Welcome to the Clinical Center Grand Rounds,
a weekly series of educational lectures for physicians and
health care professionals broadcast from the Clinical
Center at the National Institutes of Health in
Bethesda, MD.
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TODAY WE ARE HONORED TO HAVE TWO
SURGEON SCIENTISTS FROM THE
NATIONAL INSTITUTE ON DEAFNESS
AND OTHER COMMUNICATION
DISORDERS.
DR. MICHAEL HAO, PRINCIPAL
INVESTIGATOR FOR THE AUDITORY
DEVELOPMENT AND RESTORATION
PROGRAM AND ASSOCIATE PROFESSOR
IN THE DEPARTMENT OF
OTOLARYNGOLOGY AT GEORGETOWN
UNIVERSITY AND DR. WADE CHIEN,
PROFESSOR IN THE DEPARTMENT OF
OTOLARYNGOLOGY AT JOHNS HOPKINS.
OUR FIRST SPEAKER IS DR. HAO.
DR. HAO RECEIVED HIS BACHELOR'S
AND MEDICAL DEGREE FROM BOSTON
UNIVERSITY.
HE THEN COMPLETED HIS
OTOLARYNGOLOGY HEAD NECK SURGERY
RESIDENCY TRAINING AT THE WAYNE
STATE UNIVERSITY FOLLOWED BY A
NEURON WILLING TO FELLOWSHIP IN
LOS ANGELES.
DR. HAO ALSO PURSUED A T32
USC/HRI HEARING AND
COMMUNICATIONS NEUROSCIENE
POSTDOCTORAL RESEARCH TRAINING
FELLOWSHIP PRIOR TO JOINING
NIDCD.
HE JOINED THE NIDCD'S
OTOLARYNGOLOGY SURGEON SCIENTIST
PROGRAM IN 2013.
IN ADDITION TO HIS ROLE AS
ASSOCIATE PROFESSOR THE A
GEORGETOWN, DR. HAO ALSO SERVES
AS MEDICAL DIRECTOR FOR THE
MEDSTAR GEORGETOWN UNIVERSITY
HOSPITAL COCHLEAR IMPLANT
PROGRAM.
DR. HAO'S LAB IDENTIFIES NEW
TREATMENTS FOR HEARING
INSTABILITY DISORDERS, WHICH
INCLUDE MOST NOTABLY MENIERE'S
DISEASE, IDEOPATHIC SUDDEN
NEURAL CENTER HEARING LOSS AND
AUTOIMMUNE EAR DISEASE.
HIS LABORATORY USES A
COMBINATION OF MICROBIOLOGY AND
BIOINFORMATICS INCLUDING SINGLE
KREM MRNA SEQUENCING UTILIZING
ADULT MAMMALIAN SYSTEMS.
DR. HAO HAS AUTHORED OR
CO-AUTHORED OVER 60 PUBLICATIONS
AND BOOK CHAPTERS.
DR. HAO IS THE CHAIRMAN OF THE
RESEARCH COMMITTEE FOR THE
AMERICAN COCHLEAR IMPLANT
ALLIANCE, AND IS A MEMBER OF
NUMEROUS MEDICAL SOCIETIES
INCLUDING THE AMERICAN
NEUROONTOLOGY SOCIETY, THE
AMERICAN ACADEMY OF
OTOLARYNGOLOGY, HEAD NECK
SURGERY, THE ASSOCIATION FOR
RESEARCH IN OTOLARYNGOLOGY, THE
NORTH AMERICAN SKULL BASE
SOCIETY AND DR. HAO IS BOARD
CERTIFIED IN OTOLARYNGOLOGY HEAD
AND NECK SURGERY, OWE WILLING TD
NEUROONTOLOGY.
OUR SECOND SPEAKER IS DR. WADE
CHIEN, NIDCD, AND DEPARTM PROVEE
JOHNS HOPKINS SCHOOL OF
MEDICINE.
DR. CHIN RECEIVED HIS BACHELOR'S
IN BIOLOGY AND MUSIC FROM
BRANDEIS UNIVERSITY, AND THEN
EARNED HIS MEDICAL DEGREE AT THE
UNIVERSITY OF SOUTHERN
CALIFORNIA, KECK SCHOOL OF
MEDICINE.
HE COMPLETED RESIDENCY TRAINING
AT THE HARVARD COMBINED
OTOLARYNGOLOGY TRAINING PROGRAM
AND A NEUROONTOLOGY FELLOWSHIP
AT JOHNS HOPKINS HOSPITAL.
ALUMNUS OF THE HOWARD HUGHES
MEDICAL INSTITUTE NIH RESEARCH
SCHOLARS PROGRAM, DURING WHICH
HE WORKED WITH DR. GERALD
FISHBACH.
ALSO RECIPIENT OF A T32 TRAINING
GRANT IN RESIDENCY.
DR. CHIEN JOINED NIDCD AS AN
INVESTIGATOR IN THE
OTOLARYNGOLOGY SURGEON SCIENTIST
PROGRAM IN 2011 AND NOW HIS LAB
STUDIES THE APPLICATION OF INNER
EAR GENE THERAPY AS A TREATMENT
FOR HEARING LOSS AND DIZZINESS.
A NATIONALLY AND INTERNATIONALLY
RECOGNIZED SPEAKER, DR. CHIEN IS
A DIPLOMAT FOR THE AMERICAN
BOARD OF OTOLARYNGOLOGY AND
AMERICAN BOARD OF NEUROONTOLOGY
SKULL BASE SURGERY.
HE IS A MEMBER OF THE AMERICAN
OAT LOGICAL SOCIETY AND A FELLOW
OF THE -- AND SERVES ON THE
COMMITTEES OF NUMEROUS NATIONAL
PROFESSIONAL NEURON WILLING TO
ORGANIZATIONS.
SO NOW PLEASE WELCOME OUR
SPEAKERS, DR. HAO AND DR. CHIEN,
FOR THEIR TALKS ENTITLED HEARING
INSTABILITY DISORDERS, MOUSE AND
HUMAN CORRELATES, AND THE
DEVELOPMENT OF GENE THERAPY --
INNER EAR GENE THERAPY FOR USHER
SYNDROME.
>>THANK YOU, EVERYONE, FOR
YOUR ATTENTION TODAY.
SO I HAVE NO DISCLOSURES.
AND THESE ARE THE LEARNING
OBJECTIVES FOR TODAY.
I HOPE THAT I CAN LEAVE YOU WITH
SOME SENSE FOR AN AWARENESS OF
HEARING INSTABILITY DISORDERS
INCLUDING MENIERE'S DISEASE, AND
TO GIVE YOU A SENSE FOR THE
DIAGNOSTIC UTILITY OF MRI IN
DIAGNOSING ENDO LYMPHATIC
HYDROPS, ONE OF THE HISTOLOGICAL
HALLMARKS FOR THIS DISEASE.
SO THIS IS A BRIEF OUTLINE OF
WHAT I'M GOING TO BE TALKING
ABOUT TODAY.
AND I'LL GET RIGHT INTO IT.
BUT TAKING A STEP BACK, HEARING
INSTABILITY DISORDERS ARE
DISEASES WHERE PATIENTS
EXPERIENCE EITHER FLUCTUATION IN
OR SUDDEN CHANGES IN THEIR
HEARING, AND OPERATIONALLY, WE
DEFINE THIS AS EITHER A
20-DECIBEL CHANGE OVER ONE OR
MORE FREQUENCIES OR A 15-DECIBEL
CHANGE AT TWO OR MORE
FREQUENCIES.
THIS IS KIND OF BASED ON THE
AMERICAN SPEECH AND HEARING
ASSOCIATION'S DEFINITION OF
SIGNIFICANT CHANGE IN HEARING.
SO MY LAB'S MISSION IS TO FIND
TREATMENTS FOR HEARING
INSTABILITY DISORDERS, AND ONE
OF THE MAIN HYPOTHESES OF MY LAB
IS THAT ION HOMEOSTATIC
DYSFUNCTION CONTRIBUTES TO
HEARING INSTABILITY.
WE'VE USED THE COMBINATION OF
MOUSE MODELS AND TRANSCRIPTOME
STUDIES OF THE STRIA VAST CHAIRS
TVASCULARIS,AND WE'VE ALSO -- OT
WORK INVOLVES BOTH AN EFFORT TO
LINK ION HOMEOSTATIC DYSFUNCTION
WITH HUMAN HEARING INSTABILITY,
AND A DEEP PHENOTYPING CLINICAL
PROTOCOL FOR THESE PATIENTS WITH
AN AIM IN THE FUTURE TO IDENTIFY
REPURPOSEFUL AND NOVEL
THERAPEUTICS FOR THESE DISEASES.
SO HEARING INSTABILITY
DISORDERS, WHICH INCLUDE MOST
COMMONLY MENIERE'S DISEASE AND
IDEOPA TICK SUDDEN SENSORINEURAL
HEARING LOSS AMONG OTHERS,
REMAIN INCOMPLETELY UNDERSTOOD,
AND REALLY HAVE FEW IF ANY
EFFECTIVE TREATMENTS FOR HEARING
LOSS.
NOW, HERE YOU SEE AN EXAMPLE OF
A PATIENT WITH MENIERE'S
DISEASE, AND THEY HAVE A
HISTOLOGICAL HALLMARK CALLED
ENDO LYMPHATIC HYDROPS, THE ENDO
LYMPH CONTAINING COMPARTMENT OF
THE COCHLEA.
