN
ITS LEVELS ARE CHANGED IT
AFFECTS FUNCTION OF PRIMARY
CILIA. SO MAX WORKED ON THIS A
WILD BACK WHERE WE WANTED TO
START EXPLORING IS THIS A CCRK
SPECIFIC THING OR IS THE ROLE OF
SAY CCRK ULTIMATELY LEADING TO
HIGH LEVELS OF IL 6, IS THAT
JUST SOMETHING THAT IS PRIMARY
CILIA BUT WE SEE THAT THROUGH
CCRK BECAUSE OF THE AFFECT ON
THE PRIMARY CILIA. SO WHAT WE
DID IS THIS EXPERIMENT WHERE WE
SAID OKAY, LET'S LOOK AT CCRK
KNOCK DOWNS, THIS IS A STABLE
ANTIVIRAL BASE KNOCK DOWN. WE
CAN LOOK AND SEE ARE PRIMARY
CILIAING WITH MADE. WE SEE THAT
THERE IS REDUCTION IN THAT
PRIMARY CILIA PHENOTYPE. WE CAN
ASK WHAT HAPPENS TO IL 6 LEVELS
WHEN WE DO THAT. MOREOVER SO YOU
CAN SEE HERE THAT WE ARE LOSING
THE IL 6. SORRY WE ARE LOSING
PRIMARY CILIA, WHEN WE DO ELISA
FOR IL 6 WITH CCRK IL 6 IS DOWN
QUITE A BIT SO CCRK DOES SEEM TO
HELP TO DRIVE IL 6 PRODUCTION AS
FOLKS STUDYING HEPATOCELLULAR
CARCINOMA FOUN FOUND. WHAT WE DD
HERE IS SAY WHAT IF WE TOOK TWO
OTHER PROTEINS THAT ARE KNOWN TO
BE SPECIFIC AND ESSENTIAL FOR
PRIMARY CILIA MORPHOLOGY, THAT
ARE UNRELATED TO CCRK? SO WE DID
THE CAI KNEE SIN MOTOR KIF 3A
THOUGHT TO BUILD THE -- IT IS A
FORWARD MOTOR THAT TAKES THE
MICROTUBULES AND STACKS THEM ON
TOP OF EACH OTHER SO YOU CAN
BUILD A PRIMARY CILIA. THEN IFT
88 IS ONE OF THE MANY INTRAFLAY
CELLULAR PROTEINS THAT ATTACHES
AND BRINGS THE CONSIDER GO UP
NEEDED TO BUILD PRIMARY SILL
YUM. SO THESE ARE VERY DIFFERENT
CLASSES OF MOLECULES UNRELATED
SO THE QUESTION WE ARE TRYING TO
PROBE IS, IS THIS REALLY A CCRK
THING OR IS THIS A CILIA THING
MORE BROADLY? SO WHAT YOU CAN
SEE IS REGARDLESS OF WHICH
METHOD WE USE TO BLOCK CILIA
FORMATION, IL 6 LEVELS GO DOWN
NONETHELESS. SO TO US THIS LOOKS
LIKE MORE OF A CILIA MECHANISM
AS OPPOSED TO SOMETHING THAT IS
CCRK. SO THEN WE CAN START TO
BUILD ON OUR MODEL AS WE GO
ALONG SO NOW WE KNOW THAT THE
PRIMARY CILIA ARE PRESENT AND
BECAUSE WE ALREADY KNOW THIS
DOWNSTREAM STEP WE TALKED ABOUT
WHERE THE EVs ARE FORMED, IL 6
ARE PROBABLY NECESSARY, IN THIS
PROCESS AS WELL TO CONVERT THESE
NAIVE MONOCYTES TO MDSC AND THE
REST YOU KNOW. SO THIS GIVES US
TESTABLE HYPOTHESES. SO BEFORE I
DO THAT, I WANT TO SHOW YOU THIS
IMAGE COMES FROM PATIENTS THAT
