20-Cyclopropyl-cholecalciferol vitamin D3 analogs: a unique class of potent inhibitors of proliferation of human prostate, breast and myeloid leukemia cell lines.

We have synthesized and studied the ability of a series of nine novel 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] analogs to inhibit clonal growth of myeloid leukemic cells (HL,60), prostate (LNCaP, PC-3 and DU-145) and breast (MCF-7) cancers cells. DU-145 cells were actively resistant to compounds (cmpd) with all of these modifications, but when we removed C-19 (E, 1,25-Dihydroxy-23E-ene-26,27-hexafluoro-19-nor-20-cyclopropy l- cholecalciferol) an analog resulted that was inhibitory against all three prostate cell lines, breast and HL-60 cell lines. Further analysis showed that pulse exposure (3 days, 10(-7) M) to this analog was enough to inhibit clonal growth of PC-3 cell by 50%. Furthermore, cmpd E increased the number of PC-3 cells in G1 and decreased the number in S phase. 1,25(OH)2D3 mediates its biological activities through specific binding to the vitamin D3 receptor (VDR) and subsequent association with vitamin D3 response elements (VDRE) in genes modulated by 1,25(OH)2D3. Several novel vitamin D3 cmpds have recently been identified which have 5- to 1000-fold greater abilities to induce differentiation and to inhibit proliferation of prostate cancer, breast cancer and HL-60 leukemic blast cells as compared to the parental 1,25(OH)2D3. To clarify the mechanism by which nine of these vitamin D3 analogs mediate their remarkably potent biological activities, we have investigated their abilities in PC-3 prostate cancer cells to transactivate a chroramphenicol acetyl transferase (CAT) reporter gene containing a VDRE from the human osteocalcin gene attached to a thymidine kinase minimal promoter. Dose-response studies of Cmpd E showed that in serumless culture conditions, transactivation of the VDRE-CAT was stronger than cmpd J [1,25(OH)2D3]. Then, we investigated the effects of vitamin D3 cmpd J in mice. Our data showed the growth inhibitory action of the vitamin D3 cmpd E in prostate cancer cell line (PC-3) was stastically superior to the non-treatment group in terms of tumor size and tumor weight in mice. In summary, this is the first report of a potent series of 20-cyclopropyl-cholecalciferol vitamin D3 analogs with the ability to inhibit proliferation of LNCaP, PC-3, DU-145, MCF-7 and HL-60 cell lines. These cmpds may mediate their potent anti-proliferative activities through a cell cycle arrest pathway.

The ATM gene and susceptibility to childhood T-cell acute lymphoblastic leukaemia.

Ataxia-telangiectasia (A-T) is a multisystem recessive disease characterized by cerebellar ataxia, oculocutaneous telangiectasias, immunodeficiency and increased risk of cancer. The ATM gene, responsible for A-T, was recently cloned at human chromosome band 11q22-23, a region of frequent alterations in childhood acute lymphoblastic leukaemia (ALL). Children with A-T frequently develop T-ALL. We investigated 18 T-ALL samples for ATM mutations and loss of heterozygosity (LOH) at the ATM locus. No mutations of ATM were found within the coding region in the 18 T-ALL samples, and LOH at the ATM locus was detected in three. The ATM gene appears to be an infrequently altered tumour suppressor gene in childhood T-ALL.

Frequent loss of heterozygosity on the long arm of chromosome 6: identification of two distinct regions of deletion in childhood acute lymphoblastic leukemia.

Cytogenetic analysis of childhood acute lymphoblastic leukemia (ALL) identified nonrandom chromosomal abnormalities of the long arm of chromosome 6. Most of the alterations are deletions that are thought to be indicative of the presence of a tumor suppressor gene that is mutated on the remaining allele. These observations led us to consider whether 6q loss may contribute to the pathogenesis of childhood ALL. To define further a region containing this gene, we analyzed the loss of heterozygosity (LOH) of chromosome 6 in 113 primary ALL samples with matched normal DNA using 34 highly informative microsatellite markers. LOH was found in 17 (15%) samples at one or more of the loci, and partial or interstitial deletions of 6q were detected in 11 of these tumors. On the basis of these results, we performed a detailed deletional map and identified two distinct regions of deletion. The first region is flanked by D6S283 and D6S302 loci at 6q21-22. The second region is flanked by D6S275 and D6S283 loci at 6q21. Clinical analysis determined that LOH of 6q was demonstrated both in precursor-B cell ALLs (15 of 93; 16%) and in T cell ALLs (2 of 19; 11%). In addition, 19 patients have been studied at diagnosis and relapse; 18 showed the same 6q21-22 structural abnormality at relapse (normal, 16 patients; LOH, 2 patients) as their initial presentation, suggesting, albeit with a small patient sample size, that 6q21-22 deletions may be an initial event in leukemogenesis and may occur less frequently during the progression of childhood ALL. These data suggest the presence of putative tumor suppressor genes on chromosome arm 6q that are important in the development of both T and precursor-B childhood ALLs. Our map provides important information toward cloning putative ALL tumor suppressor genes.