Purpose: These studies were designed to determine whether ritonavir exhibits inhibitory activity against non-small cell lung cancer (NSCLC) lines in vitro and whether inhibition of receptor kinase pathways or promotion of apoptotic pathways can potentiate ritonavir activity. Reduction of survivin mRNA has been demonstrated to potentiate chemotherapy activity and therefore, its potential role in promoting ritonavir inhibition of NSCLC was examined. Experimental Design: Ritonavir was tested for inhibition of NSCLC growth and promotion of apoptosis in the NSCLC lines A549, H460, H522 and H23. Effects of siRNA targeting c-Src, STAT3 and survivin on ritonavir inhibition of NSCLC cell proliferation were studied. Results: Ritonavir inhibits growth of A549, H460, H522 and H23 lines, exhibiting IC50 values of 35, 48, 42 and 44 microM, respectively. The lines A549 and H522 lines were studied in more detail for mechanism of ritonavir sensitivity. Ritonavir down-regulates CDK2, 4, and 6, and cyclin D1 and dephosphorylates Rb, associated with G1 arrest. Ritonavir also induces apoptosis, but Akt and ERK phosphorylation may be increased or decreased depending upon the line, suggesting that this drug may inhibit other NSCLC survival pathways. Ritonavir reduced phospho-c-Src, phospho-STAT3 and survivin levels. siRNA species that target c-Src, STAT3 and survivin markedly reduce the proliferation of the A549 and H522 lines and induce apoptosis, confirming that all three lines are dependent upon these signaling proteins. To develop a therapeutic strategy to effectively use ritonavir for NSCLC inhibition, it was determined whether c-Src, STAT3 or survivin siRNA potentiates ritonavir inhibition. Survivin siRNA reduces the ritonavir IC50 to 25 μM for both lines, while c-Src and STAT3 siRNA had no effect, indicating a specific role for survivin in resistance to ritonavir. Over-expression of survivin increases the ritonavir IC50 in both lines further suggesting that ritonavir resistance may be mediated through the survivin. Conclusions: These results indicate that ritonavir inhibits NSCLC growth, in part, by inhibiting CDK regulation of cell cycle progression, in part, by inhibition of c-Src-, STAT3- and survivin-mediated proliferation and survival. Ritonavir may be of interest for clinical development for NSCLC therapeutics and its efficacy may be potentiated by strategies that target survivin.
[Proc Amer Assoc Cancer Res, Volume 47, 2006]