THIS IS IN DISTINCTION TO WHAT A
NORMAL COCHLEA WOULD LOOK LIKE
IN TERMS OF THAT CENTRAL
COMPARTMENT, WHICH IS CALLED THE
SCALA MEDIA SO IT LOOKS MORE
TRIANGULAR.
WE DON'T KNOW WHETHER THIS IS --
IT REMAINS KIND OF A POINT OF
ACTIVE DEBATE ABOUT WHETHER THIS
IS AN ACTUAL CAUSE OF THE
DISEASE OR WHETHER IT'S A
REFLECTION OF SOME UNDERLYING
PROCESS.
REGARDLESS, IT'S THOUGHT THAT
THERE'S SOME DYSFUNCTIONAL ENDO
LYMPH HOMEOSTATIC PROCESS THAT'S
OCCURRING IN THE COCHLEA THAT'S
SOMEHOW CONNECTED WITH THESE
DISEASES.
AND FURTHERMORE, STRIA
VASCULARIS PATHOLOGY HAS BEEN
NOTED IN THESE DISEASES, AND
HERE IS JUST AN EXAMPLE OF KIND
OF THIS ABNORMALLY THIN STRIA
VASCULARIS, YOU SEE A THICKER
ONE IN AN AGE MATCHED NORMAL
HEARING INDIVIDUAL.
IT'S BEEN SHOWN THAT THE STRIA
IN THESE DISEASES IS MORE
ATROPHIED, IN PARTICULAR IN
MENIERE'S DISEASE VERSUS KIND OF
AGE MATCHED CONTROLS.
SO MENIERE'S DISEASE, A MORE
COMMON FORM OF HEARING
INSTABILITY, IS BROADLY
CHARACTERIZED BY E EPISODIC
VERTIGO, SENSORINEURAL HEARING
LOSS AND FLU FLUCTUATING SYMPTOS
SUCH AS TINNITUS, FULLNESS, EAR
PRESSURE OR HEARING LOSS.
THE DIAGNOSTIC CRITERIA
SEPARATES MENIERE'S DISEASE INTO
TWO CATEGORIES: DEFINITE AND
PROBABLE MENIERE'S DISEASE.
THE PRIMARY DISTINCTION IS THAT
IN DEFINITE MENIERE'S DISEASE,
THERE'S AN AUDIOMETRICLY
DOCUMENTED LOW TO MEDIUM
FREQUENCY SENSORINEURAL HEARING
LOSS IN AT LEAST ONE EAR ON AT
LEAST ONE OCCASION BEFORE,
DURING AND AFTER AN EPISODE OF
VERTIGO.
SO THIS DISEASE IN PARTICULAR
REMAINS VERY MUCH A CLINICAL
DIAGNOSIS, BUT THESE ARE
DISEASES WITH KIND OF FAIRLY
CHALLENGING -- THAT ARE FAIRLY
CHALLENGING IN TERMS OF HEARING
LOSS AND THE TREATMENT OF THAT.
SO TO POINT YOU TO THE STRIA
VASCULARIS FUNCTION, THE STRIA
IS COMPOSED OF THREE MAIN LAYERS
OF CELLS INCLUDING MARGINAL,
INTERMEDIATE, AND BASAL CELLS,
AS WELL AS RARER CELL TYPES.
JUST TO TAKE A STEP BACK, THE
STRIA VASCULARIS IS HOUSED IN
THE LATERAL WALL OF THE COCHLEA,
AND IT FUNCTIONS TO GENERATE
WHAT'S CALLED THE ENDO COCHLEAR
POTENTIAL AS WELL AS TO REGULATE
ION CONCENTRATION HERE, AND
WITHOUT IT, THE HAIR CELLS WOULD
NOT BE CAPABLE OF FUNCTIONING
AND IT ESSENTIALLY SERVES AS A
BATTERY.
SO THIS THREE-LAYERED STRUCTURE
IS COMPOSED OF THESE MARGINAL
CELLS WHICH FACE THE END LINTH H
FOLLOWED BY THE INTERMEDIATE
CELLS HERE IN GREEN FOLLOWED BY
THE BASAL CELLS HERE IN BLUE.
NOTICE IN THIS INTERMEDIATE CELL
LAYER, THERE ARE ALSO
ENDOTHELIAL CELLS, PERICYTES AND
THERE ARE ALSO MACROPHAGES HERE.
SO THESE THREE LAYERS WORK
TOGETHER TO GENERATE THE
ENDOCOCHLEAR POTENTIAL AND
HOMEOSTEA TIS AND IT REALLY
REMAINS TO BE DEFINED ABOUT
WHETHER CHANGES IN THE STRUCTURE
OR THE FUNCTION OF THIS TISSUE
RESULTS IN HEARING INSTABILITY.
BUT IT IS A GOOD PLACE TO START
TO UNDERSTAND HOW ION
HOMEOSTASIS OCCURS IN THE
COCHLEA.
SO WHILE ITS CAUSE REMAINS
UNKNOWN, MENIERE'S DISEASE HAS
BEEN ASSOCIATED WITH SEVERAL
GENETIC VARIANTS OVER TIME, AND
WITH THIS IN MIND, WE
SYSTEMATICALLY REVIEWED THE
LITERATURE ON GENETIC VARIANTS
IN MENIERE'S DISEASE.
AND CROSS-REFERENCED THESE WITH
SINGLE CELL RNA SEQUENCING DATA
GENERATED BY MY LAB.
WHAT I SEE HERE IS A HEAT MAP OF
THE MAJOR STRIAL CELL TYPES
ARRAYED HORIZONTALLY, SO THE
MARGINAL CELLS IN RED,
INTERMEDIATE CELLS IN GREEN AND
BASAL CELLS IN BLUE HERE.
AND THE GENES ASSOCIATED WITH
THE GENETIC VARIANTS CONNECTED
TO MENIERE'S DISEASE ARE ALONG
THE VERTICAL ACCESS.
WHAT WE FOUND IS A SIGNIFICANT
SUBSET ARE EXPRESSED BY MARGINAL
AND INTERMEDIATE CELLS.
NOW, THIS IS CIRCUMSTANTIAL
EVIDENCE FOR A POTENTIAL ROLE OF
THESE CELL TYPES IN MENIERE'S
DISEASE.
SO WITH THAT BACKGROUND, I'M
GOING TO GO INTO THE FIRST
PROJECT.
WHICH IS THE CLINICAL PROTOCOL
INVOLVING DEEP PHENOTYPING OF
PATIENTS WITH THESE DISORDERS.
SO THIS IS AN ACTIVELY
RECRUITING PROTOCOL HERE, AND
THE GOAL IS TO UTILIZE DEEP
PHENOTYPING MEASURES TO
EVENTUALLY IDENTIFY NOVEL
THERAPIES FOR THESE PATIENTS.
DEEP PHENOTYPING IN THIS CASE
INVOLVES THE USE OF CONTRAST
ENHANCED DELAYED FLAIR MRI,
AUDIO NE TAUDIOMETRIC AND VESTIR
MEASURES AND IMMUNOPHENOTYPING.
THIS IS A LONGITUDINAL STUDY
THAT IS DESIGNED TO TRY TO
IDENTIFY GROUPS OF PATIENTS THAT
MIGHT HAVE COMMON FEATURES.
AND ONE OF OUR CHALLENGES IN
THIS PATIENT POPULATION IS, IT'S
THOUGHT THAT PATIENTS WITH THESE
DISEASES PROBABLY CONSTITUTE
SEVERAL DIFFERENT DISORDERS, SO
THIS PROTOCOL IS DESIGNED TO TRY
TO HELP SORT THAT OUT.
SO AS I MENTIONED, ONE OF THE
HALLMARKS OF THIS DISEASE IS
SOMETHING CALLED END LYMPHATIC
HYDROPS.
IT'S REALLY AN EXPANSION OF THE
ENDO LYMPH CONTAINING SPACES OF
THE INNER EAR, SPECIFICALLY OF
THE VESTIBULE HERE IN GREEN, AND
THE COCHLEA HERE IN YELLOW.
SO YOU CAN SEE THAT EXPANSION.
PREVIOUSLY WE HAD REALLY ONLY
BEEN ABLE TO DO THIS BY
EXAMINING TEMPORAL BONES THAT
HAD BEEN DONATED TO US AFTER
PATIENTS HAD DIED.
BUT NOW THE USE OF CON TRANS
ENHANCED FLAIR MRI INVOLVES
GADOLINIUM INJECTIONS AND
WAITING FOR ABOUT 4 TO 8 HOURS
AFTER THE INJECTION.
THIS ALLOWS THE GADOLINIUM TO
FILL AND THAT'S DEMONSTRATED BY
THE WHITE SPACES ON THIS FLAIR
MRI.
THIS IS AN AXIAL FLAIR, DELAYED
FLAIR IMAGE OF A PATIENT WITH
RIGHT-SIDED MENIERE'S DISEASE,
SO THE WHITE SPACES ARE THE
PERILYMPH AND THE DARK SPACES
WHAT YOU SEE WITH THE RED ARROWS
ARE THE END LYMPH.
NORMALLY THE ENDOLYMPH IS HARD
TO SEE SO WHEN YOU ACTUALLY SEE
THESE DARK SPACES, THAT'S AN
INDICATOR OF ENDOLYMPHATIC
HYDROPS.
WE HAVE A VERY SKILLED
NEURORADIOLOGIST HERE THAT WE
WORK WITH, DR. JOHN BUCKMAN, WHO
I'LL ACKNOWLEDGE AT THE END BUT
HIS EXPERTISE HERE IS CRITICAL.
