Background Vitamin D plays a role in cancer development and acts through the vitamin D receptor (VDR). Although African-Americans have the lowest levels of serum vitamin D, there is a dearth of information on VDR gene polymorphisms and breast cancer among African-Americans and Hispanics. This study examines whether VDR gene polymorphisms are associated with breast cancer in these cohorts. Methods Blood was collected from 232 breast cancer patients (Cases) and 349 non-cancer subjects (Controls). Genotyping for four polymorphic variants of VDR (FokI, BsmI, TaqI and ApaI) was performed using the PCR-RFLP method. Results An increased association of the VDR-Fok1 f allele with breast cancer was observed in African-Americans (OR = 1.9, p = 0.07). Furthermore, the FbTA, FbtA and fbtA haplotypes were associated with breast cancer among African-Americans (p<0.05). Latinas were more likely to have the VDR-ApaI alleles (Aa or aa) (p = 0.008). The VDR-ApaI aa genotype was significantly associated with poorly-differentiated breast tumors (p = 0.04) in combined Cases. Kaplan-Meier survival analysis showed decreased 5-year disease-free-survival (DFS) in breast cancer patients who had the VDR-Fok1 FF genotype (p<0.05). The Cox regression with multivariate analysis revealed the independent predictor value of the VDR-FokI polymorphism for DFS. The other three variants of VDR (BsmI, TaqI and ApaI) were not associated with disease outcome. Conclusions VDR haplotypes are associated with breast cancer in African-Americans, but not in Hispanic/Latinas. The VDR-FokI FF genotype is linked with poor prognosis in African-American women with breast cancer.
Metastasis is the main cause of death for lung cancer patients. The ex vivo 4D acellular lung model has been shown to mimic this metastatic process. However, the main concern is the model's lack of cellular components of the tumor's microenvironment. In this study, we aim to determine if the intact lung microenvironment will still allow lung cancer metastasis to form. We harvested a heart-lung block from a rat and placed it in a bioreactor after cannulating the pulmonary artery, trachea and tying the right main bronchus for 10–15 days without any tumor cells as a control group or with NSCLC (A549, H1299 or H460), SCLC (H69, H446 or SHP77) or breast cancer cell lines (MCF7 or MDAMB231) through the trachea. We performed lobectomy, H&E staining and IHC for human mitochondria to determine the primary tumor's growth and formation of metastatic lesions. In addition, we isolated circulating tumor cells (CTC) from the model seeded with GFP tagged cells. In the control group, no gross tumor nodules were found, H&E staining showed hyperplastic cells and IHC showed no staining for human mitochondria. All of the models seeded with cancer cell lines formed gross primary tumor nodules that had microscopic characteristics of human cancer cells on H&E staining with IHC showing staining for human mitochondria. CTC were isolated for those cells labeled with GFP and they were viable in culture. Finally, all cell lines formed metastatic lesions with cells stained for human mitochondria. The cellular ex vivo 4D model shows that human cancer cells can form a primary tumor, CTC and metastatic lesions in an intact cellular environment. This study suggests that the natural matrix scaffold is the only necessary component to drive metastatic progression and that cellular components play a role in modulating tumor progression.
Supplementary Methods, Figure Legends 1-4 from Global Methylation Pattern of Genes in Androgen-Sensitive and Androgen-Independent Prostate Cancer Cells
The mechanisms that could explain the poor sensitivity to 5-FU in certain colorectal cancer (CRC) cells were investigated and whether or not cotreatment with low doses of selenium would offer a therapeutic benefit was explored.
