WO2021158814A1 - Gene therapy systems and related methods for treatment of hearing loss - Google Patents
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61N—ELECTROTHERAPY; MAGNETOTHERAPY; RADIATION THERAPY; ULTRASOUND THERAPY
- A61N1/00—Electrotherapy; Circuits therefor
- A61N1/18—Applying electric currents by contact electrodes
- A61N1/32—Applying electric currents by contact electrodes alternating or intermittent currents
- A61N1/36—Applying electric currents by contact electrodes alternating or intermittent currents for stimulation
- A61N1/36036—Applying electric currents by contact electrodes alternating or intermittent currents for stimulation of the outer, middle or inner ear
- A61N1/36038—Cochlear stimulation
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P27/00—Drugs for disorders of the senses
- A61P27/16—Otologicals
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
- C12N15/86—Viral vectors
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61N—ELECTROTHERAPY; MAGNETOTHERAPY; RADIATION THERAPY; ULTRASOUND THERAPY
- A61N1/00—Electrotherapy; Circuits therefor
- A61N1/02—Details
- A61N1/04—Electrodes
- A61N1/05—Electrodes for implantation or insertion into the body, e.g. heart electrode
- A61N1/0526—Head electrodes
- A61N1/0541—Cochlear electrodes
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2750/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssDNA viruses
- C12N2750/00011—Details
- C12N2750/14011—Parvoviridae
- C12N2750/14111—Dependovirus, e.g. adenoassociated viruses
- C12N2750/14141—Use of virus, viral particle or viral elements as a vector
- C12N2750/14143—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
Definitions
- Various embodiments of the present disclosure relate generally to gene therapy systems and methods useful in the treatment and/or prevention of hearing loss. Exemplary embodiments described herein are directed to systems and related methods for preventing the further decline in a patient’s hearing loss. More specifically, embodiments taught in this present disclosure relate to gene therapy systems, and related methods, useful for treating and/or preventing deafness caused by genetic mutation of the TMPRSS3 gene or the LOXHD1 gene. These systems and methods may utilize a combination of gene therapy (e.g., molecular therapeutics) for hearing loss caused by a genetic mutation together with implantation of a cochlear implant.
- gene therapy e.g., molecular therapeutics
- Cochlear implant is a common procedure with a large associated healthcare cost, over $1,000,000 lifetime cost per patient (Mohr PE, et al.
- Hereditary hearing loss and deafness may be conductive, sensorineural, or a combination of both; syndromic (associated with malformations of the external ear or other organs or with medical problems involving other organ systems) or nonsyndromic (no associated visible abnormalities of the external ear or any related medical problems); and prelingual (before language develops) or postlingual (after language develops) (Richard JH Smith, MD, A Eliot Shearer, Michael S Hildebrand, PhD, and Guy Van Camp, PhD, Deafness and Hereditary Hearing Loss Overview, GeneReviews Initial Posting: February 14, 1999; Last Revision: January 9, 2014.
- DFN for DeaFNess
- Loci are named based on mode of inheritance: DFNA (Autosomal dominant), DF B (Autosomal recessive) and DFNX (X-linked).
- DFNA Autosomal dominant
- DF B Autosomal recessive
- DFNX X-linked
- SNHL Sensorineural hearing loss
- AAV adeno associated viral vectors
- CGF166 The first human clinical trial to address deafness and hearing loss using a gene therapy was (CGF166) initiated on June of 2014 and completed in December of 2019.
- the Principal Investigator for CGF166 was Dr. Hinrich Staecker and the trial was sponsored by Novaris. (https://clinicaltrials.gov/ct2/show/NCT02132130).
- An ideal disease target for translational research in this domain is a recessive genetic hearing loss that affects a defined group of cells within the inner ear and occurs postnatally after the development of speech. Prevalence of the mutation is an additional consideration.
- TMPRSS3 is a fairly common cause of hearing loss that is severe enough to warrant cochlear implantation.
- patients with mutations in TMPRSS3 may not respond to cochlear implantation as well as patients with other mutations (Shearer et al. , 2017). This presents the opportunity of targeting TMPRSS3, or other genes such as LOXHD1 , as a stand-alone therapeutic or in combination with other therapeutic agents and/or cochlear implantation to improve implant outcomes for this disorder.