SO CURRENT GRADING SCHEMES OR
SYSTEMS FOR ENDOLYMPHATIC
HYDROPS ARE HIGHLY SUBJECTIVE,
AND YOU CAN SEE EXAMPLES OF A
COCHLEAR GRADING SCHEME AND WHAT
YOU'RE SEEING TO THE RIGHT ARE
THE -- THE IMAGES ABOVE ARE
IMAGES THAT SHOW TOTAL FLUID AND
IMAGES BELOW ARE IMAGES THAT ARE
THE FLAIR IMAGES THAT SHOW US
BEING ABLE TO IDENTIFY HYDROPS.
HERE IS AN EXAMPLE WHERE THERE'S
NONE AND HERE IS AN EXAMPLE
WHERE THERE IS A GRADE
2 ENDOLYMPHATIC HYDROPS IN THE
COCHLEA.
SIMILARLY, THERE'S A SYSTEM,
THIS IS AN EXAMPLE OF A SYSTEM
THAT'S USED TO DIAGNOSE HYDROPS
IN THE VESTIBULE, OR THE BALANCE
PART OF THE INNER EAR.
SO OUR GOAL REALLY, GIVEN THE
SUBJECTIVITY OF THESE GRADING
SYSTEMS, IS TO ACCURATELY
QUANTIFY ENDOLYMPHATIC HYDROPS.
AND BEING ABLE TO DO THIS COULD
CHANGE THE UTILITY OF THIS
IMAGING MODALITY, AS WELL AS OUR
ABILITY TO ASSESS CHANGES IN THE
INNER EAR.
NOW, WE USE SEVERAL MRI
SEQUENCES.
THE FIRST ONE BEING SHORT TAU
INVERSION RECOVERY OR STIR
SEQUENCES ON MRI, WHICH GIVE US
THE TOTAL FLUID IN THE INNER
EAR.
AND HERE YOU SEE AN EXAMPLE OF
THAT.
AND THEN FLUID ATTENUATED
INVERSION RECOVERY SEQUENCES OR
FLAIR SEQUENCES ON DELAYED
SEQUENCING ALLOW US TO DETERMINE
PERILYMPH SIGNAL, AND THAT'S
WHAT YOU SEE HERE.
THEN THOSE TWO IMAGES ARE
UTILIZED TO SUBTRACT ONE FROM
THE OTHER TO IDENTIFY THE
ENDOLYMPH SPACE YOU SEE IN THIS
DIFFERENCE IMAGE, AND THEN WE'RE
OVERLAYING THE TWO MASKS OF THE
DIFFERENT FLUIDS SO THE
PERILYMPH IN GREEN AND THE
ENDOLYMPH IN PINK, OR RED.
SO USING THIS METHODOLOGY, WE'RE
CAPABLE -- WE CAN IDENTIFY
EVIDENCE OF HYDROPS AND IDENTIFY
KIND OF THE ENDOLYMPH CONTAINING
SPACES OF THE COCHLEA AND
VESTIBULE.
SO THIS IS A BRIEF OVERVIEW OF
OUR CURRENT EFFORTS AT
QUANTIFYING IN A PEOP PIPELINE
FASHION BOTH ENDOLYMPH AND
PERILYMPH FASHIONS.
CURRENTLY IT'S SUPERVISED BUT
THE CURRENT GOAL IS TO DEVELOP
METHODS TO OUGHT MADE ALL THE
STEPS INCLUDING SEGMENTATION,
AND WE'RE ACTIVELY RECRUITING
PATIENTS TO BUILD THE DATASETS
TO TRAIN THE NECESSARY MODELS TO
ACCOMPLISH THIS TASK
SO NOW I KIND OF WANT TO TALK
ABOUT A FEW EXAMPLES TO GIVE YOU
A FLAVOR OF THIS KIND OF
IMAGING.
THIS IS ONE PATIENT WHOSE
HEARING IN THIS CASE BECAME
BETTER AND WORSE DURING TWO
SEGMENTS OF THE PROTOCOL.
THIS PATIENT HAD EXHIBITED
EVIDENCE OF HEARING INSTABILITY
PRIOR TO BEING RECRUITED TO THE
CLINICAL CENTER HERE.
SO HERE'S AN EXAMPLE OF THE
KINDS OF RECONSTRUCTIONS WE'RE
DOING IN TERMS OF IMAGING
ENDOLYMPHATIC HYDROPS.
THE PERILYMPH SIGNALS IN GREEN
HERE AND THE ENDOLYMPH SIGNAL IS
IN PINK.
AND WE'RE QUANTIFYING THE RATIO
OF ENDOLYMPH TO PERILYMPH
VOLUMES, AND IN THIS CASE, THIS
PATIENT'S VESTIBULAR VOLUME
RIGHT HERE STARTED OUT LARGER
AND THEN BECAME SMALLER, AND
THIS APPEARS TO CORRELATE WITH
THE PATIENT'S HEARING
IMPROVEMENT.
THE COCHLEA IS MORE DIFFICULT
FOR US TO VISUALIZE AND
QUANTIFY, BUT WE ARE WORKING ON
THAT CURRENTLY TOO.
SO THESE ARE ANOTHER COUPLE
PATIENTS FROM THE PROTOCOL WHO
HAVE COMPLETED THEIR ENTIRE
PROTOCOL.
WE'VE BEEN ABLE TO LOOK AT THEIR
DATA OVER TIME, AND PATIENT 1 IS
A PATIENT WITH HEARING
INSTABILITY WHILE ON PROTOCOL.
BOTH OF THESE PATIENTS HAVE A
HISTORY OF HEARING INSTABILITY,
BUT THE PATIENT, PATIENT 2, HAD
RELATIVELY STABLE HEARING WHILE
ON PROTOCOL.
AND IN THIS CASE, PATIENT 1 HAD
EVIDENCE OF ENDOLYMPHATIC
HYDROPS AND PATIENT 2 DID NOT
HAVE EVIDENCE OF ENDOLYMPHATIC
HYDROPS AS KIND OF ASCERTAINED
BY DR. BUCKMAN ON HIS READS OF
THE SEQUENTIAL MRIs OF THESE
PATIENTS OVER TIME.
SO ONE OF THE THINGS THAT WE SEE
USING SPECTRAL FLOW CYTOMETRY TO
PERFORM CELLULAR IMMUNOPROFILING
IS THAT PATIENT 1, WHO HAS
EVIDENCE OF HEARING INSTABILITY,
DEMONSTRATES A GREATER
PROPORTION OF IMMUNE CELL
POPULATIONS THAT ARE INVOLVED IN
INNATE IMMUNITY.
AND THOSE INCLUDE MONOCYTES,
DENDRITIC CELLS AND NK CELLS.
AND THE T-CELL POPULATIONS ARE
NOT SIGNIFICANTLY CHANGED.
SO FURTHERMORE, CYTOKINE
PROFILING ON THESE TWO PATIENTS
AS AN EXAMPLE SUGGESTS THE
POSSIBILITY OF AN INCREASE IN
PRO INFLAMMATORY CYTOKINES IN
THE SETTING OF HEARING
INSTABILITY.
WHAT WE DID HERE WAS, WE
PERFORMED -- STIMULATED THE
PERIPHERAL BLOOD MONO NUCLEAR
CELLS AND COMPARED THEM TO
UNSTIMULATED CONTROLS, AND WHAT
WE FOUND IS THAT THE PATIENT
WITH HEARING INSTABILITY IN THIS
CASE DEMONSTRATES A
GREATER DEGREE OF PRO
INFLAMMATORY CYTOKINES AFTER
STIMULATION, COMPARED TO THE
PATIENT WITH RELATIVELY STABLE
HEARING IN ORANGE HERE.
SO TO SUMMARIZE KIND OF WHAT
WE'VE STARTED TO LEARN IN THIS
PROTOCOL, CONTRAST ENHANCED
DELAYED FLAIR MRI CAN DIAGNOSE
ENDOLYMPHATIC HYDROPS IN
PATIENTS WITH HEARING
INSTABILITY.
PRELIMINARY DATA FROM OUR GROUP
SUGGESTS THAT SOME PATIENTS WITH
HEARING INSTABILITY DEMONSTRATE
ALTERATIONS IN INNATE IMMUNE
CELL POPULATIONS AND AN
UNDERLYING PREDISPOSITION
TOWARDS THE PRODUCTION OF PRO
INFLAMMATORY CYTOKINES.
FUTURE WORK IS FOCUSING ON
CREATING AN AI-BASED AUTOMATED
METHOD TO OBJECTIVELY QUANTIFY
ENDOLYMPHATIC HYDROPS AND WILL
IDENTIFY POTENTIAL IMMUNOLOGIC
AND NON-IMMUNOLOGIC BIOMARKERS
THAT MAY SERVE AS THERAPEUTIC
TARGETS.
SO GOING ON TO OUR WORK WITH THE
MOUSE MODELS, WE'VE BEEN
INVESTIGATING THE MECHANISMS OF
HEARING INSTABILITY IN THE
SETTING OF STRIA VASCULARIS
DYSFUNCTION USING A MOUSE MODEL
DEVELOPED HERE AT THE NIDCD.
IT'S THE INSUFFICIENCY MOUSE
MODEL.
WHILE I DON'T HAVE TIME TO
DESCRIBE ALL THE DETAILS ABOUT
OUR INVESTIGATIONS, I'M GOING TO
DESCRIBE THIS MOUSE MODEL AND
WHY I THINK IT'S A GOOD MOUSE
MODEL TO STUDY THESE DISEASES.
