Genome-wide association studies have identified thousands of loci for common diseases, but, for the majority of these, the mechanisms underlying disease susceptibility remain unknown. Most associated variants are not correlated with protein-coding changes, suggesting that polymorphisms in regulatory regions probably contribute to many disease phenotypes. Here we describe the Genotype-Tissue Expression (GTEx) project, which will establish a resource database and associated tissue bank for the scientific community to study the relationship between genetic variation and gene expression in human tissues

Sentinel lymph node (SLN) metastasis is the first step in the spreading of cancer in many malignancies. Tumor-reactive lymphadenopathy in SLNs has been observed for decades, but alterations of the lymphatic channels and vasculature in these nodes before the arrival of metastatic tumor cells remain unexplored. Using animal models, we show here that, before the establishment of metastasis in the SLN, there are reorganizations of the lymphatic channels and the vasculature. The node becomes a functional blood vessel-enriched and lymph vessel/sinus-enriched organ before metastasis. The enlargement of the lymph sinuses is correlated with the primary tumor weight. The newly emerged functional blood vessels develop from high endothelial venules (HEV), in which the proliferation rate of the endothelial cells is also significantly increased. Similar alterations of the HEVs are also characterized in the axillary lymph nodes from human breast cancer patients without the evidence of metastasis. These findings support the hypothesis that modification of the microenvironment for a secondary tumor (i.e., vasculature reorganization in the SLN) can be initiated by a primary tumor before and independent of the physical presence of metastatic cancer cells. (Cancer Res 2006; 66(21): 10365-76)

BackgroundPrognostic markers and molecular breast cancer subtypes reflect underlying biological tumor behavior and are important for patient management. Compared to Western countries, women in North Africa are less likely to be prognosticated and treated based on well-characterized markers such as the estrogen receptor (ER), progesterone receptor (PR) and Her2. We conducted this study to determine the prevalence of breast cancer molecular subtypes in the North African country of Egypt as a measure of underlying biological characteristics driving tumor manifestations.MethodsTo determine molecular subtypes we characterized over 200 tumor specimens obtained from Egypt by performing ER, PR, Her2, CK5/6, EGFR and Ki67 immunohistochemistry.ResultsOur study demonstrated that the Luminal A subtype, associated with favorable prognosis, was found in nearly 45% of cases examined. However, the basal-like subtype, associated with poor prognosis, was found in 11% of cases. These findings are in sharp contrast to other parts of Africa in which the basal-like subtype is over-represented.ConclusionsEgyptians appear to have favorable underlying biology, albeit having advanced disease at diagnosis. These data suggest that Egyptians would largely profit from early detection of their disease. Intervention at the public health level, including education on the benefits of early detection is necessary and would likely have tremendous impact on breast cancer outcome in Egypt.

Animal models greatly facilitate understanding of cancer and importantly, serve pre-clinically for evaluating potential anti-cancer therapies. We developed an invasive orthotopic human glioblastoma multiforme (GBM) mouse model that enables real-time tumor ultrasound imaging and pre-clinical evaluation of antineoplastic drugs such as 17-(allylamino)-17-demethoxy geldanamycin (17AAG). Clinically, GBM metastasis rarely happen, but unexpectedly most human GBM tumor cell lines intrinsically possess metastatic potential. We used an experimental lung metastasis assay (ELM) to enrich for metastatic cells and three of four commonly used GBM lines were highly metastatic after repeated ELM selection (M2). These GBM-M2 lines grew more aggressively orthotopically and all showed dramatic multifold increases in IL6, IL8, MCP-1 and GM-CSF expression, cytokines and factors that are associated with GBM and poor prognosis. DBM2 cells, which were derived from the DBTRG-05MG cell line were used to test the efficacy of 17AAG for treatment of intracranial tumors. The DMB2 orthotopic xenografts form highly invasive tumors with areas of central necrosis, vascular hyperplasia and intracranial dissemination. In addition, the orthotopic tumors caused osteolysis and the skull opening correlated to the tumor size, permitting the use of realtime ultrasound imaging to evaluate antitumor drug activity. We show that 17AAG significantly inhibits DBM2 tumor growth with significant drug responses in subcutaneous, lung and orthotopic tumor locations. This model has multiple unique features for investigating the pathobiology of intracranial tumor growth and for monitoring systemic and intracranial responses to antitumor agents.