- Table 1 (adapted from (Miyagawa, Nishio, & Usami, 2016)) demonstrates that mutations in TMPRSS3 may be the most common cause of postlingual recessive hearing loss that has a fairly limited distribution within the cochlea and, due to the size of the gene, may be built into existing AAV vectors.
- Table 1 Incidence of different mutations in 176 adult cochlear implant patients.
- TMPRSS3 The human transmembrane protease, serine 3 (TMPRSS3; also referred to as DFNB10, DFNB8, ECHOS1 , TADG12; Acc: HGNC:11877) was identified by its association with both congenital (present at birth) and childhood onset autosomal recessive deafness. Mutations in the TMPRSS3 gene are associated with Autosomal Recessive Nonsyndromic Flearing Impairment type DFNB8 and 10.
- TMPRSS3 is a 1646 base pair gene that codes for a serine protease and is associated with DFNA 8/10 and may make up to 1-5% of patients with hearing loss undergoing cochlear implantation (Weegerink et al. , 2011). Loss of function of this gene appears to result in a broad spectrum of hearing phenotypes depending on the site of the mutation. Both congenital and adult onset progressive hearing loss have been associated with the loss of this gene.
- DFNB8 hearing loss The onset of DFNB8 hearing loss is postlingual (age 10-12 years), while the onset of DFNB10 hearing loss is prelingual (congenital). This phenotypic difference reflects a genotypic difference.
- the DFNB8 causing variant is a splice site variant, suggesting that inefficient splicing is associated with a reduced amount of normal protein that is sufficient to prevent prelingual deafness but not sufficient to prevent eventual hearing loss.
- TMPRSS3 mutations on chromosome 21 known to cause hearing loss are described in Table 2.
- the lipoxygenase homology domains 1 gene (LOXHD1 ; also referred to as LH2D1 , DFNB77, FLJ32670; OMIM: 613072; Acc:HGNC:26521) encodes a highly conserved protein consisting entirely of PLAT (polycystin/lipoxygenase/alpha-toxin) domains, thought to be involved in targeting proteins to the plasma membrane.
- PLAT polycystin/lipoxygenase/alpha-toxin domains
- DFNB77 is caused by homozygous mutation in the LOXFID1 gene (613072) on chromosome 18q21.
- Loxhdl was expressed in the developing mouse inner ear at embryonic days 13.5 and 16, but not in any other tissue. At postnatal day 4, expression was detected in cochlear and vestibular hair cells, with highest concentration in the nucleus. Loxhdl progressively localized to the cytoplasm, and in the adult, Loxhdl was expressed in hair cells along the length of stereocilia.
- ENU N-ethyl-N-nitrosourea
- samba was a mutation in the mouse Loxhdl gene that destabilized the beta-sandwich structure of PLAT domain 10.
- the mutation did not alter mRNA or protein stability or localization of Loxhdl protein along the length of stereocilia.
- some hair cells showed morphologic defects with fused stereocilia and membrane ruffling at the apical cell surface.
- Profound degenerative changes were obvious by postnatal day 90, including hair cell loss and a reduction in spiral ganglion neurons. Grillet et al. (2009) hypothesized that the degeneration of spiral ganglion neurons was likely secondary to perturbations in the function and maintenance of hair cells.
- LOXHD1 mutations on chromosome 18 known to cause hearing loss are described in Table 3.
- U.S. Application Publication No. 2013/0095071 describes gene therapy methods for restoring age- related hearing loss using mutated tyrosine adeno-associated viral vectors to deliver the X-linked inhibitor of apoptosis protein (XIAP) to the round window membrane of the inner ear.
- XIAP X-linked inhibitor of apoptosis protein
- the publication does not contemplate the delivery of a nucleic acid sequence encoding functional TMPRSS3 or LOXHD1 to prevent or delay the onset of or restore hearing loss or deafness caused by genetic mutation of the TMPRSS3 or LOXHD1 gene, as disclosed herein.
- cochlear implantation is one common method of treatment of choice for addressing hearing loss ranging from severe to profound.