ARE RESECTED WITH GLIOBLASTOMA
WE CAN BRING TO THE LAB AND
STAIN. THESE ARE REAL THINGS
THAT YOU CAN SEE. SO WE HAVE
CONFOCAL MICROSCOPE IN THE LAB
AND YOU CAN SEE PERICENTRIN
WHICH STAINS THE BASAL BODY AND
YOU CAN SEE ACETYLATED ALPHA
TUBULIN. YOU CAN SEE THE
90-DEGREE RELATIONSHIP, YOU CAN
SEE THE EXINANENE GROWING OUT OF
THE CELL BODY WITH THE NUCLEUS
AND THAT IS WHAT I'M CALLING A
PRIMARY CILIUMM. WE TOOK A
SIMILAR APPROACH TO SAY IF WE
DID WHAT WE DID BEFORE FOR IL 6
YOU CANEE CCRK AND SEE THE CILIA
FREQUENCY, THE FREQUENCY IS
DOWN. SO THE QUESTION BECOMES WE
KNOW WHERE WE BLOCK CILIA IL 6
GOES DOWN BUT WHAT HAPPENS TO
THE MYELOID DERIVED SUPPRESSER
CELLS? ? DO THOSE GO DOWN? WHAT
YOU CAN SEE IS YES. WHEN YOU
BLOCK THE PRIMARY CILIA
REGARDLESS OF THE METHOD USED TO
BLOCK IT, YOU CAN SEE A HUGE
REDUCTION IN THE MDSC
POPULATION. HERE WE ARE USING
RADIATION BECAUSE WE KNOW THAT
WHEN YOU TAKE THESE CELLS, THESE
BRAIN TUMOR CELLS YOU RADIATE
THEM YOU CAN SEE AT BASELINE IN
THE CONTROL GROUP A BIG
INDUCTIONS IN THE MDSC
POPULATION. SO EASIER TO STUDY
AND SEE THE DIFFERENCE IF THERE
IS ONE. SO THAT'S WHY WE DID
THAT OVER HERE. YOU CAN SEE
THERE'S HUGE REDUCTION IN THAT
UPREGULATION OF MDSCs IN
RESPONSE TO TUMOR CELLS. THERE
IS A REDUCTION IN THAT IF YOU
BLOCK CILIA REGARDLESS HOW YOU
DO THIS GENETICALLY. THEN
IMPORTANTLY THEN, IF WE SAY THAT
THE MDSCs ARE SUPPOSED TO
BLOCK THE T-CELLS, IN IN TERMS
OF PROLIFERATION OF FUNCTION WE
CAN THEN DO -- WE CAN THEN DO
ASSAYS TO ASK ARE THE LEVELS OF
THE T-CELLS RESTORED? SO AGAIN
YOU CAN SEE SAME IDEA HERE THAT
IN THE CONTROL SETTING YOU ADD
THE TUMOR, T-CELLS PROLIFERATION
FALLS OFF BUT IF YOU BLOCK CILIA
T-CELL PROLIFERATION IS
RESTORED, DOESN'T MATTER WHETHER
CCRK OR UNRELATED STRUCTURAL
NON-KINASE PROTEIN. SO THEN THE
QUESTION WAS IS THIS ACTUALLY
MEDIATED THROUGH IL 6? IS THIS
HAPPENING BECAUSE IL 6 IS LOW
AND WHAT HAPPENS IF WE BLOCK THE
CILIA BUT ACTUALLY RESTORE IL 6
LEVELS? SO THIS IS EXACTLY WHAT
WE ARE TRYING TO DO HERE. THE
MORAL OF THIS STORY IS THAT EVEN
FOLLOWING BLOCKADE OF CILIA, IF
YOU RESTORE WITH EXOGENOUS IL 6
YOU CAN TO SOME DEGREE
CIRCUMVENT THAT REDUCTION IN THE
MDSCs AND EVEN IN ABSENCE OF