We hypothesized that signaling through multiple mitogenactivated protein kinase (MAPK) kinase (MKK) pathways is essential for the growth and vascularization of softtissue sarcomas, which are malignant tumors derived from mesenchymal tissues. We tested this using HT-1080, NCI, and Shac fibrosarcoma-derived cell lines and anthrax lethal toxin (LeTx), a bacterial toxin that inactivates MKKs. Western blots confirmed that LeTx treatment reduced the levels of phosphorylated extracellular signal-regulated kinase and p38 MAPK in vitro. Although short treatments with LeTx only modestly affected cell proliferation, sustained treatment markedly reduced cell numbers. LeTx also substantially inhibited the extracellular release of angioproliferative factors including vascular endothelial growth factor, interleukin-8, and basic fibroblast growth factor. Similar results were obtained with cell lines derived from malignant fibrous histiocytomas, leiomyosarcomas, and liposarcomas. In vivo, LeTx decreased MAPK activity and blocked fibrosarcoma growth. Growth inhibition correlated with decreased cellular proliferation and extensive necrosis, and it was accompanied by a decrease in tumor mean vessel density as well as a reduction in serum expression of angioproliferative cytokines. Vital imaging using high-resolution ultrasound enhanced with contrast microbubbles revealed that the effects of LeTx on tumor perfusion were remarkably rapid (<24 h) and resulted in a marked reduction of perfusion within the tumor but not in nontumor tissues. These results are consistent with our initial hypothesis and lead us to propose that MKK inhibition by LeTx is a broadly effective strategy for targeting neovascularization in fibrosarcomas and other similar proliferative lesions. [Mol Cancer Ther 2008;7(3):648 -58]

The inappropriate expression of the c-MET cell surface receptor in many human solid tumors necessitates the development of companion diagnostics to identify those patients who could benefit from c-MET targeted therapies. Tumor tissues are formalin-fixed and paraffin embedded (FFPE) for histopathological evaluation, making the development of an antibody against c-MET that accurately and reproducibly detects the protein in FFPE samples an urgent need. We have developed a monoclonal antibody, designated MET4, from a panel of MET-avid monoclonal antibodies, based on its specific staining pattern in FFPE preparations of normal human prostate tissues. The accuracy of MET4 immunohistochemistry (MET4-IHC) was assessed by comparing MET4-IHC in FFPE cell pellets with immunoblotting analysis. The technical reproducibility of MET4-IHC possessed a percentage coefficient of variability (%CV) of 6.25% in intra-assay and inter-assay testing.Comparison with other commercial c-MET antibody detection reagents demonstrated equal specificity and increased sensitivity for c-MET detection in prostate tissues. In two cohorts of ovarian cancers and gliomas, MET4 reacted with ovarian cancers of all histological subtypes (strong staining in 25%) and with 63% of gliomas. In addition, MET4 bound c-Met on the surfaces of cultured human cancer cells and tumor xenografts. In summary, the MET4 monoclonal antibody accurately and reproducibly measures c-MET expression by IHC in FFPE tissues and can be used for molecular imaging in-vivo. These properties encourage further development of MET4 as a multipurpose

BACKGROUND. The objective of this study was to evaluate the effect of pericyte coverage (PC) of differentiated tumor microvessels on the prognosis of patients with clear cell renal cell carcinoma (CCRCC). METHODS. Samples from 2 cohorts of patients with CCRCC (101 Asian patients and 524 US patients) were prepared using 2 different histologic approaches: routine sectioning versus tissue microarray. Then, the samples were immunohistochemically doubled-stained for a pericyte marker (alpha smooth muscle actin [a-SMA]) and a differentiated vessel marker (cluster of differentiation 34 [CD34]), followed by multispectral image capturing and computerized image analyses to quantify the microvessel density (MVD) and the PC of differentiated vessels. The correlations of PC and the MVD:PC ratio with clinicopathologic characteristics were analyzed. RESULTS. There was an inverse correlation between differentiated MVD and PC. Higher PC correlated with more aggressive clinicopathologic characteristics of CCRCC in both cohorts, including more advanced T-classification, higher pathologic grades, and the occurrence of tumor necrosis. The MVD:PC ratio was an independent favorable prognostic factor for overall and recurrence-free survival in the Asian cohort and for recurrence-free survival in the US cohort. PC also was an independent prognostic factor, with higher PC predicting a poorer outcome. The combination of PC and MVD was better at distinguishing the outcome of patients with CCRCC. PC combined with differentiated MVD or with the MVD:PC ratio provided additional, independent prognostic information to the Leibovich risk model, and that information was used to generate improved risk models. CONCLUSIONS. The authors consistently observed that higher PC was correlated with more aggressive clinicopathologic characteristics. PC was an independent unfavorable prognostic factor. The authors concluded that pericytes should be considered for therapeutic targeting. Cancer 2013;119:313-34.

Purpose: Anthrax Lethal Toxin (LeTx), composed of protective antigen and lethal factor, catalytically cleaves mitogen-activated protein kinase (MAPK) kinases and inhibits the MAPK signaling pathways. The majority of metastatic melanomas possess the V599E BRAF mutation, which constitutively activates MAPK1/2 signaling. LeTx is cytotoxic to BRAF mutant melanoma cell lines in vitro, whereas most normal cells are resistant to this toxin. In this study, we determine the in vivo potency and safety of systemically administered LeTx. Experimental Design: A s.c. xenograft melanoma model in athymic nude mice was treated with different i.p. doses of LeTx. Results: In this study, we show that in vivo systemic LeTx treatment of s.c. xenograft melanoma tumors in athymic nude mice yields partial and complete tumor regressions with minor toxicity to mice. When animal toxicity was observed, we did not find any histologic evidence of tissue damage. Conclusions: LeTx is one of the rare targeted agents to produce complete remissions of human melanomas in an animal model and thus warrants further preclinical development.