Transcriptomic analysis of hepatocellular carcinoma reveals molecular features of disease progression and tumor immune biology.

Hepatocellular carcinoma (HCC) develops in the context of chronic inflammatory liver disease and has an extremely poor prognosis. An immunosuppressive tumor microenvironment may contribute to therapeutic failure in metastatic HCC. Here, we identified unique molecular signatures pertaining to HCC disease progression and tumor immunity by analyzing genome-wide RNA-Seq data derived from HCC patient tumors and non-tumor cirrhotic tissues.

The molecular landscape of high-risk early breast cancer: comprehensive biomarker analysis of a phase III adjuvant population.

Breast cancer is a heterogeneous disease and patients are managed clinically based on ER, PR, HER2 expression, and key risk factors. We sought to characterize the molecular landscape of high-risk breast cancer patients enrolled onto an adjuvant chemotherapy study to understand how disease subsets and tumor immune status impact survival. DNA and RNA were extracted from 861 breast cancer samples from patients enrolled onto the United States Oncology trial 01062.

Development of a robust RNA-based classifier to accurately determine ER, PR, and HER2 status in breast cancer clinical samples.

Breast cancers are categorized into three subtypes based on protein expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2/ERBB2). Patients enroll onto experimental clinical trials based on ER, PR, and HER2 status and, as receptor status is prognostic and defines treatment regimens, central receptor confirmation is critical for interpreting results from these trials. Patients enrolling onto experimental clinical trials in the metastatic setting often have limited available archival tissue that might better be used for comprehensive molecular profiling rather than slide-intensive reconfirmation of receptor status.

Considerations for the successful co-development of targeted cancer therapies and companion diagnostics.

As diagnostic tests become increasingly important for optimizing the use of drugs to treat cancers, the co-development of a targeted therapy and its companion diagnostic test is becoming more prevalent and necessary. In July 2011, the US Food and Drug Administration released a draft guidance that gave the agency's formal definition of companion diagnostics and introduced a drug-diagnostic co-development process for gaining regulatory approval. Here, we identify areas of drug-diagnostic co-development that were either not covered by the guidance or that would benefit from increased granularity, including how to determine when clinical studies should be limited to biomarker-positive patients, defining the diagnostically selected patient population in which to use a companion diagnostic, and defining and clinically validating a biomarker signature for assays that use more than one biomarker.

An industry statistician's perspective on PHC drug development.

In the past decade, the cost of drug development has increased significantly. The estimates vary widely but frequently quoted numbers are staggering-it takes 10-15 years and billions of dollars to bring a drug to patients. To a large extent this is due to many long, expensive and ultimately unsuccessful drug trials.

Phosphoinositide 3-kinase (PI3K) pathway alterations are associated with histologic subtypes and are predictive of sensitivity to PI3K inhibitors in lung cancer preclinical models.

Purpose: Class 1 phosphatidylinositol 3-kinase (PI3K) plays a major role in cell proliferation and survival in a wide variety of human cancers. Here, we investigated biomarker strategies for PI3K pathway inhibitors in non-small-cell lung cancer (NSCLC).

Experimental Design: Molecular profiling for candidate PI3K predictive biomarkers was conducted on a collection of NSCLC tumor samples.

Widespread potential for growth-factor-driven resistance to anticancer kinase inhibitors.

Mutationally activated kinases define a clinically validated class of targets for cancer drug therapy. However, the efficacy of kinase inhibitors in patients whose tumours harbour such alleles is invariably limited by innate or acquired drug resistance. The identification of resistance mechanisms has revealed a recurrent theme—the engagement of survival signals redundant to those transduced by the targeted kinase.

Analysis of tumor burden versus progression-free survival for Phase II decision making.

Purpose: There have been recent recommendations to use percentage change in tumor burden (dTB) as a primary endpoint in randomized Phase II trials. We assessed whether dTB is better for the decision to start a Phase III trial than is progression-free survival (PFS).

Methods: We repeatedly sampled patients from six large randomized trials to obtain simulated Phase II trials.

Statistical techniques to construct assays for identifying likely responders to a treatment under evaluation from cell line genomic data.