SO THIS MOUSE MODEL IS THE ONLY
MOUSE MODEL THAT DOESN'T RELY ON
A SURGICAL PROCEDURE OR
PERSISTENT DRUG ADMINISTRATION
TO PRODUCE PERSISTENT
INTERMITTENTLY -- INTERMITTENT
HEARING FLUCTUATION OVER TIME.
AND AGAIN, THIS MOUSE MODEL WAS
DEVELOPED HERE AS A
COLLABORATION BETWEEN THE
GRIFFITH AND FRIEDMAN LAB AT
NIDCD, AND IT CONSISTS OF TWO
TRANSGENIC MOUSE LINES, AN
EFFECTOR AND RESPONDER MOUSE
LINE BRED ON TO THE BACKGROUND
OF A PENDRIN NULL MOUSE.
NOW THE EFFECTOR AND RESPONDER
TRANSGENIC LINES ALLOW US TO USE
DOXYCYCLINE TO INDUCE THE
EXPRESSION OF PENDRIN.
AND WE DO THIS BETWEEN E0 AND
E17.5 TO ALLOW FOR THE INNER EAR
TO DEVELOP NORMALLY.
AFTER THIS TIME, DOC DOXYCYCLINS
STOPPED AND THESE MICE ARE
TESTED BETWEEN P30 OR ONE MONTH
AND TWO MONTHS, AND THESE
PATIENTS -- OR THESE MICE
EXHIBIT HEARING FLUCTUATION AS
PREVIOUSLY DESCRIBED BY THE
GRIFFITH LAB.
AND THE EXPERIMENTAL MICE ARE
CALLED -- WITH THE TWO TRANS
GENES ON THE PENDRIN NULL
BACKGROUND ARE CALLED DE17.5
MICE AND THE MICE HETEROSIGH
GUESS FOR PENDRIN SERVE AS AN
ADDITIONAL CONTROL.
SO YOU SEE THAT HERE.
SO APPROXIMATELY 60% OF THE
DE17.5 MICE DEMONSTRATE HEARING
INSTABILITY AND YOU CAN SEE THAT
HERE.
AND OF THE MICE THAT
FLUCTUATFLUCTUATE,THE DE17.5 MIN
EXHIBIT GREATER CUMULATIVE
THRESHOLD SHIFTS THAN THEIR
NON-FLUCTUATING COUNTERPARTS.
FURTHERMORE, THE DE17.5 MICE
ALSO DEMONSTRATE STRIA
VASCULARIS DYSFUNCTION IN THE
FORM OF ENDOCOCHLEAR POTENTIAL
REDUCTIONS, AND ALSO DEMONSTRATE
EVIDENCE OF IMMUNE DYSFUNCTION
OR DYSREGULATION, SPECIFICALLY
MICE WITH FLUCTUATION
DEMONSTRATE INCREASED NUMBERS OF
LATERAL WALL AND STRIA
VASCULARIS COCHLEAR MACROPHAGES.
SO ONE OF OUR HYPOTHESES HERE IS
THAT EITHER THE REDUCED ABILITY
TO RESPOND TO INFLAMMATION
AND/OR -- AND FORM OF IONIC
HOMEOSTATIC DYSFUNCTION RESULTS
IN THIS PREDISPOSITION TO
HEARING INSTABILITY IN THE MOUSE
MODEL.
AND WE THINK THAT MIGHT BE
TRANSLATABLE TO HUMANS TOO.
FINALLY, THIS MOUSE MODEL
DEMONSTRATES A HISTOLOGIC
HALLMARK FOR HEARING INSTABILITY
DISORDERS.
NAMELY, ENDOLYMPHATIC HYDROPS.
AND HERE YOU SEE PLASTIC
SECTIONS FROM MICE WITH AND
WITHOUT HEARING INSTABILITY OVER
TIME, AND THE MICE WITH HEARING
INSTABILITY SEEM TO DEMONSTRATE
A GREATER EXTENT OF
ENDOLYMPHATIC HYDROPS.
OF GREATER DEGREE, THAT IS.
THAT'S THAT EXPANSION OF THIS
CENTRAL COMPARTMENT.
SO IN SUMMARY, HERE, THE PENDRIN
A INSUFFICIENCY MOUSE MODEL MAY
SERVE AS A REASONABLE MODEL FOR
HEARING INSTABILITY DISORDERS.
CURRENTLY WE'RE UTILIZING SINGLE
CELL AND SINGLE NUCLEAR
TRANSCRIPTIONAL PROFILING TO
INVESTIGATE THE UNDERLYING
MECHANISMS OF HEARING
INSTABILITY IN THIS MOUSE MODEL
AND TO IDENTIFY POTENTIALLY
REPURPOSABLE THERAPEUTICS FOR
THESE DISEASES IN THE MOUSE
MODEL.
AND IN THE FUTURE, AND REALLY
CURRENTLY, WE'RE FOCUSING ON
UTILIZING THIS MOUSE MODEL TO
DEVELOP PRELIMINARY DATA TO
SUPPORT THE REPURPOSING OF
FDA-APPROVED MEDICATIONS TO
TREAT HEARING INSTABILITY.
AND WITH THAT, I'D LIKE TO THANK
YOU FOR YOUR ATTENTION, AND I'D
LIKE TO ACKNOWLEDGE THE MEMBERS
OF MY LAB, WHOSE WORK YOU'VE
SEEN HERE TODAY, AND ALSO THE
NIDCD CLINICAL RESEARCH TEAM AND
THE AUDIOLOGY UNIT, WHO HAVE
BEEN VERY SUPPORTIVE IN GETTING
MY CLINICAL PROTOCOL UP AND
RUNNING AND RECRUITING LIKE IT
IS NOW.
AND WITH THAT, THANK YOU AGAIN
FOR YOUR ATTENTION.
>>GOOD AFTERNOON, EVERYONE.
I WANT TO THANK DR. BURKLOW FOR
THE INVITATION TO PRESENT IN
TODAY'S GRAND ROUNDS WITH
DR. HAO.
SO OVER THE NEXT 25 MINUTES OR
SO, I'D LIKE TO TALK TO YOU
ABOUT THE DEVELOPMENT OF INNER
EAR GENE THERAPY AS THE
TREATMENT FOR HEARING LOSS AND
DIZZINESS, AND MORE
SPECIFICALLY, I'D LIKE TO TALK
ABOUT SOME OF THE WORK WE'RE
DOING IN THE LAB TO TRY TO APPLY
INNER EAR GENE THERAPY TO USHER
SYNDROME.
I HAVE NOTHING TO DISCLOSE.
SO AS MANY OF YOU KNOW, HEARING
LOSS IS A MAJOR DISABILITY
AFFECTING THE WORLD'S POPULATION
TODAY.
WE KNOW THAT ABOUT HALF OF THE
POPULATION 75 YEARS AND OLDER
HAVE HEARING LOSS.
HEARING LOSS IS NOT JUST A
DISEASE PROCESS, IT AFFECTS THE
ELDERLY POPULATION.
IT ALSO AFFECTS MANY NEWBORNS AS
WELL.
WE KNOW THAT ABOUT 3 OUT OF
1,000 LIVE BIRTHS ARE AFFECTED
WITH HEARING LOSS.
SO IT IS TRULY A DISEASE PROCESS
THAT AFFECTS A WIDE RANGE OF
POPULATION.
CURRENTLY, PATIENTS WITH HEARING
LOSS ARE TYPICALLY TREATED WITH
HEARING AIDS IF THEIR HEARING
LOSS IS MILD TO MODERATE IN
SEVERITY, AND FOR THOSE THAT
HAVE SEVERE TO PROFOUND HEARING
LOSS, A COCHLEAR IMPLANT MAY BE
AN OPTION.
THE TECHNICAL ADVANCES OF
HEARING AID AND COCHLEAR IMPLANT
TECHNOLOGIES, SIGNIFICANT
DRAWBACKS TO BOTH OF THESE
THERAPEUTIC OPTIONS.
SIMILARLY, DIZZINESS CAUSED BY
VESTIBULAR DYSFUNCTION IS ALSO A
MAIL JUROR CAUSE OF DISABILITY
IN THE WORLD POPULATION TODAY
FROM EPIDEMIOLOGIC STUDIES, WE
KNOW THAT ABOUT 40% OF THE
AMERICAN ADULTS SEEK MEDICAL
HELP FOR DIZZINESS EVERY YEAR.
VESTIBULAR DYSFUNCTION CAN ALSO
AFFECT CHILDREN AND IN FACT
CHILDREN WITH VESTIBULAR
DYSFUNCTION CAN DEVELOP LEARNING
DISABILITIES AND DELAYED MOTOR
SCIELS SCILS.
TREATMENT OPTIONS FOR THOSE
PATIENTS WITH VESTIBULAR
DYSFUNCTION ARE EVEN MORE
LIMITED THAN THOSE AVAILABLE FOR
PATIENTS WITH HEARING LOSS.
AND IT IS BECAUSE OF THE LIMITED
NUMBER OF EFFECTIVE TREATMENT
OPTIONS FOR BOTH HEARING LOSS
AND DIZZINESS THAT HAVE REALLY
INSPIRED PEOPLE LIKE MYSELF AND
MANY OTHERS IN THE FIELD TO TRY
TO DEVELOP NEWER THERAPIES TO
HELP THESE PATIENTS.
