1 Introduction
Hearing loss is among the most common sensory disorders, affecting more than 430 million people worldwide and imposing a substantial social and economic burden
[1].. A large proportion of congenital and early-onset cases arise from monogenic defects, positioning the inner ear as a compelling target for gene-based therapeutic intervention. The genetic landscape of hereditary deafness is highly heterogeneous: more than 200 genes have been implicated in syndromic and non-syndromic forms of hearing loss, with mutations in
GJB2,
OTOF,
SLC26A4, and
TMC1 accounting for a substantial fraction of inherited cases
[2,
3]. Among these,
OTOF mutations represent one of the most frequent causes of autosomal recessive auditory neuropathy spectrum disorder (ANSD), disrupting synaptic vesicle exocytosis at the inner hair cell (IHC) ribbon synapse
[4,
5].
The cochlea possesses several biological features that make it uniquely suited—and simultaneously challenging—for gene therapy
[6,
7]. It is highly compartmentalized, relatively immune-privileged, and composed of discrete cell populations with specialized auditory functions. These characteristics create an attractive environment for localized, long-lasting gene delivery, yet they also demand precise vector distribution, efficient tissue penetration, and stringent cell-type specificity
[8]. Adeno-associated viruses (AAVs) have emerged as the leading platform for cochlear gene therapy due to their favorable safety profile, ability to support stable expression in post-mitotic cells, and compatibility with microsurgical delivery
[9]. Advances in capsid engineering, including rational design, directed evolution, and
in vivo selection, have yielded next-generation AAV variants with markedly enhanced transduction of IHCs and, increasingly, outer hair cells and supporting cells
[10-
13].
Momentum in the field has accelerated substantially with the recent clinical translation of dual-AAV therapies for
OTOF. Multiple phase 1/2 trials initiated by Akouos (Eli Lilly, NCT05821959), Decibel Therapeutics (Regeneron, NCT05788536), Sensorion (CTIS2023-504466-28-00/NCT06370351), and others have reported encouraging early outcomes
[3]. Several Chinese organizations have also conducted investigator-initiated clinical trials for the same indication. For example, Otovia Therapeutics (NCT05901480), Eye and ENT Hospital of Fudan University (NCT06722170), Refreshgene (ChiCTR2200063181), Shanghai Ninth People’s Hospital and EmayGene (ChiCTR2400091517), and others
[3]. These trials demonstrated favorable safety profiles and, critically, evidence of meaningful auditory recovery in children, including restored auditory brainstem responses and measurable improvements in sound perception. These first-in-human results provide compelling proof that AAV vectors can efficiently transduce human IHCs, drive expression of large genes through dual-vector strategies, and restore auditory function
[14,
15]. The success of
OTOF gene therapy marks a pivotal milestone for the field, validating decades of preclinical work and establishing a clinical precedent for targeting additional genetic forms of hearing loss
[16].
As gene therapy moves closer to broader clinical adoption, new challenges and opportunities have emerged. These include the need for capsids with improved specificity and adult transduction efficiency, scalable manufacturing for dual-vector systems, optimized delivery strategies that minimize surgical variability, and regulatory frameworks suited to inner ear interventions. This review synthesizes recent advances in AAV vector development for the inner ear, examines insights from early clinical studies, and outlines future directions toward safe, precise, and durable gene therapy for a broad spectrum of hereditary auditory disorders.