Background: Developing the right drugs for the right patients has become a mantra of drug development. In practice, it is very difficult to identify subsets of patients who will respond to a drug under evaluation. Most of the time, no single diagnostic will be available, and more complex decision rules will be required to define a sensitive population, using, for instance, mRNA expression, protein expression or DNA copy number.

Reversing HOXA9 oncogene activation by PI3K inhibition: epigenetic mechanism and prognostic significance in human glioblastoma.

HOXA genes encode critical transcriptional regulators of embryonic development that have been implicated in cancer. In this study, we documented functional relevance and mechanism of activation of HOXA9 in glioblastoma (GBM), the most common malignant brain tumor. Expression of HOXA genes was investigated using reverse transcription-PCR in primary gliomas and glioblastoma cell lines and was validated in two sets of expression array data.

Characterization of a naturally occurring breast cancer subset enriched in epithelial-to-mesenchymal transition and stem cell characteristics.

Metaplastic breast cancers (MBC) are aggressive, chemoresistant tumors characterized by lineage plasticity. To advance understanding of their pathogenesis and relatedness to other breast cancer subtypes, 28 MBCs were compared with common breast cancers using comparative genomic hybridization, transcriptional profiling, and reverse-phase protein arrays and by sequencing for common breast cancer mutations. MBCs showed unique DNA copy number aberrations compared with common breast cancers.

Co-amplified genes at 8p12 and 11q13 in breast tumors cooperate with two major pathways in oncogenesis.

Co-amplification at chromosomes 8p11-8p12 and 11q12-11q14 occurs often in breast tumors, suggesting possible cooperation between genes in these regions in oncogenesis. We used high-resolution array comparative genomic hybridization (array CGH) to map the minimal amplified regions. The 8p and 11q amplicons are complex and consist of at least four amplicon cores at each site.

Improving melanoma classification by integrating genetic and morphologic features.

Background: In melanoma, morphology-based classification systems have not been able to provide relevant information for selecting treatments for patients whose tumors have metastasized. The recent identification of causative genetic alterations has revealed mutations in signaling pathways that offer targets for therapy. Identifying morphologic surrogates that can identify patients whose tumors express such alterations (or functionally equivalent alterations) would be clinically useful for therapy stratification and for retrospective analysis of clinical trial data.

High-resolution genomic and expression analyses of copy number alterations in breast tumors.

Analysis of recurrent DNA amplification can lead to the identification of cancer driver genes, but this process is often hampered by the low resolution of existing copy number analysis platforms. Fifty-one breast tumors were profiled for copy number alterations (CNAs) with the high-resolution Affymetrix 500K SNP array. These tumors were also expression-profiled and surveyed for mutations in selected genes commonly mutated in breast cancer (TP53, CDKN2A, ERBB2, KRAS, PIK3CA, PTEN).

Integration of genomic analysis and in vivo transfection to identify sprouty 2 as a candidate tumor suppressor in liver cancer.

Unlabelled: Hepatocellular carcinoma (HCC) is 1 of the leading causes of cancer-related deaths worldwide, yet the molecular genetics underlying this malignancy are still poorly understood. In our study, we applied statistical methods to correlate human HCC gene expression data obtained from complementary DNA (cDNA) microarrays and corresponding DNA copy number variation data obtained from array-based comparative genomic hybridization. We have thus identified 76 genes that are up-regulated and show frequent DNA copy number gain, and 37 genes that are down-regulated and show frequent DNA copy loss in human HCC samples.

Protein biomarker identification in the CSF of patients with CNS lymphoma.

Purpose: Elucidation of the CSF proteome may yield insights into the pathogenesis of CNS disease. We tested the hypothesis that individual CSF proteins distinguish CNS lymphoma from benign focal brain lesions.

Methods: We used a liquid chromatography/mass spectrometry-based method to differentially quantify and identify several hundred CSF proteins in CNS lymphoma and control patients.

Evaluation of whole genome amplification protocols for array and oligonucleotide CGH.

Genome-based technologies such as genomic arrays and next generation sequencing are poised to make significant contributions to clinical oncology. However, translation of these technologies to the clinic will require that they produce high-quality reproducible data from small archived tumor specimens and biopsies. Herein, we report on a systematic and comprehensive microarray analysis comparing multiple whole genome amplification methods using a variety of difficult clinical specimens, including formalin-fixed and paraffin-embedded tissues.