CILIA, RESTORING IL 6 CAN
INCREASE YOUR MDSC LEVELS NOT
QUITE BACK TO NORMAL BUT
INCREASE THEM. UP HERE AS I
SHOWED YOU, ULTIMATELY PDL 1 IS
THE CONNECTOR PD 1 AND PDL 1 IS
THE CONNECTOR TO ALLOW MDSCs
TO HAVE THE IMPACT ON T-CELLS SO
WE ARE TRYING TO UNDERSTAND AND
WE HAVE SHOWN PREVIOUSLY IN
PRIOR PUBLICATION IF YOU BLOCK
PDL 1 MDSC LEVELS DECREASE AS
WELL SO WE WANTED TO UNDERSTAND
WHAT HAPPENS IF YOU ARE ALSO
BLOCKING PDL 1 AS YOU ARE
BLOCKING IL 6. YOU CAN SEE THAT
THAT COMBINATION LEADS TO THE
LOWEST REDUCTION HERE IN THE
MDSC LEVELS. ALL RIGHT. SO THIS
IS ALL COOL BUT SO WHAT? IS THIS
SOMETHING THAT'S RELEVANT TO THE
PATIENT WE ARE LOOKING AT IN
THIS STUDY? AND THE ANSWER IS
YES. SO WHAT WE DID HERE, WE
WENT TO THE BLOOD BANK HERE, AND
WERE ABLE THE GET BLOOD THEN
COMPARE THIS TO BLOOD THAT I
TAKE FROM PATIENTS IN THE
OPERATING ROOM. AND ASK WELL,
WHAT IF WE LOOKED FOR MDSCs
USING FLOW CYTOMETRY? ARE WE
SEEING THESE MDSCs IN THE
BLOOD OF OUR PATIENT?
THIS IS SYSTEMIC BLOOD, NOT
BLOOD FROM THE BRAIN. YOU CAN
SEE IN OUR PATIENT THERE IS IS
ACTIVE MDSCs THAT ARE PRETTY
ABUNDANT AND MEASURABLE. THE
OTHER QUESTION, WHAT ABOUT THE
MECHANISMS AND THESE MACHINERY
SUCH AS KIF 3 AND CCRK, CAN WE
SEE THOSE IN THE TUMOR?
THIS THE IMMUNOHISTOCHEMISTRY
DONE BY PJ, THAT WE CAN ACTUALLY
SEE THESE MACHINE AND SAY THEY
ARE THERE AND THEY ARE PRESENT
AND WE HAVE THIS THAT IS
INCREASED. SO WITH THAT BEING
SAID, WE THEN SAID LET'S LOOK AT
A MOUSE MODEL OF THE DISEASE. T
SO DISEASE. SOIF YOU TAKE -- YOE
WITH IMMUNE SYSTEM TO STUDY
IMPACT OF TUMOR ON IMMUNE
SYSTEM. SO WE HAVE A MOUSE MODEL
OF GLIOBLASTOMAS USING GL 261
CELLS AND WE CAN DO ORTHO TOPIC
TRANSPLANTATION INTO THE MOUSE.
SO WE TAKE THE BRAIN TUMOR
CELLS, INJECT INTO THE MOUSE.
WHEN WE DO THAT YOU CAN SEE JUST
-- THIS IS THE MOUSE WITH NO
TUMOR JUST A SHUNT SURGERY, SO
WE NORMALIZE THAT TO ONE HERE
BUT WHEN WE INJECT TUMOR CELLS
YOU CAN SEE THIS MDSC
SPECIFICALLY THIS MONOSITIC MDSC
POPULATION GOES UP OVER TWOFOLD.
BUT IF IN THE CELLS YOU ACTUALLY
BLOCK ANY ONE OF THOSE
COMPONENTS NECESSARY FOR PRIMARY
CILIA FORMATION, PRIOR TO THE
IMPLANTATION, COMPARED TO A
SCRAMBLE CONTROL YOU CAN SEE THE
MDSC POPULATION GOES DOWN.
