Development and Clinical Validation of a 90-Gene Expression Assay for Identifying Tumor Tissue Origin.

The accurate identification of tissue origin in patients with metastatic cancer is critical for effective treatment selection but remains a challenge. The aim of this study is to develop a gene expression assay for tumor molecular classification and integrate it with clinicopathologic evaluations to identify the tissue origin for cancer of uncertain primary (CUP). A 90-gene expression signature, covering 21 tumor types, was identified and validated with an overall accuracy of 89.8% (95% CI, 0.87-0.92) in 609 tumor samples. More specifically, the classification accuracy reached 90.4% (95% CI, 0.87-0.93) for 323 primary tumors and 89.2% (95% CI, 0.85-0.92) for 286 metastatic tumors, with no statistically significant difference (P = 0.71). Furthermore, in a real-life cohort of 141 CUP patients, predictions by the 90-gene expression signature were consistent or compatible with the clinicopathologic features in 71.6% of patients (101/141). Findings suggest that this novel gene expression assay could efficiently predict the primary origin for a broad spectrum of tumor types and support its diagnostic utility of molecular classification in difficult-to-diagnose metastatic cancer. Additional studies are ongoing to further evaluate the clinical utility of this novel gene expression assay in predicting primary site and directing therapy for CUP patients.

90-gene signature assay for tissue origin diagnosis of brain metastases.

BACKGROUND: Brain metastases (BM) are the most common intracranial tumors. 2-14% of BM patients present with unknown primary site despite intensive evaluations. This study aims to evaluate the performance of a 90-gene expression signature in determining the primary sites for BM samples. METHODS: The sequence-based gene expression profiles of 708 primary brain tumors (PBT) collected from The Cancer Genome Atlas (TCGA) database were analyzed by the 90-gene expression signature, with a similarity score for each of 21 common tumor types. We then used Optimal Binning algorithm to generate a threshold for separating PBT from BM. Eighteen PBT samples were analyzed to substantiate the reliability of the threshold. In addition, the performance of the 90-gene expression signature for molecular classification of metastatic brain tumors was validated in a cohort of 48 BM samples with the known origin. For each BM sample, the tumor type with the highest similarity score was considered tissue of origin. When a sample was diagnosed as PBT, but the similarity score below the threshold, the second prediction was considered as the primary site. RESULTS: A threshold of the similarity score, 70, was identified to discriminate PBT from BM (PBT: > 70, BM: ≤ 70) with an accuracy of 99% (703/708, 95% CI 98-100%). The 90-gene expression signature was further validated with 18 PBT and 44 BM samples. The results of 18 PBT samples matched reference diagnosis with a concordance rate of 100%, and all similarity scores were above the threshold. Of 44 BM samples, the 90-gene expression signature accurately predicted primary sites in 89% (39/44, 95% CI 75-96%) of the cases. CONCLUSIONS: Our findings demonstrated the potential that the 90-gene expression signature could serve as a powerful tool for accurately identifying the primary sites of metastatic brain tumors.

Activation of Nrf2/HO-1 signal with Myricetin for attenuating ECM degradation in human chondrocytes and ameliorating the murine osteoarthritis.

BACKGROUND: Osteoarthritis (OA), one of the prevailing joint degenerative disorders, contributes to the disability around the world. However, no effective therapeutic was introduced currently. Myricetin was reported to possess the function of anti-inflammatory, anti-diabetic and anti-cancer. Thus, we investigate the protection role of myricetin in OA progression and the potential molecular mechanism in present study. METHODS: Quantitative realtime PCR and western blotting were performed to evaluate the expression of MMP-13, Aggrecan, iNOS, and COX-2 at both gene and protein levels. An enzyme-linked immunosorbent assay was used to evaluate the levels of inflammatory factors (PGE2, TNF-α, and IL-6). The PI3K/AKT, Nrf2/HO-1 and nuclear factor kappa B (NF-κB) signaling pathways were analyzed by western blotting, and immunofluorescence was used to assess the expression of Nrf2, Collagen II and MMP13. The in vitro effect of myricetin was evaluated by intragastric administration into a mouse osteoarthritis model induced by destabilization of the medial meniscus. RESULTS: Myricetin not only inhibited the generation of inflammatory mediators and cytokines such as nitric oxide (NO), prostaglandin E2 (PGE2), TNF-α and IL-6, but also suppressed the production of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in human chondrocytes under IL-1ß stimulation. Moreover, Metalloproteinase 13 (MMP13) and thrombospondin motifs 5 (ADAMTS5), which resulted in the degradation of cartilage, were also suppressed in chondrocytes with the treatment of myricetin. To explore the potential mechanism, we found out that myricetin suppressed NF-κB signaling pathway through Nrf2/HO-1 axis in human chondrocytes. Besides, myricetin regulated the Nrf2 signaling pathway through PI3K/Akt pathway. In addition, in vivo study demonstrated that myricetin could ameliorated the progression of OA in mice DMM model through PI3K/Akt mediated Nrf2 signaling pathway. CONCLUSION: Taken together, our data first demonstrated that myricetin possesses the therapeutic potential on OA through PI3K/Akt mediated Nrf2/HO-1 signaling pathway.

