HPRT1

Gene Summary

Gene:HPRT1; hypoxanthine phosphoribosyltransferase 1
Aliases: HPRT, HGPRT
Location:Xq26.2-q26.3
Summary:The protein encoded by this gene is a transferase, which catalyzes conversion of hypoxanthine to inosine monophosphate and guanine to guanosine monophosphate via transfer of the 5-phosphoribosyl group from 5-phosphoribosyl 1-pyrophosphate. This enzyme plays a central role in the generation of purine nucleotides through the purine salvage pathway. Mutations in this gene result in Lesch-Nyhan syndrome or gout.[provided by RefSeq, Jun 2009]
Databases:OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:hypoxanthine-guanine phosphoribosyltransferase
Source:NCBIAccessed: 29 August, 2019

Ontology:

What does this gene/protein do?
Show (35)
Pathways:What pathways are this gene/protein implicaed in?
Show (1)

Cancer Overview

Research Indicators

Publications Per Year (1994-2019)
Graph generated 29 August 2019 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • fas Receptor
  • Telomere
  • Thymidine Kinase
  • Smoking
  • p53 Protein
  • Y Chromosome
  • Cervical Cancer
  • Xeroderma Pigmentosum
  • Remission Induction
  • Hypoxanthine Phosphoribosyltransferase
  • Bladder Cancer
  • Wilms Tumour
  • Restriction Fragment Length Polymorphism
  • Xanthine Oxidase
  • Mutation
  • Vesicular stomatitis Indiana virus
  • Tioguanine
  • Plasmacytoma
  • Vaginal Smears
  • Repetitive Sequences, Nucleic Acid
  • alpha 1-Antitrypsin
  • X Chromosome
  • Whole-Body Irradiation
  • Topoisomerase II Inhibitors
  • RNA Splicing
  • Skin Cancer
  • Proto-Oncogenes
  • X-Rays
  • Oncogenes
  • Xeroderma Pigmentosum Group A Protein
  • Polymorphism
  • Polymerase Chain Reaction
  • Pyrenes
  • RNA-Binding Proteins
  • Species Specificity
  • Ultraviolet Rays
  • Risk Factors
  • Sex Chromosomes
  • T-Cell Antigen Receptors
  • Transferases
Tag cloud generated 29 August, 2019 using data from PubMed, MeSH and CancerIndex

Specific Cancers (4)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: HPRT1 (cancer-related)

Rot S, Taubert H, Bache M, et al.
Low HIF-1α and low EGFR mRNA Expression Significantly Associate with Poor Survival in Soft Tissue Sarcoma Patients; the Proteins React Differently.
Int J Mol Sci. 2018; 19(12) [PubMed] Free Access to Full Article Related Publications
In various tumors, the hypoxia inducible factor-1α (

Feng L, Yang Y, Li M, et al.
Systems biology analysis of the lung cancer‑related secretome.
Oncol Rep. 2018; 40(2):1103-1118 [PubMed] Related Publications
Tumorigenesis is closely and highly associated with developmental biology. The present study aimed to discover and identify marker proteins strongly associated with the occurrence and development of non‑small cell lung cancer (NSCLC) in humans and to provide new ideas for investigating lung cancer markers by combining biological analyses of embryonic development. We established primary cultures for samples of tumor and control tissues from 9 patients with NSCLC and collected conditioned medium (CM). Subsequently, we used liquid chromatography and linear ion trap (LTQ) mass spectrometry to isolate and identify proteins in CM samples. Data mining of free proteins was conducted using the analogous analysis strategy in systems biology to obtain important lung cancer‑associated proteins (plasma markers). Proteins with significant plasma enrichment in lung cancer patients were detected via enzyme‑linked immunosorbent assay (ELISA). We identified 987 high‑confidence proteins and established a primary database of free proteins associated with lung cancer. Furthermore, 511 high‑confidence proteins were present in CM from primary tissue cultures from at least 2 of the 9 examined cases of lung cancer. Analysis using Gene Set Enrichment Analysis (GSEA) software revealed significant enrichment for 197 proteins from the CM of lung cancer samples in maternal‑placental interface expression profiles for a mid‑term placenta with strong invasiveness relative to RNA expression profiles for a human full‑term placenta after delivery. ELISA results demonstrated that hypoxanthine phosphoribosyltransferase 1 (HPRT1) was associated with worse rates of disease‑free survival (DFS) and overall survival (OS). The biological behaviors of embryonic implantation are similar to those of tumor invasion and metastasis, and the information obtained regarding developmental biology could facilitate the interpretation of tumor invasion and metastasis. Therefore, similar biological behaviors combined with analyses at different molecular levels from the perspective of systems biology will provide new ideas for tumor research.

Janik ME, Szwed S, Grzmil P, et al.
RT-qPCR analysis of human melanoma progression-related genes - A novel workflow for selection and validation of candidate reference genes.
Int J Biochem Cell Biol. 2018; 101:12-18 [PubMed] Related Publications
The objective of this study was to identify a normalizer or combination of normalizers for quantitative evaluation of the expression of a target gene of interest during melanoma progression. Adult melanocytes, uveal primary melanoma cells and cutaneous primary and metastatic melanoma cells were used to construct a panel of 14 experimental models reflecting cancer promotion and progression. Hypoxanthine phosphoribosyltransferase 1 (HPRT1), glucuronidase beta (GUSB), ribosomal protein S23 (RPS23), phosphoglycerate kinase 1 (PGK1) and small nuclear ribonucleoprotein progression. Adult melanocytes, uveal primary melanoma cells and cutaneous primary and metastatic melanoma cells were used to construct a panel of 14 experimental models reflecting cancer promotion and progression. Hypoxanthine phosphoribosyltransferase 1 (HPRT1), glucuronidase beta (GUSB), ribosomal protein S23 (RPS23), phosphoglycerate kinase 1 (PGK1) and small nuclear ribonucleoprotein polypeptide A (SRNPA) were chosen as candidate housekeeping genes. NormFinder software was used to identify the best reference gene or pair of reference genes from five candidate housekeeping genes, on the basis of expression stability in a given experimental model. The suitability of references was validated by normalizing the transcriptional activities of E-cadherin (CDH1), N-cadherin (CDH2) and endoplasmic reticulum aminopeptidase 1 (ERAP1) target genes. It has been shown that the relative expression of CDH2 and ERAP1 target genes in a given cell line may vary between experimental models, leading to biological misinterpretation. In view of this, we devised a strategy for improved selection of the best stable reference and for obtaining biologically consistent results. This strategy avoided experimental model- and normalizer-dependent conclusions concerning the relative expression of target gene, in the examined cell lines.