Acquired genomic aberrations associated with methotrexate resistance vary with background genomic instability.

Tumors vary widely in chromosomal level genome instability. To gain a better understanding of the underlying defects which foster specific types of aberrations, we investigated the response of cells of related genetic backgrounds to challenge with methotrexate. We studied mismatch repair deficient HCT116 cells, two derivatives also deficient in XRCC5 (HCT116 Ku86+/-) or BLM (HCT116 BLM-/-), and mismatch repair competent HCT116+chr3 cells.

Amplification of PVT1 contributes to the pathophysiology of ovarian and breast cancer.

Purpose: This study was designed to elucidate the role of amplification at 8q24 in the pathophysiology of ovarian and breast cancer because increased copy number at this locus is one of the most frequent genomic abnormalities in these cancers.

Experimental Design: To accomplish this, we assessed the association of amplification at 8q24 with outcome in ovarian cancers using fluorescence in situ hybridization to tissue microarrays and measured responses of ovarian and breast cancer cell lines to specific small interfering RNAs against the oncogene MYC and a putative noncoding RNA, PVT1, both of which map to 8q24.

Results: Amplification of 8q24 was associated with significantly reduced survival duration.

Aging impacts transcriptomes but not genomes of hormone-dependent breast cancers.

Introduction: Age is one of the most important risk factors for human malignancies, including breast cancer; in addition, age at diagnosis has been shown to be an independent indicator of breast cancer prognosis. Except for inherited forms of breast cancer, however, there is little genetic or epigenetic understanding of the biological basis linking aging with sporadic breast cancer incidence and its clinical behavior.

Methods: DNA and RNA samples from matched estrogen receptor (ER)-positive sporadic breast cancers diagnosed in either younger (age or= 70 years) Caucasian women were analyzed by array comparative genomic hybridization and by expression microarrays.

Novel risk stratification of patients with neuroblastoma by genomic signature, which is independent of molecular signature.

Human neuroblastoma remains enigmatic because it often shows spontaneous regression and aggressive growth. The prognosis of advanced stage of sporadic neuroblastomas is still poor. Here, we investigated whether genomic and molecular signatures could categorize new therapeutic risk groups in primary neuroblastomas.

FBXW7 and DNA copy number instability.

SKP1-cullin-F-box protein (SCF) type ubiquitin ligases degrade proteins controlling the G1/S transition. Deficiency for FBXW7 (also known as hCDC4), which encodes the F-box protein of the SCF type ubiquitin ligase is associated with genomic instability. Here, we investigated the association of FBXW7 deficiency with chromosomal instability in breast cancer.

Genome position and gene amplification.

Background: Amplifications, regions of focal high-level copy number change, lead to overexpression of oncogenes or drug resistance genes in tumors. Their presence is often associated with poor prognosis; however, the use of amplification as a mechanism for overexpression of a particular gene in tumors varies. To investigate the influence of genome position on propensity to amplify, we integrated a mutant form of the gene encoding dihydrofolate reductase into different positions in the human genome, challenged cells with methotrexate and then studied the genomic alterations arising in drug resistant cells.

Phase I study of intraventricular administration of rituximab in patients with recurrent CNS and intraocular lymphoma.

Purpose: We previously determined that intravenous administration of rituximab results in limited penetration of this agent into the leptomeningeal space. Systemic rituximab does not reduce the risk of CNS relapse or dissemination in patients with large cell lymphoma. We therefore conducted a phase I dose-escalation study of intrathecal rituximab monotherapy in patients with recurrent CNS non-Hodgkin's lymphoma (NHL).

Lack of germ-line promoter methylation in BRCA1-negative families with familial breast cancer.

Hereditary breast cancer accounts for about 10% of breast cancer in the United States, but high-penetrance, germ-line mutations in BRCA1 and BRCA2 are responsible for less than half of these high-risk families. Epigenetic modification of DNA by promoter methylation can result in a potentially heritable epimutation that silences the gene. Using a highly sensitive technique, we assayed the BRCA1 gene for promoter methylation among 41 BRCA1- and BRCA2-negative women whose personal and family histories indicated a high risk of BRCA mutations (median prior likelihood = 60%) using the BRCAPro model.