Kinjal Shah

Characterizing the cancer genome in lung adenocarcinoma

Somatic alterations in cellular DNA underlie almost all human cancers. The prospect of targeted therapies and the development of high-resolution, genome-wide approaches are now spurring systematic efforts to characterize cancer genomes. Here we report a large-scale project to characterize copy-number alterations in primary lung adenocarcinomas. By analysis of a large collection of tumours (n = 371) using dense single nucleotide polymorphism arrays, we identify a total of 57 significantly recurrent events. We find that 26 of 39 autosomal chromosome arms show consistent large-scale copy-number gain or loss, of which only a handful have been linked to a specific gene. We also identify 31 recurrent focal events, including 24 amplifications and 7 homozygous deletions. Only six of these focal events are currently associated with known mutations in lung carcinomas. The most common event, amplification of chromosome 14q13.3, is found in ∼12% of samples. On the basis of genomic and functional analyses, we identify NKX2-1 (NK2 homeobox 1, also called TITF1), which lies in the minimal 14q13.3 amplification interval and encodes a lineage-specific transcription factor, as a novel candidate proto-oncogene involved in a significant fraction of lung adenocarcinomas. More generally, our results indicate that many of the genes that are involved in lung adenocarcinoma remain to be discovered.

Assessing the significance of chromosomal aberrations in cancer: Methodology and application to glioma

Comprehensive knowledge of the genomic alterations that underlie cancer is a critical foundation for diagnostics, prognostics, and targeted therapeutics. Systematic efforts to analyze cancer genomes are underway, but the analysis is hampered by the lack of a statistical framework to distinguish meaningful events from random background aberrations. Here we describe a systematic method, called Genomic Identification of Significant Targets in Cancer (GISTIC), designed for analyzing chromosomal aberrations in cancer. We use it to study chromosomal aberrations in 141 gliomas and compare the results with two prior studies. Traditional methods highlight hundreds of altered regions with little concordance between studies. The new approach reveals a highly concordant picture involving ≈35 significant events, including 16–18 broad events near chromosome-arm size and 16–21 focal events. Approximately half of these events correspond to known cancer-related genes, only some of which have been previously tied to glioma. We also show that superimposed broad and focal events may have different biological consequences. Specifically, gliomas with broad amplification of chromosome 7 have properties different from those with overlapping focalEGFR amplification: the broad events act in part through effects on MET and its ligand HGF and correlate with MET dependence in vitro. Our results support the feasibility and utility of systematic characterization of the cancer genome.

Subtype-specific genomic alterations define new targets for soft tissue sarcoma therapy

Soft-tissue sarcomas, which result in approximately 10,700 diagnoses and 3,800 deaths per year in the United States, show remarkable histologic diversity, with more than 50 recognized subtypes. However, knowledge of their genomic alterations is limited. We describe an integrative analysis of DNA sequence, copy number and mRNA expression in 207 samples encompassing seven major subtypes. Frequently mutated genes included TP53 (17% of pleomorphic liposarcomas), NF1 (10.5% of myxofibrosarcomas and 8% of pleomorphic liposarcomas) and PIK3CA (18% of myxoid/round-cell liposarcomas, or MRCs). PIK3CA mutations in MRCs were associated with Akt activation and poor clinical outcomes. In myxofibrosarcomas and pleomorphic liposarcomas, we found both point mutations and genomic deletions affecting the tumor suppressor NF1. Finally, we found that short hairpin RNA (shRNA)-based knockdown of several genes amplified in dedifferentiated liposarcoma, including CDK4 and YEATS4, decreased cell proliferation. Our study yields a detailed map of molecular alterations across diverse sarcoma subtypes and suggests potential subtype-specific targets for therapy.

Sensitive mutation detection in heterogeneous cancer specimens by massively parallel picoliter reactor sequencing

The sensitivity of conventional DNA sequencing in tumor biopsies is limited by stromal contamination and by genetic heterogeneity within the cancer. Here, we show that microreactor-based pyrosequencing can detect rare cancer-associated sequence variations by independent and parallel sampling of multiple representatives of a given DNA fragment. This technology can thereby facilitate accurate molecular diagnosis of heterogeneous cancer specimens and enable patient selection for targeted cancer therapies. NOTE: In the version of this article initially published, it should have been acknowledged that Jan F. Simons, in addition to Roman K. Thomas and Elizabeth Nickerson, contributed equally to this work. The error has been corrected in the HTML and PDF versions of the article.

Predicting drug susceptibility of non–small cell lung cancers based on genetic lesions

Somatic genetic alterations in cancers have been linked with response to targeted therapeutics by creation of specific dependency on activated oncogenic signaling pathways. However, no tools currently exist to systematically connect such genetic lesions to therapeutic vulnerability. We have therefore developed a genomics approach to identify lesions associated with therapeutically relevant oncogene dependency. Using integrated genomic profiling, we have demonstrated that the genomes of a large panel of human non-small cell lung cancer (NSCLC) cell lines are highly representative of those of primary NSCLC tumors. Using cell-based compound screening coupled with diverse computational approaches to integrate orthogonal genomic and biochemical data sets, we identified molecular and genomic predictors of therapeutic response to clinically relevant compounds. Using this approach, we showed that v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations confer enhanced Hsp90 dependency and validated this finding in mice with KRAS-driven lung adenocarcinoma, as these mice exhibited dramatic tumor regression when treated with an Hsp90 inhibitor. In addition, we found that cells with copy number enhancement of v-abl Abelson murine leukemia viral oncogene homolog 2 (ABL2) and ephrin receptor kinase and v-src sarcoma (Schmidt-Ruppin A-2) viral oncogene homolog (avian) (SRC) kinase family genes were exquisitely sensitive to treatment with the SRC/ABL inhibitor dasatinib, both in vitro and when it xenografted into mice. Thus, genomically annotated cell-line collections may help translate cancer genomics information into clinical practice by defining critical pathway dependencies amenable to therapeutic inhibition.

Non-small-cell lung cancer and Ba/F3 transformed cells harboring the ERBB2 G776insV_G/C mutation are sensitive to the dual-specific epidermal growth factor receptor and ERBB2 inhibitor HKI-272.

Mutation-specific cancer therapy has shown promising clinical efficacy. In non-small-cell lung cancer (NSCLC), the presence of mutations in the epidermal growth factor receptor (EGFR) tyrosine kinase correlates with clinical response to small-molecule tyrosine kinase inhibitors. Here, we show that cells harboring the G776insV_G/C mutation in the related ERBB2 tyrosine kinase (also known as HER2 or Neu), present in a small percentage of NSCLCs, are sensitive to HKI-272, an irreversible dual-specific kinase inhibitor targeting both EGFR and ERBB2. In the ERBB2-mutant NCI-H1781 cell line, HKI-272 treatment inhibited proliferation by induction of G(1) arrest and apoptotic cell death. Furthermore, HKI-272 abrogated autophosphorylation of both ERBB2 and EGFR. Finally, Ba/F3 murine pro-B cells, engineered to express mutant ERBB2, became independent of interleukin-3 and sensitive to HKI-272. Thus, the subset of NSCLC patients with tumors carrying the ERBB2 G776insV_G/C mutation may benefit from treatment with HKI-272.

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.