Venkatraman E. Seshan

Gene expression-based survival prediction in lung adenocarcinoma: A multi-site, blinded validation study

Although prognostic gene expression signatures for survival in early-stage lung cancer have been proposed, for clinical application, it is critical to establish their performance across different subject populations and in different laboratories. Here we report a large, training–testing, multi-site, blinded validation study to characterize the performance of several prognostic models based on gene expression for 442 lung adenocarcinomas. The hypotheses proposed examined whether microarray measurements of gene expression either alone or combined with basic clinical covariates (stage, age, sex) could be used to predict overall survival in lung cancer subjects. Several models examined produced risk scores that substantially correlated with actual subject outcome. Most methods performed better with clinical data, supporting the combined use of clinical and molecular information when building prognostic models for early-stage lung cancer. This study also provides the largest available set of microarray data with extensive pathological and clinical annotation for lung adenocarcinomas.

A CXCL1 Paracrine Network Links Cancer Chemoresistance and Metastasis

Metastasis and chemoresistance in cancer are linked phenomena, but the molecular basis for this link is unknown. We uncovered a network of paracrine signals between carcinoma, myeloid, and endothelial cells that drives both processes in breast cancer. Cancer cells that overexpress CXCL1 and 2 by transcriptional hyperactivation or 4q21 amplification are primed for survival in metastatic sites. CXCL1/2 attract CD11b(+)Gr1(+) myeloid cells into the tumor, which produce chemokines including S100A8/9 that enhance cancer cell survival. Although chemotherapeutic agents kill cancer cells, these treatments trigger a parallel stromal reaction leading to TNF-α production by endothelial and other stromal cells. TNF-α via NF-kB heightens the CXCL1/2 expression in cancer cells, thus amplifying the CXCL1/2-S100A8/9 loop and causing chemoresistance. CXCR2 blockers break this cycle, augmenting the efficacy of chemotherapy against breast tumors and particularly against metastasis. This network of endothelial-carcinoma-myeloid signaling interactions provides a mechanism linking chemoresistance and metastasis, with opportunities for intervention.

Extrapleural pneumonectomy versus pleurectomy/decortication in the surgical management of malignant pleural mesothelioma: results in 663 patients.

OBJECTIVE The optimal procedure for resection of malignant pleural mesothelioma is controversial, partly because previous analyses include small numbers of patients. We performed a multi-institutional study to increase statistical power to detect significant differences in outcome between extrapleural pneumonectomy and pleurectomy/decortication. METHODS Patients with malignant pleural mesothelioma who underwent extrapleural pneumonectomy or pleurectomy/decortication at 3 institutions were identified. Survival and prognostic factors were analyzed by the Kaplan-Meier method, log-rank test, and Cox proportional hazards analysis. RESULTS From 1990 to 2006, 663 consecutive patients (538 men and 125 women) underwent resection. The median age was 63 years (range, 26-93 years). The operative mortality was 7% for extrapleural pneumonectomy (n = 27/385) and 4% for pleurectomy/decortication (n = 13/278). Significant survival differences were seen for American Joint Committee on Cancer stages 1 to 4 (P < .001), epithelioid versus non-epithelioid histology (P < .001), extrapleural pneumonectomy versus pleurectomy/decortication (P < .001), multimodality therapy versus surgery alone (P < .001), and gender (P < .001). Multivariate analysis demonstrated a hazard rate of 1.4 for extrapleural pneumonectomy (P < .001) controlling for stage, histology, gender, and multimodality therapy. CONCLUSION Patients who underwent pleurectomy/decortication had a better survival than those who underwent extrapleural pneumonectomy; however, the reasons are multifactorial and subject to selection bias. At present, the choice of resection should be tailored to the extent of disease, patient comorbidities, and type of multimodality therapy planned.

