HIV-1 Tat Peptide Immunoconjugates Differentially Sensitize Breast Cancer Cells to Selected Antiproliferative Agents That Induce the Cyclin-Dependent Kinase Inhibitor p21~(WAF-1/CIP-1)

摘要

In this study,we evaluated the ability of anti-p21 antibodies conjugated to 17-mer peptides [GRKKRRQRRRP-PQGYGC] harboring the membrane-translocating and nuclear import sequences [underlined] of HIV-1 tat protein to inhibit the cyclin-dependent kinase inhibitor,p21~(WAF-1/Cip-1) (p21) and differentially sensitize MDA-MB-468 and MCF-7 human breast cancer (BC) cells to the antiproliferative effects of treatments that induce or do not induce p21.BC cells were treated with increasing concentrations of epidermal growth factor (EGF;0.5-10 nM),the topoisomerase I inhibitor,camptothecin (CPT;0.1-4 mu M),or increasing doses of gamma-radiation (2-20 Gy).Western blot was used to evaluate p21 expression.The effect of treatment on cell cycle distribution was studied.Growth inhibition was measured by the WST-1 assay.Expression of p21 was increased in MDA-MB-468 cells treated with EGF or CPT but not by gamma-irradiation.MCF-7 cells exhibited p21 upregulation following exposure to CPT and y-radiation but not EGF.EGF caused cell cycle arrest in G_1 phase for MDA-MB-468 cells.CPT caused G_1-phase arrest in MDA-MB-468 cells and prolonged S phase in MCF-7 cells.gamma-Radiation caused an increase in cells in G_2/M phase for MDA-MB-468 and MCF-7.MDA-MB-468 cells were growth-inhibited by EGF,CPT,and y-radiation.MCF-7 cells were growth-stimulated by EGF and inhibited by CPT and y-radiation.Combining EGF with tat-anti-p21 immunoconjugates (ICs) amplified the growth-inhibitory effect on MDA-MB-468 cells 1.2-fold to 2.3-fold,but had no effect on the growth stimulation of MCF-7 cells by EGF.Tat-anti-p21 ICs sensitized MCF-7 cells 1.4-fold to gamma-radiation but had no effect on the growth of gamma-irradiated MDA-MB-468 cells.Tat-anti-p21 ICs sensitized both MDA-MB-468 and MCF-7 cells 1.7-fold to CPT.We conclude that tat-anti-p21 ICs are promising sensitizers for cytotoxic cancer therapies and that their sensitization is dependent on treatment-related p21 expression.This general approach could potentially be extended to other growth-regulatory molecules that are associated with tumor growth and progression.