Abstract B181: A lentivirus-encoding shRNA targeting a noncoding mitochondrial RNA inhibits melanoma tumor growth and metastasis in a mouse model in vivo.

Abstract

Abstract We have described a family of non-coding mitochondrial RNAs (ncmtRNA) differentially expressed in normal and tumor cells. Normal proliferating cells express high levels of both sense and antisense ncmtRNAs. In contrast, nondividing cells express low levels of these transcripts. Interestingly, tumor cells express sense ncmtRNA and down-regulate antisense ncmtRNA. Previously, we have shown that transfection with oligonucleotides complementary to any of these ncmtRNAs induce death in different cancer cells type in vitro, but not in normal cells, raising the possibility of using interference to target these transcripts as a potential cancer treatment. The aim of this study was to determine whether in vivo administration of a lentivirus encoding shRNA against both ncmtRNA could inhibit melanoma tumor growth and metastasis in a mouse model. Non-replicative lentiviral vectors encoding shRNA against ncmtRNA (Lv-ncmtRNA) and luciferase (Lv-control) were constructed and their effects were evaluated in vitro by transducing B16F10 melanoma cells. A 6-fold decrease in relative expression of sense and antisense ncmtRNA was observed in cells transduced with Lv-ncmtRNA in comparison to cells transduced with Lv-control as measured by real time quantitative PCR. In addition, Lv-ncmtRNA transduction induces apoptotic cell death determined by cell surface exposure of phosphatidylserine and DNA fragmentation. The ability of this strategy to inhibit growth of solid tumors and metastasis of B16F10 melanoma cells was evaluated in vivo. C57BL/6 mice bearing subcutaneous B16F10 tumors received five intratumoral injections with 4×107 TU of Lv-ncmtRNA or Lv-control every two days. Mice treated with Lv-ncmtRNA showed a significant (p=0.0007) decrease in tumor volume (458 × 83 mm3) compared to mice treated with Lv-control (1167 × 154 mm3) 20 days after cell inoculation. For the metastasis assay, C57BL/6 mice were injected intravenously with 4×105 B16F10 cells and four days later treated with four intravenous administrations of 5×107 TU Lv-ncmtRNA or Lv-control every three days. Mice were sacrificed two weeks after cell injection for analysis of lung metastasis. Mice treated with Lv-ncmtRNA showed a significant (p=0,0005) decrease in the number of metastatic foci (77 × 9) compared to mice treated with Lv-control (286 × 18) as well as in lung weight (393,4 × 26,2 mg v/s 290,2 × 9,2 mg, p = 0,0005). In addition, hematoxylin and eosin-stained tissue sections also showed that the lung treated with Lv-ncmtRNA had only a few small nodules. In contrast, metastasis nodules occupied most of the lung when mice were treated whit Lv-control. These results suggest that intratumoral and systemic administration of shRNA targeting ncmtRNA represents a promising strategy towards the development of potential cancer therapies. Financed by CONICYT PFB-16; UNAB DI-11-11/I. MV is supported by a CONICYT doctoral fellowship. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B181.