IntroductionHJURP (Holliday Junction Recognition Protein) is a newly discovered gene reported to function at centromeres and to interact with CENPA. However its role in tumor development remains largely unknown. The goal of this study was to investigate the clinical significance of HJURP in breast cancer and its correlation with radiotherapeutic outcome.MethodsWe measured HJURP expression level in human breast cancer cell lines and primary breast cancers by Western blot and/or by Affymetrix Microarray; and determined its associations with clinical variables using standard statistical methods. Validation was performed with the use of published microarray data. We assessed cell growth and apoptosis of breast cancer cells after radiation using high-content image analysis.ResultsHJURP was expressed at higher level in breast cancer than in normal breast tissue. HJURP mRNA levels were significantly associated with estrogen receptor (ER), progesterone receptor (PR), Scarff-Bloom-Richardson (SBR) grade, age and Ki67 proliferation indices, but not with pathologic stage, ERBB2, tumor size, or lymph node status. Higher HJURP mRNA levels significantly decreased disease-free and overall survival. HJURP mRNA levels predicted the prognosis better than Ki67 proliferation indices. In a multivariate Cox proportional-hazard regression, including clinical variables as covariates, HJURP mRNA levels remained an independent prognostic factor for disease-free and overall survival. In addition HJURP mRNA levels were an independent prognostic factor over molecular subtypes (normal like, luminal, Erbb2 and basal). Poor clinical outcomes among patients with high HJURP expression were validated in five additional breast cancer cohorts. Furthermore, the patients with high HJURP levels were much more sensitive to radiotherapy. In vitro studies in breast cancer cell lines showed that cells with high HJURP levels were more sensitive to radiation treatment and had a higher rate of apoptosis than those with low levels. Knock down of HJURP in human breast cancer cells using shRNA reduced the sensitivity to radiation treatment. HJURP mRNA levels were significantly correlated with CENPA mRNA levels.ConclusionsHJURP mRNA level is a prognostic factor for disease-free and overall survival in patients with breast cancer and is a predictive biomarker for sensitivity to radiotherapy.

Malignant Pleural Effusions (MPE) may be useful as a model to study hierarchical progression of cancer and/or intratumoral heterogeneity. To strengthen the rationale for developing the MPE-model for these purposes, we set out to find evidence for the presence of cancer stem cells (CSC) in MPE and demonstrate an ability to sustain intratumoral heterogeneity in MPE-primary cultures. Our studies show that candidate lung CSC-expression signatures (PTEN, OCT4, hTERT, Bmi1, EZH2 and SUZ12) are evident in cell pellets isolated from MPE, and MPE-cytopathology also labels candidate-CSC (CD44, cMET, MDR-1, ALDH) subpopulations. Moreover, in primary cultures that use MPE as the source of both tumor cells and the tumor microenvironment (TME), candidate CSC are maintained over time. This allows us to live-sort candidate CSC-fractions from the MPE-tumor mix on the basis of surface markers (CD44, c-MET, uPAR, MDR-1) or differences in xenobiotic metabolism (ALDH). Thus, MPE-primary cultures provide an avenue to extract candidate CSC populations from individual (isogenic) MPE-tumors. This will allow us to test whether these cells can be discriminated in functional bioassays. Tumor heterogeneity in MPE-primary cultures is evidenced by variable immunolabeling, differences in colony-morphology, and differences in proliferation rates of cell subpopulations. Collectively, these data justify the ongoing development of the MPE-model for the investigation of intratumoral heterogeneity, tumor-TME interactions, and phenotypic validation of candidate lung CSC, in addition to providing direction for the pre-clinical development of rational therapeutics.

We previously cloned and identified the esophageal cancer related gene 1 (ECRG1), a novel candidate tumor suppressor gene, from human esophageal cells. A single nucleotide polymorphism (Arg290Gln) was identified in the coding region of ECRG1 and might play a role in susceptibility to esophageal squamous cell carcinoma (ESCC). To examine this hypothesis, we analyzed 998 ESCC patients and 1252 controls in a hospital-based, case-control study in a Chinese population for this polymorphism. We observed a statistically significantly increased risk of ESCC associated with the ECRG1 290Arg/Gln and 290Gln/Gln genotypes compared with the 290Arg/Arg [odds ratio (OR)=1.23, 95% confidence interval (CI)=1.03-1.46; P<0.05]. A greater than multiplicative joint effect between the ECRG1 polymorphism and tobacco smoking exposure was also observed (OR=1.95, 95% CI=1.48-2.56; P<0.001). Furthermore, the elevated risk of ESCC associated with the ECRG1 polymorphism was increased consistently with cumulative smoking dose. ORs (95% CI) for 290Arg/Gln and 290Gln/Gln genotypes among non-smokers and smokers who smoked27 pack-years were 1.03 (0.78-1.35), 1.91 (1.36-2.67) and 2.08 (1.48-2.92), respectively (P trend test<0.001). Taken together, our results indicate that the ECRG1 290Gln variant allele might be a genetic susceptibility factor for developing ESCC, especially in the smoking population.

The esophageal cancer-related gene 2 (ECRG2) is a novel gene that shows sequence similarity to KAZAL-type serine protease inhibitor. In this study, the migration and invasion of PG cancer cells were inhibited by ectopic expression of ECRG2 in vitro, and metastases decreased after injecting PG/pcDNA3.1-ECRG2 cells into the tail veins of nude mice. Control mice were injected with PG/pcDNA3.1 cells. To test the hypothesis that ECRG2 interacts with proteases and inactivates extracellular matrix degradation, binding affinity and co-immunoprecipitation experiments were performed using serum-free conditioned medium. The results showed that ECRG2 bound to two species of urokinase-type plasminogen activator (uPA) with molecular weights of 55 and 33 kDa. Furthermore, analysis of the uPA/plasmin activity showed that expression of ECRG2 reduced proteolysis of the plasmin substrate D-Val-Phe-Lys-p-nitroanilide, which was seen by a decrease of absorbance at 405 nm. Taken together, these results suggested that ECRG2 inhibits aggressiveness of cancer cell, possibly through the down-regulation of uPA/plasmin activity.

Ideal bone tissue engineering scaffolds should be biocompatible, biodegradable, and mechanically robust and have the ability to regulate cell function.