Abstract 4878: Transcriptional response to rocaglate translation inhibitors in lymphoma shows cellular adaptation through induction of stress and survival pathways
Authors: Manara Paola , Cunningham Tyler Andrew , Ho Jr Jyun David , Lightfuss Olivia B. , Alaoui Abdessamad Youssfi , Schatz Jonathan Harry 
Published: June 2023

Abstract
Background: 40% of diffuse large B-cell lymphoma (DLBCL) patients do not achieve cure in response to the standard immunochemotherapy. Relapsed and refractory tumors respond poorly to targeted signaling inhibitors due to pathway redundancies in this genetically heterogenous disease. Pharmacologic disruption of the cap-initiation complex eIF4F is an emerging clinical treatment strategy to bypass resistance. In preclinical experiments, B-lymphomas show high susceptibility to rocaglates, natural and synthetic cap-dependent translation inhibitors. Among these compounds, eFT226 (zotatifin), is currently in phase I/II clinical evaluation for solid tumors. Rocaglate mechanisms remain incompletely defined for use in B-cell lymphoma. Recent findings in our laboratory revealed translationally mediated adaptation that increased synthesis of many targets. Proteins (i.e.CD98hc) that paradoxically increase in production contribute to persistence of cells that survive treatment. By contrast, transcriptional responses to rocaglates remain minimally studied. Here we sought to fill this gap through unbiased transcriptomics.
Method: RNA sequencing (RNA-seq) was performed on zotatifin treated DLBCL cells (SU-DHL10) vs DMSO controls. Genes with a false discovery rate p-value (FDR) ≤0.05, logarithm of counts per million reads (logCPM) ≥ 0 and P value ≤0.05 were considered differentially expressed. Log2 fold change (FC) ≥ +/-2-fold for a specific gene was considered significantly differentially expressed between the two groups. Pathway analysis was performed using Gene Set Enrichment Analysis (GSEA).
Results: We found 4450 differentially expressed genes, 12 significantly upregulated, and 42 significantly downregulated in response to zotatifin (FC ≥ 2). Further pathways analyses showed 4/25 gene sets significantly upregulated, and 3/22 gene sets significantly downregulated (FDR &lt;0.25). We found significantly upregulated genes related to mechanisms stress and survival pathways including NF-kB induction by TNF signaling (normalized enriched score (NES)=2.28; p-value &lt;0.0001), genes induced in response to hypoxia translational stress responses (NES=1.85; nominal p-value=0.008).
Conclusions: These results implicate induction of specific survival pathways that induce transcriptionally mediated adaptation to the stress of rocaglate therapy, but it is unclear if these responses can result in altered protein production in the context of eIF4A inhibition. Parallel assessment of the translatome currently under analysis will provide a more complete picture of overall adaptation and provide rational combination targets for rocaglates in lymphoma as clinical development progresses.
Citation Format: Paola Manara, Tyler Andrew Cunningham, Jr Jyun David Ho, Olivia B. Lightfuss, Abdessamad Youssfi Alaoui, Jonathan Harry Schatz. Transcriptional response to rocaglate translation inhibitors in lymphoma shows cellular adaptation through induction of stress and survival pathways. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4878.