BackgroundIn non-small cell lung cancer (NSCLC) KRAS-mutant status is a negative prognostic and predictive factor. Nitrogen-containing bisphosphonates inhibit prenylation of small G-proteins (e.g. Ras, Rac, Rho) and thus may affect proliferation and migration. In our preclinical work, we investigated the effect of an aminobisphosphonate compound (zoledronic acid) on mutant and wild type KRAS-expressing human NSCLC cell lines.ResultsWe confirmed that zoledronic acid was unable to inhibit the prenylation of mutant K-Ras unlike in the case of wild type K-Ras. In case of in vitro proliferation, the KRAS-mutant human NSCLC cell lines showed resistance to zoledronic acid wild-type KRAS-cells proved to be sensitive. Combinatory application of zoledronic acid enhanced the cytostatic effect of cisplatin. Zoledronic acid did not induce significant apoptosis. In xenograft model, zoledronic acid significantly reduced the weight of wild type KRAS-EGFR-expressing xenograft tumor by decreasing the proliferative capacity. Futhermore, zoledronic acid induced VEGF expression and improved in vivo tumor vascularization.Materials and methodsMembrane association of K-Ras was examined by Western-blot. In vitro cell viability, apoptotic cell death and migration were measured in NSCLC lines with different molecular background. The in vivo effect of zoledronic acid was investigated in a SCID mouse subcutaneous xenograft model.ConclusionsThe in vitro and in vivo inhibitory effect of zoledronic acid was based on the blockade of cell cycle in wild type KRAS-expressing human NSCLC cells. The zoledronic acid induced vascularization supported in vivo cytostatic effect. Our preclinical investigation suggests that patients with wild type KRAS-expressing NSCLC could potentially benefit from aminobisphosphonate therapy.

In non-small cell lung cancer (NSCLC) KRAS-mutant status is a negative prognostic and predictive factor of both classic and target based therapy. In human malignancies bisphosphonate drugs are applied to treat or prevent formation of bone metastases. Nitrogen-containing bisphosphonates inhibit the posttranslational modification of small G-proteins (e.g. Ras, Rac, Rho) and thus may affect proliferation and migration. In our preclinical work, we investigated the effect of an aminobisphosphonate compound (zoledronic acid) on mutant and wild type KRAS-expressing human NSCLC cell lines. In vitro cell viability was measured by MTT assay in H358, LCLC-103H, H1650 and H1975 cells. Apoptotic cell death was quantified by flow cytometric detection of sub-G1-fraction in propidium-iodide stained cultures. Furthermore, we investigated the in vivo effect of zoledronic acid in a SCID mouse subcutaneous xenograft model. The extracted tumors were examined by traditional histological methods and the proportion of proliferating tumor cells was determined by Ki67 labeling. KRAS-mutant H358 human NSCLC cell line showed relative resistance to zoledronic acid on proliferation, while human lung cancer cell lines carried wild type KRAS proved to be sensitive. Zoledronic acid improved the cytostatic effect of traditional anti-cancer agent cisplatin. Zoledronic acid did not induce significant apoptosis in NSCLC cell lines in vitro, rather the blockade of the cell cycle appeared. In in vivo subcutaneous animal model, zoledronic acid significantly reduced the weight of LCLC-103H xenograft tumor harboring wild type KRAS, furthermore, the treatment decreased the number of Ki67-labeled tumor. Zoledronic acid induced the expression of VEGF in LCLC-103H cells and stimulated in vivo angiogenesis. We concluded that in human NSCLC cells the antiproliferative effect of zoledronic acid dependended on KRAS-status (wild-type or mutant). The inhibitory effect of zoledronic manifested through the in vitro and in vivo blockade of cell cycle. The in vivo angiogenic effect of zoledronic acid supported cytostatic effect of cisplatin. Our preclinical investigation suggests that patients with wild type KRAS-expressing NSCLC could potentially benefit from aminobisphosphonate therapy. Citation Format: István Kenessey, Krisztina Kói, Mihály Cserepes, Judit Dobos, Balázs Hegedűs, József Tóvári, József Tímár. KRAS-mutation dependent effect of zoledronic acid in human NSCLC preclinical models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 47. doi:10.1158/1538-7445.AM2015-47