- a cochlear implant is a small, complex electronic device that can help to provide a sense of sound to a person who is profoundly deaf or severely hard-of-hearing.
- the implant consists of an external portion that sits behind the ear and a second portion that is surgically placed under the skin.
- Embodiments of the present disclosure relate to, among other things, gene therapy systems and methods useful in treating and/or preventing hearing loss.
- Systems and methods described herein relate to combination gene therapy with cochlear implantation to repair and/or rescue degenerating hair cells and/or degenerating spiral ganglion cells depending on the time of intervention.
- Each of the embodiments disclosed herein may include one or more of the features described in connection with any of the other disclosed embodiments.
- an expression vector including the nucleic acid sequence of SEQ ID NO: 1 or SEQ ID NO:2, or a nucleic acid sequence having at least 90% sequence identity to the nucleic acid of SEQ ID NO:1 or SEQ ID NO:2, wherein the nucleic acid sequence is operatively linked to a promoter.
- a pharmaceutical composition for use in a method for the treatment or prevention of hearing loss that includes an expression vector having the nucleic acid sequence of SEQ ID NO:1 or SEQ ID NO:2, or a nucleic acid sequence having at least 90% sequence identity to the nucleic acid of SEQ ID NO:1 or SEQ ID NO:2, wherein the nucleic acid sequence is operatively linked to a promoter.
- the nucleic acid sequence has at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO: 1 or SEQ ID NO:2.
- the expression vector is selected from an adeno-associated viral vector, an adenoviral vector, a herpes simplex viral vector, a vaccinia viral vector, a helper dependent adenoviral vector or a lentiviral vector.
- the vector is an adeno- associated viral vector selected from AAV2, AAV2/Anc80, AAV5, AAV6, AAV6.2, AAV7, AAV8, AAV9, AAVrh8, AAVrhIO, AAVrh39, AAVrh43AAV1 , AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, Anc80, or a synthetic version of an adeno associated viral vector serotype.
- the adeno-associated viral vector is AAV2, Anc80, or a synthetic version of an adeno associated viral vector serotype.
- the promoter is selected from any hair cell promoter that drives the expression of an operably linked nucleic acid at early development and maintains expression throughout the life, for example, TMPRSS3 promoters, human cytomegalovirus (HCMV) promoters, cytomegalovirus/chicken beta-actin (CBA) promoters, Myo7a promoters or Pou4f3 promoters.
- TMPRSS3 promoters human cytomegalovirus (HCMV) promoters, cytomegalovirus/chicken beta-actin (CBA) promoters, Myo7a promoters or Pou4f3 promoters.
- the nucleic acid sequence has at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO:1 or SEQ ID NO:2.
- the cell is a stem cell.
- the stem cell is an induced pluripotent stem cell.
- nucleic acid sequence has at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO:1 or SEQ ID NO:2.
- the expression vector is selected from an adeno-associated viral vector, an adenoviral vector, a herpes simplex viral vector, a vaccinia viral vector, a helper dependent adenoviral vector or a lentiviral vector.
- the vector is an adeno-associated viral vector selected from AAV2, AAV2/Anc80, AAV5, AAV6, AAV6.2, AAV7, AAV8, AAV9, AAVrh8, AAVrhIO, AAVrh39, AAVrh43, AAV1 , AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8 or Anc80, or a synthetic version of an adeno associated viral vector serotype.
- the adeno-associated viral vector is AAV2, Anc80, or a synthetic version of an adeno associated viral vector serotype.
- the promoter is selected from any hair cell promoter that drives the expression of an operably linked nucleic acid sequence at early development and maintains expression throughout the life, for example, TMPRSS3 promoters, human cytomegalovirus (HCMV) promoters, cytomegalovirus/chicken beta-actin (CBA) promoters, Myo7a promoters or Pou4f3 promoters.
- the expression vector is administered into the inner ear of the subject, for example, by injection.
- the delivery method is selected from cochleostomy, round window membrane, canalostomy or any combination thereof (see, Erin E. Leary Swan, et al. (2008) Inner Ear Drug Delivery for Auditory Applications. Adv Drug Deliv Rev. 60(15): 1583-1599).