Gene Expression Profiling for Diagnosis of Triple-Negative Breast Cancer: A Multicenter, Retrospective Cohort Study.

Background: Triple-negative breast cancer (TNBC) accounts for 12-20% of all breast cancers. Diagnosis of TNBC is sometimes quite difficult based on morphological assessment and immunohistochemistry alone, particularly in the metastatic setting with no prior history of breast cancer. Methods: Molecular profiling is a promising diagnostic approach that has the potential to provide an objective classification of metastatic tumors with unknown primary. In this study, performance of a novel 90-gene expression signature for determination of the site of tumor origin was evaluated in 115 TNBC samples. For each specimen, expression profiles of the 90 tumor-specific genes were analyzed, and similarity scores were obtained for each of the 21 tumor types on the test panel. Predicted tumor type was compared to the reference diagnosis to calculate accuracy. Furthermore, rank product analysis was performed to identify genes that were differentially expressed between TNBC and other tumor types. Results: Analysis of the 90-gene expression signature resulted in an overall 97.4% (112/115, 95% CI: 0.92-0.99) agreement with the reference diagnosis. Among all specimens, the signature correctly classified 97.6% of TNBC from the primary site (41/42) and lymph node metastasis (41/42) and 96.8% of distant metastatic tumors (30/31). Furthermore, a list of genes, including AZGP1, KRT19, and PIGR, was identified as differentially expressed between TNBC and other tumor types, suggesting their potential use as discriminatory markers. Conclusion: Our results demonstrate excellent performance of a 90-gene expression signature for identification of tumor origin in a cohort of both primary and metastatic TNBC samples. These findings show promise for use of this novel molecular assay to aid in differential diagnosis of TNBC, particularly in the metastatic setting.

Identification and validation of a 44-gene expression signature for the classification of renal cell carcinomas.

BACKGROUND: Renal cancers account for more than 3% of all adult malignancies and cause more than 23,400 deaths per year in China alone. The four most common types of kidney tumours include clear cell, papillary, chromophobe and benign oncocytoma. These histological subtypes vary in their clinical course and prognosis, and different clinical strategies have been developed for their management. Some kidney tumours can be very difficult to distinguish based on the pathological assessment of morphology and immunohistochemistry. METHODS: Six renal cell carcinoma microarray data sets, including 106 clear cell, 66 papillary, 42 chromophobe, 46 oncocytoma and 35 adjacent normal tissue samples, were subjected to integrative analysis. These data were combined and used as a training set for candidate gene expression signature identification. In addition, two independent cohorts of 1020 RNA-Seq samples from The Cancer Genome Atlas database and 129 qRT-PCR samples from Fudan University Shanghai Cancer Center (FUSCC) were analysed to validate the selected gene expression signature. RESULTS: A 44-gene expression signature derived from microarray analysis was strongly associated with the histological differentiation of renal tumours and could be used for tumour subtype classification. The signature performance was further validated in 1020 RNA-Seq samples and 129 qRT-PCR samples with overall accuracies of 93.4 and 93.0%, respectively. CONCLUSIONS: A 44-gene expression signature that could accurately discriminate renal tumour subtypes was identified in this study. Our results may prompt further development of this gene expression signature into a molecular assay amenable to routine clinical practice.

Cellular senescence occurred widespread to multiple selective sites in the fetal tissues and organs of mice.

Cellular senescence protects multicellular organisms from tissue overgrowth including cancer, and contributes to tissue ageing. With stable cell cycle arrests, cellular senescence has been mostly studied in the adult tissues of mammals. In the present study, we report widespread cellular senescence within certain time windows of late-phase normal development of mouse embryos. Using in situ senescence-associated ß-galactosidase (SA-ß-gal) staining, we showed SA-ß-gal activity in selected cell populations of the brain, stomach, interdigital webs, tail, ear, limbs and nasal mouth area on gestation day 14.5 of the mouse embryos. On day 18.5 of gestation, selected cells in the intestines and bone developmental areas showed SA-ß-gal activity. The chondrocytes in ossification zones were significantly marked by the activities of SA-ß-gal, p21, p15 and Hp1Y, suggesting activation of the cell cycle checkpoint by the p53 and Rb pathways, and development of senescence-associated heterochromatic foci. Throughout gestation days 14.5-18.5, the trophoblast cells in the labyrinth layer of the placentas also showed strong activities of SA-ß-gal, p53 and p21. Increased expressions of p19, p16 and Rb of the p16/Rb pathway, and reduced expressions of Ki67 were also observed in the placentas. Taken together, the present findings suggest that cellular senescence represents an essential mechanism at multiple sites including the fetal bone forming zones and placenta during mammalian embryonic development, playing potential roles in the full embryonic development of tissue growth and organ formation.