A forward genetic screen identifies modifiers of rocaglate responsiveness

Leo Shen,Lauren Pugsley,John A Porco,David Langlais,Regina Cencic,Jerry Pelletier,Geneviève Morin,Sai Kiran Naineni,Wenhan Zhang,Ananya Sahni,Hanchen Wang,Francis Robert,Anastasia Nijnik,Sidong Huang

doi:10.1038/s41598-021-97765-8

Leo Shen, Lauren Pugsley + Show 12 more

Open Access

https://doi.org/10.1038/s41598-021-97765-8

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Abstract

Rocaglates are a class of eukaryotic translation initiation inhibitors that are being explored as chemotherapeutic agents. They function by targeting eukaryotic initiation factor (eIF) 4A, an RNA helicase critical for recruitment of the 40S ribosome (and associated factors) to mRNA templates. Rocaglates perturb eIF4A activity by imparting a gain-of-function activity to eIF4A and mediating clamping to RNA. To appreciate how rocaglates could best be enabled in the clinic, an understanding of resistance mechanisms is important, as this could inform on strategies to bypass such events as well as identify responsive tumor types. Here, we report on the results of a positive selection, ORFeome screen aimed at identifying cDNAs capable of conferring resistance to rocaglates. Two of the most potent modifiers of rocaglate response identified were the transcription factors FOXP3 and NR1I3, both of which have been implicated in ABCB1 regulation—the gene encoding P-glycoprotein (Pgp). Pgp has previously been implicated in conferring resistance to silvestrol, a naturally occurring rocaglate, and we show here that this extends to additional synthetic rocaglate derivatives. In addition, FOXP3 and NR1I3 impart a multi-drug resistant phenotype that is reversed upon inhibition of Pgp, suggesting a potential therapeutic combination strategy.

Highlights

Rocaglates are a class of eukaryotic translation initiation inhibitors that are being explored as chemotherapeutic agents
Eukaryotic initiation factor 4F resides at this nexus and its activity dictates the spectrum of mRNAs that enter into the initiation phase and that are subsequently translated6. eIF4F consists of (1) an eIF4E subunit which recognizes cap structures, (2) an eIF4A DEAD-box RNA helicase, and (3) a multi-domain scaffolding protein, eIF4G
Despite this profound impact on global translation, a therapeutic index is achievable in the clinic and is thought to result from Chronic myeloid leukemia (CML) being addicted to oncogenic drivers that have short half-lives (e.g. MYC, MCL-1, cyclin D1) which become rapidly depleted in transformed cells upon inhibition of translation[19]

Summary

Introduction

Rocaglates are a class of eukaryotic translation initiation inhibitors that are being explored as chemotherapeutic agents. The normal cellular constraints on translation regulation mediated at the ribosome recruitment step of initiation are removed, and dysregulated protein synthesis ensues to support remodeling of the tumor cell p roteome[2,3,4] This dysregulation promotes increased proliferation, resistance to apoptosis, and enhanced metastatic p otential[5]. Rocaglates (a class of natural products characterized by the densely functionalized cyclopenta[b]benzofuran skeleton) exhibit potent activity (Fig. 1a) They impart onto eIF4A and eIF4F gain-of-function properties. Validation by inducing clamping between said proteins and RNA to block ribosome recruitment and s canning[12,13,14] These compounds exhibit anti-neoplastic activity towards a large range of cancer types in pre-clinical mouse models[5], and this activity is linked to eIF4A target engagement and inhibition[14,15,16].

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