- the expression vector is delivered into the scala media via the endolymphatic sac (Colletti V, et al. (2010) Evidence of gadolinium distribution from the endolymphatic sac to the endolymphatic compartments of the human inner ear. Audiol Neurootol. 15(6):353-63; Marco Mandala, MD, et al.
- the subject has one or more genetic risk factors associated with hearing loss.
- one of the genetic risk factors is a mutation in the TMPRSS3 gene.
- the mutation in the TMPRSS3 gene is selected from any one or more TMPRSS3 mutations known to cause hearing loss (see, for example, Table 2).
- one of the genetic risk factors is a mutation in the LOXHD1 gene.
- the mutation in the LOXHD1 gene is selected from any one or more LOXHD1 mutations known to cause hearing loss (see, for example, Table 3).
- the subject does not exhibit any clinical indicators of hearing loss.
- an expression vector described herein is administered as a combination therapy with one or more expression vectors comprising other nucleic acid sequences and/or with one or more other active pharmaceutical agents for treating hearing loss.
- a combination therapy may include a first expression vector that has the nucleic acid sequence of SEQ ID NO:1 and a second expression vector that has the nucleic acid sequence of SEQ ID NO:2, wherein both expression vectors are administered to a subject as part of a combination therapy to treat hearing loss.
- transgenic mouse having a human TMPRSS3 gene with a mutation selected from any one or more TMPRSS3 mutation known to cause hearing loss (see, for example, Table 2).
- TMPRSS3 TMPRSS3 mutation known to cause hearing loss
- transgenic mouse having a human LOXHD1 gene with a mutation selected from any one or more LOXHD1 mutation known to cause hearing loss (see, for example, Table 3).
- Figure 1 shows a cDNA sequence encoding wild-type human TMPRSS3 (GenBank Accession No. BC074847.2).
- Figure 2 shows the wild-type human TMPRSS3 amino acid sequence encoded by the cDNA in Figure 1 .
- Figure 3 shows a cDNA sequence encoding wild-type human LOXFIDI (GenBank Accession No. AK057232.1 ).
- Figure 4 shows the wild-type human LOXFIDI amino acid sequence encoded by the cDNA in Figure 3.
- Figure 5 shows TMPRSS3 immunohistochemistry in the adult mouse cochlea.
- Figure 6 shows an exemplary cochlear implant and the corresponding anatomy of the inner human, according to an aspect of the present disclosure.
- Figure 7 shows an exemplary TMPRSS3 plasmid map beginning at “ORI” and including an initial “AAV2 ITR” vector, a “CMV enhancer”, a “CMV promoter”, a “h- TMPRSS3”, a “bGH poly(A) signal, and a closing “AAV2 ITR” vector.
- Figure 8 illustrates proof of concept by graphically comparing hearing recovery of a disease model mouse receiving gene therapy treatment (treated) vs a disease model mouse not receiving treatment (untreated) by way of Auditory Brainstem Response (ABR) testing.
- ABR Auditory Brainstem Response
- Figure 9 illustrates proof of concept by graphically comparing hearing recovery of a disease model mouse receiving gene therapy treatment (treated) vs a disease model mouse not receiving treatment (untreated) by way of Distortion Product Otoacoustic Emissions (DPOAE) testing.
- DPOAE Distortion Product Otoacoustic Emissions
- Figure 10 graphically illustrates proof of concept by graphically comparing auditory neuronal function recovery of a disease model mouse receiving gene therapy treatment (treated) vs a disease model mouse not receiving treatment (untreated) by way of WAVEI amplitude testing.
- FIG 11 illustrates the location of the Round Window Membrane (RWM) within the human ear as an exemplary drug delivery site for delivering one or more of the gene therapies taught herein.
- RWM Round Window Membrane
- the present disclosure is drawn to gene therapy systems, and related methods, useful for treating and/or preventing deafness caused by genetic mutation.
- Examples of two genes that can mutate to cause deafness are the TMPRSS3 gene or the LoxHDI gene.
