This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Elstner, E. Articles by Koeffler, H. P. Search for Related Content PubMed PubMed Citation Articles by Elstner, E. Articles by Koeffler, H. P.

20-epi-Vitamin D₃ Analogues: A Novel Class of Potent Inhibitors of Proliferation and Inducers of Differentiation of Human Breast Cancer Cell Lines¹

Divisions of Hematology-Oncology [E. E., S. d. V., H. P. K.] and Rheumatology [M. L-I.], Departments of Medicine, Pathology [J. S., I. P. S.], and Pediatrics [T. U.], University of California at Los Angeles, School of Medicine and Cedars-Sinai Medical Center; Clinical Nutrition, University of California at Los Angeles, School of Medicine, Los Angeles, California 90048 [D. H.]; Department of Biology, Leo Pharmaceutical Products, Ballerup, Denmark DK-2750 [L. B.]; and Hoffman-LaRoche, Nutley, New Jersey 07110-1199 [M. U.]

We have studied the in vitro biological activities and mechanism of action of 1,25-dihydroxyvitamin D₃ (1,25D₃) and four potent 1,25D₃ analogues [20-epi-22oxa-24a,26a,27a-tri-homo-1,25(OH)₂D₃ (KH 1060); 20-epi-1,25(OH)₂D₃; 1,25(OH)₂-16ene-D₃; and 1,25(OH)₂-16ene-23yne-D₃] on proliferation and differentiation of estrogen receptor-negative (MDA-MB-436, BT-20, SK-BR-3, and MDA-MB-231), estrogen receptor-weakly positive (BT474), and estrogen receptor-positive (MCF-7) breast cancer cell lines. Dose-response studies showed that KH 1060 was the most potent analogue, because it was able to induce differentiation in all seven breast cancer cell lines (measured by lipid staining) and to suppress more than 50% clonal proliferation (ED₅₀) at 10^-10 M in all cell lines, except MDA-MB-436 and BT-20. To explore how these compounds mediated antiproliferative actions, their effects on the cell cycle, on expression of bcl-2 and p53, and on apoptosis were assessed. Five of six cell lines have a mutant p53 gene, whereas MCF-7 has wild-type p53. Immunohistochemical staining showed that the p53 protein was predominantly localized in the nucleus in each of the breast cancer cell lines except for MCF-7, which expressed the protein predominantly in the cytoplasm. After incubation with KH 1060 (3 days; 10^-7 M), expression of bcl-2 protein as determined by immunohistochemical localization was markedly decreased in BT-474, MCF-7, and MDA-MB-231; these same cells were profoundly inhibited in their clonal proliferation and arrested in the G₀/G₁ phase of the cell cycle when cultured with KH 1060. In contrast, BT-20 and MDA-MB-436 cells that were refractory to the antiproliferative effect of KH 1060 (ED₅₀<10^-6 M) had no down-regulation of their bcl-2 expression and no cell cycle changes after exposure to KH 1060. MCF-7 showed morphological changes and DNA fragmentation, indicative of apoptosis after 48 h incubation with KH 1060 (10^-6 M), during which time p53 protein accumulated in the nucleus and decreased in the cytoplasm. In contrast, no apoptosis was detected in three other breast lines (MDA-MB-231, SK-BR-3, and BT-474) that had a mutated p53. In conclusion, the data indicate that KH 1060 is an extremely potent 1,25D₃ analogue inducing differentiation of all six breast cancer lines and potently inhibiting clonal growth of four of them with concomitant decreased bcl-2 and cell cycle arrest at G₀/G₁. Only one (MCF-7) of six breast cancer cell lines underwent apoptosis; these cells have a wild-type p53 that translocated from the cytoplasm to the nucleus during culture with KH 1060, probably allowing p53 to become a functional nuclear transcriptional activator.

¹ Supported in part by Grant in Aid for Scientific Research from the Ministry of Education, Science, and Culture, Deutsche Forschungsgemeinschaft (to E. E.), Support for this study also came from the NIH Grants CA 43277, CA 42710, and CA 26038, as well as the Concern Foundation and the Parker Hughes Leukemia Foundation.

² To whom requests for reprints should be addressed, at Cedars-Sinai Medical Center/UCLA School of Medicine, 8700 Beverly Blvd., D5033, Los Angeles, CA 90048.

