Letourneau, E. | Search bioRxiv

bioRxiv2026

Novel Replication Stress-Associated High-Grade Serous Ovarian Cancer Models Recapitulate Diverse Tumour Ecosystems and Enable Preclinical Studies

Abstract Background: High-grade serous ovarian cancer (HGSC) is characterized by extensive genomic heterogeneity and frequent alterations that promote replication stress (RS). Although RS has emerged as a promising therapeutic vulnerability, it remains unclear whether distinct RS-associated genomic alterations generate similar tumour states or drive unique biological programs and therapeutic dependencies. Methods: We developed and characterized a panel of syngeneic HGSC models representing recurrent RS-associated alterations, including overexpression of Ccne1, Brd4, Myc, Ndrg1, Pik3ca, and Brca1 heterozygosity, alongside previously established Pten/Nf1-deficient and Myc overexpressing models. Tumour fitness, transcriptomic programs, histopathology, extracellular matrix composition, immune organization, and therapeutic vulnerabilities were assessed using subcutaneous (s.c.) and intraperitoneal (i.p.) allografts, RNA sequencing (RNAseq), mass spectrometry (MS), cyclic-immunofluorescence (Cyc-IF), and preclinical therapeutic studies. Results: RS-associated genotypes exhibited marked differences in tumour fitness and generated distinct tumour ecosystem states despite sharing a common genetic background. Tumours harbouring BRCA1 heterozygosity displayed reduced tumour fitness, mesenchymal features, extracellular matrix remodelling, stromal expansion, and immune-excluded phenotypes. In contrast, tumours co-expressing Ccne1 and Brd4 exhibited the highest RS level together with strong activation of DNA damage response pathways and limited immune infiltration. Immune characterization identified immune-desert, immune-suppressed, and immune-excluded states across models, demonstrating substantial genotype-dependent immune heterogeneity. Elevated CD47 expression emerged as a defining feature of the Ccne1/Brd4 tumour state and was confirmed in human HGSC specimens with high CCNE1 and BRD4 protein expression. Therapeutic targeting of ATR and CD47 in intraperitoneal Ccne1/Brd4 allografts reduced tumour burden and induced histopathological responses compared with either monotherapy alone, supporting the translational relevance of this RS-high tumour state. Conclusions: Distinct RS-associated alterations do not converge on a common HGSC phenotype but instead generate diverse tumour ecosystem states with unique effects on tumour fitness, stromal remodelling, immune organization, and therapeutic vulnerabilities. These findings highlight the importance of considering the genetic origin of RS when developing therapeutic strategies and establish a framework for investigating genotype-specific vulnerabilities in HGSC.

cancer biology↗

bioRxiv2026

Highly effective modulator therapy hinders the emergence of hyperbiofilm variants of Pseudomonas aeruginosa PA14 grown in a cystic fibrosis lung model

Pseudomonas aeruginosa is an opportunistic pathogen that often adopts persistent phenotypes -- such as biofilm formation-- that are associated with chronic infections including those observed in the cystic fibrosis (CF) lung Recently, highly effective modulator therapy (HEMT) such as elexacaftor/tezacaftor/ivacaftor (ETI) has significantly improved the quality of life of people with CF (pwCF). Yet a potential direct impact of ETI on the physiology of P. aeruginosa during growth to a remodeled CF lung environment has remained unexplored. To address this, we conducted an experimental evolution using P. aeruginosa PA14 grown in CF-like conditions in the presence or absence of ETI. We observed a marked reduction in biofilm formation and in the number of small colony variants (SCVs) for P. aeruginosa populations evolved under ETI treatment. Also, sequencing of specific evolved clones exhibiting distinct morphotypes revealed two major observations: (i) P. aeruginosa-evolved communities exposed to ETI retained a wild type-like morphotype and, (ii) P. aeruginosa populations evolved in the absence of ETI adopted a SCV-like phenotype with mutations acquired in the Wsp chemosensory pathway. Furthermore, analysis of evolved populations revealed that ETI treatment likely modulates c-di-GMP pools by driving mutations in an enzyme catalyzing the degradation of this second messenger. Overall, our work suggests that ETI has the potential to hinder the acute to chronic biofilm transition of P. aeruginosa thereby limiting the emergence of variants typically associated with long-term CF lung colonization.

microbiology↗