Synergistic Effects of Sanglifehrin-Based Cyclophilin Inhibitor NV651 with Cisplatin in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC), commonly diagnosed at an advanced stage, is the most common primary liver cancer. Owing to a lack of effective HCC treatments and the commonly acquired chemoresistance, novel therapies need to be investigated. Cyclophilins-intracellular proteins with peptidyl-prolyl isomerase activity-have been shown to play a key role in therapy resistance and cell proliferation. Here, we aimed to evaluate changes in the gene expression of HCC cells caused by cyclophilin inhibition in order to explore suitable combination treatment approaches, including the use of chemoagents, such as cisplatin. Our results show that the novel cyclophilin inhibitor NV651 decreases the expression of genes involved in several pathways related to the cancer cell cycle and DNA repair. We evaluated the potential synergistic effect of NV651 in combination with other treatments used against HCC in cisplatin-sensitive cells. NV651 showed a synergistic effect in inhibiting cell proliferation, with a significant increase in intrinsic apoptosis in combination with the DNA crosslinking agent cisplatin. This combination also affected cell cycle progression and reduced the capacity of the cell to repair DNA in comparison with a single treatment with cisplatin. Based on these results, we believe that the combination of cisplatin and NV651 may provide a novel approach to HCC treatment.

Novel Cyclophilin Inhibitor Decreases Cell Proliferation and Tumor Growth in Models of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC), the most common primary liver cancer, is usually diagnosed in its late state. Tyrosine kinase inhibitors such as sorafenib and regorafenib are one of the few treatment options approved for advanced HCC and only prolong the patient's life expectancy by a few months. Therefore, there is a need for novel effective treatments. Cyclophilins are intracellular proteins that catalyze the cis/trans isomerization of peptide bonds at proline residues. Cyclophilins are known to be overexpressed in HCC, affecting therapy resistance and cell proliferation. In the present study, we explored the potential of cyclophilin inhibitors as new therapeutic options for HCC in vitro and in vivo. Our results showed that the novel cyclophilin inhibitor, NV651, was able to significantly decrease proliferation in a diverse set of HCC cell lines. The exposure of HCC cells to NV651 caused an accumulation of cells during mitosis and consequent accumulation in the G2/M phase of the cell cycle. NV651 reduced tumor growth in vivo using an HCC xenograft model without affecting the body weights of the animals. The safety aspects of NV651 were also confirmed in primary human hepatocytes without any cytotoxic effects. Based on the results obtained in this study, we propose NV651 as a potential treatment strategy for HCC.

Structurally distinct cyclosporin and sanglifehrin analogs CRV431 and NV556 suppress established HCV infection in humanized-liver mice.

We previously reported that the non-immunosuppressive cyclophilin inhibitors (CypIs)-cyclosporin A analog CRV431 and sanglifehrin analog NV556-efficiently inhibit HCV replication in vitro. In this study, we asked whether they can also reduce HCV replication in vivo. We found that a single oral administration of CRV431 and NV556 to HCV-infected humanized-liver mice drastically reduced HCV blood levels. The antiviral effect was observed when CRV431 or NV556 were each individually administered with HCV, 3, 6 weeks or even 3 months post-infection when viral replication is robust. These results were confirmed in chimeric mice implanted with human hepatocytes isolated from three distinct donors. Remarkably, no viral rebound was observed 5 months after a single dose administration of 50 mg/kg of CRV431 or NV556 four weeks post-HCV infection, indicating the possibility of suppression of an established viral infection. Since we recently demonstrated that both CRV431 and NV556 also inhibit the development of liver fibrosis and hepatocellular carcinoma in nonviral-induced non-alcoholic steatohepatitis mouse models, our present data suggest that the two entirely structurally different CypIs-CRV431 and NV556-derived from unrelated natural products, represent attractive partners to current direct-acting agent (DAA) regimens for the treatment of hepatitis C and liver diseases.

Cyclophilin Inhibitor NV556 Reduces Fibrosis and Hepatocellular Carcinoma Development in Mice With Non-Alcoholic Steatohepatitis.