Townsend MH, Robison RA, O'Neill KL
A review of HPRT and its emerging role in cancer.
Med Oncol. 2018; 35(6):89 [PubMed] Related Publications
Hypoxanthine guanine phosphoribosyltransferase (HPRT) is a common salvage housekeeping gene with a historically important role in cancer as a mutational biomarker. As an established and well-known human reporter gene for the evaluation of mutational frequency corresponding to cancer development, HPRT is most commonly used to evaluate cancer risk within individuals and determine potential carcinogens. In addition to its use as a reporter gene, HPRT also has important functionality in the body in relation to purine regulation as demonstrated by Lesch-Nyhan patients whose lack of functional HPRT leads to significant purine overproduction and further neural complications. This regulatory role, in addition to an established connection between other salvage enzymes and cancer development, points to HPRT as an emerging influence in cancer. Recent work has shown that not only is the enzyme upregulated within malignant tumors, it also has significant surface localization within some cancer cells. With this is mind, HPRT has the potential to become a significant biomarker not only for the characterization of cancer, but also for its potential treatment.

Ahmed EM, Bandopadhyay G, Coyle B, Grabowska A
A HIF-independent, CD133-mediated mechanism of cisplatin resistance in glioblastoma cells.
Cell Oncol (Dordr). 2018; 41(3):319-328 [PubMed] Free Access to Full Article Related Publications
PURPOSE: Glioblastoma (GBM) is the commonest brain tumour in adults. A sub-population of cells within these tumours, known as cancer stem cells (CSCs), is thought to mediate their chemo-/radiotherapy resistance. CD133 is a cell surface marker that is used to identify and isolate GBM CSCs. However, its functional significance, as well as the relevant microenvironment in which to study CD133, have so far remained unknown. Here, we examined the effect of hypoxia on the expression of CD133 and on that of the hypoxia-related factors HIF-1α and HIF-2α, and the potential functional significance of CD133 expression on the acquisition of chemo-resistance by GBM cells.
METHODS: CD133, HIF-1α, HIF-2α, VEFG and (control) HPRT mRNA expression analyses were carried out on GBM cells (U251, U87 and SNB19; 2D or 3D cultures) under both normoxic and hypoxic conditions, using qRT-PCR. siRNA was used to downregulate CD133, HIF-1α and HIF-2α expression in the GBM cells, which was confirmed by flow cytometry and qRT-PCR, respectively. Drug sensitivity-related IC50 values were established using an Alamar Blue cell viability assay in conjunction with the Graphpad prism software tool.
RESULTS: We found that the expression of CD133 was upregulated under hypoxic conditions in both the 2D and 3D GBM cell culture models. In addition, an increased resistance to cisplatin, temozolomide and etoposide was observed in the GBM cells cultured under hypoxic conditions compared to normoxic conditions. siRNA-mediated knockdown of either HIF-1α or HIF-2α resulted in a reduced CD133 expression, with HIF-2α having a more long-term effect. We also found that HIF-2α downregulation sensitized the GBM cells to cisplatin to a greater extent than HIF-1α, whereas CD133 knockdown had a more marked effect on cisplatin sensitisation than knockdown of either one of the HIFs, suggesting the existence of a HIF-independent cisplatin resistance mechanism mediated by CD133. This same mechanism does not seem to be involved in temozolomide resistance, since we found that HIF-1α downregulation, but not HIF-2α or CD133 downregulation, sensitized GBM cells to temozolomide.
CONCLUSIONS: From our data we conclude that the mechanisms underlying hypoxia-induced CD133-mediated cisplatin resistance may be instrumental for the design of new GBM treatment strategies.

Zhao H, Ma TF, Lin J, et al.
Identification of valid reference genes for mRNA and microRNA normalisation in prostate cancer cell lines.
Sci Rep. 2018; 8(1):1949 [PubMed] Free Access to Full Article Related Publications
RT-qPCR offers high sensitivity, for accurate interpretations of qPCR results however, normalisation using suitable reference genes is fundamental. Androgens can regulate transcriptional expression including reference gene expression in prostate cancer. In this study, we evaluated ten mRNA and six non-protein coding RNA reference genes in five prostate cell lines under varied dihydrotestosterone (DHT) treatments. We validated the effects of DHT-treatments using media containing charcoal-stripped serum prior to DHT stimulation on the test samples by Western blot experiments. Reference gene expression stability was analysed using three programs (geNorm, NormFinder and BestKeeper), and the recommended comprehensive ranking is provided. Our results reveal that ACTB and GAPDH, and miR-16 and miR-1228-3p are the most suitable mRNA and miRNA reference genes across all cell lines, respectively. Considering prostate cancer cell types, ACTB/GAPDH and ACTB/HPRT1 are the most suitable reference gene combinations for mRNA analysis, and miR-16/miR-1228-3p and RNU6-2/RNU43 for miRNA analysis in AR+, and AR- and normal cell lines, respectively. Comparison of relative target gene (PCA3 and miR-141) expression reveals different patterns depending on reference genes used for normalisation. To our knowledge, this is the first report on validation of reference genes under different DHT treatments in prostate cancer cells. This study provides insights for discovery of reliable DHT-regulated genes in prostate cells.