OVER THE PAST FEW YEARS, OUR LAB
HAS BEEN FOCUS ON INNER EAR GENE
THERAPY AS WAY TO TREAT HEARING
LOSS AND DIZZINESS.
THE IDEA OF GENE THERAPY WAS
FIRST PROPOSED IN THE 1970s BY
ROBLIN AND FRIEDMANN AND AT THAT
TIME THERE WAS VERY LITTLE KNOWN
ABOUT THE HUMAN GENOME.
DESPITE THERE WAS NOT A LOT
KNOWN ABOUT THE HUMAN GENOME,
THEY PROPOSED THE RADICAL IDEA
THAT IF THE DISEASE PROCESS IS
CAUSED BY A MUTATION OF A
CERTAIN GENE OF INTEREST, FOR
EXAMPLE, AND THE MUTATION CAUSES
DYSFUNCTION IN AN ORGAN SYSTEM,
COULD WE POTENTIALLY DELIVER
NORMAL COPIES OF THE CDNA OF
THAT PARTICULAR GENE OF INTEREST
INTO THE DISEASE ORGAN SYSTEM TO
TRY TO RESTORE FUNCTION.
SO TO APPLY THIS TO THE INNER
EAR, HEARING LOSS AND DIZZINESS
IS CAUSE BID MUTATIONS AFFECTING
A GENE THAT IS IMPORTANT FOR THE
DEVELOPMENT OF THE
MECHANOSENSORY HAIR CELLS, WHICH
ARE THE VERY IMPORTANT CELL TYPE
FOR SOME PERCEPTION, THEY ARE
THE MAIN SENSORS FOR AUDITORY
PERCEPTION IN THE INNER EAR,
THEN COULD WE POTENTIALLY
DELIVER NORMAL COPIES OF THIS
PARTICULAR GENE CDNA INTO THE
INNER EAR TO TRY TO RESTORE
FUNCTION IN THE INNER EAR.
OVER THE PAST FEW YEARS, WE HAVE
FOCUSED ON USING PRIMARILY THE
ADENOASSOCIATED VIRUS OR THE AAV
AS THE VIRAL VECTOR OF CHOICE
FOR GENE DELIVERY IN VARIOUS
MOUSE MODELS WE'RE WORKING ON IN
THE LAB.
FIRST OF ALL, AAV IS
NON-PATHOGENIC IN HUMANS AND
ALSO IN MICE, THEREFORE, IT IS A
VERY SAFE VIRUS TO USE.
IT HAS BEEN SHOWN TO BE ABLE TO
INFECT MULTIPLE CELL TYPES AND
BY ENGINEERING THE -- WE CAN
ACTUALLY ACHIEVE FAIRLY CELL
TYPE SPECIFIC TRANSDUCTION.
AND BECAUSE OF THESE FAVORABLE
PROPERTIES, AAV IS ONE OF THE
MOST COMMONLY USED VIRAL VECTORS
IN HUMAN GENE THERAPY CLINICAL
TRIALS TODAY.
ONE DRAWBACK FOR AAV IS THE FACT
THAT THE CARRYING CAPACITY OF
THE SIZE THAT THE CDNA CAN CARRY
IS ABOUT 4.5 KB AND THEREFORE,
IF ONE IS INTERESTED IN
DELIVERING A LARGER CDNA, THAT
COULD POTENTIALLY BECOME
CHALLENGING AND WE'LL TALK ABOUT
THAT TODAY.
SO ONE COMMON CAUSE OF
HEREDITARY HEARING LOSS IS USHER
SYNDROME.
USHER SYNDROME IS THE MOST
COMMON CAUSE OF DEAFNESS AND
BLINDNESS IN THE WORLD.
IT IS INHERITED IN AN AUTOSOMAL
RECESSIVE MANNER.
AND USHER SYNDROME CAN BE
CLASSIFIED INTO THREE DIFFERENT
TYPES, DEPENDING ON THE DISEASE
PHENOTYPE, THE S SEVERITY OF THE
PHENOTYPE AND ALSO THE ONSET OF
THE SYMPTOMS.
SO THE TIME 1 PATIENTS HAVE THE
MOST SEVERE DISEASE PHENOTYPE,
THEY ARE USUALLY BORN WITH
PROFOUND HEARING LOSS AT BIRTH,
THEY DEVELOP VISUAL LOSS AS A
RESULT OF RETINITIS PIGMENTOSA
AT AN EARLY AGE AND THEY ALSO
HAVE VESTIBULAR DYSFUNCTION AS
WELL.
THE TYPE 2 PATIENTS ARE
TYPICALLY BORN WITH MODERATE TO
SEVERE HEARING LOSS AT BIRTH,
AND THEIR VISUAL LOSS APPEARS A
LITTLE LATER IN LIFE COMPARED TO
THE TYPE 1 PATIENTS, AND THEY
TYPICALLY HAVE FAIRLY NORMAL
BALANCE FUNCTIONS.
THE TYPE 3 PATIENTS, THEY
DEVELOP PROGRESSIVE HEARING LOSS
USUALLY DURING CHILDHOOD OR
EARLY TEENS, AND THEY HAVE
VARIABLE ONSET OF VISUAL LOSSES
AND THEY TYPICALLY HAVE NORMAL
TO NEAR NORMAL BALANCE FUNCTIONS
DURING CHILDHOOD AND THEY MAY
DEVELOP BALANCE FUNCTION ISSUES
DOWN THE ROAD.
AND OVER HERE ON THE LEFT-HAND
SIDE IS A LIST OF THE GENES THAT
ARE INVOLVED WITH -- THAT HAVE
BEEN SHOWN TO CAUSE USHER SIM
DROME LISTED RIGHT HERE.
SO WE WANTED TO IDENTIFY A
SUITABLE TYPE OF USHER SYNDROME
TO APPLY INNER EAR GENE THERAPY
TO.
SO WE DECIDED TO TRY TO STUDY
USHER SYNDROME TYPE 1B.
THE USHER SYNDROME TYPE 1B IS
ONE OF THE MORE COMMON CAUSES OF
USHER SYNDROME.
SO THIS FIGURE HERE WAS TAKEN
FROM A META-ANALYSIS WHERE THEY
ANALYZED THE GENETIC DATA FROM
PATIENTS WITH USHER SYNDROME AND
WHAT THEY FOUND WAS THAT IN
ABOUT 21% OF PATIENTS, USHER
SYNDROME IS ACTUALLY CAUSED BY
MUTATIONS IN MYO7A, THE
CAUSATIVE GENE FOR USHER
SYNDROME TYPE 1B.
THE SECOND REASON WE WANTED TO
STUDY TYPE 1B IS THE FACT THAT
THE TYPE 1 PATIENTS HAVE THE
MOST SEVERE DISEASE PHENOTYPE
AND, THEREFORE, WE FELT THE
POTENTIAL TO DEVELOP AN
EFFECTIVE TREATMENT OPTION FOR
THEIR HEARING LOSS AND DIZZINESS
WOULD POTENTIALLY HAVE THE
BIGGEST IMPACT ON THEIR QUALITY
OF LIFE.
SO WE TURNED TO THE LITERATURE
TO TRY TO IDENTIFY A SUITABLE
ANIMAL MOUSE MODEL FOR STUDYING
US H-1B AND WE DECIDED TO USE
THE 4626SB MOUSE MUTANT.
THE REASON IS THIS MOUSE
ESSENTIALLY IS A NULL, HAS A
NULL MUTATION, AND THIS MOUSE
HAS PROFOUND HEARING LOSS
THROUGHOUT LIFE SIMILAR TO
PATIENTS WITH US H-1B.
IN ADDITION, THIS PARTICULAR
MUTANT MOUSE HAS SIGNIFICANT
VESTIBULAR DYSFUNCTION AS WELL
AS EVIDENCED BY THE CIRCLING
BEHAVE ARE YO, HEAD BOBBING AND
HYPERACTIVITIES, VERY SIMILAR TO
THOSE PATIENTS WITH US H-1B.
SO WE'LL CALL THIS MOUSE MODEL
THE SHAKER1 MOUSE THROUGHOUT THE
REST OF THE TALK.
WHEN THE INVESTIGATORS ANALYZED
THE INARE EAR TISSUES FROM THE
SHAKER1 MICE, WHAT THEY FOUND
WAS THAT THE HAIR CELLS IN THE
COCHLEA AND THOSE IN THE
VESTIBULAR ORGANS HAVE
SIGNIFICANT STEREOCILIA BUNDLE,
THESE HAIRLIKE PROJECTIONS ON
THE MECHANOSENSORY HAIR CELLS
CRITICAL FOR THE DETECTION OF
SOUND INFORMATION COMING INTO
THE INNER EAR.
IN ADDITION THE COCHLEAR HAIR
CELLS UNDERGO RAPID DEGENERATION
AFTER BIRTH.
SO ONE CHALLENGE OF DELIVERING
THIS INTO THE MOUSE INNER EAR IS
THAT THE MOUSE IS QUITE LARGE,
ABOUT 6.7, 6.8KB IN SIZE,
THEREFORE IT EXCEEDS THE
CARRYING CAPACITY OF A SINGLE
AAV.
SO TO OVERCOME THIS CHALLENGE,
INVESTIGATORS IN THE FIELD HAVE
USED WHAT'S CALLED A DUO AAV
APPROACH TO DELIVER LARGE
CDNAs INTO THE CELL TYPES.
SO THE WAY THIS WORKS IS ONE CAN
ACTUALLY BREAK UP A LARGE CDNA
IN TWO DIFFERENT FRAGMENTS, THE
5 PRIME AND 3 PRIME FRAGMENT AND
ONE CAN USE TWO AAV TO DELIVER
THEM INTO THE CELL TYPES.
