BackgroundLung, 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.MethodsWe 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.ResultsWe 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.ConclusionsOur 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.Electronic supplementary materialThe online version of this article (10.1186/s12935-018-0633-9) contains supplementary material, which is available to authorized users.

Hypoxanthine Guanine Phosphoribosyltransferase (HPRT) is a housekeeping enzyme involved in the purine synthesis of guanine and inosine in the salvage pathway. While other salvage pathway enzymes, such as TK1, have been established as biomarkers for both cancer cell proliferation and cancer development, little research been done to evaluate whether HPRT has the same potential as a cancer biomarker. We designed this study to determine if HPRT has value as an identifier of malignancy within the most common types of cancer. We utilized histological samples from lung, colon, prostate, and breast cancer with additional normal tissue to evaluate whether there was any elevation of HPRT within malignant samples. In addition, we also assessed general HPRT expression within patient"s samples from The Cancer Genome Atlas (TCGA) to confirm clinical relevance. We found that within a subset of patients, there was significant elevation of HPRT when compared to normal tissue controls. This elevation was seen in 33-55% of the malignant samples and appears to have no dependence on cancer stage. There were slight differences in staining patterns among all the organ types, but overall each organ displayed the same pattern of "HPRT high" and "HPRT low" populations within malignant samples. We found that in our TCGA samples, there was a similar elevation of HPRT that was significant when compared to normal controls. Overall, as an upregulated enzyme that does not directly correlate with stage, HPRT could become a valuable marker in the early diagnosis of a variety of solid tumors.

The aim of this study is to examine the gene expression of the purine salvage pathway enzyme Hypoxanthine Guanine Phosphribosyltransferase (HPRT) in malignant and normal tissue to determine potential upregulation. Due to the critical role HPRT plays in the cell cycle, it was hypothesized that in a rapidly proliferating malignant tissue there may be differential gene expression. In breast, colon, lung, and prostate cancer the 5-year survival rates decrease by nearly 72% when diagnosed in stage 3 or 4 as opposed to an early stage diagnosis. We focused our investigation on evaluating if HPRT could serve as an early stage diagnostic agent for the four most commonly diagnosed cancers (lung, colorectal, breast, and prostate cancer) which together make up 42.5% of all cancer diagnoses. Initially, we evaluated differences in HPRT expression levels via RNA-sequencing data in 3,147 tumor and 316 normal samples from The Cancer Genome Atlas (TCGA). Samples from 1119 breast invasive carcinoma (p-value= 1.66x10-42), 483 colon adenocarcinoma (p-value= 9x10-18), 541 lung adenocarcinoma (p-value=3.16x10-32), 502 lung squamous carcinoma (p-value= 1.49x10-59), and 502 prostate adenocarcinoma (p-value= 1.53x10-4) patients were compared to healthy individuals and showed significant HPRT over-expression shifts in malignant tumors. To continue this investigation we obtained histological tissue from 52 breast cancer patients, 54 lung cancer patients, 100 colorectal cancer patients, and 56 prostate cancer patients with a varying level of cancer stage and tumor type. Healthy tissue, margins of carcinoma, and pre-cancerous tissues were also stained to determine stage dependence of HPRT expression. Briefly, tissues were treated with a monoclonal anti-HPRT antibody along with a GAPDH positive control and an isotope control. Tissues were incubated with an HRP-polymer conjugated anti-HPRT antibody and followed by a diaminobenzidine (DAB) substrate which, when oxidized, results in antigen labelling. Tissues were imaged and analyzed using ImageJ software, which converted images to a grayscale. From there, we set a parameter for tissues with “HPRT high” expression, and “HPRT low” expression. Overall, we found 33-55% of malignant tissues to have a significant upregulation of HPRT (Lung-33%, Breast-55%, Colon-33%, Prostate-47%). These findings were consistent with our examination of HPRT expression in TCGA and our findings also indicated that the protein over-expression is not dependent on stage. These findings indicate that HPRT may potentially be a valuable biomarker in detecting early cases of malignancy in all of the four major cancers. Citation Format: Michelle H. Passey, Abigail M. Felsted, Zachary E. Ence, Stephen R. Piccolo, Kim L. O'Neill, Richard A. Robison. Unique HPRT1 upregulation in malignant tissue: Potential use as diagnostic biomarker [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2536.

Command-line software plays a critical role in biology research. However, processes for installing and executing software differ widely. The Common Workflow Language (CWL) is a community standard that addresses this problem. Using CWL, tool developers can formally describe a tool’s inputs, outputs, and other execution details. CWL documents can include instructions for executing tools inside software containers. Accordingly, CWL tools are portable—they can be executed on diverse computers—including personal workstations, high-performance clusters, or the cloud. CWL also supports workflows, which describe dependencies among tools and using outputs from one tool as inputs to others. To date, CWL has been used primarily for batch processing of large datasets, especially in genomics. But it can also be used for analytical steps of a study. This article explains key concepts about CWL and software containers and provides examples for using CWL in biology research. CWL documents are text-based, so they can be created manually, without computer programming. However, ensuring that these documents conform to the CWL specification may prevent some users from adopting it. To address this gap, we created ToolJig, a Web application that enables researchers to create CWL documents interactively. ToolJig validates information provided by the user to ensure it is complete and valid. After creating a CWL tool or workflow, the user can create ‘input-object’ files, which store values for a particular invocation of a tool or workflow. In addition, ToolJig provides examples of how to execute the tool or workflow via a workflow engine. ToolJig and our examples are available at https://github.com/srp33/ToolJig.

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