Lung Adenocarcinoma: Modification of the 2004 Who Mixed Subtype to Include the Major Histologic Subtype Suggests Correlations Between Papillary and Micropapillary Adenocarcinoma Subtypes, egfr Mutations and Gene Expression Analysis

The histologic heterogeneity of lung adenocarcinoma creates a variety of complex challenges to pathologists in analyzing the various subtypes. Current classification schemas do not take into account the major subtype. We analyzed 100 cases for clinical, pathologic, and molecular features using a modification of the 2004 World Health Organization (WHO) classification to record the major component in the mixed subtype tumors. The tumors were analyzed for KRAS mutation and epidermal growth factor receptor (EGFR) by mutation, chromogenic in situ hybridization, and immunohistochemistry for EGFR and phosphorylated EGFR. Gene expression analysis was performed using HG-U133A Affymetrix oligonucleotide microarrays revealing 3 gene clusters. The most common major histologic subtype was papillary (37%) followed by acinar (30%), solid (25%) and bronchioloalveolar (7%) carcinoma (BAC), although no pure BACs were seen. Sixteen tumors harbored EGFR mutations and 14 KRAS mutations. Papillary adenocarcinoma strongly correlated with EGFR mutation (P<0.001) and gene profile Cluster 1 (P=0.006) with weaker correlations with low grade (P=0.038) and favorable behavior in Stage 1 patients (P=0.047). Micropapillary subtype correlated strongly with EGFR mutation (P<0.001) and weakly with Cluster 1 (P=0.030). Solid adenocarcinoma strongly correlated with gene profile Cluster 3 (P=0.001) and worse survival (P=0.001). BAC as a major component strongly correlated with gene Cluster 2 (P=0.001). Cluster 1 correlated strongly with less smoking (P<0.001), papillary histology (P<0.001), and EGFR mutations (P<0.001). Cluster 3 strongly correlated with heavier smoking (P<0.001), larger tumor size (P<0.001), solid subtype (P<0.001), and poor grade (P=0.004); weak correlations were found with KRAS mutation (P=0.025). No correlation was found with BAC and EGFR by mutation, chromogenic in situ hybridization or immunohistochemistry. Higher stage (P<0.001), grade (P<0.001), and solid subtype (P=0.001) correlated with shorter survival. Our data suggest that EGFR mutations are associated with papillary adenocarcinoma and gene profile Cluster 1. We discovered this only because we used a comprehensive approach examining in detail all histologic subtypes and we modified the 2004 WHO mixed subtype to include the major histologic subtype. As we do not know the major genetic pathways of 30% to 70% of lung adenocarcinomas, the comprehensive histologic subtyping we propose gives advantage for recognition of unanticipated histologic-genetic correlations that might not be detected using classification systems that focus primarily on specific aspects of adenocarcinomas such as BAC or EGFR mutations. Such an approach should be considered in future studies for validation in other datasets.

Detectable clonal mosaicism from birth to old age and its relationship to cancer

We detected clonal mosaicism for large chromosomal anomalies (duplications, deletions and uniparental disomy) using SNP microarray data from over 50,000 subjects recruited for genome-wide association studies. This detection method requires a relatively high frequency of cells with the same abnormal karyotype (>5–10%; presumably of clonal origin) in the presence of normal cells. The frequency of detectable clonal mosaicism in peripheral blood is low (<0.5%) from birth until 50 years of age, after which it rapidly rises to 2–3% in the elderly. Many of the mosaic anomalies are characteristic of those found in hematological cancers and identify common deleted regions with genes previously associated with these cancers. Although only 3% of subjects with detectable clonal mosaicism had any record of hematological cancer before DNA sampling, those without a previous diagnosis have an estimated tenfold higher risk of a subsequent hematological cancer (95% confidence interval = 6–18).

