Biology subjects

Ivanchenko, M. V.

Publications and source records attributed to Ivanchenko, M. V..

5 recordsLinked to original sources

bioRxiv2025

Mini-Pcdh15b Gene Therapy Rescues Visual Deficits in a Zebrafish Model of Usher Syndrome Type 1F

Usher syndrome type 1F (USH1F) is a severe inherited disorder caused by mutations in PCDH15, resulting in congenital deafness, vestibular dysfunction, and progressive retinal degeneration leading to blindness. While cochlear implantation can restore hearing, no therapeutic interventions currently exist for vision loss. Gene augmentation therapy represents a promising approach; however, the PCDH15 coding sequence ([~]5.3 kb) exceeds the packaging capacity of adeno-associated virus (AAV) vectors. To overcome this limitation, we previously engineered shortened "mini-PCDH15" constructs that retain key structural and functional domains while fitting within a single AAV. Among these, the mini-PCDH15-V4 variant successfully restored hearing in Pcdh15-deficient mice. Here, we investigated the ability of a cone-targeted mini-pcdh15b-V4 transgene to rescue vision in a zebrafish model of USH1F. Untreated pcdh15b-deficient zebrafish exhibited severe structural and functional defects of the retina, including disorganized and shortened photoreceptor outer segments, disrupted calyceal processes, and markedly reduced electroretinogram (ERG) and optokinetic response (OKR) performance. Targeted expression of mini-pcdh15b-V4 recapitulated typical localization of Pcdh15b to calyceal processes and outer segment membranes, rescued photoreceptor architecture, and re-established both structural organization and functional output. Treated mutants exhibited improved visual tracking behavior and full recovery of ERG a-wave and b-wave amplitudes, indicating restoration of photoreceptor and synaptic function. Importantly, mini-pcdh15b-V4 expression produced no adverse effects in wild-type or heterozygous fish, supporting the safety of cone-specific expression. Together, these findings demonstrate that mini-pcdh15b-V4 can restore both photoreceptor structure and visual function in pcdh15b-deficient zebrafish. This work establishes the pcdh15b mutant zebrafish as a powerful preclinical model for studying USH1F retinopathy and supports the translational potential of rationally engineered mini-PCDH15 constructs as a feasible gene therapy approach for preventing or reversing vision loss in individuals with USH1F.

neuroscience↗

bioRxiv2024

Cell-specific delivery of GJB2 restores auditory function in mouse models of DFNB1 deafness and mediates appropriate expression in NHP cochlea

Mutations in the GJB2 gene cause the most common form of human hereditary hearing loss, known as DFNB1. GJB2 is expressed in two cell groups of the cochlea--epithelial cells of the organ of Corti and fibrocytes of the inner sulcus and lateral wall--but not by sensory hair cells or neurons. Attempts to treat mouse models of DFNB1 with AAV vectors mediating nonspecific Gjb2 expression have not substantially restored function, perhaps because inappropriate expression in hair cells and neurons could compromise their electrical activity. Here, we used genomic chromatin accessibility profiling to identify candidate gene regulatory elements (GREs) that could drive cell-type-specific expression of Gjb2 in the cochlea. HA-tagged GJB2, delivered to a conditional knockout model in an AAV vector with GRE control of expression, was localized to the appropriate cell types, prevented the cochlear degeneration observed in untreated knockout mice, and partially rescued hearing sensitivity. In a Gjb2 partial knockdown mouse model, such exogenous GJB2 prevented degeneration and completely restored hearing sensitivity. We tested control of expression by these GREs in nonhuman primate cochleas and found that vector-delivered human GJB2.HA was located in the appropriate cell types and caused little or no reduction in hearing sensitivity. Together, these findings suggest that GRE-mediated expression of GJB2 could prevent hearing loss in DFNB1 patients.

neuroscience↗

bioRxiv2024

Elasticity and Thermal Stability are Key Determinants of Hearing Rescue by Mini-Protocadherin-15 Proteins

Protocadherin-15 is a core protein component of inner-ear hair-cell tip links pulling on transduction channels essential for hearing and balance. Protocadherin-15 defects can result in non-syndromic deafness or Usher syndrome type 1F (USH1F) with hearing loss, balance deficits, and progressive blindness. Three rationally engineered shortened versions of protocadherin-15 (mini-PCDH15s) amenable for gene therapy have been used to rescue function in USH1F mouse models. Two can successfully or partially rescue hearing, while another one fails. Here we show that despite varying levels of hearing rescue, all three mini-PCDH15 versions can rescue hair-cell mechanotransduction. Negative-stain electron microscopy shows that all three versions form dimers like the wild-type protein, while crystal structures of some engineered fragments show that these can properly fold and bind calcium ions essential for function. In contrast, simulations predict distinct elasticities and nano differential scanning fluorimetry shows differences in melting temperature measurements. Our data suggest that elasticity and thermal stability are key determinants of sustained hearing rescue by mini-PCDH15s.

bioengineering↗

bioRxiv2023

Hexafluoro slows retinal degeneration and improves visual function in zebrafish models of Usher syndrome 1F

Usher syndrome is the leading genetic cause of deafblindness, affecting hundreds of thousands of people worldwide. The deafness can be addressed with hearing aids or cochlear implants, but there is currently no treatment for the vision loss, which is due to progressive degeneration of retinal photoreceptors. Studies in animal models of Usher syndrome have shown that photoreceptor degeneration is exacerbated by exposure to bright light, and other studies have shown that light-induced photostress reduces mitochondrial function. We previously synthesized hexafluoro and showed that it is a potent Sirt3 activator that promotes mitochondrial respiration. Here we examined the efficacy of hexafluoro as a potential therapeutic for treatment of vison loss in a zebrafish model of Usher syndrome type 1F, which exhibits early and severe vision defects along with vestibular dysfunction as seen in Usher type 1 pathology. We find that hexafluoro improves visual function, reduces photoreceptor degeneration, and protects the retina against exposure to bright light in this USH1F model.

neuroscience↗

bioRxiv2023

PCDH15 Dual-AAV Gene Therapy for Deafness and Blindness in Usher Syndrome Type 1F

Usher syndrome type 1F (USH1F), resulting from mutations in the protocadherin-15 (PCDH15) gene, is characterized by congenital lack of hearing and balance, and progressive blindness in the form of retinitis pigmentosa. In this study, we explore a novel approach for USH1F gene therapy, exceeding the single AAV packaging limit by employing a dual adeno-associated virus (AAV) strategy to deliver the full-length PCDH15 coding sequence. We demonstrate the efficacy of this strategy in mouse USH1F models, effectively restoring hearing and balance in these mice. Importantly, our approach also proves successful in expressing PCDH15 in clinically relevant retinal models, including human retinal organoids and non-human primate retina, showing efficient targeting of photoreceptors and proper protein expression in the calyceal processes. This research represents a major step toward advancing gene therapy for USH1F and the multiple challenges of hearing, balance, and vision impairment.

neuroscience↗