Hepatocellular carcinoma (HCC), the third major cause of cancer mortality, can result from non-alcoholic steatohepatitis (NASH). Due to limited efficacy of drugs approved for HCC and no drug available yet for NASH, identification of new effective treatments is crucial. Here, we investigated whether NV556, a cyclophilin inhibitor derived from sanglifehrins, would decrease the development of NASH and HCC in a preclinical mouse model. For our experiment, male mice were administered streptozotocin to disrupt pancreatic cells and nourished with high-fat diet since 3 weeks of age. Afterward, NV556 or vehicle was orally administered daily for 6 weeks before the 14-week-old time point for the development of NASH, or 10 weeks before the 30-week-old time point for the establishment of HCC. Body weight, blood glucose level, and liver weight were recorded. Moreover, for NASH, livers were histologically examined for inflammation and steatosis. Collagen was measured by Sirius Red staining of hepatic tissues. Systemic cytokine levels in serum were detected by multiplex assays. For HCC, nodules of livers were measured and scored according to a developed system with number and size of nodules as criteria. NV556 significantly decreased collagen deposition (p = 0.0281), but did not alter inflammation, steatosis, body and liver weight, and systemic cytokine production compared to control mice with NASH symptoms. For HCC, NV556 statistically reduced the number (p = 0.0091) and diameter of tumorous nodules (p = 0.0264), along with liver weight (p = 0.0026) of mice.Our study suggests NV556 as a promising candidate for treatment of NASH-derived fibrosis and HCC.

Bioengineering and semisynthesis of an optimized cyclophilin inhibitor for treatment of chronic viral infection.

Inhibition of host-encoded targets, such as the cyclophilins, provides an opportunity to generate potent high barrier to resistance antivirals for the treatment of a broad range of viral diseases. However, many host-targeted agents are natural products, which can be difficult to optimize using synthetic chemistry alone. We describe the orthogonal combination of bioengineering and semisynthetic chemistry to optimize the drug-like properties of sanglifehrin A, a known cyclophilin inhibitor of mixed nonribosomal peptide/polyketide origin, to generate the drug candidate NVP018 (formerly BC556). NVP018 is a potent inhibitor of hepatitis B virus, hepatitis C virus (HCV), and HIV-1 replication, shows minimal inhibition of major drug transporters, and has a high barrier to generation of both HCV and HIV-1 resistance.

Bioequivalence and tolerability assessment of a novel intravenous ciclosporin lipid emulsion compared to branded ciclosporin in Cremophor ® EL.

BACKGROUND: Ciclosporin is used as an immunosuppressant in current clinical practice but recent research implies novel indications for the drug, such as neuro- and cardioprotection. The intravenous formulation currently on the market, Sandimmune(®) Injection (Sandimmune(®)), uses Cremophor(®) EL as emulsifying excipient. Cremophor(®) EL is known to cause hypersensitivity reactions in some patients, ranging from skin reactions to potentially fatal anaphylactic shock. OBJECTIVES: The primary objective was to assess if CicloMulsion(®), a Cremophor(®) EL-free lipid emulsion of ciclosporin for intravenous administration, is bioequivalent to Sandimmune(®), and the secondary objective was to compare the tolerability profiles of the two preparations. METHODS: This was a single-centre, open-label, subject-blind, laboratory-blind, single-dose, randomized, two-treatment, two-period, two-sequence crossover study of the pharmacokinetics of two formulations of intravenous ciclosporin. Fifty-two healthy volunteer subjects were administered 5 mg/kg of each of the two formulations of ciclosporin as a 4-h intravenous infusion. The last blood sample was acquired 48 h after the end of the infusion. Bioequivalence assessments according to current guidelines were performed. RESULTS: The geometric mean ratios for CicloMulsion(®)/Sandimmune(®) (90 % confidence interval [CI]) were 0.90 (0.88, 0.92) for AUC(0-last) (area under the blood concentration-time curve from time zero to time of last measurable concentration) and 0.95 (0.92, 0.97) for C(max) (maximum blood concentration). For all additional variables analysed, the 90 % CIs were also within the accepted bioequivalence range of 0.80-1.25. One anaphylactoid and one anaphylactic reaction, both classified as serious adverse events, were reported after treatment with Sandimmune(®). No serious adverse events were recorded after treatment with CicloMulsion(®). CONCLUSION: We have assessed the pharmacokinetics and tolerability of a new Cremophor(®) EL-free lipid emulsion of ciclosporin, CicloMulsion(®), compared to Sandimmune(®). The proportion of adverse events was significantly higher for the Cremophor(®) EL-based product Sandimmune(®). We conclude that CicloMulsion(®) is bioequivalent to Sandimmune(®) and exhibits fewer adverse reactions.