Overexpression and surface localization of HPRT in prostate cancer provides a potential target for cancer specific antibody mediated cellular cytotoxicity.

We chose to evaluate Hypoxanthine Guanine Phosphoribosyltransferase (HPRT) as a possible biomarker for prostate cancer due to its involvement in nucleotide synthesis and cell cycle progression. We utilized two prostate cancer cell lines (PC3 and DU145) along with patient tissue and knockdowns to evaluate overall HPRT expression. The surface localization of HPRT was determined utilizing flow cytometry, confocal microscopy, and scanning electron microscopy followed by ADCC to evaluate targeting potential. We found significant upregulation of HPRT within malignant samples with approximately 47% of patients had elevated levels of HPRT compared to normal controls. We also observed a significant association between HPRT and the plasma membrane of DU145 cells (p = 0.0004), but found no presence on PC3 cells (p = 0.14). This was confirmed with scanning electron microscopy and confocal microscopy. ADCC experiments were performed to determine whether HPRT could be used as a target antigen for selective cell-mediated killing. We found that DU145 cells treated with HPRT antibodies had a significantly higher incidence of cell death than both isotype treated samples and PC3 cells treated with the same concentrations of HPRT antibody. Finally, we determined that p53 had a significant impact on HPRT expression both internally and on the surface of cancer cells. These results suggest HPRT as a possible biomarker target for the treatment of patients with prostate cancer.

Potential new biomarkers for endometrial cancer.

BACKGROUND: Incidence of endometrial cancer are rising both in the United States and worldwide. As endometrial cancer becomes more prominent, the need to develop and characterize biomarkers for early stage diagnosis and the treatment of endometrial cancer has become an important priority. Several biomarkers currently used to diagnose endometrial cancer are directly related to obesity. Although epigenetic and mutational biomarkers have been identified and have resulted in treatment options for patients with specific aberrations, many tumors do not harbor those specific aberrations. A promising alternative is to determine biomarkers based on differential gene expression, which can be used to estimate prognosis. METHODS: We evaluated 589 patients to determine differential expression between normal and malignant patient samples. We then supplemented these evaluations with immunohistochemistry staining of endometrial tumors and normal tissues. Additionally, we used the Library of Integrated Network-based Cellular Signatures to evaluate the effects of 1826 chemotherapy drugs on 26 cell lines to determine the effects of each drug on HPRT1 and AURKA expression. RESULTS: Expression of HPRT1, Jag2, AURKA, and PGK1 were elevated when compared to normal samples, and HPRT1 and PGK1 showed a stepwise elevation in expression that was significantly related to cancer grade. To determine the prognostic potential of these genes, we evaluated patient outcome and found that levels of both HPRT1 and AURKA were significantly correlated with overall patient survival. When evaluating drugs that had the most significant effect on lowering the expression of HPRT1 and AURKA, we found that Topo I and MEK inhibitors were most effective at reducing HPRT1 expression. Meanwhile, drugs that were effective at reducing AURKA expression were more diverse (MEK, Topo I, MELK, HDAC, etc.). The effects of these drugs on the expression of HPRT1 and AURKA provides insight into their role within cellular maintenance. CONCLUSIONS: Collectively, these data show that JAG2, AURKA, PGK1, and HRPT1 have the potential to be used independently as diagnostic, prognostic, or treatment biomarkers in endometrial cancer. Expression levels of these genes may provide physicians with insight into tumor aggressiveness and chemotherapy drugs that are well suited to individual patients.

Correction to: Membrane expression of thymidine kinase 1 and potential clinical relevance in lung, breast, and colorectal malignancies.

[This corrects the article DOI: 10.1186/s12935-018-0633-9.].

Membrane expression of thymidine kinase 1 and potential clinical relevance in lung, breast, and colorectal malignancies.

BACKGROUND: Lung, breast, and colorectal malignancies are the leading cause of cancer-related deaths in the world causing over 2.8 million cancer-related deaths yearly. Despite efforts to improve prevention methods, early detection, and treatments, survival rates for advanced stage lung, breast, and colon cancer remain low, indicating a critical need to identify cancer-specific biomarkers for early detection and treatment. Thymidine kinase 1 (TK1) is a nucleotide salvage pathway enzyme involved in cellular proliferation and considered an important tumor proliferation biomarker in the serum. In this study, we further characterized TK1's potential as a tumor biomarker and immunotherapeutic target and clinical relevance. METHODS: We assessed TK1 surface localization by flow cytometry and confocal microscopy in lung (NCI-H460, A549), breast (MDA-MB-231, MCF7), and colorectal (HT-29, SW620) cancer cell lines. We also isolated cell surface proteins from HT-29 cells and performed a western blot confirming the presence of TK1 on cell membrane protein fractions. To evaluate TK1's clinical relevance, we compared TK1 expression levels in normal and malignant tissue through flow cytometry and immunohistochemistry. We also analyzed RNA-Seq data from The Cancer Genome Atlas (TCGA) to assess differential expression of the TK1 gene in lung, breast, and colorectal cancer patients. RESULTS: We found significant expression of TK1 on the surface of NCI-H460, A549, MDA-MB-231, MCF7, and HT-29 cell lines and a strong association between TK1's localization with the membrane through confocal microscopy and Western blot. We found negligible TK1 surface expression in normal healthy tissue and significantly higher TK1 expression in malignant tissues. Patient data from TCGA revealed that the TK1 gene expression is upregulated in cancer patients compared to normal healthy patients. CONCLUSIONS: Our results show that TK1 localizes on the surface of lung, breast, and colorectal cell lines and is upregulated in malignant tissues and patients compared to healthy tissues and patients. We conclude that TK1 is a potential clinical biomarker for the treatment of lung, breast, and colorectal cancer.

