General MM

Targeted re-sensitization of lenalidomide resistant cell lines using cereblon replacement, inhibition of STAT3 or IRF4 molecular targeting

Immunomodulatory drugs (IMiDs), such as lenalidomide, thalidomide, pomalidomide, and CC-220, play a crucial role in the treatment of newly diagnosed patients with multiple myeloma (MM). Early diagnosis is critical as most patients will eventually develop resistance to therapy and experience a relapsing disease state.1 The primary target of IMiDs has been identified as the cereblon (CRBN) protein, with the binding of IMiDs altering substrate specificity and affecting the recruitment and degradation of proteins which in turn are involved in the regulation of survival, immune response, and tumor proliferation.2

Dr. Yuan Xiao Zhu, from the Division of Hematology, Mayo Clinic, AZ, USA, and colleagues generated lenalidomide resistant human multiple myeloma cell lines (HMCLs) by culturing the lenalidomide-sensitive isogenic cell lines MM.1S, KMS11, XG1, and OPM2, in the presence of increasing doses of lenalidomide (5 µM to 50 µM) for an extended period of time until resistant cells were generated.3

Key findings:
  • CRBN alterations are associated with resistance to IMiDs in MM cell lines
  • The isogenic cell lines were resistant to lenalidomide, pomalidomide, and CC-220, but not the proteasome inhibitor bortezomib
  • The resistant cell lines MM.1SLenRes, KMS11LenRes, and OPM2LenRes had a significant decrease or absence of CRBN compared to the isogenic cell lines
    • Re-introduction of wild-type CRBN restored sensitivity to lenalidomide
  • Mutational analysis of the resistant cell lines
    • MM.1SLenRes contains a deletion and mutation in the remaining copy of CRBN
    • KMS11LenRes contains a CRBN deletion
  • Expression of CRBN was abundant in XG1LenRes following the acquisition of lenalidomide resistance
    • Introduction of exogenous CRBN did not restore sensitivity to lenalidomide
  • Downregulation of full-length IRF4 was found in XG1 and XG1LenRes cell lines, with truncated IRF4 also downregulated in XG1 cell lines
  • In the XG1LenRes cell line, knockdown of CD147 did not reduce cell proliferation or restore sensitivity to lenalidomide
  • Interleukin 6 (IL6) and STAT3 were among the most up-regulated genes in resistant cell lines compared to sensitive isogenic cell lines
    • IL6 supplemented XG1 cells developed lenalidomide resistance, with activated STAT3 expression
    • PB-1-102, a selective STAT3 inhibitor, re-sensitized XG1LenRes to lenalidomide, inducing synergistic downregulation of IRF4 and MYC, and anti-myeloma activity
  • In XG1LenRes, SGC-CBP30 reduced autocrine IL6 production in the absence or presence of lenalidomide and inhibited STAT3 activation
  • In the three IMiD-sensitive cell lines, SGC-CBP30 increased lenalidomide sensitivity

The authors concluded that a deficiency or mutation in CRBN is the most common mechanism of acquired resistance to IMiD therapy in MM cells. Furthermore, the researchers identified a novel mechanism of resistance involving the activation of STAT3 and IL6. In addition to this, the study illustrated a potential strategy to overcome IMiD resistance with inhibition of IRF and CBP/EP300.

References
  1. Raza S. et al. Immunomodulatory drugs (IMiDs) in multiple myeloma. Curr Cancer Drug Targets. 2017; 17(9): 846–857. DOI: 10.2174/1568009617666170214104426.
  2. Ito T. & Handa H. Cereblon and its downstream substrates as molecular targets of immunomodulatory drugs. Int J Hematol. 2016 Sep; 104(3): 293–299. DOI: 10.1007/s12185-016-2073-4.
  3. Zhu Y.Z. et al. Identification of lenalidomide resistance pathways in myeloma and targeted resensitization using cereblon replacement, inhibition of STAT3 or targeting of IRF4. Blood Cancer J. 2019 Feb 11; 9(2): 19. DOI: 10.1038/s41408-019-0173-0.
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