Mitochondrial genomics in the cancer cell line encyclopedia and a scoring method to effectively pair cell lines for cytoplasmic hybridization.

Mitochondrial sequence variants have been associated with many human diseases, including cancer. A well-established experimental strategy to assess the impact of mitochondrial sequence variants is to generate cytoplasmic hybrids (cybrids). Cybridization facilitates the study of mitochondrial DNA (mtDNA) mutations under a controlled nuclear genetic background. However, in generating cybrids, it is important to select most suitable mtDNA donor and recipient cells so that any change in recipient's cellular phenotype may be attributed to the introduction, through the donor, of mutations in the gene of interest. Here we catalog mitochondrial sequence variants, gene expression, and haplogroups across 937 publicly available cancer cell lines curated by the Cancer Cell Line Encyclopedia (CCLE). We then present DoReMi (Donor, Recipient Mitochondrial DNA matching) as a method to score candidate donor and recipient cell lines based on their mtDNA mutational profiles, including their heteroplasmy levels. DoReMi will allow researchers to design successful cybrid experiments for querying the role of mutations in their mitochondrial-encoded gene of interest. Researchers may also apply DoReMi to study their own cell lines. DoReMi and the other resources provided by this study help optimize cybridization experiments. This software is available for use at http://mendel.gene.cwru.edu/laframboiselab/software.php.

Genomic determinants of chronic myelomonocytic leukemia.

The biology, clinical phenotype and progression rate of chronic myelomonocytic leukemia (CMML) are highly variable due to diverse initiating and secondary clonal genetic events. To determine the effects of molecular features including clonal hierarchy in CMML, we studied whole-exome and targeted next-generation sequencing data from 150 patients with robust clinical and molecular annotation assessed cross-sectionally and at serial time points of disease evolution. To identify molecular lesions unique to CMML, we compared it to the related myeloid neoplasms (N=586), including juvenile myelomonocytic leukemia, myelodysplastic syndromes (MDS) and primary monocytic acute myeloid leukemia and discerned distinct molecular profiles despite similar pathomorphological features. Within CMML, mutations in certain pathways correlated with clinical classification, for example, proliferative vs dysplastic features. While most CMML patients (59%) had ancestral (dominant/co-dominant) mutations involving TET2, SRSF2 or ASXL1 genes, secondary subclonal hierarchy correlated with clinical phenotypes or outcomes. For example, progression was associated with acquisition of new expanding clones carrying biallelic TET2 mutations or RAS family, or spliceosomal gene mutations. In contrast, dysplastic features correlated with mutations usually encountered in MDS (for example, SF3B1 and U2AF1). Classification of CMML based on hierarchies of ancestral and subclonal mutational events may correlate strongly with clinical features and prognosis.

Selected mitochondrial DNA landscapes activate the SIRT3 axis of the UPRmt to promote metastasis.

By causing mitochondrial DNA (mtDNA) mutations and oxidation of mitochondrial proteins, reactive oxygen species (ROS) leads to perturbations in mitochondrial proteostasis. Several studies have linked mtDNA mutations to metastasis of cancer cells but the nature of the mtDNA species involved remains unclear. Our data suggests that no common mtDNA mutation identifies metastatic cells; rather the metastatic potential of several ROS-generating mutations is largely determined by their mtDNA genomic landscapes, which can act either as an enhancer or repressor of metastasis. However, mtDNA landscapes of all metastatic cells are characterized by activation of the SIRT/FOXO/SOD2 axis of the mitochondrial unfolded protein response (UPRmt). The UPRmt promotes a complex transcription program ultimately increasing mitochondrial integrity and fitness in response to oxidative proteotoxic stress. Using SOD2 as a surrogate marker of the UPRmt, we found that in primary breast cancers, SOD2 is significantly increased in metastatic lesions. We propose that the ability of selected mtDNA species to activate the UPRmt is a process that is exploited by cancer cells to maintain mitochondrial fitness and facilitate metastasis.

The major lung cancer-derived mutants of ERBB2 are oncogenic and are associated with sensitivity to the irreversible EGFR/ERBB2 inhibitor HKI-272.

Mutations in the ERBB2 gene were recently found in approximately 2% of primary non-small cell lung cancer (NSCLC) specimens; however, little is known about the functional consequences and the relevance to responsiveness to targeted drugs for most of these mutations. Here, we show that the major lung cancer-derived ERBB2 mutants, including the most frequent mutation, A775insYVMA, lead to oncogenic transformation in a cellular assay. Murine cells transformed with these mutants were relatively resistant to the reversible epidermal growth factor receptor (EGFR) inhibitor erlotinib, resembling the resistant phenotype found in cells carrying the homologous mutations in exon 20 of EGFR. However, the same cells were highly sensitive to the irreversible dual-specificity EGFR/ERBB2 kinase inhibitor HKI-272, as were those overexpressing wild-type ERBB2. Finally, the NSCLC cell line, Calu-3, overexpressing wild-type ERBB2 owing to a high-level amplification of the ERBB2 gene were highly sensitive to HKI-272. These results provide a rationale for treatment of patients with ERBB2-mutant or ERBB2-amplified lung tumors with HKI-272.