Prognostic and therapeutic implications of EGFR and KRAS mutations in resected lung adenocarcinoma

Background: Somatic mutations in EGFR (exons 19 and 21) and KRAS (exon 2) are found in lung adenocarcinomas and have potential prognostic value in patients with advanced disease. These mutations also have therapeutic significance, as they predict for sensitivity and resistance, respectively, to EGFR tyrosine kinase inhibitor therapy. Whether EGFR and KRAS mutations also have an impact on survival in patients who undergo lung resection for curative intent in the absence of targeted therapy has not been established. Methods: We analyzed the clinical characteristics and outcomes data for 296 patients who underwent resection at our institution for stage I–III lung adenocarcinoma. Tumors were assessed for both EGFR and KRAS mutations by established methods. Results: EGFR and KRAS mutations were found in tumors from 40 (14%) and 50 (17%) patients, respectively. Patients with EGFR mutant tumors were more likely to be never smokers (48%), present with stage I disease (88%), and had a 90% (95% confidence interval [CI] 70–97%) 3-year overall survival, whereas patients with KRAS mutant tumors were more likely to be former/current smokers (92%), present with locally advanced disease (40%), and had a 66% (95% CI 48–79%) 3-year overall survival. Conclusions: EGFR and KRAS mutations define distinct molecular subsets of resected lung adenocarcinoma. Because EGFR and KRAS mutations also predict whether tumors are sensitive or resistant, respectively, to EGFR tyrosine kinase inhibitors, they can readily be used in clinical trials to help guide the administration of specific types of adjuvant therapy.

Breast Cancer Methylomes Establish an Epigenomic Foundation for Metastasis

Breast cancer methylomes contribute to metastatic potential, modulate the metastasis transcriptome, and predict disease outcome. Meditating on Breast Cancer People of diverse faiths and backgrounds have gained new mindsets when contemplating “Om” (or Aum)—a meditation symbol that represents the universe in its entirety. The concept of examining existence from a global perspective has begun to take hold in cancer research as well. Indeed, researchers have created their own “omes”: the genome, the transcriptome, the proteome. Here, Fang et al. examine the methylome of breast cancer and find a signature that may predict metastasis. The authors used genome-wide analysis to examine methylome signatures in breast cancers with various metastatic behaviors and found a signature that was associated with low metastatic risk and improved rates of survival. This breast CpG island methylator phenotype (B-CIMP) tracked with reduced metastasis independently of other breast cancer markers [such as estrogen receptor/progesterone receptor (ER/PR) and human epidermal growth factor receptor 2 (HER2) status] and was shared by multiple human malignancies, including glioma and colon cancer. However, altered methylation status may not just be a marker of metastasis: Methylation of B-CIMP signature genes correlated with transcriptional diversity among breast cancers with different prognoses. Thus, the B-CIMP phenotype may thus play a mechanistic role in metastatic risk, and future meditation on the methylome may improve breast cancer prognosis and therapy. Cancer-specific alterations in DNA methylation are hallmarks of human malignancies; however, the nature of the breast cancer epigenome and its effects on metastatic behavior remain obscure. To address this issue, we used genome-wide analysis to characterize the methylomes of breast cancers with diverse metastatic behavior. Groups of breast tumors were characterized by the presence or absence of coordinate hypermethylation at a large number of genes, demonstrating a breast CpG island methylator phenotype (B-CIMP). The B-CIMP provided a distinct epigenomic profile and was a strong determinant of metastatic potential. Specifically, the presence of the B-CIMP in tumors was associated with low metastatic risk and survival, and the absence of the B-CIMP was associated with high metastatic risk and death. B-CIMP loci were highly enriched for genes that make up the metastasis transcriptome. Methylation at B-CIMP genes accounted for much of the transcriptomal diversity between breast cancers of varying prognosis, indicating a fundamental epigenomic contribution to metastasis. Comparison of the loci affected by the B-CIMP with those affected by the hypermethylator phenotype in glioma and colon cancer revealed that the CIMP signature was shared by multiple human malignancies. Our data provide a unifying epigenomic framework linking breast cancers with varying outcome and transcriptomic changes underlying metastasis. These findings significantly enhance our understanding of breast cancer oncogenesis and aid the development of new prognostic biomarkers for this common malignancy.

