Lessons learned for public health workforce development: An evaluation of the centers for disease control and prevention's laboratory leadership service fellowship.

The Centers for Disease Control and Prevention launched the Laboratory Leadership Service (LLS) Fellowship Program in July 2015 to develop public health laboratory (PHL) leaders who will improve PHL quality and safety. This article describes a retrospective, summative evaluation to determine the extent to which LLS has met its short-term goals for PHL workforce development. The evaluation relied on existing data from routine LLS data collection and reporting, supplemented with a new alumni survey. The purpose of the design was threefold: 1) to reduce data collection burden on program staff and participants, 2) to assess the value and limits of routine fellowship data for comprehensive public health workforce development program evaluation, and 3) to identify ways to improve LLS's routine data collections for program evaluation. We used descriptive statistics, qualitative analysis, and participatory methods (i.e., a data party) to analyze and interpret data. Results show LLS short-term outcome achievement and highlight opportunities for program improvement, particularly related to the design of certain training requirements and for future evaluations. Overall, the evaluation contributes to lessons learned for PHL workforce development efforts, including how routine data collections can contribute to comprehensive public health workforce development evaluations.

MiRNA 26a expression in a novel panel of African American prostate cancer cell lines.

INTRODUCTION: African American men have disproportionately high incidence and mortality rates of prostate cancer when compared to other ethnic groups in the United States. The identification of molecular factors that contribute to this disparity could improve diagnosis and therapeutic intervention. Therefore, the purpose of this study was to determine the miRNA 26a expression profile in novel African American and Caucasian prostate cell lines at each clinical stage of prostate cancer progression. METHODS: The miR-26a expression profile was investigated using novel African American and Caucasian prostate cell lines representing each pathological stage: non-malignant, malignant, and metastatic tumors. Relative miRNA expression was determined by qRT-PCR. RESULTS: Our data showed a 2.25 fold increase for miR-26a in the non-malignant, a 13.3 fold increase in malignant and 2.38 fold increase in metastatic tumors, when comparing African American and Caucasian prostate cell lines of similar clinical stage and pathological grade. African American malignant prostate cancer cell lines showed the most significant fold difference in expression among all cell lines tested. Furthermore, there was a general increase in miR-26a expression toward the more aggressive cell lines in both African American and Caucasian prostate cell lines. CONCLUSION: To date, we are unaware of any studies that compare the miRNA profile at different stages of prostate cancer among two racial groups. Although a gene target for miR-26a has not been identified, our data show a possible role for miRNA regulation of gene expression in prostate cancer progression. Furthermore, this study suggests that miRNAs could possibly contribute to the aggressiveness associated in African American patients with prostate cancer.

Transcriptional repressor Kaiso promotes epithelial to mesenchymal transition and metastasis in prostate cancer through direct regulation of miR-200c.

The loss of miR-200 family, through DNA methylation, results in cancer cells undergoing an epithelial to mesenchymal transition (EMT), and metastasis. In this study, we established that the transcriptional repressor Kaiso directly binds methylated regions of the miR-200 family, and this is reversed with 5-aza treatment. sh-Kaiso PC-3 cells display increased miR-200-a/b/c, miR-141, and miR-429 expression, with miR-200c demonstrating the most significant increase. Interestingly, overexpression of EGFR or treatment with EGF decreases miR-200c expression and this is reversed after treatment with EGFR specific kinase inhibitor PD153035. However, EGF did not have a significant effect on miR-200c in sh-Kaiso DU-145 or PC-3 cell lines, suggesting Kaiso silences miR-200c through the activation of EGFR signaling. Overexpression of Kaiso in LNCaP cells results in decreased expression of miR-200-a/b/c, miR-141, and miR-429, along with increased expression of ZEB1, p-EGFR and total EGFR levels. Overexpression of miR200c in PC-3 cells results in decreased expression of EGFR, ZEB1, ERK1/2 and Kaiso. Additionally, sh-Kaiso PC-3 demonstrates reduced in vivo tumor formation and metastasis. Thus, our data suggests that EGFR signaling regulates the silencing of miR-200 family through Kaiso binding to methylated regions in the promoter.

Nuclear localization of Kaiso promotes the poorly differentiated phenotype and EMT in infiltrating ductal carcinomas.

