Chemical dissection of eIF4A-mediated translation

The ribosome recruitment phase of translation initiation is the rate-limiting step of protein synthesis and involves binding of the 40S ribosomal subunit and associated factors to the 5'end of mRNAs. Ribosome recruitment to mRNA templates is catalyzed by the eIF4 class of initiation factors comprising eIF4F, a multisubunit complex that binds to the 5'end of mRNAs and prepares transcripts to receive ribosomes, as well as eIF4B and eIF4H, which help eIF4F in its task. mRNA preparation by eIF4F is thought to be accomplished by the eIF4A subunit of the eIF4F complex, an RNA helicase believed to unwind secondary structure close to the cap structure to facilitate ribosome binding.Deregulated ribosome binding has been shown to contribute to cellular transformation. The ribosome recruitment phase of initiation therefore emerged as an attractive chemotherapeutic target, however a limited number of approaches have been described that target this process in vivo. This prompted our laboratory to conduct a high throughput screening campaign aimed at identifying novel small molecule inhibitors of the ribosome recruitment phase of translation initiation. We identified three types of natural products; pateamine, hippuristanol and cyclopenta[b]benzofurans, that were all found to inhibit translation initiation in vitro and in vivo, and to target the eIF4A subunit of the eIF4F complex.Characterization of the small molecules identified in the high throughput screen revealed that although these compounds target the same initiation factor, their mechanism of action is different. Given the role of the ribosome recruitment phase of translation initiation in tumorigenesis and chemoresistance, the effects of cyclopenta[b]benzofurans were tested in a mouse lymphoma model of drug resistance mediated by activation of the PI3K/mTOR pathway, and one compound, silvestrol, was found to reverse chemoresistance in this system. Characterization of the eIF4A modulators described herein also revealed that these compounds can be used to study cellular processes involving translation. The small molecules described in this thesis therefore represent potential drug leads for cancer therapy as well as a powerful arsenal of small molecules to evaluate the role of eIF4A and translation initiation in various cellular processes