Adjusting HIV Prevalence Estimates for Non-participation: an Application to Demographic Surveillance

Introduction: HIV testing is a cornerstone of efforts to combat the HIV epidemic, and testing conducted as part of surveillance provides invaluable data on the spread of infection and the effectiveness of campaigns to reduce the transmission of HIV. However, participation in HIV testing can be low, and if respondents systematically select not to be tested because they know or suspect they are HIV positive (and fear disclosure), standard approaches to deal with missing data will fail to remove selection bias. We implemented Heckman-type selection models, which can be used to adjust for missing data that are not missing at random, and established the extent of selection bias in a population-based HIV survey in an HIV hyperendemic community in rural South Africa.

Methods: We used data from a population-based HIV survey carried out in 2009 in rural KwaZulu-Natal, South Africa. In this survey, 5565 women (35%) and 2567 men (27%) provided blood for an HIV test. We accounted for missing data using interviewer identity as a selection variable which predicted consent to HIV testing but was unlikely to be independently associated with HIV status. Our approach involved using this selection variable to examine the HIV status of residents who would ordinarily refuse to test, except that they were allocated a persuasive interviewer. Our copula model allows for flexibility when modelling the dependence structure between HIV survey participation and HIV status.

Results: For women, our selection model generated an HIV prevalence estimate of 33% (95% CI 27–40) for all people eligible to consent to HIV testing in the survey. This estimate is higher than the estimate of 24% generated when only information from respondents who participated in testing is used in the analysis, and the estimate of 27% when imputation analysis is used to predict missing data on HIV status. For men, we found an HIV prevalence of 25% (95% CI 15–35) using the selection model, compared to 16% among those who participated in testing, and 18% estimated with imputation. We provide new confidence intervals that correct for the fact that the relationship between testing and HIV status is unknown and requires estimation.

Conclusions: We confirm the feasibility and value of adopting selection models to account for missing data in population-based HIV surveys and surveillance systems. Elements of survey design, such as interviewer identity, present the opportunity to adopt this approach in routine applications. Where non-participation is high, true confidence intervals are much wider than those generated by standard approaches to dealing with missing data suggest.

The CXCR2 antagonist, SCH-527123, shows antitumor activity and sensitizes cells to oxaliplatin in preclinical colon cancer models

Colorectal cancer is the second most common cause of cancer-related death in the United States. Recent studies showed that interleukin-8 (IL-8) and its receptors (CXCR1 and CXCR2) are significantly upregulated in both the tumor and its microenvironment, and act as key regulators of proliferation, angiogenesis, and metastasis. Our previous study showed that IL-8 overexpression in colorectal cancer cells triggers the upregulation of the CXCR2-mediated proliferative pathway. The aim of this study was to investigate whether the CXCR2 antagonist, SCH-527123, inhibits colorectal cancer proliferation and if it can sensitize colorectal cancer cells to oxaliplatin both in vitro and in vivo. SCH-527123 showed concentration-dependent antiproliferative effects in HCT116, Caco2, and their respective IL-8-overexpressing variants colorectal cancer cell lines. Moreover, SCH-527123 was able to suppress CXCR2-mediated signal transduction as shown through decreased phosphorylation of the NF-κB/mitogen-activated protein kinase (MAPK)/AKT pathway. These findings corresponded with decreased cell migration and invasion, while increased apoptosis in colorectal cancer cell lines. In vivo results verified that SCH-527123 treatment decreased tumor growth and microvessel density when compared with vehicle-treated tumors. Importantly, these preclinical studies showed that the combination of SCH-527123 and oxaliplatin resulted in a greater decrease in cell proliferation, tumor growth, apoptosis, and angiogenesis that was superior to single-agent treatment. Taken together, these findings suggest that targeting CXCR2 may block tumor proliferation, migration, invasion, and angiogenesis. In addition, CXCR2 blockade may further sensitize colorectal cancer to oxaliplatin treatment.

Inhibition of dUTPase induces synthetic lethality with thymidylate synthase-targeted therapies in non-small cell lung cancer