AT THE END, THESE CDNA
FRAGMENTS, THE SEQUENCES CAN
ACTUALLY HELP THE TWO FRAGMENTS
RECOMBINE TO PRODUCE A FULL
LENGTH PROTEIN PRODUCT.
SO THIS WORK WAS DONE BY
DR. BOYE'S GROUP AT UNIVERSITY
OF FLORIDA.
THEY APPLIED THE DUAL-AAV
TECHNIQUE TO DELIVER THE HUMAN
MYO7A CDNA, AND THEY WERE ABLE
TO FIND FOLDING PROTEIN IN THESE
CELLS.
THEY THEN INJECTED THE DUAL AAV
VECTORS CARRYING THE HUMAN MOUSE
RETINA AND THEY FOUND IN THE
INJECTED MICE, HUMAN MYO7A
PROTEIN WAS EXPRESSED SHOWN IN
GREEN IN THE MOUSE RETINA.
IN A SEPARATE STUDY THAT WAS
PUBLISHED AROUND THE SAME TIME,
THEY ALSO APPLY THE SAME DUAL
AAV APPROACH TO TRY TO DELIVER
THE HUMAN MYO7A CDNA INTO FIRST
A CELL LINE SHOWN RIGHT HERE,
AND AGAIN ON THE WESTERN BLOT
ANAL SEIZE, THEY WERE ABLE TO
SHOW EXPRESSION OF THE FULL
LENGTH MYO7A PROTEIN.
AND WHAT THEY THEN DID IN THE
STUDY WAS THEY INJECTED THEIR
DUAL AAV VECTORS CARRYING THE
HUMAN MYO7A CDNA INTO THE
SHAKER1 MOUSE, THE SAME MOUSE
WE'RE USING IN OUR STUDY.
IN THE TREATED MICE, THEY WERE
ABLE TO INCREASE THE MYO7A
PRODUCTION UP TO ABOUT 20% OF
THE WILD TYPE LEVELS.
SO WE WANTED TO SEE IF WE COULD
POTENTIALLY ALSO APPLY THE DUAL
AAV APPROACH TO DELIVER THE
HUMAN MYO7A CDNA INTO THE
SHAKER1 MOUSE INNER EAR TO HELP
TO IMPROVE ITS AUDITORY AND
VESTIBULAR FUNCTION.
SO WE COLLABORATED WITH BOYE'S
GROUP.
WE THEN INJECTED THESE VECTORS
INTO THE NEONATAL SHAKER1 MICE
INNER EARS, WE THEN ASSESSED
THEIR AUDITORY AND VESTIBULAR
FUNCTION ABOUT ONE MONTH AFTER
INJECTION.
WE FIRST TURN OUR ATTENTION TO
THE COCHLEA.
IN THE WILD TYPE, WE CAN SEE THE
MYO7A SHOWN IN GREEN EXPRESSED
BY BOTH THE INNER AND OUTER HAIR
CELLS.
MYO7A IS A COMMONLY USED MARKER
FOR MARKING THE HAIR CELLS IN
THE INNER EAR.
IN THE SHAKER1 MOUSE THAT DIDN'T
UNDERGO GENE THERAPY TREATMENT,
THERE WAS NO EXPRESSION AT ALL,
ONLY HAIR CELLS.
WHEN WE INJECTED THE DUAL-AAV
VECTORS INTO THE SHAKER1 MICE
WHAT WE FOUND WAS THAT IN SOME
OF THE INNER HAIR CELLS AND
OUTER HAIR CELLS, THEY STARTED
TO EXPRESS MYO7A.
WE THEN QUANTIFIED THE
TRANSDUCTION EFFICIENCY OF THE
DUAL AAV VECTORS IN THE TREATED
SHAKER 1 MICE AND WHAT WE FOUND
WAS THAT THE TRANSDUCTION
EFFICIENCY WAS CLOSE TO 80% OF
THE COCHLEAR HAIR CELLS AT THE
APEX OF THE COCHLEA AND THIS
DECREASED TOWARDS THE BASE OF
THE COCHLEA DOWN TO ABOUT 40%
FOR THE INNER HAIR CELLS AND
DOWN TO ABOUT 20% FOR THE OUTER
HAIR CELLS.
THE TRANSDUCTION EFFICIENCY WAS
HIGHER AT THE APEX OF THE
COCHLEA THAN THE BASE OF THE
COCHLEA.
WE WANTED TO SEE IF THE
TREATMENT OF DUAL AAV GENE
THERAPY CAN POTENTIALLY IMPROVE
THE HAIR CELL SURVIVAL IN THE
SHAKER 1 MICE.
REMEMBER I MENTIONED EARLIER
THAT THESE -- IN THESE MUTANT
MICE, THE COCHLEAR HAIR CELLS
UNDERGO RAPID DEGENERATION AFTER
BIRTH.
WHAT WE FOUND IS IN THE TREATED
SHAKER1 MICE THERE WAS A
SIGNIFICANT INCREASE COMPARED TO
THE UNTREATED SHAKER1 MICE SHOWN
IN BLUE HERE.
AGAIN NOTICE THAT THE
IMPROVEMENT IN HAIR CELL
SURVIVAL WAS THE BEST AT THE
APEX OF THE COCHLEA COMPARED TO
THE BASE OF THE COCHLEA AND THIS
CORRELATED WITH THE TRANSDUCTION
EFFICIENCY PATTERN AS WELL.
AS I MENTIONED BEFORE, THESE
ANIMALS, THESE SHAKER1 MICE HAVE
THESE PROJECTIONS ON THE HAIR
CELLS.
THIS IS WHAT THE STEREOCILIA
BUNDLES LOOK LIKE.
THEY'RE FAIRLY ORGANIZED IN THE
INNER AND OUTER HAIR CELLS.
IN THE UNTREATED, YOU ACTUALLY
SEE A FAIR AMOUNT OF HAIR CELL
DEGENERATION, PARTICULARLY AT
THE OUTER HAIR CELLS.
IN A TREATED SHAKER1 MICE WE
DIDN'T SEE ANY IMPROVEMENT IN
THE STEREOCILIA BUNDLES.
YOU CAN SEE THE BUNDLES FOR THE
INNER AND OUTER HAIR CELLS WERE
STILL FAIRLY DISORGANIZED.
SO WE SENSE THAT WITH THE DUAL
AAV GENE THERAPY -- WE ARE GOING
TO SEE EXPRESSION OF MYO7A AND
ALSO IMPROVEMENT IN HAIR CELL
SURVIVAL OF STEREOCILIA BUNDLES
THAT ARE STILL FAIRLY
DISORGANIZED.
WE THEN TESTED THE AUDITORY
FUNCTION OF THESE ANIMALS USING
AUDITORY BRAINSTEM RESPONSE
TESTING AND WHAT WE FOUND IS
THAT IN THE TREATED SHAKER1 MICE
THERE WAS NO MEASURABLE AAV
WAVEFORMS EVEN AT THE MAXIMUM
LEVELS OF 90 DB.
SO WE THEN TURN OUR ATTENTION TO
THE VESTIBULAR ORGANS AND THESE
ARE IMAGES TAKEN FROM THE MOUSE
K --EXPRESSED BY THE VESTIBULARR
CELLS SHOWN IN GREEN RIGHT HERE.
WHEREAS IN THE UNTREATED SHAKERE
ANY MYO7A EXPRESSION AT ALL.
IN THE TREATED SHAKER 1 MICE --
SHOWN IN GREEN RIGHT HERE.
WE WANTED TO SEE WHAT A
TRANSDUCTION EFFICIENCY OF THE
VESTIBULAR HAIR CELLS WERE SO WE
CALCULATED A NUMBER OF
VESTIBULAR HAIR CELLS THAT
EXPRESS MYO7A AND WHAT WE FOUND
WAS THAT TRANSDUCTION EFFICIENCY
WAS ABOUT 80% IN THE VESTIBULAR
ORGANS.
ONE INTERESTING THING THAT WE
FOUND WAS THAT AS OPPOSED TO THE
COCHLEA, WHERE THE HAIR CELLS
UNDERWENT RAPID DEGENERATION,
THE VESTIBULAR HAIR CELLS IN THE
SHAKER1 MICE SEEMED TO REMAIN
VIABLE FOR BOTH THE WILD TYPE
ANIMALS AND HETEROZYGOUS LITTER
MATES, AS YOU CAN SEE THE
VESTIBULAR HAIR CELL COUNT WERE
FAIRLY SIMILAR SO THE VESTIBULAR
HAIR CELLS IN SHAKER1 MICE DO
NOT -- THIS POTENTIALLY
INDICATES THEY'RE STILL VIABLE
TO POTENTIALLY BENEFIT FROM THE
DUAL AAV GENE THERAPY.
AS I MENTIONED BEFORE, THE HAIR
CELLS IN THE SHAKER 1 MICE HAD
HIGHLY DISORGANIZED STEREOCILIA
BUNDLES.
M THE BUNDLES IN THE HAIR CELLS
WERE VERY DISORGANIZED.
WHEN WE TREATED THE SHAKER1 MICE
WITH THE DUAL AAV GENE THERAPY,
WE SAW THE STER STEREOCILIA WERH
MORE ORGANIZED SIMILAR TO THE
WILD TYPE ANIMALS.
WE WANTED TO SEE IF THIS
IMPROVEMENT MORPHOLOGY
TRANSLATED INTO IMPROVEMENT IN
VESTIBULAR FUNCTION.