2 Structural and Molecular Characteristics of Adeno-Associated Virus
AAV is a non-enveloped, single-stranded DNA virus belonging to the family Parvoviridae. The AAV virion is an icosahedral particle with a diameter of approximately 20–25 nm, composed of three viral capsid proteins (VP1, VP2, and VP3) assembled in a molar ratio of approximately 1:1:10, which together form a symmetric capsid shell of 60 polypeptide copies
[17]. The AAV genome consists of a single-stranded DNA (ssDNA) molecule of approximately 4.7 kilobases in length, flanked by two inverted terminal repeats (ITRs) at its 5' and 3' ends. These ITR sequences serve as critical cis-acting elements for viral replication and also function as promoter and regulatory elements essential for stable transgene expression. Between the ITRs lie two open reading frames (ORFs): the rep gene, which encodes replication-associated proteins (Rep78 and Rep68) involved in viral DNA replication and transcriptional regulation, and the cap gene, which encodes the three structural capsid proteins
[18]. This compact genomic organization determines AAV’s critical packaging capacity limitation—typically accommodating only about 4.7 kb of foreign genetic material—a constraint that necessitates dual-AAV strategies for delivering large therapeutic genes such as
OTOF (which exceeds 6 kb)
[5].
From a structural biology perspective, the AAV capsid is dominated by β-barrel and β-sandwich fold structures that assemble into highly organized tertiary configurations
[18]. The capsid surface contains multiple functional domains, including receptor-binding sites (such as the heparan sulfate proteoglycan interaction region), immunogenic epitopes recognized by the host immune system, and signal sequences involved in membrane penetration and nuclear localization
[18,
19]. These surface characteristics of the capsid fundamentally determine the cellular tropism, tissue specificity, and immunogenicity of different AAV serotypes
[18-
20]. Through rational design, directed evolution, deep mutational scanning, peptide insertion, and high-throughput screening, researchers can systematically engineer capsid protein surfaces—by inserting peptide sequences, substituting critical amino acid residues, or reconstructing ancestral capsid sequences—to generate novel AAV variants with improved transduction efficiency, enhanced cell-type specificity, and superior penetration into adult or resistant tissues. These engineered capsids maintain AAV’s inherent safety advantages (non-integrating, non-pathogenic) while overcoming many limitations of natural serotypes, establishing AAV as an ideal gene delivery vehicle for inner ear gene therapy applications
[17].
3 Biological and Technical Challenges in Inner Ear Gene Delivery
The inner ear presents a unique combination of opportunities and technical barriers for gene therapy. Although its compartmentalized fluid spaces allow for localized vector delivery with minimal systemic exposure, the cochlea’s intricate microanatomy limits vector distribution
[21]. The round-window membrane (RWM), the most commonly targeted entry point in clinical practice, restricts passive diffusion, creating a gradient in which the basal turn is often more effectively transduced than the apical region
[22]. Additionally, the tight junctions between epithelial cells in the organ of Corti restrict lateral spread of viral particles, further constraining access to certain target populations
[23]. Cellular heterogeneity adds another layer of complexity. IHCs and spiral ganglion neurons are generally more permissive to AAV transduction, whereas OHCs, supporting cells (such as Deiters cells and Hensen cells), and the stria vascularis are substantially more resistant
[24]. The superior transduction of IHCs compared with OHCs reflects both differences in receptor expression and cellular accessibility. Achieving efficient and homogeneous transduction across these diverse cell types remains an unmet need, especially for disorders requiring OHC targeting or coordinated multicellular correction
[25]. Notably, many syndromic and non-syndromic hearing loss genes are expressed in multiple cochlear cell types, necessitating pan-cochlear or multi-target strategies
[13,
26].
Developmental stage is another major determinant of transduction success
[26]. Studies in rodents showed that neonatal cochleae exhibit markedly higher AAV transduction rates than adult tissue, partly due to immature barrier structures, more permissive extracellular spaces, and differences in receptor expression patterns
[13,
27]. These developmental differences complicate translation to human patients, whose treatment windows may not align with rodent developmental timelines. Clinical programs have typically focused on very young children (age < 18 years old) for auditory neuropathy, reflecting the superior transduction observed in neonates. However, extending AAV therapy to older children and adults remains a critical translational challenge
[14,
15].
Finally, safety must be carefully balanced with efficacy
[28]. High doses of AAV have been associated with dose-dependent ototoxicity in preclinical studies, particularly at levels required for widespread OHC transduction or synaptic targeting. Long-term expression, potential immune responses to AAV capsid proteins, and the risk of off-target expression in non-auditory tissues must be carefully considered as clinical use expands. The narrow therapeutic window between efficacious dosing and toxicity—a phenomenon well-documented in the inner ear—demands rigorous optimization of vector production, formulation, and delivery parameters.