- the systems and methods described herein may utilize a combination of gene therapy (e.g., molecular therapeutics) for hearing loss caused by a genetic mutation together with implantation of a cochlear implant. It can be appreciated that while the systems and methods are in view of gene mutations caused by either the TMPRSS3 gene or the LoxHDI gene, other gene mutations may be targeted for repair that have been found to cause deafness or hearing loss.
- Gene therapy may refer to when DNA is introduced into a patient to treat a genetic disease.
- the new DNA usually contains a functioning gene to correct the effects of a disease-causing mutation in the existing gene.
- Gene transfer either for experimental or therapeutic purposes, relies upon a vector or vector system to shuttle genetic information into target cells.
- the vector or vector system is considered the major determinant of efficiency, specificity, host response, pharmacology, and longevity of the gene transfer reaction.
- the terms “treat,” “treating,” and “treatment” encompass a variety of activities aimed at desirable changes in clinical outcomes.
- the term “treat”, as used herein encompasses any activity aimed at achieving, or that does achieve, a detectable improvement in one or more clinical indicators or symptoms of hearing loss, as described herein.
- LOXHD1 gene for example, as detected in a genetic diagnostic test
- the present invention provides methods for therapeutic intervention during the period of gradual regression of hearing.
- the methods of the present invention can be commenced prior to such time period.
- the methods of treating hearing loss provided by the invention include, but are not limited to, methods for preventing or delaying the onset of hearing loss or the progression of clinical indicators or symptoms of hearing loss.
- hearing loss is used to describe the reduced ability to hear sound, and includes deafness and the complete inability to hear sound.
- an effective amount refers to an amount of an active agent as described herein that is sufficient to achieve, or contribute towards achieving, one or more desirable clinical outcomes, such as those described in the "treatment” description above.
- An appropriate “effective” amount in any individual case may be determined using standard techniques known in the art, such as a dose escalation study.
- active agent refers to a molecule (for example, an AAV vector described herein) that is intended to be used in the compositions and methods described herein and that is intended to be biologically active, for example, for the purpose of treating hearing loss.
- composition refers to a composition comprising at least one active agent as described herein or a combination of two or more active agents, and one or more other components suitable for use in pharmaceutical delivery such as carriers, stabilizers, diluents, dispersing agents, suspending agents, thickening agents, excipients, and the like.
- subject or “patient” as used interchangeably herein encompass mammals, including, but not limited to, humans, non-human primates, rodents (such as rats, mice and guinea pigs), and the like. In some embodiments of the invention, the subject is a human.
- vector may refer to a virus capable of transferring the desired gene into cells, but not capable of taking over or harming cells.
- adenovirus adeno-associated virus
- herpes simplex virus vaccinia virus
- vaccinia virus vaccinia virus
- retrovirus helper dependent adenovirus
- lentivirus adeno associated virus
- AAV can effectively transfect inner hair cells, a critical feature if one hopes to correct genetic defects due to hair cell-specific mutations.
- a number of different AAV subtypes have been used with success for cochlear gene delivery, demonstrating little if any damage to the organ of Corti.
- a recent report studying AAV serotypes 1 , 2, 5, 6 and 8 demonstrated successful gene expression in hair cells, supporting cells, the auditory nerve and spiral ligament, with hair cells being the most effectively transduced (Lawrence R. Lustig, MD and Omar Akil, PhD (2012) Cochlear Gene Therapy. Curr Opin Neurol. 25(1): 57-60).
- AAV vectors that can be administered to the inner ear are further described in U.S. Patent Application No. 2013/0095071, which is incorporated herein by reference in its entirety.
- Cochlear implants function by bypassing the function of hair cells and directly stimulate spiral ganglion cells. Hair cells are the sensory receptors of both the auditory system and the vestibular system in the ears of all vertebrates. Through mechanotransduction, hair cells detect movement in their environment.
- the spiral (cochlear) ganglion is the group of nerve cells that serve the sense of hearing by sending a representation of sound from the cochlea to the brain.
- the cell bodies of the spiral ganglion neurons are found in the modiolus, the conical shaped central axis in the cochlea. Therefore, having a functional spiral ganglion is vital for having a cochlear implant function optimally.
