Investigating the Mechanism and Species-Specificity of Thalidomide Derivatives

Abstract

Thalidomide, once infamous for its teratogenic effects, and its derivatives lenalidomide and pomalidomide have found new clinical utility as highly effective treatments for multiple myeloma, other B cell neoplasms, and myelodysplastic syndrome (MDS) with del(5q). These drugs bind to cereblon (CRBN), the substrate adaptor of the CRL4CRBN E3 ubiquitin ligase and induce the recruitment and ubiquitination of specific protein targets, ultimately resulting in their degradation by the proteasome. Prior work identified lenalidomide-induced ubiquitination and degradation of the lymphoid transcription factors Ikaros (IKZF1) and Aiolos (IKZF3). Degradation of IKZF1 and IKZF3 kills multiple myeloma cells and increases IL-2 release by T cells, explaining two aspects of lenalidomide’s clinical efficacy, but does not account for lenalidomide’s therapeutic window in MDS with del(5q). Here, we demonstrated that lenalidomide induces the ubiquitination of casein kinase 1A1 (CK1α) by CRL4CRBN, resulting in CK1α degradation. CK1α is encoded by a gene, CSNK1A1, within the common deleted region for del(5q) MDS, and its haploinsufficiency sensitizes cells to lenalidomide therapy. Thus, lenalidomide-induced degradation of CK1α provides a mechanistic basis for lenalidomide’s therapeutic window in del(5q) MDS. Despite their widespread clinical use, studies of thalidomide derivatives have been limited by their lack of effect in mouse models. While characterizing CK1α as a lenalidomide-induced substrate, we found that mouse cells are resistant to thalidomide-derivatives because of species-specific differences in CRBN. To enable in vivo study of this class of drugs, we developed a knock-in mouse model with a single amino acid change in Crbn, CrbnI391V, which shows thalidomide-induced degradation of Ikzf1, Ikzf3, and Ck1α. Using this model, we demonstrated that haploinsufficiency for Csnk1a1 confers lenalidomide sensitivity in vivo and that both lenalidomide-induced selection and Trp53-mediated resistance occur at the level of hematopoietic stem cells. We also demonstrated that CrbnI391V is sufficient to confer thalidomide-induced fetal loss in mice. Further study of the CrbnI391V model will provide valuable insights into the in vivo efficacy and toxicity of this class of drugs, as well as the opportunities to identify tissue-specific substrates which may contribute to efficacy in other conditions.