Chemotherapies that target thymidylate synthase (TS) continue to see considerable clinical expansion in non-small cell lung cancer (NSCLC). One drawback to TS-targeted therapies is drug resistance and subsequent treatment failure. Novel therapeutic and biomarker-driven strategies are urgently needed. The enzyme deoxyuridine triphosphate nucleotidohydrolase (dUTPase) is reported to protect tumor cells from aberrant misincorporation of uracil during TS inhibition. The goal of this study was to investigate the expression and significance of dUTPase in mediating response to TS-targeted agents in NSCLC. The expression of dUTPase in NSCLC cell lines and clinical specimens was measured by quantitative real-time reverse transcriptase PCR and immunohistochemistry. Using a validated RNA interference approach, dUTPase was effectively silenced in a panel of NSCLC cell lines and response to the fluoropyrimidine fluorodeoxyuridine (FUdR) and the antifolate pemetrexed was analyzed using growth inhibition and clonogenic assays. Apoptosis was analyzed by flow cytometry. Significant variation in the quantity and cellular expression of dUTPase was observed, including clear evidence of overexpression in NSCLC cell line models and tumor specimens at the mRNA and protein level. RNA interference-mediated silencing of dUTPase significantly sensitized NSCLC cells to growth inhibition induced by FUdR and pemetrexed. This sensitization was accompanied by a significant expansion of intracellular dUTP pools and significant decreases in NSCLC cell viability evaluated by clonogenicity and apoptotic analyses. Together, these results strongly suggest that uracil misincorporation is a potent determinant of cytotoxicity to TS inhibition in NSCLC and that inhibition of dUTPase is a mechanism-based therapeutic approach to significantly enhance the efficacy of TS-targeted chemotherapeutic agents.

Novel opportunities for thymidylate metabolism as a therapeutic target

For over 40 years, the fluoropyrimidine 5-fluorouracil (5-FU) has remained the central agent in therapeutic regimens employed in the treatment of colorectal cancer and is frequently combined with the DNA-damaging agents oxaliplatin and irinotecan, increasing response rates and improving overall survival. However, many patients will derive little or no benefit from treatment, highlighting the need to identify novel therapeutic targets to improve the efficacy of current 5-FU-based chemotherapeutic strategies. dUTP nucleotidohydrolase (dUTPase) catalyzes the hydrolysis of dUTP to dUMP and PPi, providing substrate for thymidylate synthase (TS) and DNA synthesis and repair. Although dUTP is a normal intermediate in DNA synthesis, its accumulation and misincorporation into DNA as uracil is lethal. Importantly, uracil misincorporation represents an important mechanism of cytotoxicity induced by the TS-targeted class of chemotherapeutic agents including 5-FU. A growing body of evidence suggests that dUTPase is an important mediator of response to TS-targeted agents. In this article, we present further evidence showing that elevated expression of dUTPase can protect breast cancer cells from the expansion of the intracellular uracil pool, translating to reduced growth inhibition following treatment with 5-FU. We therefore report the implementation of in silico drug development techniques to identify and develop small-molecule inhibitors of dUTPase. As 5-FU and the oral 5-FU prodrug capecitabine remain central agents in the treatment of a variety of malignancies, the clinical utility of a small-molecule inhibitor to dUTPase represents a viable strategy to improve the clinical efficacy of these mainstay chemotherapeutic agents.

Plastin Polymorphisms Predict Gender- and Stage-Specific Colon Cancer Recurrence after Adjuvant Chemotherapy

Tumor recurrence after curative resection remains a major problem in patients with locally advanced colorectal cancer treated with adjuvant chemotherapy. Genetic single-nucleotide polymorphisms (SNP) may serve as useful molecular markers to predict clinical outcomes in these patients and identify targets for future drug development. Recent in vitro and in vivo studies have demonstrated that the plastin genes PLS3 and LCP1 are overexpressed in colon cancer cells and play an important role in tumor cell invasion, adhesion, and migration. Hence, we hypothesized that functional genetic variations of plastin may have direct effects on the progression and prognosis of locally advanced colorectal cancer. We tested whether functional tagging polymorphisms of PLS3 and LCP1 predict time to tumor recurrence (TTR) in 732 patients (training set, 234; validation set, 498) with stage II/III colorectal cancer. The PLS3 rs11342 and LCP1 rs4941543 polymorphisms were associated with a significantly increased risk for recurrence in the training set. PLS3 rs6643869 showed a consistent association with TTR in the training and validation set, when stratified by gender and tumor location. Female patients with the PLS3 rs6643869 AA genotype had the shortest median TTR compared with those with any G allele in the training set [1.7 vs. 9.4 years; HR, 2.84; 95% confidence interval (CI), 1.32-6.1; P = 0.005] and validation set (3.3 vs. 13.7 years; HR, 2.07; 95% CI, 1.09-3.91; P = 0.021). Our findings suggest that several SNPs of the PLS3 and LCP1 genes could serve as gender- and/or stage-specific molecular predictors of tumor recurrence in stage II/III patients with colorectal cancer as well as potential therapeutic targets.

