Brief communicationVitamin D protects acute lymphoblastic leukemia cells from dexamethasone
Introduction
In addition to its classical actions on intestine and bone, vitamin D exerts other diverse effects regulating cellular proliferation, differentiation, apoptosis, autophagy, hematopoiesis, immune function and angiogenesis. The vitamin D receptor which mediates a majority of these actions is expressed in several tissues including bone, kidney, prostate, breast, blood, intestine and cells of the immune system. Vitamin D deficiency has been proposed as a link between cancer and lack of sun exposure [1]. This hypothesis is supported by large prospective studies which show that vitamin D deficiency is associated with increased risk of several types of cancer [2], and by in vitro studies, which show that vitamin D and its analogues suppress the proliferation of many cancer cell types. Furthermore, calcitriol (1α,25(OH)2D3), the active form of vitamin D, potentiates the antitumor effects of several chemotherapeutic agents on various cancer types in vitro and in xenograft models [3].
Although vitamin D receptors are expressed in hematopoietic precursors and malignant T and B lymphocyte derived cell lines [4], the effects of calcitriol on the most common childhood cancer, acute lymphoblastic leukemia (ALL) have to our knowledge not been reported. Although survival rates of childhood ALL have improved dramatically over the prior decades, relapse continues to be a problem. Leukemia cell resistance to steroids is a significant problem, which likely contributes to relapse. In the present study, we hypothesized that calcitriol would: (1) suppress the growth of B precursor acute lymphoblastic leukemia (Pre B ALL) cells, and (2) act synergistically with the steroid chemotherapy dexamethasone to augment its cytotoxicity against ALL cells.
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Materials and methods
Human ALL cell lines were purchased from ATCC (RS4;11, Sup-B15) and DSMZ (SD-1, RCH-ACV), and cultured under standard conditions. Viable cells were counted at 72 h time points using trypan blue exclusion. Apoptosis was measured using Annexin V and 7-AAD staining with flow cytometry on a Becton Dixon FACscan. Gene expression of the vitamin D receptor, glucocorticoid receptor, and Bim was measured using quantitative rtPCR. Primers used included: human β-actin-forward (5′-ACA GAG CCT CGC CTT TGC
Results
Contrary to our first hypothesis, we found that various doses of calcitriol (Sigma) had no effect on proliferation of four different human ALL cell lines in vitro (Fig. 1A). To see whether calcitriol would augment the cytotoxic effect of the steroids on ALL, we cultured two dexamethasone-sensitive ALL cell lines with 100 nM (RS4;11) or 60 nM (Sup-B15) dexamethasone for 72 h, with and without calcitriol. Again contrary to our hypothesis, we found that calcitriol had a small but consistent effect to
Discussion
A number of epidemiological studies have found that calcitriol has a protective role against many types of cancer [2]. In this study we found that calcitriol, different from its effect on other types of cancer cells, does not impair the proliferation/survival of ALL cells. In fact, calcitriol had a small but consistent effect to protect ALL cells from dexamethasone. Although this was a small effect in vitro, it raises the concern that very high doses of vitamin D could potentially have adverse
Conflict of interest
The authors declare no conflict of interest.
Acknowledgements
This study was supported in part by a grant from the National Institutes of Health National Cancer Institute (CA139060).
Contributions. RA designed and performed the experiments and prepared the manuscript; RA, XS, EAE, EN, RP, and BI performed experiments and collected data; LK performed and interpreted the FACS analysis; SDM designed experiments, analyzed results, and prepared the manuscript.
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