1-alpha-24-dihydroxyvitamin-d3 and Leukemia--Myeloid

The seco-steroid hormone, 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] inhibits proliferation and induces differentiation of malignant cells including those of the hematopoietic system. The 24-oxo metabolite of 1,25(OH)(2)D(3) also has prominent antiproliferative activities against various cancer cells. We chemically synthesized five novel 24-oxo vitamin D(3) analogues and evaluated their abilities both to inhibit clonal growth and induce differentiation of myeloid leukemia cells and to cause hypercalcemia. The 1alpha,25-dihydroxy-16-ene-D(3) [1,25(OH)(2)-16-ene-D(3)] and 1alpha,25-dihydroxy-16-ene-19-nor-D(3) [1,25(OH)(2)-16-ene-19-nor-D(3)] and their 24-oxo metabolites showed greater potency than 1,25(OH)(2)D(3) in their abilities to inhibit clonal proliferation of HL-60, NB4, and U937 leukemic cell lines as measured by methylcellulose soft-gel assay. Their inhibition of clonal growth was irreversible as analyzed by pulse exposure studies. The synthetic analogues also had greater potency than 1,25(OH)(2)D(3) to induce differentiation of HL-60 and NB4 cells as measured by generation of superoxide, nonspecific esterase production, and induction of CD11b and CD14 cell surface antigens and to increase the proportion of these cells in the G(0)-G(1) phase of the cell cycle. For most assays, the 24-oxo metabolite was slightly more potent than the unmodified analogue, and 50% activity was usually found in the nanomolar range. These analogues and their 24-oxo metabolites also inhibited fresh leukemic cell clonal proliferation. Expression of p27(KIP1), a cyclin-dependent kinase inhibitor that plays an important role in blocking the cell cycle, was found by Western blot analysis to be induced by the analogues and their 24-oxo metabolites in both HL-60 and U937 cells, suggesting a possible mechanism by which these analogues inhibit leukemic growth. Notably, the calcemic activity tested by injections of 1alpha,25-dihydroxy-16-ene-24-oxo-19-nor-D(3) in mice was at least 12-fold less than 1alpha,25(OH)(2)-16-ene-19-nor-D(3). Taken together, chemically synthesized 24-oxo metabolites of 1alpha,25(OH)(2)-16-ene-D(3) and 1alpha,25(OH)(2)-16-ene-19-nor-D(3) irreversibly inhibited proliferation and induced differentiation of acute myeloid leukemia cells with minimal toxicity; these compounds may have a role in the maintenance phase of therapy for acute myeloid leukemia.

Topics: Calcium; Cell Cycle; Cell Cycle Proteins; Cell Differentiation; Cell Division; Cholecalciferol; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; Dihydroxycholecalciferols; Dose-Response Relationship, Drug; Growth Inhibitors; HL-60 Cells; Humans; Leukemia, Myeloid; Microtubule-Associated Proteins; Tumor Suppressor Proteins

The hormonally active form of vitamin D, 1 alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3], is an efficient stimulator of intestinal calcium absorption (ICA) and bone calcium mobilization (BCM) in humans and experimental animals and, as well, has been shown to be effective in inducing differentiation and inhibiting proliferation of leukemia cells. Thus, it has been proposed that analogues of 1,25(OH)2D3 could be synthesized which might allow for separation of biological functions, i.e., promote a differentiation of leukemia cells without a significant stimulation of ICA or BCM, both biological effects which can cause hypercalcemia in humans. Here we report the results of an evaluation of four analogues of the previously studied (Zhou et al., Blood, 74:82-92, 1989) 1 alpha,25-dihydroxy-16-ene-23-yne-vitamin D3 [1,25(OH)2-16-ene-23-yne-D3]; these analogues allowed evaluation of the consequences of (a) the presence or absence of six deuterium atoms on carbons 26 and 27 of the side chain and (b) the deletion or substitution by a fluorine atom of the 1 alpha-hydroxyl group on the A-ring. The 1,25(OH)2-16-ene-23-yne-D3 analogue was found to be 7-fold more potent than the parent 1,25(OH)2D3 with respect to (a) inhibition of clonal proliferation of HL-60 cells as well as (b) induction of differentiation of HL-60 promyelocytes. Variants of this analogue which possessed the six deuterium atoms on carbons 26 and 27 were slightly less active than the 1,25(OH)2-16-ene-yne-D3. However, replacement of the 1 alpha-hydroxyl group by a 1-fluoro group, or the absence of the 1-hydroxyl group, resulted in analogues that were somewhat less effective than the parent 1,25(OH)2D3 in achieving these biological responses but more potent as inhibitors of the renal mitochondrial 25-OH-D3-1 alpha-hydroxylase, the site of endogenous production of 1,25(OH)2D3. ICA and BCM were assessed in vivo in vitamin D-deficient chickens, and each of the analogues was markedly less potent than the standard 1,25(OH)2D3. The analogue 1,25(OH)2-16-ene-23-yne-D3 had 2% of the ICA and 3% of the BCM activity of the parent 1,25(OH)2D3. Absence of the 1 alpha-hydroxyl group or substitution of the 1-fluoro group for the 1-hydroxyl group significantly diminished both the ICA and BCM activity in comparison to 1,25(OH)2-16-ene-23-yne-D3. Receptor binding studies indicated that 1,25(OH)2-16-ene-23-yne-D3 competed about 75% as effectively as 1,25(OH)2D3 for 1,25(OH)2D3 receptors present in both chick intestinal cells

Topics: Bone and Bones; Calcification, Physiologic; Calcium; Cell Differentiation; Cell Division; Cholestanetriol 26-Monooxygenase; Dihydroxycholecalciferols; Humans; Intestinal Absorption; Leukemia, Experimental; Leukemia, Myeloid; Steroid Hydroxylases; Stimulation, Chemical; Structure-Activity Relationship; Tumor Cells, Cultured; Vitamin D