- these spiral ganglion cells may be susceptible to genetic mutation that result in hearing impairment or hearing loss. Hair cells, as mentioned, may also be susceptible to genetic mutation that may also result in hearing loss or impairment.
- delivery of a native copy of the TMPRSS3 gene (or any other suitable gene), via a viral vector may be used to treat either hair cells and/or spiral ganglion cells depending on the vector and the promoters used. Depending on the level of deterioration of the hair cells and/or spiral ganglion cells
- TMPRSS3 has the potential to rescue degenerating hair cells and/or degenerating spiral ganglion cells.
- TMPRSS3 gene therapy may enhance implant function by preserving spiral ganglion function and preventing further degeneration thereby allowing the implant to function optimally given the underlying cellular substrate.
- TMPRSS3 is a fairly common cause of hearing loss that is severe enough to warrant cochlear implantation. Additionally, patients with mutations in TMPRSS3 may not respond to cochlear implantation as well as patients with other mutations (Shearer et al. , 2017).
- TMPRSS3 may be the most common cause of postlingual recessive hearing loss that has a fairly limited distribution within the cochlea and, due to the size of the gene, may be built into existing AAV vectors.
- U.S. Application Publication No. 2013/0095071 describes gene therapy methods for restoring age-related hearing loss using mutated tyrosine adeno-associated viral vectors to deliver the X- linked inhibitor of apoptosis protein (XIAP) to the round window membrane of the inner ear.
- XIAP X- linked inhibitor of apoptosis protein
- the publication does not contemplate the delivery of a nucleic acid sequence encoding functional TMPRSS3 or LOXHD1 to prevent or delay the onset of or restore hearing loss or deafness caused by genetic mutation of the TMPRSS3 or LOXHD1 gene, as disclosed herein.
- the therapeutic treatment may be delivered through the round window membrane (RMW) of the inner ear using a catheter or port in the cochlear implant, as depicted in Figure 11.
- RMW round window membrane
- the round window membrane (RMW) within the human inner ear may serve as a potential drug delivery site.
- Figure 11 is an annotated version of an image of the anatomy of the human ear, available at https://commons.wikimedia.Org/wiki/File:Blausen_0328_EarAnatomy.png. See Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014”. WikiJournal of Medicine 1 (2).
- TMPRSS3 or LOXHD1 are currently no approved therapeutic treatments for preventing or treating hearing loss or deafness and there is a lack of useful preclinical animal models for testing such treatments.
- the present disclosure therefore describes systems and methods for viral vector gene delivery of TMPRSS3 or LOXHD1 into the inner ear to restore activity of a mutated TMPRSS3 or LOXHD1 gene, promote hair cell survival and restore hearing in patients suffering from hearing loss or deafness, and cell-based and animal-based models for testing such compositions and methods, while also combining treatment with cochlear implantation.
- Tmprss3 Expression of mouse Tmprss3 was evaluated in 1 month old C57BI5 mice using antibody anti-TMPRSS3 (1:100, ab167160, Abeam, Cambridge, MA). Labelling was seen in inner and outer hair cells, the stria vascularis and in about 50% of spiral ganglion cells (Fig5). This suggests that loss of TMPRSS3 function could additionally result in loss of strial function although no changes in endocochlear potential were seen in the Fasttle mouse model (Fasetti et al., 2011 ).
- TMPRSS3 genotype-phenotype studies demonstrate a wide range of different forms of hearing loss ranging from profound congenital to adult onset progressive hearing losses (Chung et al. , 2014; Gao et al. , 2017; Weegerink et al. , 2011 ). Studies suggest that hearing loss due to TMPRSS3 mutations may make up 2 to 5% of patients undergoing adult cochlear implantation (Jolly et al., 2012; Miyagawa, Nishio, & Usami, 2016; Sloan-Fleggen et al., 2016). Many of the patients with these mutations have significant amounts of residual hearing.
- an object of the present disclosure is to provide opportunities for using a combination the gene therapy techniques described above together with with cochlear implantation.