Received 1/19/95. Accepted 5/ 2/95.

This article has been cited by other articles:

				F. Demichelis, H. Greulich, J. A. Macoska, R. Beroukhim, W. R. Sellers, L. Garraway, and M. A. Rubin SNP panel identification assay (SPIA): a genetic-based assay for the identification of cell lines Nucleic Acids Res., April 1, 2008; 36(7): 2446 - 2456. [Abstract] [Full Text] [PDF]

				C. M. Banwell, D. P. MacCartney, M. Guy, A. E. Miles, M. R. Uskokovic, J. Mansi, P. M. Stewart, L. P. O'Neill, B. M. Turner, K. W. Colston, et al. Altered nuclear receptor corepressor expression attenuates vitamin d receptor signaling in breast cancer cells. Clin. Cancer Res., April 1, 2006; 12(7): 2004 - 2013. [Abstract] [Full Text] [PDF]

				M. Teng, M. Wolf, M. N. Ofsthun, J. M. Lazarus, M. A. Hernan, C. A. Camargo Jr, and R. Thadhani Activated Injectable Vitamin D and Hemodialysis Survival: A Historical Cohort Study J. Am. Soc. Nephrol., April 1, 2005; 16(4): 1115 - 1125. [Abstract] [Full Text] [PDF]

				T. M. Beer and A. Myrthue Calcitriol in cancer treatment: From the lab to the clinic Mol. Cancer Ther., March 1, 2004; 3(3): 373 - 381. [Abstract] [Full Text]

				T. Kumagai, T. Ikezoe, D. Gui, J. O'Kelly, X.-J. Tong, F. J. Cohen, J. W. Said, and H. P. Koeffler RWJ-241947 (MCC-555), A Unique Peroxisome Proliferator-Activated Receptor-{gamma} Ligand with Antitumor Activity against Human Prostate Cancer in Vitro and in Beige/Nude/ X-Linked Immunodeficient Mice and Enhancement of Apoptosis in Myeloma Cells Induced by Arsenic Trioxide Clin. Cancer Res., February 15, 2004; 10(4): 1508 - 1520. [Abstract] [Full Text] [PDF]

				J. Hayakawa, C. Depatie, M. Ohmichi, and D. Mercola The Activation of c-Jun NH2-terminal Kinase (JNK) by DNA-damaging Agents Serves to Promote Drug Resistance via Activating Transcription Factor 2 (ATF2)-dependent Enhanced DNA Repair J. Biol. Chem., May 30, 2003; 278(23): 20582 - 20592. [Abstract] [Full Text] [PDF]

				A. L. M. Sutton and P. N. MacDonald Vitamin D: More Than a "Bone-a-Fide" Hormone Mol. Endocrinol., May 1, 2003; 17(5): 777 - 791. [Abstract] [Full Text] [PDF]

				A. Krones-Herzig, E. Adamson, and D. Mercola Early growth response 1 protein, an upstream gatekeeper of the p53 tumor suppressor, controls replicative senescence PNAS, March 18, 2003; 100(6): 3233 - 3238. [Abstract] [Full Text] [PDF]

				V. Vegesna, J. O'Kelly, M. Uskokovic, J. Said, N. Lemp, T. Saitoh, T. Ikezoe, L. Binderup, and H. P. Koeffler Vitamin D3 Analogs Stimulate Hair Growth in Nude Mice Endocrinology, November 1, 2002; 143(11): 4389 - 4396. [Abstract] [Full Text] [PDF]

				J. Lin, X. Jin, K. Rothman, H.-J. Lin, H. Tang, and W. Burke Modulation of Signal Transducer and Activator of Transcription 3 Activities by p53 Tumor Suppressor in Breast Cancer Cells Cancer Res., January 1, 2002; 62(2): 376 - 380. [Abstract] [Full Text] [PDF]

				H. You, W. Yu, B. G. Sanders, and K. Kline RRR-{alpha}-Tocopheryl Succinate Induces MDA-MB-435 and MCF-7 Human Breast Cancer Cells to Undergo Differentiation Cell Growth Differ., September 1, 2001; 12(9): 471 - 480. [Abstract] [Full Text] [PDF]

