Biology subjects

Dupuy, A. J.

Publications and source records attributed to Dupuy, A. J..

4 recordsLinked to original sources

bioRxiv2025

Co-targeting the PI3K-Akt pathway improves response to MEK inhibition in low-grade serous ovarian cancer cell lines

Low grade serous ovarian cancer (LGSOC) accounts for [~]5% of all ovarian cancers, with >1,000 new cases diagnosed each year in the United States. Currently, there is no established standard-of-care treatment for this subtype, and chemotherapy response rates are only [~]4%. One defining characteristic of LGSOC is a high prevalence of mutations within the MAPK pathway, which has motivated studies using MEK inhibitors to treat this disease. A recent phase II/III clinical trial using trametinib to treat patients with recurrent LGSOC improved the response rate to 26%; however, all patients eventually progressed due to the emergence of therapeutic resistance. Ongoing clinical trials are investigating combined inhibition of MEK and FAK with avutometinib plus defactinib for LGSOC patients, with preliminary results demonstrating response rates near 50%. While these results are highly encouraging, a significant subset of patients still fails to respond to existing targeted treatments. Additionally, development of therapeutic resistance in initial responders remains a major obstacle. Here, we sought to characterize molecular responses of LGSOC to MEK inhibition, both acutely and during the development of spontaneous drug resistance, with a goal of identifying adaptive mechanisms LGSOC cells utilize to withstand MAPK inhibition. Our research identified significant upregulation of the PI3K-Akt pathway in response to MEK inhibition. While targeted inhibition of AKT had minimal impact on its own, combination with MEK inhibitors produced strong synergistic suppression of proliferation in LGSOC cells. This combination strategy could potentially be used to prevent or reverse the emergence of MEK inhibitor resistance in LGSOC patients.

cancer biology↗

bioRxiv2022

In vivo functional genomics identifies essentiality of potassium homeostasis in medulloblastoma

The identification of cancer maintenance genes--driver genes essential to tumor survival--is fundamental for developing effective cancer therapy. Transposon-based insertional mutagenesis screens can identify cancer driver genes broadly but not discriminate maintenance from progression or initiation drivers, which contribute to cancer phenotypes and tumorigenesis, respectively. We engineered a nested, double-jumping transposon system to first dysregulate gene expression during tumorigenesis and then restore gene expression following tumor induction, allowing for genome-wide screening of maintenance essentiality in vivo. In a mouse model of medulloblastoma, the most common pediatric malignancy, insertion and remobilization of this nested transposon uncovers potassium channel genes as recurrent maintenance drivers. In human medulloblastoma, KCNB2 is the most overexpressed potassium channel across Group 3, Group 4, and SHH subgroups, and Kcnb2 knockout in mice diminishes the replicative potential of medulloblastoma-propagating cells to mitigate tumor growth. Kcnb2 governs potassium homeostasis to regulate plasma membrane tension-gated EGFR signaling, which drives proliferative expansion of medulloblastoma-propagating cells. Thus, our novel transposon system reveals potassium homeostasis as essential to tumor maintenance through biomechanical modulation of membrane signaling.

cancer biology↗

bioRxiv2019

A simplified transposon mutagenesis method to perform phenotypic forward genetic screens in cultured cells

The introduction of genome-wide shRNA and CRISPR libraries has facilitated cell-based screens to identify loss-of-function mutations associated with a phenotype of interest. Approaches to perform analogous gain-of-function screens are less common, although some reports have utilized arrayed viral expression libraries or the CRISPR activation system. However, a variety of technical and logistical challenges make these approaches difficult for many labs to execute. In addition, genome-wide shRNA or CRISPR libraries typically contain of hundreds of thousands of individual engineered elements, and the associated complexity creates issues with replication and reproducibility for these methods. Here we describe a simple, reproducible approach using the Sleeping Beauty transposon system to perform phenotypic cell-based genetic screens. This approach employs only three plasmids to perform unbiased, whole-genome transposon mutagenesis. We also describe a ligation-mediated PCR method that can be used in conjunction with the included software tools to map raw sequence data, identify candidate genes associated with phenotypes of interest, and predict the impact of recurrent transposon insertions on candidate gene function. Finally, we demonstrate the high reproducibility of our approach by having three individuals perform independent replicates of a mutagenesis screen to identify drivers of vemurafenib resistance in cultured melanoma cells. Collectively, our work establishes a facile, adaptable method that can be performed by labs of any size to perform robust, genome-wide screens to identify genes that influence phenotypes of interest.

genomics↗

bioRxiv2019

Src-dependent DBL family members drive resistance to vemurafenib in human melanoma

The use of selective BRAF inhibitors (BRAFi) has produced remarkable outcomes for patients with advanced cutaneous melanoma harboring a BRAFV600E mutation. Unfortunately, the majority of patients eventually develop drug-resistant disease. We employed a genetic screening approach to identify gain-of-function mechanisms of BRAFi resistance in two independent melanoma cell lines. Our screens identified both known and unappreciated drivers of BRAFi resistance, including multiple members of the DBL family. Mechanistic studies identified a DBL/Rac1/Pak signaling axis capable of driving resistance to both current and next-generation BRAF inhibitors. However, we show that the Src inhibitor, saracatinib, can block the DBL-driven resistance. Our work highlights the utility of our straightforward genetic screening method in identifying new drug combinations to combat acquired BRAFi resistance.

cancer biology↗