CONSISTENT WITH THAT, WHAT WE
SEE IS THAT THE T-CELLS IN THE
MICE THAT HAVE TUMORS WITH A
SCRAMBLE CONTROL IS DECREASE BUT
IF YOU PRE-BLOCK PRIMARY CILIA
BEFORE IMPLANTATION THAT DOES
NOT OCCUR. SO WHAT WE ARE SEEING
IN VITRO MATCHES IN VIVO
FINDINGS HERE THAT JUST BLOCKING
THE PRIMARY CILIA IS ENOUGH TO
ACTUALLY RESTORE T-CELL FUNCTION
IN THE ANIMALS BY REDUCING THAT
MDSC POPULATION. SO THAT WAS IN
BONE MARROW. WE SEE VERY SIMILAR
THING IN THE SPLEEN. THAT I'M
NOT SHOWING HERE IS IN PATIENTS,
US AND OTHERS HAVE ALSO
PUBLISHED THAT IN A PATIENT WITH
ADVANCE GLIOBLASTOMAS, THE
THYMUS IS HYPO PLASTIC BECAUSE
THEY LOSE HAIR T-CELLS SO ONE OF
THE THINGS WE ALSO DID WAS LOOK
TO SEE IN THESE ANIMALS WHERE WE
ARE BLOCKING PRIMARY CILIA, DO
YOU SEE RESTORATION OF T-CELLS
IN THE THYMUS. AND WE DO. SO
I'M GOING TO TRY TO GET THROUGH
THE REST OF THIS RELATIVELY
QUICKLY. SO THE QUESTION HERE
FOR US WAS WELL WHY DOES THE
PRIMARY CILIA HAVE THAT ROLE? SO
PEOPLE HAVE STUDIED THAT, WELL,
YOU KNOW IF YOU LOOK AT PRIMARY
CILIA, THERE ARE VESICALES THAT
BUTT OFF OF THEM. I'M SHOWING
YOU THAT IN THE CARTOON AND ALSO
IN THAT HERE IN A T E M IMAGE OF
LONGITUDINAL SECTION THROUGH
PRIMARY CILIA. SO THIS IS THE
XENIMO BRAIN HERE, THE EXANINE
MICROTHANK YOU MULE, YOU CAN SEE
THE ECTOSOME BUDDING OFF. SO ARE
THESE EXTRA CELLULAR VESICLES WE
THINK SO IMPORTANT FOR
IMMUNOSUPPRESSION IN CANCER,
WHERE THESE VESICLE IT IS SAME
THAT ARE BUDDING FROM OUR
PRIMARY CILIA? SO THERE ARE WAYS
TO STUDY THAT AND SO THIS IS
JUST A CARTOON, THE VESICALES
TYPICALLY ARE MADE OF THESE
TETRA SPANIN PROTEINS THAT SPAN
THE MEMBRANE. AND WE CAN TEST
FOR THOSE. WHAT WE SEE FIRST IS
THAT CD 9 AND CD8 1 ARE TWO WELL
KNOWN TETRA SPANIN PROTEINS,
SPECIFIC FOR EVs WE CAN SEE
THAT REGARDLESS OF WHETHER WE
HAVE KNOCKED DOWN ANY OF THESE
THREE PROTEINS THAT WOULD
OTHERWISE MAKE PRIMARY CILIA
DISAPPEAR WE ARE STILL SEEING CD
9 AND CD8 1. MORE PROOF IS THAT
YOU CAN DETECT THE VESICLES BY
TRANSMISSION ELECTRON MICROSCOPY
SO THEY ARE THERE. SO WE CAN
THEN DO NANOPARTICLE ANALYSIS
WHICH I CAN'T SEEM TO GET VIDEO
TO PLAY BECAUSE I THINK THIS
THAT THE WAY THIS CURSOR IS. SO
BELIEVE ME, THESE LITTLE WHITE
DOTS ON THE SLIDE ARE THE
NANOPARTICLES VESICALES THAT ARE
100 NANOMETERS AND WHAT THIS IS
MEASURING IN THE GRAPHS ARE BOTH
THE SIZE DISTRIBUTION AND THE
CONCENTRATION DISTRIBUTION AND
TO OUR SURPRISE, BLOCKING THE
CILIA AFFECTED NONE OF THESE SO
SOMEHOW THE VESICALES ARE NOT
NECESSARILY THE VESICALES COMING
OFF THE CILIA, IF YOU MOVE THE
CILIA, THE VESICALES ARE STILL
THERE, CONCENTRATIONS DON'T
CHANGE AND SO WE FOUND THAT VERY
INTERESTING. SO WE ALREADY KNEW
I WENT OVER THAT PAPER THAT MAX
PUBLISHED, THAT SHOWED THAT WHEN
YOU BLOCK CILIA IL 6 GOES DOWN.
