Abstract
We reported that knockdown of the antisense noncoding mitochondrial RNAs (ASncmtRNAs) induces apoptotic death of several human tumor cell lines, but not normal cells, suggesting this approach for selective therapy against different types of cancer. In order to translate these results to a preclinical scenario, we characterized the murine noncoding mitochondrial RNAs (ncmtRNAs) and performed in vivo knockdown in syngeneic murine melanoma models. Mouse ncmtRNAs display structures similar to the human counterparts, including long double-stranded regions arising from the presence of inverted repeats. Knockdown of ASncmtRNAs with specific antisense oligonucleotides (ASO) reduces murine melanoma B16F10 cell proliferation and induces apoptosis in vitro through downregulation of pro-survival and metastasis markers, particularly survivin. For in vivo studies, subcutaneous B16F10 melanoma tumors in C57BL/6 mice were treated systemically with specific and control antisense oligonucleotides (ASO). For metastasis studies, tumors were resected, followed by systemic administration of ASOs and the presence of metastatic nodules in lungs and liver was assessed. Treatment with specific ASO inhibited tumor growth and metastasis after primary tumor resection. In a metastasis-only assay, mice inoculated intravenously with cells and treated with the same ASO displayed reduced number and size of melanoma nodules in the lungs, compared to controls. Our results suggest that ASncmtRNAs could be potent targets for melanoma therapy. To our knowledge, the ASncmtRNAs are the first potential non-nuclear targets for melanoma therapy.
Highlights
Human cells express a unique family of sense and antisense non-coding mitochondrial RNAs [1,2,3]
We reported that knockdown of the antisense noncoding mitochondrial RNAs (ASncmtRNAs) induces apoptotic death of several human tumor cell lines, but not normal cells, suggesting this approach for selective therapy against different types of cancer
How do we explain that ASK is efficient in inducing knockdown of the ASncmtRNAs since it is well known that oligonucleotides are not able to enter mitochondria in vivo [7, 8]? Previously, we have demonstrated that in normal human kidney, renal cell carcinoma, mouse testis and the murine melanoma cell line B16F10, the SncmtRNA and the ASncmtRNAs exit the mitochondria and are found localized in the cytoplasm and in the nucleus [9]
Summary
Human cells express a unique family of sense and antisense non-coding mitochondrial RNAs (ncmtRNAs) [1,2,3]. The sense transcript (SncmtRNA) is expressed in normal proliferating cells and tumor cells, but not in resting cells, suggesting a role of this transcript in cell proliferation [1,2,3]. Besides SncmtRNA, normal proliferating cells express two antisense transcripts: ASncmtRNA-1 and ASncmtRNA-2 [2]. The ASncmtRNAs are downregulated in human tumor cell lines and tumor cells present in biopsies of different types of cancer [2]. Downregulation of the ASncmtRNAs seems to be an important step in carcinogenesis and represents a new and generalized pro-tumorogenic hallmark of cancer [4]. Other mitochondrial transcripts containing stem-loop structures have been reported, their function is unknown [5]
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