Liu Y, Qin Z, Cai L, et al.
Selection of internal references for qRT-PCR assays of human hepatocellular carcinoma cell lines.
Biosci Rep. 2017; 37(6) [PubMed] Free Access to Full Article Related Publications
Selecting internal references is important for normalizing the loading quantity of samples in quantitative reverse-transcription PCR (qRT-PCR). In the present study, a systematic evaluation of reference genes among nine hepatocellular carcinoma (HCC) cell lines was conducted. After screening the microarray assay data of ten HCC cell lines, 19 candidate reference genes were preselected and then evaluated by qRT-PCR, together with

Markou A, Lazaridou M, Paraskevopoulos P, et al.
Multiplex Gene Expression Profiling of In Vivo Isolated Circulating Tumor Cells in High-Risk Prostate Cancer Patients.
Clin Chem. 2018; 64(2):297-306 [PubMed] Related Publications
BACKGROUND: Molecular characterization of circulating tumor cells (CTCs) is important for selecting patients for targeted treatments. We present, for the first time, results on gene expression profiling of CTCs isolated in vivo from high-risk prostate cancer (PCa) patients compared with CTC detected by 3 protein-based assays-CellSearch
METHODS: EpCAM-positive CTCs were isolated in vivo using the CellCollector from 108 high-risk PCa patients and 36 healthy volunteers. For 27 patients, samples were available before and after treatment. We developed highly sensitive multiplex RT-qPCR assays for 14 genes (
RESULTS: We observed high heterogeneity in gene expression in the captured CTCs for each patient. At least 1 marker was detected in 74 of 105 patients (70.5%), 2 markers in 45 of 105 (40.9%), and 3 markers in 16 of 105 (15.2%). Epithelial markers were detected in 31 of 105 (29.5%) patients, EMT markers in 46 of 105 (43.8%), and stem cell markers in 15 of 105 (14.3%) patients. EMT-marker positivity was very low before therapy (2 of 27, 7.4%), but it increased after therapy (17 of 27, 63.0%), whereas epithelial markers tended to decrease after therapy (2 of 27, 7.4%) compared with before therapy (13 of 27, 48.1%). At least 2 markers were expressed in 40.9% of patients, whereas the positivity was 19.6% for CellSearch, 38.1% for EPISPOT, and 43.8% for CellCollector-based IF-staining.
CONCLUSIONS: The combination of in vivo CTC isolation with downstream RNA analysis is highly promising as a high-throughput, specific, and ultrasensitive approach for multiplex liquid biopsy-based molecular diagnostics.

Xu W, Foster BA, Richards M, et al.
Characterization of prostate cancer cell progression in zebrafish xenograft model.
Int J Oncol. 2018; 52(1):252-260 [PubMed] Free Access to Full Article Related Publications
Early diagnosis of prostate cancer (PCa) is critical for the application of efficient treatment to PCa patients. However, the majority of PCas remains indolent from several months to several years before malignancy. Current diagnosis methods have limitations in their reliability and are inefficient in time cost. Thus, an efficient in vivo PCa cell xenograft model is highly desired for diagnostic studies in PCas. In the present study we present a standardized procedure to create a PCa cell xenograft model using zebrafish (Danio rerio) as the host. PC3-CTR cells, a cell line from adenocarcinoma with stable expression of calcitonin receptor (CRT), were subcutaneously injected into zebrafish larvae at 48 h post fertilization. The nursing conditions for the larvae were optimized with stable survival rates of post hatch and post PC3-CTR cell injection. In this system, the progression of PC3-CTR cells in vivo was evaluated by migration and proliferation of the cells. Massive migrations of PC3 cells in vivo were observed at post injection day (PID)3. The injected PC3-CTR cells eventually invaded the whole larval zebrafish at PID5. Quantification of PC3-CTR cell proliferation was done using quantitative PCR (qPCR) analysis targeting the expression profiles of two PCa housekeeping genes, TATA-binding protein (TBP) and hypoxanthine phosphoribosyltransferase 1 (HPRT1) encoding genes. The excessive proliferation of PC3 cells in vivo was detected with both qPCR assays. Expression levels of one non‑coding gene, prostate cancer associated 3 gene (pca3), and two other genes encoding transient receptor potential ion channel Melastatin 8 (trpm8) and prostate-specific membrane antigen (psma), showed a significantly enhanced aggressiveness of PC3-CTR cells in vivo. The model established in the present study provides an improved in vivo model for the diagnosis of PCas efficiently. This PCa cell xenograft model can also serve as a tool for high throughput anti-PCa drug screening in therapeutic treatments.

Freitag D, Koch A, Lawson McLean A, et al.
Validation of Reference Genes for Expression Studies in Human Meningiomas under Different Experimental Settings.
Mol Neurobiol. 2018; 55(7):5787-5797 [PubMed] Related Publications
Quantitative polymerase chain reaction (qPCR) is a sensitive technique for the quantitative analysis of gene expression levels. To compare mRNA transcripts across tumour and non-pathological tissue, appropriate reference genes are required for internal standardisation. Validation of these reference genes in meningiomas has not yet been reported. After mRNA transcription of meningioma (WHO grade I-III) and meningeal tissue from three different experimental sample types (fresh tissue, primary cell cultures and FFPE tissue), 13 candidate reference genes (ACTB, B2M, HPRT, VIM, GAPDH, YWHAZ, EIF4A2, MUC1, ATP5B, GNB2L, TUBB, CYC1, RPL13A) were chosen for quantitative expression analysis. Two statistical algorithms (GeNorm and NormFinder) were used for validation of gene expression stability. All candidate housekeepers tested for stability were checked within and across the three tissue analysis groups. Pearson correlation, the ΔC

de Campos RP, Schultz IC, de Andrade Mello P, et al.
Cervical cancer stem-like cells: systematic review and identification of reference genes for gene expression.
Cell Biol Int. 2018; 42(2):139-152 [PubMed] Related Publications
Cervical cancer is the fourth most common cancer affecting women worldwide. Among many factors, the presence of cancer stem cells, a subpopulation of cells inside the tumor, has been associated with a worse prognosis. Considering the importance of gene expression studies to understand the biology of cervical cancer stem cells (CCSC), this work identifies stable reference genes for cervical cancer cell lines SiHa, HeLa, and ME180 as well as their respective cancer stem-like cells. A literature review was performed to identify validated reference genes currently used to normalize RT-qPCR data in cervical cancer cell lines. Then, cell lines were cultured in regular monolayer or in a condition that favors tumor sphere formation. RT-qPCR was performed using five reference genes: ACTB, B2M, GAPDH, HPRT1, and TBP. Stability was assessed to validate the selected genes as suitable reference genes. The evaluation validated B2M, GAPDH, HPRT1, and TBP in these experimental conditions. Among them, GAPDH and TBP presented the lowest variability according to the analysis by Normfinder, Bestkeeper, and ΔC