				J.-i. Hisatake, J. O'Kelly, M. R. Uskokovic, S. Tomoyasu, and H. P. Koeffler Novel vitamin D3 analog, 21-(3-methyl-3-hydroxy-butyl)-19-nor D3, that modulates cell growth, differentiation, apoptosis, cell cycle, and induction of PTEN in leukemic cells Blood, April 15, 2001; 97(8): 2427 - 2433. [Abstract] [Full Text] [PDF]

				B. A. Scaglione-Sewell, M. Bissonnette, S. Skarosi, C. Abraham, and T. A. Brasitus A Vitamin D3 Analog Induces a G1-Phase Arrest in CaCo-2 Cells by Inhibiting Cdk2 and Cdk6: Roles of Cyclin E, p21Waf1, and p27Kip1 Endocrinology, November 1, 2000; 141(11): 3931 - 3939. [Abstract] [Full Text] [PDF]

				A. Bratland, K. Risberg, G. M. Mælandsmo, K. B. Gützkow, O. E. Olsen, A. Moghaddam, M.-y. Wang, C. M. Hansen, H. K. Blomhoff, J. P. Berg, et al. Expression of a Novel Factor, com1, Is Regulated by 1,25-Dihydroxyvitamin D3 in Breast Cancer Cells Cancer Res., October 1, 2000; 60(19): 5578 - 5583. [Abstract] [Full Text]

				K. El Abdaimi, N. Dion, V. Papavasiliou, P.-E. Cardinal, L. Binderup, D. Goltzman, L.-G. Ste-Marie, and R. Kremer The Vitamin D Analogue EB 1089 Prevents Skeletal Metastasis and Prolongs Survival Time in Nude Mice Transplanted with Human Breast Cancer Cells Cancer Res., August 1, 2000; 60(16): 4412 - 4418. [Abstract] [Full Text]

				S. Swami, A. V. Krishnan, and D. Feldman 1{{alpha}},25-Dihydroxyvitamin D3 Down-Regulates Estrogen Receptor Abundance and Suppresses Estrogen Actions in MCF-7 Human Breast Cancer Cells Clin. Cancer Res., August 1, 2000; 6(8): 3371 - 3379. [Abstract] [Full Text]

				K. Koli and J. Keski-Oja 1{{alpha}},25-Dihydroxyvitamin D3 and Its Analogues Down-Regulate Cell Invasion-associated Proteases in Cultured Malignant Cells Cell Growth Differ., April 1, 2000; 11(4): 221 - 229. [Abstract] [Full Text]

				Q. Wang, W. Yang, M. S. Uytingco, S. Christakos, and R. Wieder 1,25-Dihydroxyvitamin D3 and All-trans-Retinoic Acid Sensitize Breast Cancer Cells to Chemotherapy-induced Cell Death Cancer Res., April 1, 2000; 60(7): 2040 - 2048. [Abstract] [Full Text]

				H. Roh, J. Pippin, and J. A. Drebin Down-Regulation of HER2/neu Expression Induces Apoptosis in Human Cancer Cells That Overexpress HER2/neu Cancer Res., February 1, 2000; 60(3): 560 - 565. [Abstract] [Full Text]

				S. E. Blutt, T. J. McDonnell, T. C. Polek, and N. L. Weigel Calcitriol-Induced Apoptosis in LNCaP Cells Is Blocked By Overexpression of Bcl-2 Endocrinology, January 1, 2000; 141(1): 10 - 17. [Abstract] [Full Text] [PDF]

				I. S. Mathiasen, U. Lademann, and M. Jaattela Apoptosis Induced by Vitamin D Compounds in Breast Cancer Cells Is Inhibited by Bcl-2 but Does Not Involve Known Caspases or p53 Cancer Res., October 1, 1999; 59(19): 4848 - 4856. [Abstract] [Full Text] [PDF]

				R. G. Pestell, C. Albanese, A. T. Reutens, J. E. Segall, R. J. Lee, and A. Arnold The Cyclins and Cyclin-Dependent Kinase Inhibitors in Hormonal Regulation of Proliferation and Differentiation Endocr. Rev., August 1, 1999; 20(4): 501 - 534. [Abstract] [Full Text] [PDF]

