Correction: Enhanced tumorigenicity by mitochondrial DNA mild mutations.

[This corrects the article DOI: 10.18632/oncotarget.3698.].

Cisplatin resistance involves a metabolic reprogramming through ROS and PGC-1α in NSCLC which can be overcome by OXPHOS inhibition.

BACKGROUND: Platinum-based chemotherapy remains the standard of care for most lung cancer cases. However chemoresistance is often developed during the treatment, limiting clinical utility of this drug. Recently, the ability of tumor cells to adapt their metabolism has been associated to resistance to therapies. In this study, we first described the metabolic reprogramming of Non-Small Cell Lung Cancer (NSCLC) in response to cisplatin treatment. METHODS: Cisplatin-resistant versions of the A549, H1299, and H460 cell lines were generated by continuous drug exposure. The long-term metabolic changes, as well as, the early response to cisplatin treatment were analyzed in both, parental and cisplatin-resistant cell lines. In addition, four Patient-derived xenograft models treated with cisplatin along with paired pre- and post-treatment biopsies from patients were studied. Furthermore, metabolic targeting of these changes in cell lines was performed downregulating PGC-1α expression through siRNA or using OXPHOS inhibitors (metformin and rotenone). RESULTS: Two out of three cisplatin-resistant cell lines showed a stable increase in mitochondrial function, PGC1-α and mitochondrial mass with reduced glycolisis, that did not affect the cell cycle. This phenomenon was confirmed in vivo. Post-treatment NSCLC tumors showed an increase in mitochondrial mass, PGC-1α, and a decrease in the GAPDH/MT-CO1 ratio. In addition, we demonstrated how a ROS-mediated metabolism reprogramming, involving PGC-1α and increased mitochondrial mass, is induced during short-time cisplatin exposure. Moreover, we tested how cells with increased PGC-1a induced by ZLN005 treatment, showed reduced cisplatin-driven apoptosis. Remarkably, the long-term metabolic changes, as well as the metabolic reprogramming during short-time cisplatin exposure can be exploited as an Achilles' heel of NSCLC cells, as demonstrated by the increased sensitivity to PGC-1α interference or OXPHOS inhibition using metformin or rotenone. CONCLUSION: These results describe a new cisplatin resistance mechanism in NSCLC based on a metabolic reprogramming that is therapeutically exploitable through PGC-1α downregulation or OXPHOS inhibitors.

Cancer-associated fibroblasts modify lung cancer metabolism involving ROS and TGF-ß signaling.

Lung cancer is a major public health problem due to its high incidence and mortality rate. The altered metabolism in lung cancer is key for the diagnosis and has implications on both, the prognosis and the response to treatments. Although Cancer-associated fibroblasts (CAFs) are one of the major components of the tumor microenvironment, little is known about their role in lung cancer metabolism. We studied tumor biopsies from a cohort of 12 stage IIIA lung adenocarcinoma patients and saw a positive correlation between the grade of fibrosis and the glycolysis phenotype (Low PGC-1α and High GAPDH/MT-CO1 ratio mRNA levels). These results were confirmed and extended to other metabolism-related genes through the in silico data analysis from 73 stage IIIA lung adenocarcinoma patients available in TCGA. Interestingly, these relationships are not observed with the CAFs marker α-SMA in both cohorts. To characterize the mechanism, in vitro co-culture studies were carried out using two NSCLC cell lines (A549 and H1299 cells) and two different fibroblast cell lines. Our results confirm that a metabolic reprogramming involving ROS and TGF-ß signaling occurs in lung cancer cells and fibroblasts independently of α-SMA induction. Under co-culture conditions, Cancer-Associated fibroblasts increase their glycolytic ability. On the other hand, tumor cells increase their mitochondrial function. Moreover, the differential capability among tumor cells to induce this metabolic shift and also the role of the basal fibroblasts Oxphos Phosphorylation (OXPHOS) function modifying this phenomenon could have implications on both, the diagnosis and prognosis of patients. Further knowledge in the mechanism involved may allow the development of new therapies.

PGC-1alpha levels correlate with survival in patients with stage III NSCLC and may define a new biomarker to metabolism-targeted therapy.

Lung cancer remains the leading cause of cancer-related death worldwide, with one-third diagnosed with locally advanced (stage III) disease. Preoperative induction chemo-radiotherapy is key for the treatment of these patients, however conventional cisplatin based approaches has apparently reached a plateau of effectiveness. In the search for new therapies, the targeting of tumor metabolism is revealed as an interesting option to improve the patient's responses. Here we describe the importance of PGC-1alpha and GAPDH/MT-CO1 ratio levels as surrogates of the Warburg effect from a series of 28 stage III NSCLC patients, on PFS, OS and PET uptake. Moreover, our results show a great variability between tumors of different individuals, ranging from very glycolytic to more OXPHOS-dependent tumors, which compromises the success of therapies directed to metabolism. In this sense, using 3 different cell lines, we describe the relevance of Warburg effect on the response to metabolism-targeted therapies. Specifically, we show that the inhibitory effect of metformin on cell viability depends on cell's dependence on the OXPHOS system. The results on cell lines, together with the results of PGC-1alpha and GAPDH/MT-CO1 as biomarkers on patient's biopsies, would point out what type of patients would benefit more from the use of these drugs.

Functional Characterization of Three Concomitant MtDNA LHON Mutations Shows No Synergistic Effect on Mitochondrial Activity.

The presence of more than one non-severe pathogenic mutation in the same mitochondrial DNA (mtDNA) molecule is very rare. Moreover, it is unclear whether their co-occurrence results in an additive impact on mitochondrial function relative to single mutation effects. Here we describe the first example of a mtDNA molecule harboring three Leber's hereditary optic neuropathy (LHON)-associated mutations (m.11778G>A, m.14484T>C, m.11253T>C) and the analysis of its genetic, biochemical and molecular characterization in transmitochondrial cells (cybrids). Extensive characterization of cybrid cell lines harboring either the 3 mutations or the single classic m.11778G>A and m.14484T>C mutations revealed no differences in mitochondrial function, demonstrating the absence of a synergistic effect in this model system. These molecular results are in agreement with the ophthalmological characteristics found in the triple mutant patient, which were similar to those carrying single mtDNA LHON mutations.

Enhanced tumorigenicity by mitochondrial DNA mild mutations.

To understand how mitochondria are involved in malignant transformation we have generated a collection of transmitochondrial cybrid cell lines on the same nuclear background (143B) but with mutant mitochondrial DNA (mtDNA) variants with different degrees of pathogenicity. These include the severe mutation in the tRNALys gene, m.8363G>A, and the three milder yet prevalent Leber's hereditary optic neuropathy (LHON) mutations in the MT-ND1 (m.3460G>A), MT-ND4 (m.11778G>A) and MT-ND6 (m.14484T>C) mitochondrial genes. We found that 143B ρ0 cells devoid of mtDNA and cybrids harboring wild type mtDNA or that causing severe mitochondrial dysfunction do not produce tumors when injected in nude mice. By contrast cybrids containing mild mutant mtDNAs exhibit different tumorigenic capacities, depending on OXPHOS dysfunction.The differences in tumorigenicity correlate with an enhanced resistance to apoptosis and high levels of NOX expression. However, the final capacity of the different cybrid cell lines to generate tumors is most likely a consequence of a complex array of pro-oncogenic and anti-oncogenic factors associated with mitochondrial dysfunction.Our results demonstrate the essential role of mtDNA in tumorigenesis and explain the numerous and varied mtDNA mutations found in human tumors, most of which give rise to mild mitochondrial dysfunction.