- a cochlear implant may comprise: 1) a microphone, which may receive sound from the environment; 2) a speech processor, which may select and arrange sounds picked up by the microphone; 3) a transmitter and receiver/stimulator, which may be configured to receive signals from the speech processor and convert them into electric impulses; and 4) an electrode array, which is a group of electrodes that collects the impulses from the stimulator and sends them to different regions of the auditory nerve.
- the cochlear implant may be a small, complex electronic device that can help to provide a sense of sound to a person who is profoundly deaf or severely hard-of-hearing.
- the implant consists of an external portion that sits behind the ear and a second portion that is surgically placed under the skin.
- a patient that may qualify for the therapy taught herein can be either: (1 ) a current user of a cochlear implant or (2) be a candidate for a cochlear implant, but not a current user, i.e. a new cochlear implant user that desires gene therapy treatment in conjunction with a new cochlear implant installation (both done at the same time).
- Cochlear implants are designed to mimic the function of a healthy inner ear (or cochlea). They replace the function of damaged sensory hair cells inside the inner ear to help provide clearer sound than what hearing aids can provide.
- a cochlear implant may be implanted to allow a person to take in external information through their auditory nerve.
- sensorineural hearing loss which means hair cells in a person ’ s inner ear are damaged, the damaged hair cells are no longer capable of sending sounds to their auditory nerve.
- a cochlear implant bypasses or skips these damaged har ceils in the inner ear to delivery information directly to the auditory nerve.
- certain genes are susceptible to mutation that prematurely damage or deteriorate these hair cells (and/or the spiral ganglion) at birth or sometime later in the person’s life.
- TMPRSS3 and LoxHDI may have poor outcomes in cochlear implant results 1 .
- the typical TMPRSS3 mutant patient may have dysfunction in either or both of their spiral ganglion and hair cells.
- hair cells degenerated initially and was followed shortly after by the degeneration of spiral ganglion cells 3 .
- cochlear implant function declines with age, which suggests that the delayed degeneration of spiral ganglion cells also occurs in the human population 4
- TMPRSS3 may not respond to cochlear implantation as well as patients with other mutations (Shearer et al. , 2017).
- the cochlear implant may be used to bypass the defective hair cells and directly stimulate the spiral ganglion cells, and, in combination with the implant, gene therapy may be used to fix the damaged hair cells and/or the spiral ganglion cells that have either been destroyed via natural causes and/or genetic defects.
- any commercially available cochlear implant may be utilized by the systems and methods described herein.
- genetic disorders may cause defective hair cells and/or spiral ganglion at the time of birth.
- the genetic mutation that may result in partial or total hearing loss may come at a later stage in life (e.g., adolescence, adulthood, etc.).
- FIG. 7 depicts an exemplary plasmid map for a TMPRSS3 vector construct that may be utilized in gene therapy according to aspects taught herein.
- the plasmid map illustrates a “AAV-cDNA 6-hTMPRSS3” with 5,667 bp.
- Cochlear implantation, with gene therapy using the “AAV-cDNA 6-hTMPRSS3” plasmid, may be utilized to achieve one or more of the objectives prescribed in this disclosure.
- the “AAV-cDNA 6-hTMPRSS3” depicted as Figure 2 may be used to genetically treat or repair mutations of the TMPRSS3 gene.
- the modified TMPRSS3 gene may repair damaged hair cells and/or spiral ganglion caused by mutated and defective genes.
- FIG. 7 The plasmid map of Figure 7, in an exemplary embodiment, beginning at “ORI” and including an initial “AAV2 ITR” vector, a “CMV enhancer”, a “CMV promoter”, a “h- TMPRSS3”, a “bGH poly(A) signal, and a closing “AAV2 ITR” vector.
- an additional therapeutic construct “AmpR promoter’ may be used. It can be appreciated that other vectors may be utilized to achieve objectives according to aspects of the present disclosure.
- MOUSE MODEL A TRMPSS mouse model in the CBA/J background was generated. These models when bred with the CBA/J strain established the mutant line. The mutation was a knock in model point mutation. The mutation was c.916G>A(p.Ala306Thr) homozygeous mutation.
- TMPRSS3 c.916G>A has been identified in more than 10 families from Chinese, German, Dutch, and Korean deaf patients, indicating that this mutation is the main contributor to the DFNB8/DFNB10 phenotype in many ethnicities.
