Abstract 4842: The stem cell niche detoxifies chemotherapy and protects malignant hematopoietic cells via expression of cytochrome P450 enzymes
Authors: Alonso Salvador , Su Meng , Jones Richard , Ghiaur Gabriel 
Published: May 2015

The bone marrow (BM) microenvironment plays a critical role in drug resistance of malignant cells. Although both, direct contact as well as soluble factors have been proposed, the mechanisms responsible for this microenvironment-mediated protection remain unclear. We have recently shown that BM stroma protects human hematopoietic stem cells from the pro-differentiating effects of retinoid via cytochrome P450 (CYP) 26 mediated degradation (Ghiaur G et al., PNAS 2013). Here, we hypothesized that drug metabolism by BM stroma via CYP enzymes is a general cell-extrinsic mechanism of protection of malignant cells from chemotherapy. First, we have designed a qPCR expression array platform and found that human BM stroma expresses a variety of drug metabolizing enzymes to levels comparable to human hepatocytes. Next, using qRT-PCR, we confirmed that CYP3A4, a major drug metabolizing enzyme, is expressed in primary BM stroma but not in multiple myeloma (MM) or acute myeloid leukemia (AML) cells. We have found that BM stroma protects against Bortezomib mediated kill of MM cells [H929 (p&lt;0.01)] and VP16 mediated kill of AML cells [KG1 (p0.01)] (Table). Since CYP3A4 plays a major role in the metabolism of aforementioned drugs, we studied if stromal CYP3A4 contributes to this protection. We found that ketoconazole, a potent CYP3A4 antagonist, overcomes bone marrow stroma-mediated resistance to Bortezomib (p=0.01), and VP16 (p=0.03). Ketoconazole had no effects in stroma free cultures. In addition, shRNA-mediated knockdown of stromal CYP3A4 renders these cells unable to protect malignant cells from chemotherapy (p=0.01). These studies open the possibilities that the stem cell niche serves as a drug-free sanctuary, where malignant stem cells can survive conventional chemotherapy. Targeting stromal P450 enzymes hold promise of improved efficiency of stem cell directed therapies and decrease rate of relapse in hematological malignancies.

Table.Effects of VP16 and Bortezomib on cellular and clonogenic recovery of KG1 and H929 cells, respectively. “Stroma-CYP3A4” represents experiments performed on CYP3A4 knockdown stroma. Control shRNA infected stroma showed no differences compared to untransduced cells. Data are presented as mean±SEM from three independent experiments.VP16: KG1 cellular recoveryBortezomib: H929 clonogenic (CFU-C) recoveryLiquidStromaStroma+ketoconazoleStroma- CYP3A4LiquidStromaStroma +ketoconazoleStroma-CYP3A48.10 ± 1.5529.38 ± 0.515.7 ± 2.719.35 ± 0.8315.77 ± 3.289.5 ± 2.549.33 ± 860.35 ± 4.2

Citation Format: Salvador Alonso, Meng Su, Richard Jones, Gabriel Ghiaur. The stem cell niche detoxifies chemotherapy and protects malignant hematopoietic cells via expression of cytochrome P450 enzymes. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4842. doi:10.1158/1538-7445.AM2014-4842