PC-11 Select targeting of diffuse large B-cell lymphomas using MyD88 small molecule inhibitors

Abstract

Background: A recurring single amino acid somatic mutation associated with nearly 1/3 of human diffuse large B cell lymphomas (DLBCLs), over 90% of Waldenström macroglobulinemia (WM) and 54% of immunoglobulin M monoclonal gammopathy, has been identified within the myeloid differentiation factor 88 (MyD88). This gain of function single amino acid mutation correlates with tumor cell proliferation and survival involving spontaneous and sustained activation of MyD88-dependent NF-κB and Janus Kinase (JAK) signaling pathways. MyD88 acts as a central signaling adapter for mediating innate and cytokine driven inflammation for the Interleukin-1 (IL-1R) and Toll-like receptors (TLRs). In normal healthy cells, MyD88 is thought to be held in an auto-inhibitory state with its own death and TIR domains fused together in negative self-regulation until activated by appropriate receptor mediated ligand engagement, involving Toll-like or Interleukin-1 ligands. We propose the oncogenic mutation, MyD88L265P located within the TIR domain of MyD88 results in discrete structural changes which releases binding and auto-inhibition of MyD88 death domains resulting in MyDDosome assembly, spontaneous recruitment of IL-1 receptor associated kinases (IRAKs) and sustained signaling. Observation: Recently, CADD derived small molecule compounds which inhibit MyD88 dimer formation have been identified and functionally characterized to protect against Staphylococcal enterotoxin B (SEB) induced death in animal models. Hypothesis: Based on their ability to target MyD88 in protection against Staphylococcal enterotoxin B induced death we hypothesize that CADD derived MyD88 small molecule inhibitors may also be useful therapeutic agents in treating DLBCLs bearing the oncogenic mutation MYD88L265P by inhibiting MyDDosome formation and subsequent recruitment of IL-1 receptor associated kinases. Approach: Using in vitro and in vivo studies we are evaluating MyD88 specific small molecule inhibitors for the ability to inhibit tumor cell proliferation, MyDDsome assembly, IRAK recruitment and signaling in human patient cancer cells OCI Ly3 and OCI Ly19 bearing the oncogenic mutation MYD88L265P. Results: We show (1) MyD88 small molecule inhibitors are able to inhibit cell proliferation of activated human B cell lymphoma cells bearing the MyD88L265P mutation using a MTS cell proliferation assay, (2) Purified recombinant MyD88 and small molecule inhibitors exhibit unique binding chromatograms in comparison to vehicle controls using a thermal shift assay and (3) MyD88 SMIs inhibit LPS activated TLR4-blue cell NF-kB signaling in comparison to the TLR4 specific inhibitor TAK242. We are continuing to biochemically characterize MyD88 small molecule inhibitors as well as test additional human cancer cells bearing oncogenic MyD88 mutations. Conclusions: CADD derived MyD88 specific SMIs previously shown to be effective in protection again microbial infection and death may also be useful in targeting B cell lymphoma cells bearing the MyD88L265P mutation. Future studies defining the molecular mechanism of this MyD88L265P specific mutation associated with nearly one third of human patient tumor isolates will rationally inform and propel development of novel therapeutics to counteract both inflammation as well as tumor cell formation.