SO THEN WE THOUGHT WELL, IL 6
GOES DOWN MDSC IS DOWN AND WE
KNOW T-CELLS ARE COMING BACK SO
WHAT WE ALSO ANTICIPATED HERE
WAS THAT PDL 1, THE PROTEIN
BEING TARGETED WITH ALL THESE
BIG POWERFUL DRUGS LIKE ETRUDA
ALSO GO DOWN. SO WHAT WE FOUND
PARADOXICALLY IS THAT WHEN YOU
BLOCK THE CILIA PDL 1 LEVELS GO
UP. SO EVEN THOUGH THIS IS THE
PROTEIN THAT'S THOUGHT TO
MEDIATE THIS BLOCKAGE OF YOUR
T-CELLS AND WE CAN COMPLETELY
PREVENT THE BLOCKAGE OF THE
T-CELLS BY BLOCKING CILIA, PDL 1
LEVELS GO UP. SO WE CAN LOOK
THROUGH THIS MORE AND SAY WELL
IS THIS RELATED TO INTERFERON
GAMMA SIGNALING, I DIDN'T TALK
ABOUT INTERFERON GAMMA TODAY BUT
I DID TALK ABOUT INTERLEUKIN 6
SIGNALING SO INTERFERON GAMMA
AND INTERLEUKIN 6 WORK THROUGH
STAT 1 AND 3 PATHWAYS SO B WHAT
YOU CAN SEE IS WHEN YOU BLOCK
CILIA, BOTH INTERLEUKIN 6 AND
INTERFERON GAMMA PATHWAYS GO
DOWN. SO ONE THING THIS TOLD US
WAS THAT WELL, WE ARE SEEING
PROTEIN CHANGES WHEN WE BLOCK
THE CILIA, NOT WHAT WE PREDICTED
AND WE WILL TALK A LITTLE BIT
ABOUT THAT LATER. BUT MAYBE
POSSIBLE THAT WHILE THE PRIMARY
CILIA ARE IMPORTANT FOR
NECESSARILY RELEASING THE
VESICLES THEY MAY BE PERFORM FOR
ALTERING THE CONTENT OF THE
VESICALES WHICH THEN MIGHT
CHANGE WHAT HAPPENS WHEN THEY GO
OUT INTO THE SYSTEMIC
CIRCULATION. SO WHAT WE DID IS
BLOCKED THE PRIMARY CILIA, THEN
WE DID PROTEOMIC SCREEN LOOKING
AT THE NANOPARTICLES THEMSELVES,
NOT THE CELLS, BUT WHAT IS THE
ACTUAL CONTENT IN THE
NANOPARTICLES FOLLOWING THIS
BLOCKADE OF PRIMARY CILIA IN THE
CELLS FROM WHICH THEY CAME. AND
I THIS I WHAT SHOULD BE VERY
CLEAR TO ANYBODY LOOKING AT THIS
IS THAT THE CONTENT OF THE
VESICALES ARE VERY DIFFERENT. I
HIGHLIGHTED HERE ICAM 1 BECAUSE
THAT IS A PROTEIN INTRACELLULAR
ADHESION MOLECULE 1 THAT'S
IMPLICATE IN SYSTEMIC CANCERS Z
A MEDIATOR OF IMMUNOSUPPRESSION.
