Effect of Ivermectin and Atorvastatin on Nuclear Localization of Importin Alpha and Drug Target Expression Profiling in Host Cells from Nasopharyngeal Swabs of SARS-CoV-2- Positive Patients.

Nuclear transport and vesicle trafficking are key cellular functions involved in the pathogenesis of RNA viruses. Among other pleiotropic effects on virus-infected host cells, ivermectin (IVM) inhibits nuclear transport mechanisms mediated by importins and atorvastatin (ATV) affects actin cytoskeleton-dependent trafficking controlled by Rho GTPases signaling. In this work, we first analyzed the response to infection in nasopharyngeal swabs from SARS-CoV-2-positive and -negative patients by assessing the gene expression of the respective host cell drug targets importins and Rho GTPases. COVID-19 patients showed alterations in KPNA3, KPNA5, KPNA7, KPNB1, RHOA, and CDC42 expression compared with non-COVID-19 patients. An in vitro model of infection with Poly(I:C), a synthetic analog of viral double-stranded RNA, triggered NF-κB activation, an effect that was halted by IVM and ATV treatment. Importin and Rho GTPases gene expression was also impaired by these drugs. Furthermore, through confocal microscopy, we analyzed the effects of IVM and ATV on nuclear to cytoplasmic importin α distribution, alone or in combination. Results showed a significant inhibition of importin α nuclear accumulation under IVM and ATV treatments. These findings confirm transcriptional alterations in importins and Rho GTPases upon SARS-CoV-2 infection and point to IVM and ATV as valid drugs to impair nuclear localization of importin α when used at clinically-relevant concentrations.

Anticancer activity of repurposed hemostatic agent desmopressin on AVPR2-expressing human osteosarcoma.

Osteosarcoma is the most prevalent primary bone malignancy. Due to its high aggressiveness, novel treatment strategies are urgently required to improve survival of patients with osteosarcoma, especially those with advanced disease. Desmopressin (dDAVP) is a widely used blood-saving agent that has been repurposed as an adjuvant agent for cancer management due to its antiangiogenic and antimetastatic properties. dDAVP acts as a selective agonist of the vasopressin membrane receptor type 2 (AVPR2) present in the microvascular endothelium and in some cancer cells, including breast, lung, colorectal and neuroendocrine tumor cells. Despite the fact that dDAVP has demonstrated its antitumor efficacy in a wide variety of tumor types, exploration of its potential anti-osteosarcoma activity has, to the best of our knowledge, not yet been conducted. Therefore, the aim of the present study was to evaluate the preclinical antitumor activity of dDAVP in osteosarcoma. Human MG-63 and U-2 OS osteosarcoma cell lines were used to assess in vitro and in vivo therapeutic effects of dDAVP. At low micromolar concentrations, dDAVP reduced AVPR2-expressing MG-63 cell growth in a concentration-dependent manner. In contrast, dDAVP exhibited no direct cytostatic effect on AVPR2-negative U-2 OS cells. As it would be expected for canonical AVPR2-activation, dDAVP raised intracellular cAMP levels in osteosarcoma cells, and coincubation with phosphodiesterase-inhibitor rolipram indicated synergistic antiproliferative activity. Cytostatic effects were associated with increased apoptosis, reduced mitotic index and impairment of osteosarcoma cell chemotaxis, as evaluated by TUNEL-labeling, mitotic body count in DAPI-stained cultures and Transwell migration assays. Intravenous administration of dDAVP (12 µg/kg; three times per week) to athymic mice bearing rapidly growing MG-63 xenografts, was indicated to be capable of reducing tumor progression after a 4-week treatment. No major alterations in animal weight, biochemical or hematological parameters were associated with dDAVP treatment, confirming its good tolerability and safety. Finally, AVPR2 expression was detected by immunohistochemistry in 66% of all evaluated chemotherapy-naive human conventional osteosarcoma biopsies. Taking these findings into account, repurposed agent dDAVP may represent an interesting therapeutic tool for the management of osteosarcoma. Further preclinical exploration of dDAVP activity on orthotopic or metastatic osteosarcoma models are required.