Kyriakou IK, Mavridis K, Kalogianni DP, et al.
Multianalyte quantitative competitive PCR on optically encoded microspheres for an eight-gene panel related to prostate cancer.
Anal Bioanal Chem. 2018; 410(3):971-980 [PubMed] Related Publications
Nucleic acid-based tests have a profound impact in every medical discipline. Because multigene tests offer higher diagnostic accuracy and lower overall cost than single assays, they are especially useful for diseases, like prostate cancer, that present variability at the molecular level and diversity of available therapeutic interventions. We have developed a quantitative competitive PCR for an eight-gene panel, related to prostate cancer, that includes five genes of the human tissue kallikrein family (KLKs), prostate-specific membrane antigen (PSMA), prostate cancer antigen 3 (PCA3), and HPRT1 as a reference gene. Using PCR as a synthetic tool, a competitor was prepared for each target sequence containing the same primer binding sites as the target but differing in a short segment to enable discrimination by hybridization. The assay involves multiplex amplification of targets and competitors followed by a multiplex hybridization assay for the 16 amplification products. The assay was performed on optically encoded microspheres with oligonucleotide probes attached to their surface. The microspheres were analyzed rapidly (1 min) by flow cytometry. The signal ratio of the target and cognate competitor is a function of the target copy number in the sample prior to amplification. The multiplexing potential of the proposed method is much higher than real-time PCR and other end-point methods since there are 100 sets of commercially available microspheres.

Lv Y, Zhao SG, Lu G, et al.
Identification of reference genes for qRT-PCR in granulosa cells of healthy women and polycystic ovarian syndrome patients.
Sci Rep. 2017; 7(1):6961 [PubMed] Free Access to Full Article Related Publications
Comparative gene expression analysis by qRT-PCR is commonly used to detect differentially expressed genes in studies of PCOS pathology. Impaired GC function is strongly associated with PCOS pathogenesis, and a growing body of studies has been dedicated to identifying differentially expressed genes in GCs in PCOS patients and healthy women by qRT-PCR. It is necessary to validate the expression stability of the selected reference genes across the tested samples for target gene expression normalization. We examined the variability and stability of expression of the 15 commonly used reference genes in GCs from 44 PCOS patients and 45 healthy women using the GeNorm, BestKeeper, and NormFinder statistical algorithms. We combined the rankings of the three programs to produce a final ranking based on the geometric means of their stability scores. We found that HPRT1, RPLP0, and HMBS out of 15 examined commonly used reference genes are stably expressed in GCs in both controls and PCOS patients and can be used for normalization in gene expression profiling by qRT-PCR. Future gene-expression studies should consider using these reference genes in GCs in PCOS patients for more accurate quantitation of target gene expression and data interpretation.

Moltó-García R, González-Alonso V, Villaverde-Doménech ME, Novella-Maestre E
Effect of Human Fat Graft on Breast Cancer Metastasis in a Murine Model.
Plast Reconstr Surg. 2017; 139(5):1119-1128 [PubMed] Related Publications
BACKGROUND: Isolated adipose stem cells have been reported to encourage migration and early metastasis of breast cancer. Mimicking a surgical situation, the authors developed a human breast cancer model to evaluate in vivo whether human adipose tissue promotes tumor growth and invasion.
METHODS: Human adipose tissue was obtained from four patients. The MDA-MB-468 cell line was cultured with a lentiviral vector encoding a puromycin resistance gene and mCherry fluorescent protein. Virus-infected cells were selected. Animals were injected in the left renal capsule and divided into three experimental groups: group A, MDA-MB-468 cells (n = 4); group B, MDA-MB-468 cells/human adipose tissue (n = 4); and group C, Dulbecco's Modified Eagle Medium/F-12 medium (negative control, n = 4). Metastatic development was monitored using an in vivo imaging system. Small breast epithelial mucin (SBEM), human hypoxanthine-guanine phosphoribosyltransferase (HPRTh), and murine hypoxanthine-guanine phosphoribosyltransferase (HPRTm) expression were analyzed by real-time polymerase chain reaction to detect multifocal metastases in right/left renal capsule, liver, spleen, and pancreas.
RESULTS: Metastasis was observed between postinjection days 37 and 44. No significant differences were found in survival rates between groups (group A, 157 ± 42.60 days; group B, 169 ± 40.17 days). All samples expressed HPRTm. HPRTh and SBEM were expressed in left renal capsules from all group A and B mice, whereas in spleen, liver, pancreas, and right renal capsule the HPRTm and SBEM expression was not constant in all samples of group A and B mice. Differences were found between groups in HPRTh and SBEM expression but were not statistically significant.
CONCLUSION: Human adipose tissue used to restore breast defects after oncologic resection did not increase metastasis development risk when there were residual breast cancer cells in proximity.

Mertz TM, Baranovskiy AG, Wang J, et al.
Nucleotide selectivity defect and mutator phenotype conferred by a colon cancer-associated DNA polymerase δ mutation in human cells.
Oncogene. 2017; 36(31):4427-4433 [PubMed] Free Access to Full Article Related Publications
Mutations in the POLD1 and POLE genes encoding DNA polymerases δ (Polδ) and ɛ (Polɛ) cause hereditary colorectal cancer (CRC) and have been found in many sporadic colorectal and endometrial tumors. Much attention has been focused on POLE exonuclease domain mutations, which occur frequently in hypermutated DNA mismatch repair (MMR)-proficient tumors and appear to be responsible for the bulk of genomic instability in these tumors. In contrast, somatic POLD1 mutations are seen less frequently and typically occur in MMR-deficient tumors. Their functional significance is often unclear. Here we demonstrate that expression of the cancer-associated POLD1-R689W allele is strongly mutagenic in human cells. The mutation rate increased synergistically when the POLD1-R689W expression was combined with a MMR defect, indicating that the mutator effect of POLD1-R689W results from a high rate of replication errors. Purified human Polδ-R689W has normal exonuclease activity, but the nucleotide selectivity of the enzyme is severely impaired, providing a mechanistic explanation for the increased mutation rate in the POLD1-R689W-expressing cells. The vast majority of mutations induced by the POLD1-R689W are GC→︀TA transversions and GC→︀AT transitions, with transversions showing a strong strand bias and a remarkable preference for polypurine/polypyrimidine sequences. The mutational specificity of the Polδ variant matches that of the hypermutated CRC cell line, HCT15, in which this variant was first identified. The results provide compelling evidence for the pathogenic role of the POLD1-R689W mutation in the development of the human tumor and emphasize the need to experimentally determine the significance of Polδ variants present in sporadic tumors.