Surgical Capacity Building in Low- and Middle-Income Countries: Lessons for Thoracic Surgery.

There is great need for intentional investment in capacity building for thoracic surgical conditions. This article provides a brief overview of thoracic surgical capacity building for low- and middle-income countries using the Lancet framework of infrastructure, workforce, financing, and information management. The authors highlight the needs, opportunities, and challenges that are relevant for the thoracic surgical community, as it aims to increase care for patients with these conditions globally.

Falling from grace: HPRT is not suitable as an endogenous control for cancer-related studies.

HPRT is a housekeeping enzyme involved in recycling guanine and inosine in the purine salvage pathway. As a housekeeping gene, HPRT has been widely used as an endogenous control for molecular studies evaluating changes in gene expression. Yet, recent evidence has shown that HPRT exhibits high variability within malignant samples. We designed this study to determine whether this observed upregulation is consistently found, therefore rendering hprt an unsuitable normalization control in cancer. Utilizing protein and RNA-seq expression, we found that malignant and normal patient samples vary significantly both within the same tissue type and across organ sites. Upon staining for HPRT via immunohistochemistry, we found that expression is highly variable in malignant samples (Lung; 89.2-111.8, Breast; 66.7-98.3, Colon; 85.3-129.7, Prostate; 90.8-155.4, Pancreas; 74.1-132.1). Similarly, we observed high variability across cell lines via western blotting (p < 0.0001) which was further confirmed using RNA sequencing. Comparing normal and malignant patient samples, we observed consistent upregulation of HPRT expression within malignant samples relative to normal samples (p = 0.0001). These data indicate that HPRT is unsuitable as an endogenous control for cancer-related studies because its expression is highly variable and exceeds that of an appropriate control; therefore, we recommend its discontinued use as a normalization gene.

Examination of Hypoxanthine Guanine Phosphoribosyltransferase as a biomarker for colorectal cancer patients.

The aim of this study is to investigate these enzymes as possible biomarkers in two colorectal cancer cell lines: HT29, SW480, SW620, and Colo205. With 1,168,929 individuals currently diagnosed with colorectal cancer in the United States, there remains a need to find biomarkers to improve diagnosis and expand treatment options for patients. Due to their role in proliferation and cell cycle regulation, we hypothesized an increase in salvage pathway enzyme (APRT, DCK, and HPRT) expression and possible presentation within colon cancer cells. Enzyme surface localization was assessed utilizing confocal microscopy, flow cytometry, and scanning electron microscopy. General protein expression was evaluated utilizing immunohistochemistry and Western blot analysis. While we found no statistically significant presence of either APRT or DCK on the membranes of SW620, Colo205, and HT29 cells, but found significant expression of HPRT on the surface of HT29, SW480, and SW620 cells. The average population fluorescence increased by 28%, 58%, and 40% in HT29, SW620, and SW480 cells, respectively, when compared to isotype controls. Confocal microscopy images revealed direct overlap between SW620 cells stained with a membrane dye and anti-HPRT antibody, indicating co-localization on the plasma membrane. In addition, cells treated with gold labelled HPRT antibody experienced significant changes in gold weight percentage on both SW620 and HT29 cells when compared to isotype controls. When evaluating expression within normal tissue, there was insignificant levels of HPRT binding. These data collectively suggest that HPRT may be a possible biomarker target for the identification and treatment of colorectal cancer.

Metastatic colon adenocarcinoma has a significantly elevated expression of IL-10 compared with primary colon adenocarcinoma tumors.

Classical anti-inflammatory cytokines are known to play a role in both cancer progression as well as cancer elimination. We evaluated the anti-inflammatory cytokines IL-10 and TGF-ß in patients with colon adenocarcinoma and metastatic colon adenocarcinoma using immunohistochemical assays to determine the expression of the cytokines between various malignant tissues. We found tissues stained with TGF-ß showed no significant upregulation within malignant tumors when compared with normal tissue controls. We observed high levels of TGF-ß presence in most tissues similar to GAPDH expression. Within both colon adenocarcinoma and metastatic carcinomas there was a significant variability among patients in the expression of IL-10. While some patients experienced insignificant increases in the cytokine compared with controls, other patients had a clear upregulation of the protein within their tissue. In addition, there was an increase in the number of patients positive for IL-10 upregulation within metastatic tumors when compared with primary tumors. These data indicate that there is substantial variability between patients in regards to IL-10 expression, which may further aid in characterizing tumors and evaluating metastatic potential.