4 Development and Characterization of AAV Vectors for Cochlear Gene Therapy
4.1 Natural and early-generation AAV serotypes
Initial efforts in cochlear gene delivery relied heavily on natural AAV serotypes such as AAV1, AAV2, AAV6, AAV8, and AAV9. Multiple research groups demonstrated that these vectors achieved reliable inner hair cell transduction but showed limited penetration into outer hair cells and supporting cells
[22]. Early studies demonstrated that conventional AAV serotypes such as AAV2 and AAV8 achieve robust transduction of IHCs in neonatal mice, often reaching high levels of infection across the sensory epithelium
[29,
30]. However, their performance declines markedly in mature cochleae, where adult mice exhibit substantially lower IHC and OHC transduction efficiencies, with responses varying widely across studies and depending on delivery route and dose
[2,
31]. Xue et al. reported that AAV9 offers broader tropism, particularly for IHCs targeting. And an AAV9-based RNA base editor (emxABE) was delivered into the cochlea of neonatal (P0–P3)
Otof Q829X/Q829X mice via scala media injection, resulting in nearly 100% transduction of inner hair cells and about 80% adenine-to-inosine conversion at the mutant
OTOF locus
[32]. Collectively, natural AAV serotypes show inherent limitations in the inner ear, achieving reliable transduction mainly in inner hair cells but failing to efficiently target outer hair cells and supporting cells.
4.2 Engineered and directed-evolution capsids
The development of engineered and directed-evolution AAV variants has fundamentally reshaped the inner ear gene therapy landscape. Landegger et al. developed Anc80L65, an ancestral reconstructed AAV variant, which represented the first major breakthrough in engineered AAV vectors. Their work demonstrated near-complete inner hair cell transduction (> 85%) in neonatal mice and greatly improved outer hair cell targeting compared with natural serotypes
[13]. The remarkable success of Anc80L65 in preclinical models inspired further intensive capsid engineering efforts using directed evolution, rational design, peptide insertion, DNA shuffling, and phage display screening by research groups worldwide
[33-
35].
Isgrig et al. demonstrated that AAV2.7M8 achieves approximately 90% transduction efficiency in IHCs and 85%–89% in OHCs throughout the entire cochlea, significantly surpassing the performance of Anc80L65 for outer hair cell targeting
[12]. Their findings showed that AAV2.7M8 also efficiently transduces inner pillar cells and inner phalangeal cells at high levels, with minimal adverse effects on auditory and vestibular functions. The vector demonstrated no statistically significant increases in circling behavior and maintained normal ABR thresholds in injected mice, establishing AAV2.7M8 as a highly effective vector for comprehensive cochlear cell targeting
[12].
Tan and colleagues engineered AAV-inner ear (ie), a novel AAV variant designed specifically for cochlear gene therapy. Their work demonstrated that AAV-ie transduces cochlear supporting cells with remarkably high efficiency (about 90%), representing a vast improvement over conventional AAV serotypes
[10]. Subsequently, researchers identified AAV-ie-K558R, a mutant variant that achieved superior transduction efficiency in cochlear supporting cells (about 80% in the basal region) while maintaining approximately 100% transduction efficiency in both IHCs and OHCs. The exceptional supporting cell tropism of AAV-ie variants proved critical for hair cell regeneration strategies, with AAV-ie-K558R-mediated delivery of
Atoh1 successfully generating hair cell-like cells from supporting cells
[11].
György et al. and colleagues identified AAV9-PHP.B through directed evolution screening of large AAV capsid libraries
[36]. Their work demonstrated that this variant achieves near-complete OHC transduction efficiency approaching 100% at high titers. Critically, their studies in non-human primates validated the translational potential of this engineered capsid for inner ear gene therapy, supporting its adoption in clinical programs for genetic deafness
[37].