He J, Rong Z, Fu X, Xu Y
A Safety Checkpoint to Eliminate Cancer Risk of the Immune Evasive Cells Derived from Human Embryonic Stem Cells.
Stem Cells. 2017; 35(5):1154-1161 [PubMed] Related Publications
Human embryonic stem cells (hESCs) hold great promise in the regenerative therapy of many currently untreatable human diseases. One of the key bottlenecks is the immune rejection of hESC-derived allografts by the recipient. To overcome this challenge, we have established new approaches to induce immune protection of hESC-derived allografts through the coexpression of immune suppressive molecules CTLA4-Ig and PD-L1. However, this in turn raises a safety concern of cancer risk because these hESC-derived cells can evade immune surveillance. To address this safety concern, we developed a safety checkpoint so that the immune evasive hESC-derived cells in the graft can be effectively eliminated if any cellular transformation is detected. In this context, we knock-in the suicidal gene herpes simplex virus thymidine kinase (HSVTK) into the constitutive HPRT locus of CP hESCs (knock-in hESCs expressing CTLA4-Ig and PD-L1), denoted CPTK hESCs. Employing humanized mice (Hu-mice) reconstituted with human immune system, we demonstrated that the CPTK hESC-derived cells are protected from immune rejection. In addition, CPTK hESC-derived cells can be efficiently eliminated in vitro and in vivo with FDA approved TK-targeting drug ganciclovir. Therefore, this new safety checkpoint improves the feasibility to use the immune evasive hESC-derived cells for regenerative medicine. Stem Cells 2017;35:1154-1161.

Freire JM, Rego de Figueiredo I, Valle J, et al.
siRNA-cell-penetrating peptides complexes as a combinatorial therapy against chronic myeloid leukemia using BV173 cell line as model.
J Control Release. 2017; 245:127-136 [PubMed] Related Publications
Chronic myeloid leukemia (CML) is a myeloproliferative disorder caused by a single gene mutation, a reciprocal translocation that originates the Bcr-Abl gene with constitutive tyrosine kinase activity. As a monogenic disease, it is an optimum target for RNA silencing therapy. We developed a siRNA-based therapeutic approach in which the siRNA is delivered by pepM or pepR, two cell-penetrating peptides (CPPs) derived from the dengue virus capsid protein. These peptides have a dual role: siRNA delivery into cells and direct action as bioportides, i.e. intracellularly bioactive CPPs, targetting cancer-related signaling processes. Both pepM and pepR penetrate the positive Bcr-Abl

Lima L, Gaiteiro C, Peixoto A, et al.
Reference Genes for Addressing Gene Expression of Bladder Cancer Cell Models under Hypoxia: A Step Towards Transcriptomic Studies.
PLoS One. 2016; 11(11):e0166120 [PubMed] Free Access to Full Article Related Publications
Highly aggressive, rapidly growing tumors contain significant areas of hypoxia or anoxia as a consequence of inadequate and/or irregular blood supply. During oxygen deprivation, tumor cells withstand a panoply of adaptive responses, including a shift towards anaerobic metabolism and the reprogramming of the transcriptome. One of the major mediators of the transcriptional hypoxic response is the hypoxia-inducible factor 1 (HIF-1), whose stabilization under hypoxia acts as an oncogenic stimulus contributing to chemotherapy resistance, invasion and metastasis. Gene expression analysis by qRT-PCR is a powerful tool for cancer cells phenotypic characterization. Nevertheless, as cells undergo a severe transcriptome remodeling.in response to oxygen deficit, the precise identification of reference genes poses a significant challenge for hypoxic studies. Herein, we aim to establish the best reference genes for studying the effects of hypoxia on bladder cancer cells. Accordingly, three bladder cancer cell lines (T24, 5637, and HT1376) representative of two distinct carcinogenesis pathways to invasive cancer (FGFR3/CCND1 and E2F3/RB1) were used. Additionally, we have explored the most suitable control gene when addressing the influence of Deferoxamine Mesilate salt (DFX), an iron chelator often used to avoid the proteasomal degradation of HIF-1α, acting as an hypoxia-mimetic agent. Using bioinformatics tools (GeNorm and NormFinder), we have elected B2M and HPRT as the most stable genes for all cell lines and experimental conditions out of a panel of seven putative candidates (HPRT, ACTB, 18S, GAPDH, TBP, B2M, and SDHA). These observations set the molecular basis for future studies addressing the effect of hypoxia and particularly HIF-1α in bladder cancer cells.

Thumfart J, Weschke B, Ringe H, et al.
Acute renal failure unmasking Lesch-Nyhan disease in a patient with tuberous sclerosis complex.
Eur J Paediatr Neurol. 2016; 20(4):649-51 [PubMed] Related Publications
CASE REPORT: We report on a male patient with Tuberous Sclerosis Complex (TSC), which was prenatally diagnosed. At the age of 3 months the patient developed acute renal failure with excessive hyperuricemia. Kidney function improved after rehydration and application of rasburicase, however without full recovery. Due to the inappropriate high levels of uric acid compared to kidney function, screening of hypoxanthine-guanine phosphoribosyltransferase (HPRT) related diseases was initiated. Mutation analysis revealed a deletion of exon 2 and 3 of the HPRT gene confirming the diagnosis of Lesch-Nyhan Disease (LND). After initiation of allopurinol therapy renal function further improved. In the following months the patient developed clinically a typical neurological phenotype of LND and TSC with seizures, severe dystonia and developmental delay.
CONCLUSION: Acute renal failure is a rare complication of HPRT related diseases. Combination of two inherited diseases may lead to a delayed diagnosis due to a mixed and maybe misleading phenotype.

