Oncogene
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HER-2 oncogene encodes a transmembrane growth factor receptor that is overexpressed in 25-30% of patients with primary breast and ovarian cancer. A murine monoclonal antibody, 4D5, to the extracellular domain of HER-2 receptor elicits cytostatic growth inhibition of tumor cells overexpressing HER-2 protein, but clinical use of this antibody is limited by genesis of human anti-mouse antibodies. To avoid this problem, a recombinant humanized 4D5 monoclonal antibody (rhuMAb HER-2) was developed and tested using a human tumor xenograft model. ⋯ Using a direct measure of DNA repair, therapy of HER-2-overexpressing cells with rhuMAb HER-2 also blocked the removal of cisplatin-induced DNA adducts. Expression of p21/WAF1, an important mediator of DNA repair, was disrupted in breast cancer cells with HER-2 overexpression, but not in control cells, after treatment with HER-2 antibody, thus suggesting cross-communication between the HER-2 signaling and DNA repair pathways. These data demonstrate an in vivo antiproliferative effect of rhuMAb HER-2 on tumors that overexpress HER-2 receptor and a therapeutic advantage in the administration of the antireceptor antibody in combination with chemotherapeutic agents.
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EBV-immortalized lymphoblastoid B cell lines (LCLs) are a suitable in vitro model for the study of EBV-related lymphoproliferative disorders of immunosuppressed patients. We have previously shown that 9-cis-, 13-cis- and all-trans-retinoic acid (RA) powerfully inhibit LCL proliferation at concentrations corresponding to therapeutically achievable plasma levels (10(-6) M). Herein we show that RA-induced LCL accumulation in the G0/G1 phases correlated with the loss of the catalytic activity of all three G1-associated CDKs (CDK2, CDK4 and CDK6) and with increased levels of underphosphorylated pRb and, in some LCLs, p130. ⋯ In addition, RA-treated LCLs showed a marked up-regulation of the CDK inhibitor (CKI) p27Kip-1 at the protein but not mRNA level, which correlated with a progressive increase of p27Kip-1 in CDK2 complexes (more than 2.5-fold) and with a reduction in the active phosphorylated form of CDK2. p27Kip-1 may also contribute to the inhibition of CDK4 kinase activity, as the amount of CDK4-associated p27Kip-1 was increased by 50% after RA exposure. p27Kip-1 up-regulation stably persisted for more than one week after RA withdrawal concomitantly with the maintenance of the proliferative block. Moreover, neutralization of TGFbeta did not affect the growth inhibitory activity of RA, suggesting that LCL growth arrest induced by these retinoids is probably not mediated by a pathway directly involving TGFbeta. Overall, these results demonstrate that RA treatment of EBV-immortalized B lymphocytes is associated with multiple effects on G1 regulatory proteins, including p27Kip1 up-regulation, decreased levels of cyclins D2, D3 and A, and inhibition of CDK2, CDK4 and CDK6 activity, which ultimately result in reduced pRb phosphorylation and G0/G1 growth arrest.
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Dimethylsulfoxide (DMSO) was shown to inhibit the proliferation of several B cell lines including Raji, Daudi, and SKW6-CL4 but the mechanisms involved in this growth arrest are still unclear. We show that in 7TD1 mouse hybridoma cells a DMSO-induced reversible G1 arrest involves inactivation of Rb kinases, cyclin D2/CDK4 and cyclin E/CDK2. This occurs by at least three distinct mechanisms. ⋯ This in turn enables the redistribution of p27[KIP1] from cyclin D2/CDK4 to cyclin E/CDK2 complexes. In addition, the simultaneous accumulation of p21[CIP1] entails increasing association with cyclin D3/CDK4 and cyclin E/CDK2. Thus, p21[CIP1] and p27[KIP1], act in concert to inhibit cyclin E/CDK2 activity which, together with CDK4 inactivation, confers a G1-phase arrest.
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p53 is a tumour suppressor gene which functions as a transcription factor to upregulate genes for growth arrest and apoptosis following DNA damage. p53 mutations are associated with Li-Fraumeni and Li-Fraumeni like syndromes. Recently mutations of the oligomerization domain have been isolated from an LFS and an LFL family affecting respectively codon 344 (Leu to Pro) and 337 (Arg to Cys). The present study was designed to determine the affect of these mutations on the function of p53 protein. p53 344 Leu to Pro existed only in a monomeric form and could not bind to DNA. ⋯ Both mutant proteins had reduced ability to bind to MDM2, p53 337 Arg to Cys being more reduced than p53 344 Leu to Pro. These results indicate that point mutations in the oligomerization domain can disrupt p53 function. In addition, the value of LFS and LFL families for the further understanding of the biological and biochemical properties of p53 is demonstrated.
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The c-Yes proto-oncogene (pp62c-Yes) encodes a non-receptor-type protein tyrosine kinase (NRPTK) of the Src family. c-Yes activities and protein levels are elevated in human melanoma and melanocyte cell lines. Because the neurotrophins (NT) are important in the progression of melanoma to the brain-metastatic phenotype, we determined whether NT stimulate c-Yes activity in human MeWo melanoma cells and two variant sublines with opposite metastatic capabilities, 3 S 5 and 70W. The highly brain-metastatic 70W subline had an intrinsically higher c-Yes activity than parental MeWo or poorly metastatic 3 S 5 cells. c-Yes kinase was further induced by the prototypic human NT, nerve growth factor (NGF) in a dose and time-dependent manner. ⋯ Metastatic TXM-13 and TXM-18 increased c-Yes activity in response to NGF. In contrast, no increase was observed in low-metastatic TXM-40 cells. Together, these data suggest that altered c-Yes expression may play a role in the malignant progression of the human melanocyte towards the brain-metastatic phenotype and that NT enhance the activity of c-Yes in signaling penetration into the matrix of NT-rich stromal microenvironments such as the brain.