Whereas 40% of human breast carcinomas harbor mutations in the tumor suppressor protein p53, the use of tests demonstrating the presence of p53 mutations as a prognostic marker in breast cancer has not altered clinical management. Therefore, the search for new markers, especially among cell cycle-regulatory molecules, is a high priority, both in terms of prognostication and for identification of novel targets. p21 regulates the outcome of the p53 response to DNA damage, as might occur after administration of a chemotherapeutic agent, and we have shown that attenuation of p21 using an antisense oligodeoxynucleotide (ODN) inhibits cell proliferation in vitro and decreases growth of Met-1 mammary carcinomas in mice. In the current study, we extend this work to human cells and tissue. Three of eight human breast tumors that we obtained from a tissue bank show markedly increased p21 levels, variably staining in the nucleus and cytosol. All corresponding normal tissues were p21 negative. In the three p21-positive tumors, the phosphatidylinositol 3′-kinase-relevant signaling proteins p85 and PTEN were also increased. To investigate whether p21 is a feasible target for attenuation in human breast cancer, we investigated two human carcinoma cell lines. When transfected with antisense p21 ODN, both MCF7 and T47D breast cancer cells exhibit dose-dependent attenuation of p21 levels, associated with apoptosis in the absence of an additional apoptotic stimulus. Because p21 regulates the cellular repair response to damaged DNA, our work suggests that attenuation of p21 using our antisense p21 ODN may be effective in modulating the progression of breast cancer in either the presence or absence of combination chemotherapy and sets the stage for future clinical trials.