T25: Selective glucocorticoid receptor agonists as novel anti-cancer agents

Abstract

Glucocorticoids (GCs) are widely used in treatment of many cancer types due to its ability to induce apoptosis in malignant cells in blood cancer therapy, and to prevent nausea, emesis and chemotherapy-associated hepatotoxicity in case of solid tumors. However, severe dose-limiting side effects occur, including osteoporosis, diabetes and other metabolic complications. Moreover, in therapy of solid tumors GCs strongly affect microenvironment which could be associated with poor prognosis, risk of metastasis and high frequency of relapses. Biological response to GCs is mediated by glucocorticoid receptor (GR), a well-characterized transcription factor. GR controls gene expression via (1) transactivation, which requires binding of GR homodimers to glucocorticoid-responsive elements (GRE) in gene promoters and enhancers, and (2) dimerization-independent transrepression mediated via negative interaction between GR and other transcription factors including major effectors of inflammation and proliferation. Transrepression plays an important role in anti-inflammatory and anti-cancer effects of GR, including normalizing influence on microenvironment, while side effects are associated with GR transactivation. In particular, GCs induce insulin resistance in adipocytes, a major component of the mammary microenvironment, which secrete pro-inflammatory cytokines and growth factors, implicated in tumor progression. Selective GR agonists (SEGRA) that preferentially activate GR transrepression could be a better option for treatment of cancer. Dozens of candidate SEGRAs were identified, synthesized and tested by industry and academia, with some having reached clinical trials. One of the novel GR modulators is 2-(4-acetoxyphenyl)-2-chloro-N-methylethylammonium-chloride, or CpdA, synthetic analogue of aziridine precursor isolated from Namibian shrub Salsola tuberculatiformis Botschantzev. It was shown that CpdA acts as “dissociated” GR ligand: it competes with GCs for GR binding and efficiently induces GR transrepression but not transactivation. We and other authors reported recently that CpdA inhibits survival of prostate cancer cells as well as blood cancer cells in GR-dependent fashion. Furthermore, primary leukemia cells from T-ALL patients appeared to be equally sensitive to GCs and CpdA. Our further studies were concentrated on three directions: (1) GC/SEGRA-based chemotherapy. We screened biological effect of CpdA in combination with traditional agents (doxorubicin, vincristine) and newer therapeutics (Bortezomib, Carfilzomib, MLN-4924, Rapamycin). Pretreatment of lymphoma cells with proteasome inhibitor Bortezomib resulted in GR accumulation and enhanced ligand properties of CpdA. We also revealed remarkable GR-dependent cooperation between CpdA and Bortezomib in suppressing survival of lymphoma cells in vitro and in vivo. Also surprising findings were substantial cooperation in anti-cancer effect of immunosuppressant Rapamycin and CpdA in vitro, and unexpected “dissociated” effect of Rapamycin on GR signaling realized through down -regulation of REDD1, mTORC1 inhibitor. These data suggested high clinical potential of Rapamycin/GC combination in cancer treatment. (2) SEGRA list extention We used two approaches to extend SEGRA list: (1) synthesis of CpdA enantiomers and (2) its chemical derivatives. Chemical analogues of CpdA were designed by appending of bulky substituent into benzene ring, alkylation of carbon atom adjacent to chlorine atom or appending of substituents to nitrogen atom. Evaluation of biological properties of enantiomers revealed higher GR-dependent anti-cancer potential of S-CpdA. Cytotoxic and proapoptotic effects of CpdA analogues were comparable with precursor. (3) Selection of tumor types acceptable for SEGRA treatment. CpdA was selected for NCI-60 in Vitro Cell Line Screening Project providing direct support to anticancer drug discovery program. It was shown that CpdA affect viability of some adherent cancer cell lines. We demonstrated that CpdA unlike GCs did not modify microenvironment and disintegrate tight junctions between cells decreasing risk of metastasis in case of solid tumors. It demonstrates reasonability of further investigations. Overall, our data provide the rationale for novel therapy of cancer based on combination of non-steroidal GR modulators with classic and modern chemotherapeutics. Approaches to obtain more SEGRAs were elaborated.