Ovarian cancer cells that coexpress endogenous Rb and p16 are insensitive to overexpression of functional p16 protein.

Defects of the 'Rb/cyclin D1/p16 pathway' have been shown to play a critical role in the development of virtually all human malignancies assessed. To determine the contribution of G1 phase cell cycle defects to ovarian tumorigenesis, we have examined a panel of normal and tumor ovarian tissues and ovarian cancer cell lines for the expression of Rb, p16 and cyclin D1 proteins. Unlike most types of human cancer whose development involves the loss of either Rb or p16 expression, we observed the coexpression of Rb, p16 and cyclin D1 in 82% of ovarian cancer tissues and cell lines. Furthermore, the growth and cell cycle distribution profiles of three ovarian cancer cell lines (ES-2, PA-1 and NIH OVCAR-3) that coexpressed Rb and p16, were found to be unaffected by adenoviral-mediated overexpression of functional p16 protein, indicating the existence of a defect(s) downstream from p16 in these cells. By contrast overexpression of ectopic p16 in the one ovarian cancer cell line (SK-OV-3) that expressed Rb but lacked p16 protein, resulted in a G1 growth arrest. These data suggest that defects of the 'Rb/cyclin D1/p16 pathway', other than the loss of Rb or p16, may play a major role in the development of ovarian cancer.

Biphasic regulation of breast cancer cell growth by progesterone: role of the cyclin-dependent kinase inhibitors, p21 and p27(Kip1).

Depending on the tissue, progesterone is classified as a proliferative or a differentiative hormone. To explain this paradox, and to simplify analysis of its effects, we used a breast cancer cell line (T47D-YB) that constitutively expresses the B isoform of progesterone receptors. These cells are resistant to the proliferative effects of epidermal growth factor (EGF). Progesterone treatment accelerates T47D-YB cells through the first mitotic cell cycle, but arrests them in late G1 of the second cycle. This arrest is accompanied by decreased levels of cyclins D1, D3, and E, disappearance of cyclins A and B, and sequential induction of the cyclin-dependent kinase (cdk) inhibitors p21 and p27(Kip1). The retinoblastoma protein is hypophosphorylated and extensively down-regulated. The activity of the cell cycle-dependent protein kinase, cdk2, is regulated biphasically by progesterone: it increases initially, then decreases. This is consistent with the biphasic proliferative increase followed by arrest produced by one pulse of progesterone. A second treatment with progesterone cannot restart proliferation despite adequate levels of transcriptionally competent PR. Instead, a second progesterone dose delays the fall of p21 and enhances the rise of p27(Kip1), thereby intensifying the progesterone resistance in an autoinhibitory loop. However, during the progesterone-induced arrest, the cell cycling machinery is poised to restart. The first dose of progesterone increases the levels of EGF receptors and transiently sensitizes the cells to the proliferative effects of EGF. We conclude that progesterone is neither inherently proliferative nor antiproliferative, but that it is capable of stimulating or inhibiting cell growth depending on whether treatment is transient or continuous. We also suggest that the G1 arrest after progesterone treatment is accompanied by cellular changes that permit other, possibly tissue-specific, factors to influence the final proliferative or differentiative state.

An 80 Kb P1 clone from chromosome 3p21.3 suppresses tumor growth in vivo.

High frequencies of allelic loss on the short arm of chromosome 3 in small cell lung cancer (SCLC) and a number of other tumors suggest the existence of a tumor suppressor gene(s) within the deleted regions. Two small cell lung cancer lines, NCI H740 and GLC20, have been described which have homozygous deletions in the region 3p21.3. The deleted region overlaps with a 2 Mb fragment of human DNA present in the interspecies hybrid HA(3)BB9F, that suppresses tumor formation by mouse A9 fibrosarcoma cells. Human sequences from this cell hybrid were isolated using inter Alu PCR. From this starting point, a P1 contig was developed for the region of 450 Kb that is common to the homozygous deletions seen in the SCLC lines NCI H740 and GLC20 and is also present in HA(3)BB9F, the suppressed A9 hybrid. Individual P1 clones were assayed for their ability to suppress the tumorigenicity of the mouse fibrosarcoma cell line A9 as assayed by injection of transfected A9 cells into athymic nude mice. The introduction of one of the P1 clones into A9 cells resulted in suppression of tumor growth whereas two other P1 clones from the contig failed to suppress tumor formation in athymic nude mice. These data functionally delimit a tumor suppressor locus to a region of 80 kb within a P1 clone at 3p21.3.

Continuous submarginal phosphorus with broilers and the effect of preslaughter transportation: carcass defects, further-processing yields, and tibia-femur integrity.

Broiler males were given a series of feeds from 0 to 8 wk having all nutrients advocated by the NRC (1984) and were compared with birds offered feeds with available P continuously 10% below recommendation. At termination, birds in pens were divided for cooping, and coops were either subjected to 6 h of truck transportation and 4 h of preslaughter rest or held stationary for 10 h. High summer temperatures existed throughout experimentation, and low dietary P reduced body weight gain through the first 6 wk, whereas an advantage in feed conversion and mortality occurred from 6 to 8 wk. Weight loss increased when birds were subjected to transportation, regardless of P nutriture, and a portion of the loss was recovered during processing as gain in relative chilled carcass yield. Proportions of abdominal fat and skinless boneless meats from chilled carcasses were unaltered, regardless of treatment. Increased incidence of deformed drumsticks occurred because of low P as did drumstick bruising, which was further accentuated when birds had been transported. Back bruising was prominent when P was adequate and birds were held stationary, whereas the converse occurred with transportation. Tibia length was reduced as a consequence of low P, whereas the femur suffered in terms of decreased mineral density at the epiphyses and resistance to Instron-applied stress. Although transportation in itself did not affect any bone measurement, inadequate P weakened the skeleton to increase likelihood of carcass defects during preslaughter stress.