No abstract

1α,25-dihydroxyvitamin D3 (1,25D3) is a powerful differentiation inducer for acute myeloid leukemia (AML) cells. However, 1,25D3 doses required for differentiation of AML cells may cause lethal hypercalcemia in vivo. There is evidence that vitamin D2 is less toxic than vitamin D3 in animals. Here, we determined the differentiation effects of novel analogs of 1α,25-dihydroxyvitamin D2 (1,25D2), PRI-1916 and PRI-1917, in which the extended side chains of their previously reported precursors (PRI-1906 and PRI-1907, respectively) underwent further 24Z (24-cis) modification. Using four human AML cell lines representing different stages of myeloid maturation (KG-1a, HL60, U937, and MOLM-13), we found that the potency of PRI-1916 was slightly higher or equal to that of PRI-1906 while PRI-1917 was significantly less potent than PRI-1907. We also demonstrated that 1,25D2 was a less effective differentiation agent than 1,25D3 in these cell lines. Irrespective of their differentiation potency, all the vitamin D2 derivatives tested were less potent than 1,25D3 in transactivating the DR3-type vitamin D response elements. However, similar to 1,25D3, both 1,25D2 and its analogs could strongly cooperate with the plant polyphenol carnosic acid in inducing cell differentiation and inhibition of G1–S cell cycle transition. These results indicate that the 24Z modification has contrasting effects on the differentiation ability of PRI-1906 and PRI-1907 and that the addition of a plant polyphenol could result in a similar extent of cell differentiation induced by different vitamin D compounds. The enhanced antileukemic effects of the tested combinations may constitute the basis for the development of novel approaches for differentiation therapy of AML.

1α,25-dihydroxyvitamin D3 (1,25D) is known to regulate various cellular functions, including cell proliferation and differentiation. Both 1,25D and its precursor, 25-hydroxyvitamin D3 [25(OH)D], are catabolized by 24-hydroxylase encoded by the CYP24A1 gene, which is one of the most responsive vitamin D receptor (VDR) target genes, and a is candidate oncogene frequently overexpressed in various malignancies. Therefore, compounds which can inhibit CYP24A1 expression and/or activity, thereby reducing degradation of vitamin D derivatives, may potentiate their anticancer effects. We have previously shown that the plant polyphenols carnosic acid, curcumin and silibinin synergistically enhance differentiation of acute myeloid leukemia (AML) cells induced by near physiologic concentrations of 1,25D. This was associated with the transcription factor Nrf2-assisted elevation of both VDR and the retinoid X receptor alpha (RXRα) protein levels. Here, we demonstrate that despite the latter effect the polyphenols markedly inhibited 1,25D-induced CYP24A1 mRNA expression in AML cell lines but not in osteoblast-like cells. Furthermore, while potentiating 1,25D-induced transactivation of the direct repeat-3 (DR3)-type consensus vitamin D response element (VDRE) in AML cells, as measured by the luciferase (Luc) reporter gene assay, the polyphenols strongly inhibited 1,25D-dependent activation of the CYP24-Luc reporter construct containing an early (-298 bp) promoter region with two critical DR3-type VDREs (VDRE1 and VDRE2). In contrast, the polyphenols failed to inhibit and even enhanced 1,25D-induced transactivation of CYP24-Luc transfected into osteoblastic cells. Inactivating mutations of the VDRE1 or VDRE2 sequence in this construct did not abolish the inhibitory effect of polyphenols. However, overexpression of Nrf2 resulted in reduced CYP24-Luc activation by 1,25D, while dominant-negative Nrf2 had an opposite effect. Importantly, feeding of healthy Balb/c mice with standard rodent chow supplemented with carnosic acid-rich rosemary extract resulted in a marked dose- and time-dependent increase in serum levels of 25(OH)D, as compared to control animals. This was associated with a significant decrease in CYP24A1 mRNA levels in the kidneys of these mice. Collectively, the above data suggest that by downregulating CYP24A1 in a tissue-dependent manner, plant polyphenols may potentiate therapeutic and preventive effects of vitamin D derivatives. These findings may also contribute to our understanding of the beneficial effects of healthy diets. (Supported by the Israel Science Foundation grant 635/11 to M.D. and Y.S. and by the American Institute for Cancer Research grant #10A049 to G.P.S. and M.D.). Citation Format: Ehud Sharony, Ze'ev Barvish, Victoria Novik, Marina Khanin, Merav Cohen-Lahav, Doron Amichay, Yoav Sharoni, George P. Studzinski, Michael Danilenko. Plant polyphenols inhibit cellular 24-hydroxylase (CYP24A1) expression and elevate serum 25-hydroxyvitamin D levels. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4655. doi:10.1158/1538-7445.AM2015-4655