Ivanchenko et al. isolated AAV-S from random peptide display capsid libraries and demonstrated that it efficiently transduces multiple cell types in both murine and non-human primate inner ears
[38]. Their findings showed that AAV-S mediates highly efficient reporter gene expression in a variety of cochlear cells, including inner and outer hair cells, fibrocytes, and supporting cells. In disease models, AAV-S encoding
CLRN1 robustly and durably rescued hearing in a mouse model of Usher syndrome type 3A, demonstrating the therapeutic potential of this engineered variant
[38].
AAV vectors constitute the core delivery platforms in all
OTOF gene-therapy programs that have progressed to clinical evaluation. These clinical candidates primarily employ three classes of capsids: naturally occurring AAV serotypes, the engineered capsid Anc80L65, and next-generation, yet-undisclosed engineered AAV variants. Therapeutic approaches developed by Decibel/Regeneron and Refreshgene utilize a dual-AAV1 system, whereas Sensorion’s SENS-501 employs dual AAV8 vectors. Programs from Akouos/Eli Lilly and Otovia rely on dual Anc80L65 vectors to reconstitute the full-length
OTOF transcript
in vivo. In parallel, Emaygene and several emerging groups are advancing novel engineered capsids designed to further enhance cochlear tropism, transduction efficiency and cross-species translation
[3].
Collectively, these efforts highlight the prominence of dual-AAV architectures as the leading strategy to overcome the large coding sequence of OTOF, with both natural and engineered capsids demonstrating robust potential for inner-ear delivery (Table 1). As new generations of designer AAVs continue to emerge, further gains in specificity, efficiency and therapeutic breadth are anticipated, positioning AAV-mediated OTOF and other gene replacement as a compelling framework for the treatment of otoferlin-related hearing loss.
5 Delivery Routes and Determinants of Transduction
The choice of delivery route significantly influences transduction patterns, spatial distribution along the cochlear spiral, and therapeutic outcome. The RWM injection is currently the dominant clinical method, adopted across all ongoing
OTOF clinical programs worldwide
[40]. This approach benefits from its minimally invasive nature, ability to limit perilymph leakage, direct access to the cochlear scala tympani, and avoidance of cochleostomy (which carries risks of additional hearing loss and ossification)
[41]. When performed with optimized surgical technique—including controlled infusion volume (typically 10–30 μL), slow infusion rates (0.5–2 μL/min), and careful monitoring of fluid backflow—RWM delivery can achieve robust inner hair cell transduction with favorable safety profiles.
Cochleostomy-based approaches, though more invasive, enable more direct access to specific cochlear regions and offer practical advantages in large-animal studies, particularly for administration to the scala media
[41,
42]. Semicircular canal injections have been explored as alternatives for broader vestibular delivery, though their application in human therapy remains limited due to risk of vestibular dysfunction
[43]. Intracochlear perfusion systems, which provide sustained vector delivery over minutes to hours, represent an emerging strategy to overcome distribution limitations, though further validation is needed before clinical adoption.
Cerebrospinal fluid (CSF) administration via cisterna magna injection represents an emerging route with considerable promise for adult patients and offers the potential for achieving bilateral transduction with a single injection
[44,
45]. The cochlear aqueduct, a natural bony channel connecting the cochlear perilymph to the cerebrospinal fluid space, provides a physiologically relevant route for AAV vector delivery from the CSF directly into the inner ear. Blanc et al. demonstrated that various AAV serotypes (AAV8, AAV9, and Anc80L65) delivered via cisterna magna injection enable high-efficiency binaural transduction of nearly all IHCs with a basal-to-apical pattern, as well as substantial spiral ganglion neuron targeting, without affecting auditory function or cochlear structures
[46].
