B8 (new in 2nd funding period): Role of BRCC3 in the pathogenesis and treatment of myelodysplastic syndrome and acute myeloid leukemia
Lenalidomide has high clinical activity in multiple myeloma and myelodysplastic syndrome (MDS) with par-tial or complete loss of chromosome 5 (-5/del(5q)). We recently demonstrated that lenalidomide and its analogs thalidomide and pomalidomide mediate their specific effects by a previously undescribed drug mechanism. These compounds alter the substrate specificity of CRBN, the substrate adaptor of the CRL4CRBN E3 ubiquitin ligase. Lenalidomide increases the affinity of CRBN to Ikaros (IKZF1), Aiolos (IKZF3), and casein kinase 1A1 (CK1α), resulting in their ubiquitination and degradation. While depletion of the lym-phoid transcription factors Ikaros (IKZF1) and Aiolos (IKZF3) inhibits growth of multiple myeloma cells, deg-radation of casein kinase 1A1 (CK1α) is responsible for the specific sensitivity of MDS del(5q) cells to le-nalidomide.
In our quantitative proteomic-based CRBN interaction analysis we also found proteins that were removed from CRBN in the presence of lenalidomide. The BRCA1/BRCA2-containing complex subunit 3 (BRCC3), a K63-specific deubiquitinating enzyme (DUB), appeared at the top of the list of proteins whose interaction with CRBN was abrogated by lenalidomide. It is not known whether BRCC3 is an endogenous substrate or a functional component of the CRL4CRBN E3 ubiquitin ligase and how abrogation of the BRCC3-CRBN interac-tion is involved in lenalidomide activity. Recent large-scale sequencing analyses identified recurrent loss-of-function mutations in the BRCC3 gene in MDS and acute myeloid leukemia (AML), suggesting that they contribute to malignant transformation by a yet not characterized mechanism. ShRNA-mediated knockdown of BRCC3 specifically inhibited growth of lenalidomide-sensitive multiple myeloma cells while myeloid cell lines were unaffected, suggesting that BRCC3 has opposing roles in different hematopoietic tissues. We hypothesize that the oncogenic potential of loss-of-function BRCC3 mutations in myeloid malignancies de-rives from impairing ubiquitination levels of its substrate proteins that may include members of the CRBN E3 ligase complex. We will apply biochemical and quantitative proteomic analyses combined with in vitro and in vivo functional genetics studies to (1) characterize the functional interaction of BRCC3 with the CRL4CRBN E3 ubiquitin ligase and its implication for lenalidomide activity and (2) determine the pathogenetic role of BRCC3 mutations in MDS and AML. Further understanding of the role of BRCC3 in lenalidomide activity and myeloid malignancies may help to improve targeted therapy of AML and MDS.
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