Zu X, Yan R, Pan J, et al.
Aldo-keto reductase 1B10 protects human colon cells from DNA damage induced by electrophilic carbonyl compounds.
Mol Carcinog. 2017; 56(1):118-129 [PubMed] Related Publications
Electrophilic carbonyl compounds are highly cytotoxic and genotoxic. Aldo-keto reductase 1B10 (AKR1B10) is an enzyme catalyzing reduction of carbonyl compounds to less toxic alcoholic forms. This study presents novel evidence that AKR1B10 protects colon cells from DNA damage induced by electrophilic carbonyl compounds. AKR1B10 is specifically expressed in epithelial cells of the human colon, but this study found that AKR1B10 expression was lost or markedly diminished in colorectal cancer, precancerous tissues, and a notable portion of normal adjacent tissues (NAT). SiRNA-mediated silencing of AKR1B10 in colon cancer cells HCT-8 enhanced cytotoxicity of acrolein and HNE, whereas ectopic expression of AKR1B10 in colon cancer cells RKO prevented the host cells against carbonyl cytotoxicity. Furthermore, siRNA-mediated AKR1B10 silencing led to DNA breaks and activation of γ-H2AX protein, a marker of DNA double strand breaks, particularly in the exposure of HNE (10 μM). In the AKR1B10 silenced HCT-8 cells, hypoxanthine-guanine phosphoribosyl transferase (HPRT) mutant frequency increased by 26.8 times at basal level and by 33.5 times in the presence of 10 μM HNE when compared to vector control cells. In these cells, the cyclic acrolein-deoxyguanosine adducts levels were increased by over 10 times. These findings were confirmed by pharmacological inhibition of AKR1B10 activity by Epalrestat. Taken together, these data suggest that AKR1B10 is a critical protein that protects host cells from DNA damage induced by electrophilic carbonyl compounds. AKR1B10 deficiency in the colon may be an important pathogenic factor in disease progression and carcinogenesis. © 2016 Wiley Periodicals, Inc.

Choudhary A, Zachek B, Lera RF, et al.
Identification of Selective Lead Compounds for Treatment of High-Ploidy Breast Cancer.
Mol Cancer Ther. 2016; 15(1):48-59 [PubMed] Free Access to Full Article Related Publications
Increased ploidy is common in tumors but treatments for tumors with excess chromosome sets are not available. Here, we characterize high-ploidy breast cancers and identify potential anticancer compounds selective for the high-ploidy state. Among 354 human breast cancers, 10% have mean chromosome copy number exceeding 3, and this is most common in triple-negative and HER2-positive types. Women with high-ploidy breast cancers have higher risk of recurrence and death in two patient cohorts, demonstrating that it represents an important group for improved treatment. Because high-ploidy cancers are aneuploid, rather than triploid or tetraploid, we devised a two-step screen to identify selective compounds. The screen was designed to assure both external validity on diverse karyotypic backgrounds and specificity for high-ploidy cell types. This screen identified novel therapies specific to high-ploidy cells. First, we discovered 8-azaguanine, an antimetabolite that is activated by hypoxanthine phosphoribosyltransferase 1 (HPRT1), suggesting an elevated gene-dosage of HPRT1 in high-ploidy tumors can control sensitivity to this drug. Second, we discovered a novel compound, 2,3-diphenylbenzo[g]quinoxaline-5,10-dione (DPBQ). DPBQ activates p53 and triggers apoptosis in a polyploid-specific manner, but does not inhibit topoisomerase or bind DNA. Mechanistic analysis demonstrates that DPBQ elicits a hypoxia gene signature and its effect is replicated, in part, by enhancing oxidative stress. Structure-function analysis defines the core benzo[g]quinoxaline-5,10 dione as being necessary for the polyploid-specific effects of DPBQ. We conclude that polyploid breast cancers represent a high-risk subgroup and that DPBQ provides a functional core to develop polyploid-selective therapy. Mol Cancer Ther; 15(1); 48-59. ©2015 AACR.

Sharan RN, Vaiphei ST, Nongrum S, et al.
Consensus reference gene(s) for gene expression studies in human cancers: end of the tunnel visible?
Cell Oncol (Dordr). 2015; 38(6):419-31 [PubMed] Related Publications
BACKGROUND: Gene expression studies are increasingly used to provide valuable information on the diagnosis and prognosis of human cancers. Also, for in vitro and in vivo experimental cancer models gene expression studies are widely used. The complex algorithms of differential gene expression analyses require normalization of data against a reference or normalizer gene, or a set of such genes. For this purpose, mostly invariant housekeeping genes are used. Unfortunately, however, there are no consensus (housekeeping) genes that serve as reference or normalizer for different human cancers. In fact, scientists have employed a wide range of reference genes across different types of cancer for normalization of gene expression data. As a consequence, comparisons of these data and/or data harmonizations are difficult to perform and challenging. In addition, an inadequate choice for a reference gene may obscure genuine changes and/or result in erroneous gene expression data comparisons.
METHODS: In our effort to highlight the importance of selecting the most appropriate reference gene(s), we have screened the literature for gene expression studies published since the turn of the century on thirteen of the most prevalent human cancers worldwide.
CONCLUSIONS: Based on the analysis of the data at hand, we firstly recommend that in each study the suitability of candidate reference gene(s) should carefully be evaluated in order to yield reliable differential gene expression data. Secondly, we recommend that a combination of PPIA and either GAPDH, ACTB, HPRT and TBP, or appropriate combinations of two or three of these genes, should be employed in future studies, to ensure that results from different studies on different human cancers can be harmonized. This approach will ultimately increase the depth of our understanding of gene expression signatures across human cancers.

