Novel Anti-Myeloma Potential Of Cape Analogs: Downregulation Of Ikzf1-Irf4-Myc Axis

Background: Multiple myeloma (MM) is the second most common hematological malignancy and an incurable bone marrow cancer so far. The hallmark of myeloma is chromosomal translocations that transform plasma cells into malignant myeloma cells. 50-75% of myeloma patients exhibit translocations at the immunoglobulin heavy chain (IgH) locus juxtaposing oncogenes from partner chromosome under the control of strong 39 IgH enhancers. Overexpression of oncogenes such as Cyclin D1, Cyclin D3, interferon regulatory factor (IRF4) etc. occurs, depending on the translocating partner locus. We have earlier reported that elevated expression of octameric binding protein-2 (OCT2), a key transcription factor in IgH translocations as a poor prognostic factor and shown its association with reduced survival in MM patients. IRF4 is another indispensable transcription factor for plasma cell differentiation, and its juxtaposition with IgH locus leads to IRF4 overexpression mediated tumorigenesis. Both OCT2 and IRF4 are regulated by the transcription factor, nuclear factor kappa B (NF-κB). Current chemotherapeutics exhibit several side effects, and face the challenge of chemoresistance, thus warranting the need for novel therapeutics for myeloma. Caffeic acid phenethyl ester (CAPE) is an active principle of propolis from honeybee hives, with potent NF-κB inhibitory activity. Hence, we synthesized and evaluated the structure-activity relationship of twenty-one CAPE analogs for their anti-myeloma potential. Methods: Human myeloma cell lines, lenalidomide, CAPE and its analogs were used for the study. Myeloma cell growth inhibition by CAPE analogs was determined by PrestoBlue cell viability assays. Effect of CAPE analogs on IRF4, OCT2, CyclinD3, caspase-3, Ikaros (IKZF1), MYC levels were evaluated by qRT-PCR and/or immunoblotting methods; apoptosis induction detected by Annexin V-PI flow cytometry assays. Results: Cyclohexylethyl and two phenpropyl ester analogs exhibited high myeloma cell growth inhibition in comparison with CAPE and lenalidomide. IRF4 was significantly downregulated both at mRNA and protein levels by the phenpropyl ester analog that demonstrated a remarkable increase in cleaved caspase-3 levels and apoptotic cell number. For the first time, we have demonstrated the ability of CAPE analogs to decrease OCT2, IKZF1, IRF4 and MYC levels in myeloma cells. Downregulation of the IKZF1-IRF4-MYC axis by CAPE and its key analogs revealed their novel anti-myeloma mechanism of action. Furthermore, the lead analogs showed no adverse cytotoxic effect on normal human mononuclear or naive B-cells, also exhibited appropriate in silico pharmacokinetic properties and drug-likeness. Conclusion: Thus, our findings suggest the promising application of CAPE analogs to target Ikaros/IRF4 addiction, the so-called Achilles heel of myeloma, thereby leading to better therapeutic outcomes. Citation Format: Alli Murugesan, Gregoire Lasalle-Claux, Lauren Hogan, Elise Vaillancourt, Ayyoub Selka, Katie Luiker, Andy Hong, MinJi Kim, Mohamed Touaibia, Tony Reiman. Novel anti-myeloma potential of CAPE analogs: Downregulation of IKZF1-IRF4-MYC axis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6559.