ONE SIMPLE WAY OF MEASURING
VESTIBULAR FUNCTION IN MICE IS
TO LOOK AT THEIR CIRCLING
BEHAVIOR.
AND WE HAVE A SET-UP IN THE LAB
WHERE WE HAVE A CAMERA THAT CAN
RECORD THE MOVEMENT OF THESE
ANIMALS AND WE HAVE A COMPUTER
SOFTWARE THAT CAN GENERATE --
THAT I CAN TRACK THE MOVEMENT OF
THESE ANIMALS IN GENERALLY
WHAT'S CALLED A TRACK PLOT OF
THESE ANIMALS.
IN A WILD TYPE MOUSE, THEY TEND
TO WALK IN A STRAIGHT LINE MORE
OR LESS AND TENDS TO EXPLORE THE
PERIPHERY OF THE CAGE THAT IT IS
HOUSED IN.
IN THE UNTREATED SHAKER 1 MOUSE,
YOU CAN SEE THE TRACK PLOT IS
VERY MESSY BECAUSE THIS MOUSE
RAN AROUND QUITE A BIT IN
CIRCLES.
IN THE TREATED SHAKER 1 MICE,
WHAT WE FOUND WAS THAT THE TRACK
PLOT IS MUCH CLEANER SIMILAR TO
THAT OF A WILD TYPE MOUSE THAT
THE CIRCLING BEHAVIOR SEEMS TO
BE TREE DUED A ROOB D REDUCED AE
THERAPY TREATMENT.
A 2-MINUTE TIME INTERVAL IN THE
UNTREATED, THEY CIRCLED AROUND A
LITTLE BIT LESS THAN 10 CIRCLES
EVERY 2 MINUTES.
IN THE UNTREATED SHAKER 1 MICE
THEY CIRCLED ABOUT 35 ROTATIONS
IN ABOUT 2 MINUTES.
IN THE TREATED SHAKER 1 MICE, WE
SAW SIGNIFICANT REDUCTION IN IS
THE CIRCLING BEHAVIOR DOWN TO
ABOUT 15 ROTATIONS EVERY
2 MINUTES.
WE ALSO TREATED SOME OF THE
HETEROZYGOUS LITTER MATES WITH
THE GENE THERAPY AND WE DIDN'T
SEE ANY ELEVATION IN CIRCLING
BEHAVIOR.
IN ADDITION, AS ANOTHER CONTROL,
WE TREATED WITH JUST THE 5 PRIME
AAV VECTOR SO WITHOUT THE
3 PRIME VECTOR AND WHAT WE FOUND
WAS THAT THOSE ANIMALS TREATED
WITH JUST THE 5 PRIME VECTOR
STILL HAD SIGNIFICANTLY ELEVATED
CIRCLING, THAT OF THE UNTREATED
SHAKER 1 MOUSE.
NOW WHILE CIRCLING IS A GOOD
MEASURE OF VESTIBULAR FUNCTION
IT IS NOT VERY SPECIFIC FOR THE
VESTIBULAR SYSTEM AND THEREFORE
WE WANTED TO SEE IF WE CAN FIND
A DIFFERENT TEST THAT COULD
POTENTIALLY HELP US BETTER
ASSESS THE VESTIBULAR SYSTEM
SPECIFICALLY.
WE DECIDED TO USE THE VESTIBULAR
EVOKED POTENTIAL OR VSEP,
PLACING ELECTRODES ON THE SCAL
CH THESE ANIMALS' HEADS, TO
GENERATE A HEAD IMPULSE TO
STIMULATE, USING ELECTRODES TO
RECORD THE EE ELECTRICAL
RESPONSES.
IN THE WILD TYPE MOUSE WE CAN
SEE A NICE VSEP MEASURE OF
WAVEFORMS DOWN TO MINUS 90DB,
WHEREAS IN THE SHAKER 1 MICE, WE
DIDN'T SEE ANY MEASURABLE
WAVEFORMS.
IN THE SHAKER 1 MICE THAT WERE
TREATED WITH THE DUAL AAV GENE
THERAPY, THEY ACTUALLY HAD
MEASURABLE VSEP WAVEFORMS, SOME
OF THEIR THRESHOLDS ACTUALLY
DOWN TO THE LEVEL OF THE WILD
TYPE ANIMALS.
YOU CAN SEE THE THRESHOLDS ARE
LISTED RIGHT HERE.
AND AGAIN, IN THE WILD TYPE
ANIMALS, WILD TYPE LITTER MATES
AND HETEROZYGOUS LITTER MATES,
WE SEE THE VSEP THRESHOLDS ON
AVERAGE ABOUT MINUS 10DB WHEREAS
WE DIDN'T SEE ANY MEASURABLE
RESPONSES AT ALL WITH THE
UNTREATED SHAKER 1 MICE, WE
MARKED THIS ESSENTIALLY AS PLUS
7.5DB AS THE THRESHOLD.
MANY OF THE SHAKER 1 MICE WERE
TREATED WITH DUAL AAV GENE
THERAPY, WE CAN SEE MEASURABLE
VSEP WAVEFORMS.
WE ALSO TREATED SOME OF THE WILD
TYPE AND HETEROZYGOUS LITTER
MATES WITH THE DUAL AAV VECTORS,
AGAIN, WE DIDN'T SEE ANY
ELEVATION IN THE VSEP
THRESHOLDS.
WE AGAIN TREATED SOME OF THE
SHAKER 1 MICE WITH JUST A
5 PRIME VECTOR AND DIDN'T SEE
ANY IMPROVEMENT IN THE VSEP
THRESHOLDS AT ALL.
SO THESE DETAILS DEMONSTRATE
THAT WITH THE DUAL AAV GENE
THERAPY, WE SEEM TO BE ABLE TO
IMPROVE THE VESTIBULAR FUNCTION
IN THESE MUTANT MICE. SO TO
SUMMARIZE, OUR DUAL AAV GENE
THERAPY WAS ABLE TO -- WE WERE
ABLE TO USE THE DUAL AAV GENE
THERAPY TO DELIVER THE MYO7A
CDNA INTO THE SHAKER 1 MOUSE
INNER EAR AND TRANSDUCED HAIR
CELLS DID EXPRESS MYO7A.
IN THE COCHLEA, WE FOUND THAT
THE DUAL AAV GENE THERAPY WAS
ABLE TO IMPROVE THE HAIR CELL
SURVIVAL, BUT UNFORTUNATELY WE
DIDN'T SEE ANY IMPROVEMENT IN
THE STEREOCILIA BUNDLE
MORPHOLOGY AUDITORY FUNCTION.
IN THE VESTIBULAR ORGANS, WE SAW
THE THERAPY IMPROVED THE
STEREOCILIA MORPHOLOGY IN THE
SHAKER ONE VESTIBULAR HAIR CELLS
AND ALSO IMPROVED THE VESTIBULAR
FUNCTION IN THE SHAKER 1 MUTANT
MICE.
IT IS OUR HOPE THAT THESE DATA
WILL FORM A FOUNDATION TO
TRANSLATE GENE THERAPY AS A
TREATMENT FOR PATIENTS WITH
USHER SYNDROME TO IMPROVE THEIR
VESTIBULAR FUNCTION DOWN THE
ROAD.
SO I'D LIKE TO CONCLUDE BY
ACKNOWLEDGING THE LAB MEMBERS IN
MY LAB THAT DID ALL THIS WORK,
KEVIN ISGRIG, MHAMED, JIANLIANG,
YASUKO, ZAYNEP, MOAZ, SAMANTHA,
I WANT TO THANK TOM FRIEDMAN AT
NIDCD, TRACY FITZGERALD, TALAH
WAFA FOR THEIR EXPERT HELP IN
GETTING A LOT OF THIS DATA.
I WANT TO THANK SHANNON'S GROUP
AT UNIVERSITY OF FLORIDA FOR
HELPING US TO OBTAIN THESE DUAL
VECTORS, DUAL AAV VECTORS FOR
OUR STUDY.
SO WITH THAT, I'D LIKE TO
CONCLUDE MY TALK AND HAPPY TO
TAKE ANY QUESTIONS YOU MAY HAVE.
THANK YOU.
>>THANK YOU, DR. CHIEN, VERY
MUCH FOR YOUR PRESENTATION, AND
THANK YOU, DR. HAO AS WELL.
THANK YOU FOR SHARING YOUR VERY
IMPORTANT EXCITING WORK AND
UNDERSTANDING OF HEARING LOSS
AND AVENUES FOR IMPROVEMENT.
WHILE MANY OF US ARE NOT
DIRECTLY INVOFERLED WITH THE
MANAGEMENT AND TREATMENT OF
HEARING INSTABILITY AND HEARING
LOSS, MANY OF US, UP TO HALF
ACCORDING TO THE DATA SHARED BY
DR. CHIEN, CAN SEE THIS IN OUR
OWN FUTURE.
SO THIS TOPIC IS OF NOT JUST
SCIENTIFIC INTEREST BUT ALSO
PERSONAL INTEREST.
SO THANK YOU VERY MUCH.
A COUPLE QUESTIONS HAVE COME IN.
SO FIRST FOR DR. CHIEN, WITH
REGARDS TO GENE THERAPY IN
HUMANS, WHAT ARE THE CHALLENGES
IN TRANSLATING INNER GENE
THERAPY MOUSE MODELS TO
PATIENTS?
>>ABSOLUTELY.
