Biology subjects

VanBenschoten, H.

Publications and source records attributed to VanBenschoten, H..

3 recordsLinked to original sources

bioRxiv2026

A Designed Ankyrin Repeat Protein (DARPin) Targeting EGFR Inhibits Ovulation and Enables a Novel Platform for Studying Ovarian Biology and Pathophysiology

Ovarian disorders, including anovulation, primary ovarian insufficiency (POI), and polyendocrine metabolic ovarian syndrome (PMOS), affect millions of reproductive-age women worldwide; however, mechanistic studies of ovarian biology and pathophysiology remain challenging because current experimental approaches often lack selectivity, tunability, or physiological relevance. Genetically modified animal models are labor-intensive and irreversible; small molecules frequently exhibit off-target effects; and conventional antibodies have limited tissue penetration and restricted temporal control. Designed ankyrin repeat proteins (DARPins) represent a highly modular protein engineering platform with advantages in specificity, size, stability, and extracellular targeting, but their utility in reproductive biology remains largely unexplored. Here, we used epidermal growth factor receptor (EGFR)-targeting DARPins as a proof-of-concept platform to interrogate ovarian signaling during ovulation. Screening of engineered anti-EGFR DARPins identified SX-006, a bispecific tetravalent construct with robust cross-species EGFR binding and potent biological activity. Using an ex vivo murine ovulation system, SX-006 inhibited follicle rupture in a dose-dependent manner with IC50 of 1.21 M without overt cytotoxicity. Lower concentrations of SX-006 preferentially perturbed follicle rupture while largely preserving oocyte meiotic maturation and luteinization, suggesting differential sensitivity of ovulatory processes to extracellular EGFR blockade. Comparative transcriptomic analyses further revealed that extracellular EGFR blockade and small molecule-based intracellular EGFR kinase inhibition produce overlapping but also distinct transcriptional responses, supporting biologically distinct modes of ovulatory signaling pathway perturbation. Together, these findings establish DARPins as a selective, tunable, and physiologically relevant platform for studying ovarian signaling and provide proof-of-concept for extracellular receptor targeting in ovarian biology, infertility research, and non-hormonal contraceptive development. Summary sentenceAn engineered EGFR-targeting DARPin selectively inhibits ovulation through extracellular receptor blockade and establishes a versatile platform for investigating ovarian signaling and reproductive disorders.

pharmacology and toxicology↗

bioRxiv2025

A pre-menopausal single-cell atlas for ovarian drug discovery

Multi-tissue single-cell atlas efforts have transformed our understanding of cellular diversity across the human body and led to the creation of harmonized resources to advance research and therapeutic development. Ovarian biology, however, often remains underrepresented in these resources and is rarely analyzed with menopausal status as a biological variable. Here, we present the Menopause Cell Map (MenoMap), an integrated single-cell resource comprising more than 2 million cells from 13 healthy human tissues, including the ovary, from pre- and post-menopausal age donors. This harmonized atlas leverages curated samples from healthy female donors and enables transcriptomic comparisons across cell types, tissues, and organs while preserving menopausal status based on age as an interpretable variable. Using this resource, we show that menopause-associated gene expression changes are highly context dependent, with prominent remodeling in ovarian stromal, endothelial, immune, and reproductive cell populations. Within the ovary, post-menopausal remodeling rewired intercellular communication and shifted reproductive and steroidogenic programs toward collagen-integrin signaling, endothelial-to-mesenchymal transition, and senescence concentrated in the endothelial and stromal compartments. Cross-species comparison with young and aged mouse ovarian single-cell data showed these endothelial and stromal changes were conserved with age, most prominently in tumor necrosis factor-nuclear factor-kappa-beta signaling. Finally, we apply Human Protein Atlas-inspired specificity rules and fertility phenotype annotations to evaluate how menopausal age status affects tissue- and cell-type-specific gene classification and to prioritize ovary-enriched genes for downstream biological and translational investigation. Through this analysis, we find that the tissue specificity classification of several genes involved in reproductive-specific programs change in the pre- to post-menopausal transition, highlighting the need for age-aware and healthy donors in multi-tissue single-cell atlas efforts. Together, the MenoMap provides a human single-cell framework that leverages existing and standardized single-cell datasets curated for studying ovarian biology across reproductive aging, evaluating the influence of menopausal status on gene expression and tissue specificity, and nominating candidate genes for future investigation in reproductive biology, fertility, and target discovery.

bioinformatics↗

bioRxiv2025

Transcriptomic analysis of whole staged ovarian follicles reveals stage-specific folliculogenesis signatures in mice

Activation and maturation of ovarian follicles are essential for female reproduction, yet the underlying molecular and transcriptional mechanisms that govern these processes remain poorly understood. In this study, we used single follicle RNA-sequencing (RNA-seq) to identify transcriptional signatures of staged ovarian follicles, from primordial to secondary stages, to uncover the genes and pathways involved in early folliculogenesis. Our findings demonstrate that primordial follicles are transcriptionally distinct from growing follicles, with enrichment in DNA integrity and RNA processing pathways, which may play a role in preserving oocyte genomic stability and cell state during dormancy. Additionally, our analysis reveals minimal transcriptomic differences between primary and secondary follicles using traditional differential expression analysis. To better distinguish growing follicle stages, we introduce unsupervised approaches, including discrete-variable predictors of follicle stage and weighted gene co-expression analysis. We identified pathways involved in DNA integrity, meiotic arrest, and cellular metabolism that drive the transition from dormant to active follicle states, as well as pathways related to cellular growth, ECM organization, and biosynthesis in growing follicle stages. Our study offers novel insights into the molecular mechanisms governing early follicle activation and growth, providing a foundation for future research with applications in reproductive biology, contraception, and fertility preservation. Author SummaryThe development of ovarian follicles is essential for female fertility, but the molecular signals that control their growth remain unclear. In this study, we used advanced gene sequencing techniques to analyze the genetic activity of individual ovarian follicles at different stages of early development. We found that dormant follicles have unique gene expression patterns that help protect the genetic material of the egg and maintain their inactive state. In contrast, follicles that have begun to grow show increased activity in genes related to cell growth, communication, and structural changes. Interestingly, we observed that early growing follicles are more similar to each other than previously thought, prompting us to apply new analytical methods to better distinguish their developmental stages. Our findings highlight key biological pathways that regulate the transition from dormant to active follicles and uncover new genes that may play a role in this process. Understanding these mechanisms provides valuable insights into ovarian biology and could inform future research on fertility treatments, contraception, and reproductive health.

bioinformatics↗