Manzotti G, Parenti S, Ferrari-Amorotti G, et al.
Monocyte-macrophage differentiation of acute myeloid leukemia cell lines by small molecules identified through interrogation of the Connectivity Map database.
Cell Cycle. 2015; 14(16):2578-89 [PubMed] Free Access to Full Article Related Publications
The transcription factor C/EBPα is required for granulocytic differentiation of normal myeloid progenitors and is frequently inactivated in acute myeloid leukemia (AML) cells. Ectopic expression of C/EBPα in AML cells suppresses proliferation and induces differentiation suggesting that restoring C/EBPα expression/activity in AML cells could be therapeutically useful. Unfortunately, current approaches of gene or protein delivery in leukemic cells are unsatisfactory. However, "drug repurposing" is becoming a very attractive strategy to identify potential new uses for existing drugs. In this study, we assessed the biological effects of candidate C/EBPα-mimetics identified by interrogation of the Connectivity Map database. We found that amantadine, an antiviral and anti-Parkinson agent, induced a monocyte-macrophage-like differentiation of HL60, U937, Kasumi-1 myeloid leukemia cell lines, as indicated by morphology and differentiation antigen expression, when used in combination with suboptimal concentration of all trans retinoic acid (ATRA) or Vit D3. The effect of amantadine depends, in part, on increased activity of the vitamin D receptor (VDR), since it induced VDR expression and amantadine-dependent monocyte-macrophage differentiation of HL60 cells was blocked by expression of dominant-negative VDR. These results reveal a new function for amantadine and support the concept that screening of the Connectivity Map database can identify small molecules that mimic the effect of transcription factors required for myelo-monocytic differentiation.

Shinmura K, Kato H, Kawanishi Y, et al.
NEIL1 p.Gln282Stop variant is predominantly localized in the cytoplasm and exhibits reduced activity in suppressing mutations.
Gene. 2015; 571(1):33-42 [PubMed] Related Publications
Human NEIL1 protein is a DNA glycosylase known to be involved in the repair of oxidized DNA lesions. A c.C844T germline variant of the NEIL1 gene has recently been identified in the Japanese population, however, the p.Q282Stop-type protein produced from this variant gene has not yet been characterized. In this study to determine whether the NEIL1 c.C844T variant might be a defective allele, we investigated the subcellular localization of the p.Q282Stop-type protein and its ability to suppress the development of mutations in mammalian cells. In contrast to the nuclear localization of wild-type (WT) NEIL1, the p.Q282Stop-type protein tagged with GFP or FLAG was localized predominantly in the cytoplasm of human H1299 cells. Mutant forms of the putative nuclear localization signal (NLS, amino acid sequences 359 to 378) of NEIL1-GFP resulted in predominant cytoplasmic localization of the mutants, suggesting that the abnormal localization of p.Q282Stop-type NEIL1 may also be caused by a loss of the putative NLS in the protein. Next, V79 mammalian cell lines inducibly expressing WT NEIL1 or p.Q282Stop-type NEIL1 were established using the piggyBac transposon vector system, and the mutation frequency was compared between the cell lines by HPRT assay. The frequency of mutations induced by glucose oxidase, an oxidative stress inducer, was higher in the p.Q282Stop-type NEIL1-transposed cells than that in the WT NEIL1-transposed cells. Finally, the Cancer Genome Atlas (TCGA) data showed an increased number of somatic mutations in primary carcinomas containing a truncating NEIL1 mutation. These results suggest that p.Q282Stop-type NEIL1 is predominantly localized in the cytoplasm, possibly due to a loss of the NLS, and possesses a reduced ability to suppress the onset of mutations, both findings suggesting that NEIL1 c.C844T is a defective allele.

Lawlor H, Meunier A, McDermott N, et al.
Identification of suitable endogenous controls for gene and miRNA expression studies in irradiated prostate cancer cells.
Tumour Biol. 2015; 36(8):6019-28 [PubMed] Related Publications
This study aimed to to evaluate the stability of commonly used endogenous control genes for messenger RNA (mRNA) (N = 16) and miRNAs (N = 3) expression studies in prostate cell lines following irradiation. The stability of endogenous control genes expression in irradiated (6 Gy) versus unirradiated controls was quantified using NormFinder and coefficient of variation analyses. HPRT1 and 18S were identified as most and least stable endogenous controls, respectively, for mRNA expression studies in irradiated prostate cells. SNORD48 and miR16 miRNA endogenous controls tested were associated with low coefficient of variations following irradiation (6 Gy). This study highlights that commonly used endogenous controls can be responsive to radiation and validation is required prior to gene/miRNAs expression studies.

Zhang J, Wang Y, Shang D, et al.
Characterizing and optimizing human anticancer drug targets based on topological properties in the context of biological pathways.
J Biomed Inform. 2015; 54:132-40 [PubMed] Related Publications
One of the challenging problems in drug discovery is to identify the novel targets for drugs. Most of the traditional methods for drug targets optimization focused on identifying the particular families of "druggable targets", but ignored their topological properties based on the biological pathways. In this study, we characterized the topological properties of human anticancer drug targets (ADTs) in the context of biological pathways. We found that the ADTs tended to present the following seven topological properties: influence the number of the pathways related to cancer, be localized at the start or end of the pathways, interact with cancer related genes, exhibit higher connectivity, vulnerability, betweenness, and closeness than other genes. We first ranked ADTs based on their topological property values respectively, then fused them into one global-rank using the joint cumulative distribution of an N-dimensional order statistic to optimize human ADTs. We applied the optimization method to 13 anticancer drugs, respectively. Results demonstrated that over 70% of known ADTs were ranked in the top 20%. Furthermore, the performance for mercaptopurine was significant: 6 known targets (ADSL, GMPR2, GMPR, HPRT1, AMPD3, AMPD2) were ranked in the top 15 and other four out of the top 15 (MAT2A, CDKN1A, AREG, JUN) have the potentialities to become new targets for cancer therapy.

Gueugnon F, Barascu A, Mavridis K, et al.
Kallikrein-related peptidase 13: an independent indicator of favorable prognosis for patients with nonsmall cell lung cancer.
Tumour Biol. 2015; 36(7):4979-86 [PubMed] Related Publications
The KLK13 gene is dysregulated in several carcinomas, and its expression levels seem to be associated with disease prognosis. The aim of our study was to investigate the prognostic potential of KLK13 mRNA expression for patients with nonsmall cell lung cancer (NSCLC). Total RNA was isolated from cancerous and normal tissues from a cohort of 128 NSCLC patients. The KLK13 mRNA transcription levels were measured using a sensitive quantitative RT-PCR method. The results were normalized by dividing the KLK13 mRNA values with the geometric mean of mRNA expression from four reference genes: beta-actin, TATA-binding protein, hypoxanthine phosphoribosyltransferase 1, and acidic ribosomal phosphoprotein P0. The malignant tissues from the majority of patients (59.3 %) contained significantly more KLK13 mRNA transcripts than did the paired nonmalignant tissues (median difference 11.1-fold, P = 0.008). KLK13 was expressed at higher levels in females than that in males (P = 0.021). No other statistically significant association with clinicopathological data was observed. Kaplan-Meier survival analyses demonstrated that patients with KLK13-positive tumors survived significantly longer than those with KLK13-negative ones (P = 0.009). KLK13 expression was also shown to be able to stratify high-risk individuals among patients with early disease stages (P = 0.030). Multivariate Cox regression analysis showed that KLK13 expression is a favorable, independent prognostic indicator of overall survival (OS) (P = 0.024). Our results suggest that KLK13 mRNA expression constitutes a novel biomarker for the prediction of overall survival in NSCLC and that its quantitative assessment in tumor tissues can aid in treatment decision making.