				A. Agadir, G. Lazzaro, Y. Zheng, X.-K. Zhang, and R. Mehta Resistance of HBL100 human breast epithelial cells to vitamin D action Carcinogenesis, April 1, 1999; 20(4): 577 - 582. [Abstract] [Full Text] [PDF]

				O. P. Mgbonyebi, J. Russo, and I. H. Russo Roscovitine Induces Cell Death and Morphological Changes Indicative of Apoptosis in MDA-MB-231 Breast Cancer Cells Cancer Res., April 1, 1999; 59(8): 1903 - 1910. [Abstract] [Full Text] [PDF]

				H. Asou, M. Koike, E. Elstner, M. Cambell, J. Le, M. R. Uskokovic, N. Kamada, and H. P. Koeffler 19-nor Vitamin-D Analogs: A New Class of Potent Inhibitors of Proliferation and Inducers of Differentiation of Human Myeloid Leukemia Cell Lines Blood, October 1, 1998; 92(7): 2441 - 2449. [Abstract] [Full Text] [PDF]

				E. Elstner, C. Muller, K. Koshizuka, E. A. Williamson, D. Park, H. Asou, P. Shintaku, J. W. Said, D. Heber, and H. P. Koeffler Ligands for peroxisome proliferator-activated receptorgamma and retinoic acid receptor inhibit growth and induce apoptosis of human breast cancer cells in vitro and in BNX mice PNAS, July 21, 1998; 95(15): 8806 - 8811. [Abstract] [Full Text] [PDF]

				T. Koyama, M. Shibakura, M. Ohsawa, R. Kamiyama, and S. Hirosawa Anticoagulant Effects of 1alpha ,25-Dihydroxyvitamin D3 on Human Myelogenous Leukemia Cells and Monocytes Blood, July 1, 1998; 92(1): 160 - 167. [Abstract] [Full Text] [PDF]

				K. VanWeelden, L. Flanagan, L. Binderup, M. Tenniswood, and J. Welsh Apoptotic Regression of MCF-7 Xenografts in Nude Mice Treated with the Vitamin D3 Analog, EB1089 Endocrinology, April 1, 1998; 139(4): 2102 - 2110. [Abstract] [Full Text] [PDF]

				S.-H. Zhuang and K. L. Burnstein Antiproliferative Effect of 1{alpha},25-Dihydroxyvitamin D3 in Human Prostate Cancer Cell Line LNCaP Involves Reduction of Cyclin-Dependent Kinase 2 Activity and Persistent G1 Accumulation Endocrinology, March 1, 1998; 139(3): 1197 - 1207. [Abstract] [Full Text] [PDF]

				C. J. Narvaez and J. Welsh Differential Effects of 1,25-Dihydroxyvitamin D3 and Tetradecanoylphorbol Acetate on Cell Cycle and Apoptosis of MCF-7 Cells and a Vitamin D3-Resistant Variant Endocrinology, November 1, 1997; 138(11): 4690 - 4698. [Abstract] [Full Text] [PDF]

				E. B. Mawer, J. Walls, A. Howell, M. Davies, W. A. Ratcliffe, and N. J. Bundred Serum 1,25-Dihydroxyvitamin D May Be Related Inversely to Disease Activity in Breast Cancer Patients with Bone Metastases J. Clin. Endocrinol. Metab., January 1, 1997; 82(1): 118 - 122. [Abstract] [Full Text] [PDF]

				R. Yu, A. P. Heaney, W. Lu, J. Chen, and S. Melmed Pituitary Tumor Transforming Gene Causes Aneuploidy and p53-dependent and p53-independent Apoptosis J. Biol. Chem., November 17, 2000; 275(47): 36502 - 36505. [Abstract] [Full Text] [PDF]

				M. Grigorian, S. Andresen, E. Tulchinsky, M. Kriajevska, C. Carlberg, C. Kruse, M. Cohn, N. Ambartsumian, A. Christensen, G. Selivanova, et al. Tumor Suppressor p53 Protein Is a New Target for the Metastasis-associated Mts1/S100A4 Protein. FUNCTIONAL CONSEQUENCES OF THEIR INTERACTION J. Biol. Chem., June 15, 2001; 276(25): 22699 - 22708. [Abstract] [Full Text] [PDF]