The HIV-protease inhibitor saquinavir reduces proliferation, invasion and clonogenicity in cervical cancer cell lines

Abstract

Innovative therapies in cervical cancer (CC) remain a priority. Recent data indicate that human immunodeficiency virus (HIV)-protease inhibitors used in highly active antiretroviral therapy can exert direct antitumor activities also in HIV-free preclinical and clinical models. The aim of the present study was to evaluate the antineoplastic effects of various HIV-protease inhibitors (indinavir, ritonavir and saquinavir) on primary and established CC cell lines. Two CC cell lines established in our laboratory and four commercially available CC cell lines were treated with indinavir, ritonavir and saquinavir at different concentrations and for different times. Proliferation, clonogenicity and radiosensitivity were evaluated by crystal violet staining. Proteasomal activities were assessed using a cell-based assay and immunoblotting. Cell cycle was analyzed by propidium iodide staining and flow cytometric analysis. Invasion was tested with Matrigel chambers. A t-test for paired samples was used for statistical analysis. In all cell lines, saquinavir was more effective than ritonavir in reducing cell proliferation and inhibiting proteasomal activities (P≤0.05). Conversely, indinavir exerted a negligible effect. The saquinavir concentrations required to modulate the proteasome activities were higher than those observed to be effective in inhibiting cell proliferation. In HeLa cells, saquinavir was strongly effective in inhibiting cell invasion and clonogenicity (P≤0.05) at concentrations much lower than those required to perturb proteasomal activities. Saquinavir did not contribute to increase the sensitivity of HeLa cells to X-rays. In conclusion, the present results demonstrate that saquinavir is able to significantly reduce cell proliferation, cell invasion and clonogenicity in a proteasome-independent manner in in vitro models of CC, and suggest that saquinavir could be a promising CC therapeutic agent.

HIV-1 Protease Inhibitors Slow HPV16-Driven Cell Proliferation through Targeted Depletion of Viral E6 and E7 Oncoproteins

“…One interpretation of these findings is that NFV and SQV induce selective proteasome-mediated degradation of the two viral oncoproteins, which would be remarkable given that protease inhibitors were previously posited to cause inhibition of host proteasomes—an activity that, if global, would be predicted to have the opposite effect on E6 and E7 [ 49 ]. Indeed, a prior study showed SQV treatment of HPV-positive HeLa cells to increase overall levels of ubiquitinated proteins at the whole cell level [ 35 ]), albeit at levels of drug much higher than those tested in our study (>60 µM). Also unclear is what is common about E6 and E7 that would make these two proteins similarly subjected to the effects of these drugs.…”

“…Here, the complexity of HIV-HPV interactions and persistence of HPV disease in PLWH, combined with the fact that certain protease inhibitors have proven effective in treating HPV-positive cervical cancer and its precursor lesions [ 32 , 33 , 34 , 35 , 36 ], prompted us to study the specific effects of protease inhibitors on HPV16 E6 and E7 gene expression using two HPV-positive cell culture models: (1) cervical cancer-derived CaSki cells carrying more than 200 integrated copies of the high-risk HPV16 genome, and (2) normal immortalized human foreskin keratinocytes harboring extrachromosomal HPV16 DNA (NIKS16 cells) that grow in organotypic (raft) culture to form a stratified, 3-D biomimetic model of HPV-positive epithelial pre-cancer. We report the surprising finding that a subset of PIs (lopinavir, rotinavir, nelfinavir, and saquinavir) markedly reduce levels of HPV16 E6 and E7 proteins in both CaSki and NIKS16 cells correlating with suppression of HPV16-positive cell growth.…”

“…Interestingly, protease inhibitors, in particular nelfinavir, have also been studied independently of HIV-1 as anticancer agents based on their capacity to trigger cancer cell death, thought to be mediated through inhibition of the host ubiquitin-proteasome system (UBS) [29][30][31]. These benefits have previously been demonstrated in the context of HPV-positive cancers [32][33][34][35] including clinical data from Hampson and colleagues demonstrating that a combination of HIV protease inhibitors (lopinavir and ritonavir) causes regression of cervical dysplasia in HPV-positive women with high-grade cervical intraepithelial neoplasia [36].…”

Senescence as a main mechanism of Ritonavir and Ritonavir‐NO action against melanoma

“…However, Goehe et al reported that in breast and colon carcinoma cells senescence was independent of autophagy and occurred even when autophagy was suppressed. On the basis of these data we hypothesize that the primary trigger of senescence induction by Ritonavir and Ritonavir‐NO triggered is their ability to inhibit proteasome activity in mammalian cells, namely, oxidized and cross‐linked protein aggregates such as lipofuscin, age pigments, etc that are key markers of aging and senescence, accumulated due to disturbed proteasome degradation …”

“…Clonogenic assay provides proof of the irreversible proliferation arrest after a period of compound‐free growth posttreatment. Saquinavir reduced clonogenicity of HeLa cells after 96 hours of treatment . In this study, the clonogenic effect was observed after 48 hours of treatment and 5 days of cell cultivation without Ritonavir and Ritonavir‐NO, evidencing a long‐lasting effect of these compounds on colony formation.…”