Mathiesen et al. demonstrated that cerebrospinal fluid delivery via cisterna magna injection represents a novel route for inner ear gene therapy. Their work revealed that a single intracisternal injection of AAV vectors enables high-efficiency binaural transduction of inner hair cells and restores hearing in adult deaf mice by delivering the
Slc17A8 gene encoding VGLUT3, with minimal off-target expression in the brain and no detectable transgene expression in the liver. This approach circumvents the need for direct surgical manipulation of the delicate cochlear structures and offers significant potential for enabling effective gene therapy in adult patients with progressive genetic hearing loss
[47].
Transduction efficacy is exquisitely sensitive to multiple procedural parameters, including injection speed, perilymph turnover, vector dose, and formulation osmolarity. Overly rapid infusion or excessive volume risks mechanical trauma, inner ear pressure elevation, inflammation, or secondary hearing loss. The age at injection remains a critical variable—adult cochleae are consistently more difficult to transduce than neonatal tissue, representing a major translational barrier for treating late-diagnosed cases.
6 Integrated Clinical Evidence and Immunological Considerations
6.1 Summary of clinical trial outcomes
Phase 1/2 clinical trials of AAV-mediated
OTOF gene therapy have demonstrated quantifiable and clinically meaningful restoration of hearing in pediatric patients, marking the first successful reversal of a monogenic sensorineural deafness in humans
[14,
15,
48]. Table 2 provides a comparative overview of key clinical programs. These outcomes validate decades of preclinical development and confirm that AAV vectors can mediate functional recovery in the human cochlea. The clinical success of
OTOF programs establishes the feasibility of dual-AAV strategies for large-gene delivery, demonstrates the utility of engineered capsids in human subjects, and supports round-window administration as a safe and effective route for cochlear gene transfer
[3].
6.2 Immunological considerations for AAV-mediated inner ear therapy
The immune-privileged status of the cochlea, conferred by the blood-labyrinth barrier and limited lymphatic drainage, offers a relative sanctuary for AAV-mediated gene therapy. However, systemic and local immune responses to AAV vectors remain a significant concern that can impact safety, transduction efficiency, and long-term therapeutic durability. These challenges mirror broader immunogenicity issues observed in AAV gene therapy across other organs
[50].
Innate and adaptive immune activation: The immune system recognizes AAV vectors through both innate and adaptive mechanisms. Innate sensors, including Toll-like receptor 9 (TLR9) which detects unmethylated CpG motifs in the vector genome, and TLR2 which interacts with the viral capsid, can initiate inflammatory signaling. Although AAV typically elicits milder innate inflammation compared to other viral vectors, high vector doses—often required for efficient transduction in the adult or resistant cochlea—can amplify these signals, potentially leading to local inflammation, cytokine release, and complement activation
[51].
The adaptive immune system poses more persistent hurdles. Pre-existing neutralizing antibodies (NAbs) against AAV capsids, arising from prior natural infection with wild-type AAV, are prevalent in the human population. These antibodies can bind to and neutralize systemically administered vectors, reducing transduction efficiency. While the inner ear’s relative isolation may partially shield it from circulating NAbs during local delivery (e.g., via the round window), the risk of antibody-mediated inhibition remains, particularly if vector enters the systemic circulation or if pre-existing immunity is high
[52].
Capsid-specific T-cell responses: Capsid-specific CD8+ T-cell responses, primed by antigen-presenting cells that process AAV capsid proteins, pose a distinct risk. These cytotoxic T lymphocytes can eliminate transduced cells, leading to a loss of therapeutic gene expression and potential tissue inflammation. In clinical trials for liver-directed AAV therapy, such T-cell responses have been associated with transient elevations in liver enzymes
[53-
56]. Although less documented in the inner ear, the potential for capsid-specific T-cell-mediated toxicity warrants vigilance, especially with high-dose or repeat administration.
Local cochlear immune environment: The unique immunological microenvironment of the inner ear modulates these responses. Resident macrophages and other immune cells are present in the cochlea and can be activated by surgery or vector administration. Studies in mice and non-human primates have shown that intracochlear AAV delivery can induce local cellular infiltration, including macrophages, without necessarily causing systemic antibody titers to spike dramatically
[57]. This suggests that immune responses may be compartmentalized but nonetheless biologically active. The impact of such local inflammation on delicate cochlear structures, synaptic integrity, and long-term hair cell survival is an area of ongoing investigation.