Ali H, Du Z, Li X, et al.
Identification of suitable reference genes for gene expression studies using quantitative polymerase chain reaction in lung cancer in vitro.
Mol Med Rep. 2015; 11(5):3767-73 [PubMed] Related Publications
The present study aimed to examine 10 housekeeping genes (HKGs), including 18s ribosomal RNA (18S), glyceraldehyde‑3‑phosphate dehydrogenase (GAPDH), ribosomal protein large P0 (RPLP0), β‑actin (ACTB), peptidylprolyl isomerase A (PPIA), phosphoglycerate kinase‑1 (PGK1), β‑2‑microglobulin (B2M), ribosomal protein LI3a (RPL13A), hypoxanthine phosphoribosyl transferase‑1 (HPRT1) and TATA box binding protein (TBP) in order to identify the most stable and suitable reference genes for use in expression studies in non‑small cell lung cancer. The mRNA expression encoding the panel of the 10 HKGs was determined using reverse transcription‑quantitative PCR (RT‑qPCR) in human lung cancer cell lines. Three software programs, BestKeeper, NormFinder and geNorm, were used to ascertain the most suitable reference genes to normalize the RNA input. The present study examined three lung cancer cell lines (A549, NCI‑H446 and NCI‑H460). The analysis of the experimental data using BestKeeper software revealed that all 10 HKGs were stable, with GADPH, followed by 18S being the most stable genes and PPIA and HPRT1 being the least stable genes. The NormFinder software results demonstrated that PPIA followed by ACTB were the most stable and B2M and RPLP0 were the least stable. The geNorm software results revealed that ACTB and PGK1, followed by PPIA were the most stable genes and B2M and RPLP0 were identified as the least stable genes. Due to discrepancies in the ranking orders of the reference genes obtained by different analyzing software programs, it was not possible to determine a single universal reference gene. The suitability of selected reference genes requires unconditional validation prior to each study. Based on the three analyzing programs, ACTB, PPIA and PGK1 were the most stable reference genes in lung cancer cell lines.

Romani C, Calza S, Todeschini P, et al.
Identification of optimal reference genes for gene expression normalization in a wide cohort of endometrioid endometrial carcinoma tissues.
PLoS One. 2014; 9(12):e113781 [PubMed] Free Access to Full Article Related Publications
Accurate normalization is a primary component of a reliable gene expression analysis based on qRT-PCR technique. While the use of one or more reference genes as internal controls is commonly accepted as the most appropriate normalization strategy, many qPCR-based published studies still contain data poorly normalized and reference genes arbitrarily chosen irrespective of the particular tissue and the specific experimental design. To date, no validated reference genes have been identified for endometrial cancer tissues. In this study, 10 normalization genes (GAPDH, B2M, ACTB, POLR2A, UBC, PPIA, HPRT1, GUSB, TBP, H3F3A) belonging to different functional and abundance classes in various tissues and used in different studies, were analyzed to determine their applicability. In total, 100 endometrioid endometrial cancer samples, which were carefully balanced according to their tumor grade, and 29 normal endometrial tissues were examined using SYBR Green Real-Time RT-PCR. The expression stability of candidate reference genes was determined and compared by means of geNorm and NormFinder softwares. Both algorithms were in agreement in identifying GAPDH, H3F3A, PPIA, and HPRT1 as the most stably expressed genes, only differing in their ranking order. Analysis performed on the expression levels of all candidate genes confirm HPRT1 and PPIA as the most stably expressed in the study groups regardless of sample type, to be used alone or better in combination. As the stable expression of HPRT1 and PPIA between normal and tumor endometrial samples fulfill the basic requirement of a reference gene to be used for normalization purposes, HPRT1 expression showed significant differences between samples from low-grade and high-grade tumors. In conclusion, our results recommend the use of PPIA as a single reference gene to be considered for improved reliability of normalization in gene expression studies involving endometrial tumor samples at different tumor degrees.

Yu S, Yang Q, Yang JH, et al.
Identification of suitable reference genes for investigating gene expression in human gallbladder carcinoma using reverse transcription quantitative polymerase chain reaction.
Mol Med Rep. 2015; 11(4):2967-74 [PubMed] Related Publications
Reverse transcription quantitative polymerase chain reaction (RT‑qPCR) has become a frequently used strategy in gene expression studies. The relative quantification method is an important and commonly used method for the evaluation of RT‑qPCR data. The key aim of this method is to identify an applicable internal reference gene, however, there are currently no suitable reference genes for gene analysis in gallbladder carcinoma. In the present study, screening was performed using 12 common reference genes, which were selected in order to provide an experimental basis for the investigation of gene expression in gallbladder carcinoma. A total of 16 tissue samples of gallbladder carcinoma and their matched normal gallbladder tissues were used. The gene expression stability and applicability of the 12 reference gene candidates were determined using the geNorm, NormFinder and BestKeeper software programs. Following comparison of the results of the three software programs, HPRT1 was identified as the most stably expressed reference gene. In the normal gallbladder group, the relative stably expressed reference gene was PPIA and in the entire sample group, the relatively stably expressed reference gene was PPIA. The present study also demonstrated that the combination of the three reference genes was the most appropriate. The recommended combinations were PPIA + PUM1 + ACTB for the total sample group, GAPDH + PBGD + ALAS1 for the gallbladder carcinoma group and PPIA + PUM1 + TBP for the paired normal gallbladder group.

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