Abstract Increase in comorbidities such as diabetes and hypertension in cancer patients lead to poor disease free survival and overall survival. It is probable that risk factors associated with these comorbidities and cancer itself could be linked to alterations in gene polymorphisms associated with hormones and/or growth factors. In this study, we have investigated if (1) comorbidities were associated with our African American and Hispanic breast cancer subjects; and (2) if there was any association between Insulin-Like Growth Factor 1 (IGF-1), and IGF Binding Protein 3 (IGFBP3) gene polymorphisms with breast cancer and comorbidities. We studied 514 subjects (175 African Americans and 339 Hispanic/Latinas), which included 208 cases and 306 controls (no evidence of breast cancer) subjects. DNA was extracted from blood samples and IGF-1 and IGFBP-3 genotyping was performed by PCR-GeneScan and PCR-RFLP methods, respectively. Our results demonstrated that comorbidity is significantly correlated with breast cancer (OR = 2.12; 95% CI = 1.40-3.21; P-value <0.01); especially the comorbidity of hypertension (OR= 2.27; 95% CI = 1.47-3.49; P-value <0.01). Furthermore, there was a significant association between non-19/non-19 IGF-1 (CA)n polymorphisms and breast cancer (OR= 4.80; 95% CI = 1.87, 12.36; P-value <0.01). This study also found a significant association between the non-19/non-19 IGF-1 (CA)n genotype, breast cancer, and comorbidity (OR= 4.65; 95% CI = 1.57, 13.82; P-value <0.01). In summary, our study demonstrated a significant association of IGF-1 polymorphisms and breast cancer with a further link to comorbid diseases. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 878.
The extracellular matrix of epithelial tumors undergoes structural remodeling during periods of uncontrolled growth, creating regional heterogeneity and torsional stress. How matrix integrity is maintained in the face of dynamic biophysical forces is largely undefined. Here we investigated the role of fibulin-2, a matrix glycoprotein that functions biomechanically as an inter-molecular clasp and thereby facilitates supra-molecular assembly. Fibulin-2 was abundant in the extracellular matrix of human lung adenocarcinomas and was highly expressed in tumor cell lines derived from mice that develop metastatic lung adenocarcinoma from co-expression of mutant K-ras and p53. Loss-of-function experiments in tumor cells revealed that fibulin-2 was required for tumor cells to grow and metastasize in syngeneic mice, a surprising finding given that other intra-tumoral cell types are known to secrete fibulin-2. However, tumor cells grew and metastasized equally well in Fbln2-null and -wild-type littermates, implying that malignant progression was dependent specifically upon tumor cell-derived fibulin-2, which could not be offset by other cellular sources of fibulin-2. Fibulin-2 deficiency impaired the ability of tumor cells to migrate and invade in Boyden chambers, to create a stiff extracellular matrix in mice, to cross-link secreted collagen, and to adhere to collagen. We conclude that fibulin-2 is a driver of malignant progression in lung adenocarcinoma and plays an unexpected role in collagen cross-linking and tumor cell adherence to collagen.
Abstract Tumor microenvironment plays a very important role in regulating the cell behavior within a tissue, largely by interacting with the extracellular matrix (ECM). Recently, our lab has developed a novel ex vivo lung model that forms perfusable tumor nodules, which mimic human lung cancer histopathology and protease secretion patterns better than tumor cells grown on a petri dish. In this study, we have compared the genome-wide gene expression profiles (Affymetrix 430_2) and protein arrays (Reverse phase protein array or RPPA) of 2D (monolayer) culture versus our ex vivo 3D cell culture using 393p cells, derived from mice lung cancer. Expression array data showed a set 1314 gene probes differentially expressed between monolayer 2D culture and 3D ex vivo cell culture. Gene Ontology (GO) analysis of differentially expressed genes showed the upregulation of several genes associated with extracellular matrix (ECM1, BMP15, FMOD, cathepsin, MMP9, MMP10), cell signaling (VEGFA, TGFB2, FGF20), and immune response (C1, C3, MHC class II, IL1α, IL1RA, IL13, TNF receptor superfamily, CXCL2). Selected genes were also validated using real time PCR. We found 46 proteins with significant (p<0.01) differential levels in 2D vs 3D cell cultures of 393p. The proteins associated with extracellular matrix (Collagen VI, Caveolin1, Fibronectin) and cell signaling (mTOR, p38 (phosphorylated), Caspase 7 (Cleaved), BAD (phosphorylated), PI3K) were upregulated in 3D ex vivo cell culture compared to conventional 2D culture. Thus, our results showed the insightful influence of the 3D ex vivo model and its three-dimensional perfusable environment over cell growth and its behavior in terms of significant differences in gene expression and protein patterns as compared to the same cells grown on conventional 2D cell cultures. Citation Format: Dhruva K. Mishra, Chad Creighton, Yiqun Zhang, Don L. Gibbons, Jonathan M. Kurie, Min P. Kim. Gene expression analysis of perfusable nodules grown on ex vivo 3D model shows effects on extracellular matrix, cell signaling and immune response. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1508. doi:10.1158/1538-7445.AM2013-1508