Durability of transgene expression: Long-term transgene expression is a hallmark of AAV therapy in post-mitotic tissues. In the inner ear, studies in animal models have demonstrated stable expression for over 12–18 months
[58]. Early clinical data for
OTOF gene therapy also show sustained auditory improvement over follow-up periods exceeding one year, suggesting durable expression
[14,
15]. However, immune-mediated clearance of transduced cells, either via antibody-dependent mechanisms or cytotoxic T cells, remains a theoretical threat to long-term durability. The induction of regulatory T cells (Tregs), which can promote immune tolerance to the vector or transgene, has been observed in some muscle- and liver-directed AAV trials and may play a beneficial role in stabilizing expression
[59]. Whether similar tolerogenic mechanisms can be engaged in the inner ear remains unknown.
7 Translational Gaps and Remaining Challenges
The emerging clinical data illuminate persistent obstacles that must be addressed before inner-ear gene therapy can achieve broad clinical adoption.
Narrow therapeutic window and variability: A narrow therapeutic window remains a central challenge: doses below threshold yield insufficient transduction, whereas higher doses may risk ototoxicity. Inter-individual variability in vector uptake—likely influenced by round-window permeability, inner-ear fluid dynamics, and host biological factors—underscores the need for carefully structured dose-escalation paradigms
[26]. Continued safety monitoring remains essential, including vigilance for inflammatory responses, high-dose ototoxicity, and the long-term stability of transgene expression
[14,
15,
26,
48].
Adult cochlear transduction: Efficient adult cochlear transduction remains a major unmet need, particularly for disorders that are not diagnosed in infancy or for patients who present later in life. The biological barriers in the mature cochlea (e.g., tighter tissue organization, reduced receptor availability) necessitate the development of next-generation vectors with enhanced adult tropism and delivery strategies like CSF administration that may better circumvent these barriers
[46].
Expanding targetable cell types: Broadening the range of targetable cell types—including outer hair cells, supporting cells, and spiral ganglion neurons—will be necessary to address the full spectrum of hereditary and acquired hearing loss. Disorders such as Usher syndrome or
GJB2-related hearing loss may require correction in multiple cell types for complete functional restoration
[60].
Manufacturing and regulatory hurdles: At the manufacturing level, the production of high-dose AAV formulations at clinical scale poses technical and regulatory challenges, as does the development of robust analytical methods for detecting anti-AAV immunity and monitoring long-term immunological effects. Finally, evolving regulatory expectations around long-term safety, vector quality, and benefit–risk assessment for inner-ear gene therapy will shape the pathway to eventual widespread clinical use
[3,
16,
26,
61,
62].
8 Next-Generation Strategies to Advance AAV Gene Therapy
The field stands at a clinical inflection point, propelled by the success of OTOF therapy. Future efforts must focus on precision, scalability, and expansion to other forms of deafness. The following structured roadmap outlines key priorities.
8.1 Capsid and vector optimization
Advanced capsid engineering platforms leveraging deep mutational scanning, computational design, machine learning, and large-scale directed evolution with libraries approaching more than 10
8 variants promise to yield next-generation vectors optimized for distinct cochlear cell types and capable of superior performance in adult tissue
[63,
64]. Structure-guided rational design, informed by cryo-electron microscopy structures of AAV-capsid-receptor complexes, offers additional opportunities for targeted improvement
[64].
In parallel, regulatory-element engineering is gaining substantial momentum
[65]. Enhancer-promoter combinations identified through multi-omics profiling, single-cell transcriptomics, high-throughput screening platforms, or
in vivo reconstruction systems promise the possibility of precise, cell-restricted expression, thereby enhancing both safety and therapeutic relevance. In particular, promoters and regulatory elements discovered through single-cell RNA sequencing of healthy and diseased cochleae offer new opportunities for cell-type specificity
[66].