- LAYMAN EXPLANATION OF ABR TEST The ABR test measures auditory function.
- the X-axis (Florizontal) lists the Frequencies (Pitch) which are expressed in kilohertz (kh). Numbers to the left of the X-axis are low pitch (like a bass note) as you move to the right, the numbers or pitch get higher (like a flute note).
- the Y-Axis (Vertical) describes the "Threshold" of hearing or loudness (expressed in decibels or db) i.e. how loud do we have to turn up the volume until the mouse hears.
- the auditory brain response (ABR) test was utilized to measure hearing thresholds at different frequencies for mutant (untreated) mice and mutant experimental (treated) mice.
- ABR auditory brain response
- the hearing thresholds for the treated mice (12) were much lower than the hearing thresholds for the control (untreated) mouse (10).
- the treated mouse (12) hears all frequencies sooner (at a lower volume) than the untreated mouse 10.
- DPOAE is a measure of outer hair cell (OFIC) function.
- the OHCs control volume of incoming sound (i.e. the ear's volume control knob).
- the X and Y axis are same as in Figure 8.
- the X-axis is frequency or pitch and Y-axis is threshold or volume needed to hear.
- DPOAE distortion product otoacoustic emissions test
- LAYMAN EXPLANATION OF WAVE1 TEST The WAVE 1 test is an additional measurement provided by the ABR test. Wave 1 amplitudes measure neuronal activities including the synchronous firing of numerous auditory nerve fibers in the spiral ganglion cells. The (horizontal) X-axis measures the response time to a sound stimulus (click) in milliseconds. The Y-Axis (vertical) describes the "Amplitude" or intensity / sensitivity of the auditory nerve's response to the sound stimulus expressed in millivolts (mv).
- FIG. 8 shown is the auditory evoked potential as a result of acoustic stimulation, measured in millivolts, as a function of time, measured in milliseconds.
- the acoustic stimulation was at a sound pressure level (SPL) of 80dB at 32 kHz.
- SPL sound pressure level
- the neural response generates a cycle of waves of which the first wave 14 and the third wave 16 are usually considered most significant.
- WAVE1 amplitudes were measured in treated mice (12) and in untreated mice both homozygous (10) and wild type (18).
- the WAVE1 amplitudes of the treated mice (12) were significantly greater than the amplitudes for the untreated mice (10 and 18).
- the treated mice (12) nerve cells are "firing" with greater intensity and sensitivity than untreated mice (10, 18).
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JUN SUZUKI; KEN HASHIMOTO; RU XIAO; LUK H VANDENBERGHE; CHARLES LIBERMAN M: "Cochlear gene therapy with ancestral AAV in adult mice: complete transduction of inner hair cells without cochlear dysfunction", SCIENTIFIC REPORTS, vol. 7, 45524, 3 April 2017 (2017-04-03), pages 1 - 11, XP055675065 * |
LANDEGGER ET AL.: "A synthetic AAV vector enables safe and efficient gene transfer to the mammalian inner ear", NATURE BIOTECHNOLOGY, vol. 35, no. 3, 6 August 2017 (2017-08-06), pages 280 - 284, XP055675069, DOI: 10.1038/nbt.3781 * |
See also references of EP4081232A4 * |
ZINN ET AL.: "in silico reconstruction of the viral evolutionary lineage yields a potent gene therapy vector", CELL REPORTS, vol. 12, no. 6, 11 August 2016 (2016-08-11), pages 1056 - 1068, XP055524241, DOI: 10.1016/j.celrep.2015.07.019 * |
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CA3166104A1 (en) | 2021-08-12 |
EP4081232A4 (en) | 2024-02-28 |
JP2023513116A (ja) | 2023-03-30 |
KR20220137652A (ko) | 2022-10-12 |
IL295265A (en) | 2022-10-01 |
WO2021158814A9 (en) | 2022-09-01 |
BR112022015345A2 (pt) | 2022-12-06 |
AU2021216405A1 (en) | 2022-09-15 |
CN116096391A (zh) | 2023-05-09 |
EP4081232A1 (en) | 2022-11-02 |
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