Prognostic role of lemur tyrosine kinase-3 germline polymorphisms in adjuvant gastric cancer in Japan and the United States

Lemur tyrosine kinase-3 (LMTK3) was recently identified as an estrogen receptor (ER)-α modulator related to endocrine therapy resistance, and its polymorphisms rs9989661 (T>C) T/T genotype and rs8108419 (G>A) G/G or A/G genotype predicted improved outcomes in breast cancer. Because different predominant ER distributions link to breast and gastric cancer and little is known of the prognostic role of LMTK3 in gastric cancer, this study was carried out to clarify the prognostic role of these polymorphisms in gastric cancer. One-hundred and sixty-nine Japanese and 137 U.S. patients with localized gastric adenocarcinoma were enrolled. Genomic DNA was extracted from blood or tissue, and all samples were analyzed by PCR-based direct DNA sequencing. Overall, these polymorphisms were not associated with survival in both cohorts. When gender was considered, in multivariate analysis, harboring rs9989661 T/T genotype was associated with disease-free survival [HR, 4.37; 95% confidence interval (CI), 2.08-9.18; P < 0.0001] and overall survival (OS; HR, 3.69; 95% CI, 1.65-8.24; P = 0.0014) in the Japanese males and time to recurrence (HR, 7.29; 95% CI, 1.07-49.80; P = 0.043) in the U.S. females. Meanwhile, harboring rs8108419 G/G genotype was associated with OS in the Japanese females (HR, 3.04; 95% CI, 1.08-8.56; P = 0.035) and the U.S. males (HR, 3.39; 95% CI, 1.31-8.80; P = 0.012). The prognostic role of these polymorphisms may be negative in gastric cancer. These findings suggest that the estrogen pathway may play a prognostic role in patients with gastric cancer but this may be dependent on the regional differences both in physiology and genetic alterations of gastric cancer.

Abundance and diversity of sedimentary bacterial communities in a coastal productive setting in the Western Irish Sea

The bacterial community composition and biomass abundance from a depositional mud belt in the western Irish Sea and regional sands were investigated by phospholipid ester-linked fatty acid profiling, denaturing gradient gel electrophoresis and barcoded pyrosequencing of 16S rRNA genes. The study area varied by water depth (12-111 m), organic carbon content (0.09-1.57% TOC), grain size, hydrographic regime (well-mixed vs. stratified), and water column phytodetrital input (represented by algal polyunsaturated PLFA). The relative abundance of bacterial-derived PLFA (sum of methyl-branched, cyclopropyl and odd-carbon number PLFA) was positively correlated with fine-grained sediment, and was highest in the depositional mud belt. A strong association between bacterial biomass and eukaryote primary production was suggested based on observed positive correlations with total nitrogen and algal polyunsaturated fatty acids. In addition, 16S rRNA genes affiliated to the classes Clostridia and Flavobacteria represented a major proportion of total 16S rRNA gene sequences. This suggests that benthic bacterial communities are also important degraders of phytodetrital organic matter and closely coupled to water column productivity in the western Irish Sea.

Abstract B64: The role of JAK1/2-STAT3 as acute resistance mechanism to MEK inhibition in BRAF mutant colorectal cancer.

Background: Oncogenic mutations in BRAF occur in 8% of patients with advanced colorectal cancer (CRC) and have been shown to correlate with poor prognosis. In contrast to BRAF mutant (MT) melanoma, where the BRAF inhibitor PLX4032 has shown significant increases in response rates and overall survival compared to standard Dacarbazine treatment, only minor responses to PLX4032 treatment have been reported in BRAFMT CRC. Clear understanding of the vulnerabilities of BRAFMT CRC is important, and identification of druggable targets uniquely required by BRAFMT CRC tumors has the potential to fill a gap in the therapeutic armamentarium of advanced CRC. The aim of this study was to identify novel resistance mechanisms to MAPK inhibition in BRAFMT CRC.

Methods: Paired BRAFMT/WT RKO and VACO432 CRC cell line models and non-isogenic BRAFMT LIM2405, WiDR and COLO205 CRC cells were used. Changes in protein expression/activity were assessed by Western Blotting. Interaction between MEK1/2 and JAK1/2 inhibition was assessed using the MTT cell viability assays and flow cytometry. Apoptosis was measured using Western blotting for PARP, cleaved caspase 3/8 and caspase 8, 3/7 activity assays.

Results: Treatment with MEK1/2 inhibitors AZD6244, GSK1120212, UO126 and PD98059 resulted in acute increases in STAT3 activity in the BRAFMT RKO and VACO432 cells but not in their BRAFWT clones and this was associated with increases in JAK2 activity. Inhibition of JAK/STAT3 activation using gene specific siRNA or small molecule inhibitors TG101348 or AZD1480, abrogated this survival response and resulted in significant increases in cell death when combined with MEK1/2 inhibitors AZD6244 or GSK1120212 in BRAFMT CRC cells. In addition, combination of MEK1/2 and JAK/STAT3 inhibition resulted in strong synergy with CI values between 0.3 and 0.7 in BRAFMT CRC cells.

Conclusions: We have identified JAK/STAT3 activation as an important escape mechanism for BRAFMT CRC following MEK1/2 inhibition. These data provide a strong rationale for further investigation of combination of MEK1/2 and JAK/STAT3 inhibition in BRAFMT in vivo models.