8.2 Delivery and surgical refinement
Innovations in delivery technology are critical. This includes optimizing RWM injection protocols for consistency, developing less invasive alternative routes (e.g., sustained-release intracochlear devices), and fully exploring the translational potential of CSF-mediated delivery for bilateral treatment in adults
[46]. Surgical robotics and real-time imaging guidance could enhance precision and reproducibility.
8.3 Expanding the genetic therapeutic landscape
While
OTOF has led the way, the pipeline must expand.
GJB2 gene therapy represents the next major frontier. Mutations in
GJB2, encoding connexin 26, are the most common cause of hereditary hearing loss globally. Strategies are evolving from simple gene replacement for recessive loss-of-function alleles to more complex approaches like base editing to correct dominant-negative mutations, as recently demonstrated in preclinical models
[6]. Key challenges include achieving efficient transduction in the specific supporting cells of the cochlea that express
GJB2 and ensuring proper gap junction assembly. Other high-priority targets include
CDH23 (Usher syndrome type 1D),
PCDH15 (Usher syndrome type 1F), and
TMC1, each presenting unique cell tropism and biological correction challenges.
8.4 Immunological and safety hurdles
As discussed in Section 6.2, mitigating immune responses is paramount for safety and durability. Future work will focus on engineering immune-evasive capsids, developing effective yet transient immunomodulatory regimens, and establishing biomarkers for monitoring immune activation post-treatment. Long-term surveillance in clinical trials is essential to fully understand the safety profile.
8.5 Manufacturing, regulatory, and access pathways
Scalable, cost-effective manufacturing processes for high-titer, clinical-grade AAV, particularly for dual-vector systems, must be developed. Internationally harmonized regulatory pathways tailored to the unique aspects of inner ear gene therapy (e.g., endpoint definitions, surgical standards) will facilitate global development. Equitable access strategies are needed to ensure these transformative therapies reach patients worldwide.
8.6 Combinatorial and regenerative approaches
Beyond gene replacement, combinatorial approaches pairing gene correction with regenerative medicine strategies—such as AAV-mediated delivery of transcription factors (e.g., Atoh1) to convert supporting cells into hair cells—represent a promising frontier for treating not only genetic but also acquired hearing loss. The integration of human cochlear organoids and predictive animal models (e.g., non-human primates, tree shrews) into preclinical pipelines will accelerate the translation of these advanced therapies.
9 Conclusion
AAV-mediated gene therapy for hereditary hearing loss has rapidly evolved from a conceptual possibility to a clinically validated therapeutic modality. Early successes in OTOF replacement have demonstrated that precise and durable restoration of auditory function is achievable, establishing a transformative precedent for inner ear gene therapy. These advances are driven by synergistic progress in engineered AAV capsids, dual-vector systems capable of delivering large genes, increasingly refined microsurgical approaches, and rigorous clinical evaluation frameworks.
Looking ahead, the field is poised for broad expansion. Key priorities include the development of next-generation vectors with enhanced adult cochlear transduction, the extension of therapeutic strategies to additional genetic etiologies such as GJB2, CDH23 and PCDH15, and the refinement of delivery technologies that enable safe, accurate, and age-independent access to the cochlea. Integration of artificial intelligence with high-throughput capsid engineering, enhancer design, and patient-derived organoid platforms is expected to accelerate vector discovery and enable a new level of molecular precision. In parallel, scalable manufacturing capabilities and internationally harmonized regulatory pathways will be essential for translating laboratory innovation into equitable global therapies.
As the evidence base continues to grow, AAV-mediated cochlear gene therapy is positioned to become a standard component of otologic practice, offering the possibility of life-changing restoration of hearing to millions of patients with genetic deafness. The convergence of molecular engineering, neurosensory biology, and clinical innovation signals not only the maturation of a therapeutic field, but the beginning of a new era in precision hearing restoration.
The Author(s) 2025. This article is available under open access at journal.hep.com.cn.
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