An integrated genomic analysis of lung cancer reveals loss of DUSP4 in EGFR-mutant tumors

To address the biological heterogeneity of lung cancer, we studied 199 lung adenocarcinomas by integrating genome-wide data on copy number alterations and gene expression with full annotation for major known somatic mutations in this cancer. This showed non-random patterns of copy number alterations significantly linked to EGFR and KRAS mutation status and to distinct clinical outcomes, and led to the discovery of a striking association of EGFR mutations with underexpression of DUSP4, a gene within a broad region of frequent single-copy loss on 8p. DUSP4 is involved in negative feedback control of EGFR signaling, and we provide functional validation for its role as a growth suppressor in EGFR-mutant lung adenocarcinoma. DUSP4 loss also associates with p16/CDKN2A deletion and defines a distinct clinical subset of lung cancer patients. Another novel observation is that of a reciprocal relationship between EGFR and LKB1 mutations. These results highlight the power of integrated genomics to identify candidate driver genes within recurrent broad regions of copy number alteration and to delineate distinct oncogenetic pathways in genetically complex common epithelial cancers.

Phase II-I-II Study of Two Different Doses and Schedules of Pralatrexate, a High-Affinity Substrate for the Reduced Folate Carrier, in Patients With Relapsed or Refractory Lymphoma Reveals Marked Activity in T-Cell Malignancies

PURPOSE To determine the maximum-tolerated dose (MTD) and efficacy of pralatrexate in patients with lymphoma. PATIENTS AND METHODS Pralatrexate, initially given at a dose of 135 mg/m(2) on an every-other-week basis, was associated with stomatitis. A redesigned, weekly phase I/II study established an MTD of 30 mg/m(2) weekly for six weeks every 7 weeks. Patients were required to have relapsed/refractory disease, an absolute neutrophil greater than 1,000/microL, and a platelet count greater than 50,000/microL for the first dose of any cycle. RESULTS The every-other-week, phase II experience was associated with an increased risk of stomatitis and hematologic toxicity. On a weekly schedule, the MTD was 30 mg/m(2) weekly for 6 weeks every 7 weeks. This schedule modification resulted in a 50% reduction in the major hematologic toxicities and abrogation of the grades 3 to 4 stomatitis. Stomatitis was associated with elevated homocysteine and methylmalonic acid, which were reduced by folate and vitamin B12 supplementation. Of 48 assessable patients, the overall response rate was 31% (26% by intention to treat), including 17% who experienced complete remission (CR). When analyzed by lineage, the overall response rates were 10% and 54% in patients with B- and T-cell lymphomas, respectively. All eight patients who experienced CR had T-cell lymphoma, and four of the six patients with a partial remission were positron emission tomography negative. The duration of responses ranged from 3 to 26 months. CONCLUSION Pralatrexate has significant single-agent activity in patients with relapsed/refractory T-cell lymphoma.

Pattern discovery and cancer gene identification in integrated cancer genomic data

Large-scale integrated cancer genome characterization efforts including the cancer genome atlas and the cancer cell line encyclopedia have created unprecedented opportunities to study cancer biology in the context of knowing the entire catalog of genetic alterations. A clinically important challenge is to discover cancer subtypes and their molecular drivers in a comprehensive genetic context. Curtis et al. [Nature (2012) 486(7403):346–352] has recently shown that integrative clustering of copy number and gene expression in 2,000 breast tumors reveals novel subgroups beyond the classic expression subtypes that show distinct clinical outcomes. To extend the scope of integrative analysis for the inclusion of somatic mutation data by massively parallel sequencing, we propose a framework for joint modeling of discrete and continuous variables that arise from integrated genomic, epigenomic, and transcriptomic profiling. The core idea is motivated by the hypothesis that diverse molecular phenotypes can be predicted by a set of orthogonal latent variables that represent distinct molecular drivers, and thus can reveal tumor subgroups of biological and clinical importance. Using the cancer cell line encyclopedia dataset, we demonstrate our method can accurately group cell lines by their cell-of-origin for several cancer types, and precisely pinpoint their known and potential cancer driver genes. Our integrative analysis also demonstrates the power for revealing subgroups that are not lineage-dependent, but consist of different cancer types driven by a common genetic alteration. Application of the cancer genome atlas colorectal cancer data reveals distinct integrated tumor subtypes, suggesting different genetic pathways in colon cancer progression.