The expression and biological consequences of Kaiso, a novel bi-modal transcription factor, in infiltrating ductal carcinomas (IDCs) have not been widely investigated. In the present study, we determined Kaiso expression and subcellular localization in 146 normal tissues, 376 IDCs, and 85 lymph node metastases. In IDCs, there was higher Kaiso expression in both the cytoplasmic and nuclear compartments, which correlated with age <48 (cytoplasmic p < 0.0093; nuclear p < 0.0001) and moderate differentiation (cytoplasmic p < 0.0042; nuclear p < 0.0001), as determined by Chi square analysis. However, only nuclear Kaiso correlated with poor prognostic factors, i.e., race (African Americans) (p < 0.0001), poor differentiation (p < 0.0001), and metastases (p < 0.0001). Nuclear Kaiso was also associated with worse overall survival (p < 0.0019), with African American patients displaying worse survival rates relative to Caucasian patients (p < 0.029). MCF-7 (non-metastatic), MDA-MB-468 (few metastases), and MDA-MB-231 (highly metastatic) breast cancer cells demonstrated increasing Kaiso levels, with more nuclear localization in the highly metastatic cell line. Over-expression of Kaiso in MCF-7 cells increased cell migration and invasion, but treatment of MDA-MB-468 and MDA-MB-231 cells with si-Kaiso decreased cell migration and invasion and induced expression of E-cadherin RNA and protein. E-cadherin re-expression was associated with a reversal of mesenchymal associated cadherins, N-cadherin and cadherin 11, as well as decreased vitamin expression. Further, Kaiso directly bound to methylated sequences in the E-cadherin promoter, an effect prevented by 5-aza-2-deoxycytidine. Immunofluorescence co-staining of poorly differentiated IDCs demonstrated that nuclear Kaiso is associated with a loss of E-cadherin expression. These findings support a role for Kaiso in promoting aggressive breast tumors.

MicroRNA profiling of novel African American and Caucasian Prostate Cancer cell lines reveals a reciprocal regulatory relationship of miR-152 and DNA methyltranferase 1.

miRNA expression in African American compared to Caucasian PCa patients has not been widely explored. Herein, we probed the miRNA expression profile of novel AA and CA derived prostate cancer cell lines. We found a unique miRNA signature associated with AA cell lines, independent of tumor status. Evaluation of the most differentially expressed miRNAs showed that miR-132, miR-367b, miR-410, and miR-152 were decreased in more aggressive cells, and this was reversed after treatment of the cells with 5-aza-2'-deoxycytidine. Sequencing of the miR-152 promoter confirmed that it was highly methylated. Ectopic expression of miR-152 resulted in decreased growth, migration, and invasion. Informatics analysis of a large patient cohort showed that decreased miR-152 expression correlated with increased metastasis and a decrease in biochemical recurrence free survival. Analysis of 39 prostate cancer tissues with matched controls (20 AA and 19 CA), showed that 50% of AA patients had statistically significant lower miR-152 expression compared to only 35% of CA patients. Ectopic expression of miR-152 in LNCaP, PC-3, and MDA-PCa-2b cells down-regulated DNA (cytosine-5)-methyltransferase 1 (DNMT1) through direct binding in the DNMT1 3'UTR. There appeared to be a reciprocal regulatory relationship of miR-152/DNMT1 expression, as cells treated with siRNA DNMT1 caused miR-152 to be re-expressed in all cell lines. In summary, these results demonstrate that epigenetic regulation of miR-152/DNMT1 may play an important role in multiple events that contribute to the aggressiveness of PCa tumors, with an emphasis on AA PCa patients .

Establishment and characterization of a pair of non-malignant and malignant tumor derived cell lines from an African American prostate cancer patient.

Research into molecular and genetic mechanisms underlying prostate carcinogenesis in high-risk African American men would be greatly advanced by in vitro models of African American prostate tumors representing primary tumors. However, the generation of immortalized primary African American prostate cancer cells that will accurately reflect the in situ characteristics of malignant epithelium is currently limited but is greatly needed. We have successfully established immortalized cell lines of a pair of non-malignant and malignant tumors derived from an African American prostate cancer patient with HPV-16E6E7 (RC-77N/E and RC-77T/E). RC-77N/E and RC-77T/E cells are currently growing well at passage 40. Both cells exhibit epithelial morphology and are androgen sensitive. The RC-77T/E cells produced tumors in SCID mice whereas the RC-77N/E cells produced no tumor in SCID mice. These cells expressed androgen-regulated prostate-specific homobox gene, NKX 3.1, epithelial cell specific cytokeratn 8, androgen receptor (AR), prostate specific antigen (PSA), and p16. Chromosome analysis showed that both cell lines are similar; near diploid human male (XY) with most chromosome counts in the 45-48 range. However, RC-77T/E cell line has new marker chromosomes: M1B=del/t(4;?)(q28;?), M5=16q+ in addition to those observed in the RC-77N/E cell line (M1=del(4)(q28q34)+hsr in some, M1A=t(4q;?),M2=der(9?),M2A=del(M2p-),M3=iso(?), M4=der(22?)). This is the first documented case of the establishment of pair of non-malignant and malignant tumors derived from an African American prostate cancer patient. These models will provide novel tools to study the molecular and genetic mechanisms of prostate carcinogenesis, especially for high-risk African American men.