SO I THINK WHILE WE HAVE MANY
WONDERFUL MOUSE MODELS THAT WE
CAN STUDY, DIFFERENT TYPES OF
HEARING LOSS, IT'S IMPORTANT TO
REMEMBER THAT THEY ARE -- THERE
ARE DIFFERENCES BETWEEN THE
MOUSE INNER EAR AND THE HUMAN
INNER EAR.
AND THERE ARE ALSO MANY
SIMILARITIES BUT THERE ARE ALSO
SOME SIGNIFICANT DIFFERENCES TO
THINK ABOUT.
ONE OF THE MAJOR DIFFERENCES IS
THE FACT THAT IN HUMANS, WE ARE
ACTUALLY BORN WITH LEARING HEARG
WHEREAS IN MICE, HEARING DOES
NOT DEVELOP FULLY UNTIL ABOUT 12
TO 14 DAYS AFTER BIRTH.
SO IN MOST OF THE GENE THERAPY
STUDIES THAT WE PUBLISH TO TRY
TO IMPROVE THE AUDITORY FUNCTION
IN VARIOUS MOUSE MODELS OF
HEARING LOSSES, MOST OF THE TIME
YOU HAVE TO DELIVER THE THERAPY
VERY EARLY ON, BETWEEN THE FIRST
COUPLE DAYS OF LIFE IN ORDER FOR
THE THERAPY TO BE EFFECTIVE.
SO IF YOU WERE TO TRANSLATE THIS
TIMELINE INTO HUMANS, THIS WOULD
POTENTIALLY INDICATE THAT WE MAY
HAVE TO DELIVER THE GENE THERAPY
BEFORE THE ONSET OF HEARING
OCCURS IN HUMANS, WHICH WOULD
BE -- THIS WILL HAVE TO BE DONE
IN UTERO, WHICH CERTAINLY HAS
MANY TECHNICAL AND POTENTIALLY
EVEN ETHICAL CHALLENGES AS WELL
SO I THINK THAT'S ONE MAJOR
HURDLE ONE HAS TO TRY TO
OVERCOME, IS TO TRY TO SEE IF --
TO SHOW THAT GENE THERAPY IS
EFFECTIVE IN THE MATURE AND
DEVELOPED INNER EAR BEFORE
SUCCESSFULLY DONE IN HUMANS.
>>WHAT WAS THE AGE OF THE
TREATED MICE, AND CAN THE
TREATMENT BE USED IN ADULT MICE?
>>SO IN OUR STUDY, WE TREATED
THESE MICE BETWEEN T0 AND T5.
WE ARE CURRENTLY TESTING WHETHER
OUR THERAPY IS EFFECTIVE IN
ADULT MICE AND HOPEFULLY I'LL
HAVE SOME MORE DATA TO SHARE ON
THAT TOPIC NEXT TIME.
>>ONE MORE QUELL IN FOLLOW-UP.
IT WAS ENCOURAGING TO SEE THE
VESTIBULAR IMPROVEMENT IN THE
MOUSE MODEL THAT RECEIVED THE
DUAL AAV GENE THERAPY.
ARE THESE IMPROVEMENTS SUSTAINED
OR WAS THERE REGRESSION?
>>YES, THAT'S A GREAT QUESTION.
SO THAT'S SOMETHING THAT WE'RE
ALSO ASSESSING AS WELL.
SO WE ARE TRYING TO ASSESS THE
LONG TERM OUTCOME OF THESE
ANIMALS AND SEE IF WE CAN SEE
SUSTAINED VESTIBULAR IMPROVEMENT
OVER TIME.
>>AND THIS IS FOR DR. HAO.
SO WHAT EVIDENCE DO WE HAVE THAT
MENIERE'S DISEASE INVOLVES
PROBLEMS IN ION HOMEOSTASIS IN
HUMANS?
>>I THINK THE EVIDENCE THAT IT
DOES IS THAT WE CAN NOW IDENTIFY
HYDROPS IN HUMAN PATIENTS WHILE
THEY'RE LIVING, AND IN SOME
CASES, WE SEE CHANGES IN THEM.
SO THERE'S SOMETHING GOING ON IN
THE EAR THAT CONTROLS FLUID AND
LIKELY ION REGULATION.
IN MANY CASES, THESE PATIENTS
RESPOND TO MEDICATIONS THAT HAVE
A ROLE IN REGULATING ION
CHANNELS.
SO NOT JUST STEROIDS BUT OTHER
MEDICATIONS.
SO I THINK THOSE ARE THE PIECES
OF EVIDENCE THAT WE HAVE, I
THINK FOR EVIDENCE THAT, YOU
KNOW, ION HOMEOSTASIS IS
INVOLVED.
>>THANK YOU.
ARE YOU FINDING THAT THE FLAIR
MRI EVIDENCE OF THE
ENDOLYMPHATIC HYDROPS, IS THAT
AN EARLY FINDING OR LATE FINDING
WHEN COMPARED TO THE CLINICAL
SYMPTOMATOLOGY OF HEARING LOSS
AND OTHER FINDINGS?
>>THAT'S A VERY GOOD QUESTION.
I DON'T THINK WE TRULY KNOW THE
ANSWER TO THAT QUESTION YET.
ONE OF THE REASONS FOR US
WANTING TO FOLLOW ALL THESE
PATIENTS LONGITUDINALLY WAS TO
SEE HOW HYDROPS EVOLVES OVER
TIME.
AND IN SOME CASES, WE'VE SEEN
HYDROPS DECREASE WITH
IMPROVEMENT IN HEARING AND IN
SOME CASES WE'VE SEEN IT NOT
DECREASE.
SO I THINK THIS FORM OF IMAGING
MAY ALSO HELP US PHENOTYPE SOME
OF THESE PATIENTS INTO GROUPS,
INTO HOW THEIR HYDROPS MIGHT
CHANGE OVER TIME.
>>THANK YOU.
THIS QUESTION IS FOR DR. CHIEN.
DR. CHIEN, WHAT ARE THE THEORIES
ABOUT WHY THE DUAL AAV GENE
THERAPY IMPROVED VESTIBULAR
FUNCTION WHILE IT DID NOT
IMPROVE THE AUDITORY FUNCTION IN
THE SHAKER1 MUTANT MOUSE?
>>YEAH, AND THAT'S SOMETHING
THAT WE ARE THINKING QUITE A LOT
ABOUT, BUT I THINK ONE
POSSIBILITY AS I MENTIONED
BEFORE, THE COCHLEAR HAIR CELLS
ACTUALLY ON THE -- ACTUALLY THE
DEGENERATION HAS BEEN SHOWN TO
OCCUR EVEN IN UTERO, EVEN BY
EMBRYONIC DAY 17, 17.5 OR SO, SO
WHEREAS THE VESTIBULAR HAIR
CELLS ARE VIABLE AFTER BIRTH.
AND SO IT IS POSSIBLE THAT EVEN
THOUGH WE COULD RESTORE MYO7 --
IN SOME OF THESE HAIR CELLS,
MANY ARE PROBABLY ON THE WAY TO
DYING AND IT'S TOO LATE TO
INTERVENE AND TRY TO REVERSE THE
COURSE.
WHEREAS WITH VESTIBULAR HAIR
CELLS, THEY'RE STILL VIABLE AND
ABLE TO RESPOND TO BENEFICIAL
EFFECTS OF GENE THERAPY.
SO THAT'S OUR BEST GUESS AS TO
WHY IT'S MORE EFFECTIVE IN THE
VESTIBULAR ORGANS THAN COMPARED
TO THE COCHLEA.
>>THANK YOU.
AND SO ONE QUESTION COMES IN,
ARE YOU EXPLORING ANY GENE
EDITING APPROACHES?
>>YES, NOT FOR THIS PARTICULAR
MOUSE MODEL, BUT FOR SOME OF THE
OTHER MOUSE MODELS, ESPECIALLY
THOSE THAT HAVE A -- LIKE A
DOMINANT NEGATIVE MUTATION,
WE'RE TRYING TO DO GENE EDITING
AS A WAY TO IMPROVE THE FUNCTION
IN THOSE ANIMALS.
>>LOOKS LIKE WE HAVE JUST ONE
LAST QUESTION, AND THIS AGAIN IS
DR. HAO GOING BACK TO MR.
DO THE FLAIR MR FINDINGS
CORRELATE WITH THE DEGREE OF
HEARING INSTABILITY IN ANY WAY?
>>SO SOME OF OUR PRELIMINARY
DATA SUGGESTS THAT THE AMOUNT OF
HYDROPS DOES CORRELATE
PARTICULARLY FROM THE VESTIBULAR
STANDPOINT, DOES CORRELATE WITH
PURE TONE THRESHOLDS, SO THE
WORSE A PATIENT'S HEARING IS, IN
SOME CASES, THE WORSE THEIR --
OR GREATER THEIR HYDROPS IS.
>>THANK YOU VERY MUCH,
DR. CHIEN, DR. HAO, I THINK THAT
WAS THE LAST QUESTION RECEIVED.
THIS TAKES US UP TO THE TOP OF
THE HOUR.
AGAIN, I WANT TO THANK BOTH OF
YOU FOR SHARING YOUR WORK.
SINCERELY APPRECIATE THAT YOU'RE
PROVIDING SUCH HIGH QUALITY
PRESENTATIONS FOR OUR CLINICAL
CENTER GRAND ROUNDS, AND TO THE
NIH COMMUNITY AND BEYOND.
EVERYBODY, HAVE A GREAT DAY.
THANK YOU VERY MUCH.