SO WE SHOWED NOT ONLY SEE THAT
IN THE PROTEOMIC SCREEN BUT WHEN
WE DO A FLOW CYTOMETRIC SCREEN
ICAM 1 ON SURFACE OF VESSELS ARE
DOWN SO WE CAN CONFIRM THAT. SO
IN SUMMARY HERE, WE HAVE MORE
NUANCED VIEW NOW OF HOW
IMMUNOSUPPRESSION MIGHT WORK IN
GLIOBLASTOMA. SO FIRSTLY, IT IS
THE TUMOR IS CONFINED TO CENTRAL
NERVOUS SYSTEM. BUT IT IS ABLE
TO HAVE SYSTEMIC EFFECTS BECAUSE
IT RELEASES THE VESICALES. WHILE
THE VESICLES ARE NOT FROM PRIMA
CILIA, IT APPEARS THE SIGNALING
MAP IS MEDIATED THROUGH PRIMARY
CILIA, CLANGS THE CONTENTS OF
THOSE VESICLES IN A MANNER
DEPENDENT ON IL 6 SIGNALING, AND
ALLOWS FOR THAT CONTENT TO BE
SUCH THAT WHEN IT GETS THE BONE
MARROW THE SPLEEN, ET CETERA, IT
HAS IMMUNOSUPPRESSIVE EFFECT.
AND I THINK THAT THE BIG THING
FROM THIS IS THAT BILLIONS OF
DOLLARS HAS GONE INTO CREATING
IMMUNE CHECK POINT INHIBITORS
FOR THE SOLE PURPOSE OF REDUCING
CANCER MEDIATED
IMMUNOSUPPRESSION. AND WHAT WE
FOUND WAS THAT IF YOU HAVE EVEN
INCREASE IN PDL 1 IF YOUR
PRIMARY CILIA ARE NOT THERE THE
PDL 1 SIGNALING DOESN'T WORK.
BUT THE COROLLARY TO THAT IS
BLOCKING PDL 1 BY ITSELF IN
CONTEXT OF GLIOBLASTOMA MIGHT
NOT BE SUFFICIENT BECAUSE THERE
IS A PARALLEL PATHWAY. THAT BOTH
OF WHICH NEEDS TO BE INHIBITED.
SO THAT MIGHT EXPLAIN A LOT OF
THE UNSUCCESSFUL CLINICAL TRIALS
WE HAVE SEEN OF IMMUNE CHECK
POINT INHIBITORS IN THE CONTEXT
OF GLIOBLASTOMAS, THEY HAVE ALL
FAILED AND WE HAVE IDEAS WHY
THAT IS. SO AGAIN, GLIOBLASTOMA
IS A BAD DISEASE. I WANT
TRAINEES TO WALK AWAY FROM HERE
WITH THAT AND WE NEED LOT
SMARTER PEOPLE THAN ME TO BE
STUDYING THIS. PRIMARY CILIA ARE
IMPORTANT IN PATHOGENESIS OF THE
DISEASE, BECAUSE OF ALL
SIGNALING IT MEDIATES BUT ALSO
IN TERMS OF THE WAY IT INTERACTS
WITH THE IMMUNE SYSTEM. AGAIN,
PDL 1 IN IL 6 STAT 3 PATHWAY,
AND THE MEDIATION OF THOSE
PATHWAYS THROUGH PRIMARY CILIA
MIGHT BE PARALLEL AND YOU MIGHT
NEED A NUANCED STRATEGY TO GET
SOME OF THESE IMMUNE CHECK
POINTS TO WORK. OF COURSE THIS
IS A SNAP SHOT OF A LOT OF
THINGS, NONE WOULD BE POSSIBLE
WITHOUT A LOT OF PEOPLE. WE HAVE
LESS THAN A MINUTE BUT I WILL
TAKE ANY QUESTIONS OR COMMENTS
YOU HAVE NOW. THANKS.
[APPLAUSE]
>> REMIND TORE THE ONLINE
VIEWERS -- REMINDER TO THE
ONLINE VIEWERS ANY QUESTIONS
SUBMIT TO LIVE FEEDBACK BUTTON.
AND IF WE DON'T GET TO THEM WE
CAN ANSWER LATER BY EMAIL. THANK
YOU AGAIN TO OUR WONDERFUL
PRESENTERS.
>> THANK YOU.
[APPLAUSE]