Gain and overexpression of the oncostatin M receptor occur frequently in cervical squamous cell carcinoma and are associated with adverse clinical outcome.

For many oncogenes, increased expression resulting from copy number gain confers a selective advantage to cells that consequently make up the tumour bulk. To identify oncogenes of potential biological significance in cervical squamous cell carcinoma (SCC), 36 primary samples and ten cell lines were screened by array comparative genomic hybridization (CGH). The most commonly occurring regions of copy number gain that also showed amplification were 5p15.2-14.3 (59%), 5p13.3 (65%), and 5p13.2-13.1 (63%). Gene copy numbers were significantly associated with expression levels for three candidate oncogenes at these loci: OSMR (oncostatin M receptor) (p=0.03), PDZK3 (PDZ domain containing protein 3) (p=0.04), and TRIO (triple functional domain) (p=0.03). Further examination by fluorescence in situ hybridization on a tissue microarray of 110 primary cervical SCC samples revealed copy number gain frequencies of 60.9%, 57.3%, and 54.5% for OSMR, PDZK3, and TRIO, respectively, with OSMR adversely influencing overall patient survival independently of tumour stage (p=0.046). By array CGH, copy number gain of OSMR was not seen in any of 40 microdissected precursor cervical squamous intraepithelial lesions (SILs). Moreover, global mRNA expression analysis, using Affymetrix U133A 2.0 Arrays, showed no overexpression of OSMR in SILs, suggesting that OSMR gain and overexpression are relatively late steps in cervical carcinogenesis. In the cervical SCC cell lines CaSki and SW756, exogenous OSM activated downstream-signalling elements of OSMR including STAT3, p44/42 MAPK, and S6 ribosomal protein, and induced transcription of the angiogenic factor VEGF, effects that were reduced by OSMR depletion using RNA interference. We conclude that copy number gain of OSMR is frequently found in cervical SCC and is associated with adverse clinical outcome. As well as being a potential prognostic marker, OSMR is a candidate cell surface therapeutic target.

Global microRNA profiles in cervical squamous cell carcinoma depend on Drosha expression levels.

Gain of chromosome 5p is seen in over 50% of advanced cervical squamous cell carcinomas (SCCs), although the genes responsible for the selective advantage provided by this abnormality are poorly understood. In the W12 cervical carcinogenesis model, we observed that 5p gain was rapidly selected over approximately 15 population doublings and was associated with the acquisition of a growth advantage and invasiveness. The most significantly upregulated transcript following 5p gain was the microRNA (miRNA) processor Drosha. In clinically progressed cervical SCC, Drosha copy-number gain was seen in 21/36 clinical samples and 8/10 cell lines and there was a significant association between Drosha transcript levels and copy-number gain. Other genes in the miRNA processing pathway, DGCR8, XPO5 and Dicer, showed infrequent copy-number gain and over-expression. Drosha copy-number and expression were not elevated in pre-malignant cervical squamous intraepithelial lesions. Importantly, global miRNA profiling showed that Drosha over-expression in cervical SCC appears to be of functional significance. Unsupervised principal component analysis of a mixed panel of cervical SCC cell lines and clinical specimens showed clear separation according to Drosha over-expression. miRNAs most significantly associated with Drosha over-expression are implicated in carcinogenesis in other tissues, suggesting that they regulate fundamental processes in neoplastic progression. Our evidence suggests that copy-number driven over-expression of Drosha and consequent changes in miRNAs are likely to be important contributors to the selective advantage provided by 5p gain in cervical neoplastic progression.

Heme protein dynamics revealed by geminate nitric oxide recombination in mutants of iron and cobalt myoglobin.

Nitric oxide myoglobin (MbNO) at 300 K was photodissociated with 405 nm pulses. The NO recombination in several mutants of iron and cobalt myoglobins was investigated at a time resolution of ca. 70 fs. The geminate recombination of NO was nonexponential on sub-nanosecond time scales. For both metals, the change of the detailed structure of the heme pocket (position 68 mutations) caused significant changes in the rates of recombination; however, the metal substitution influenced the recombination much less than did amino acid substitution. The results indicate a primary role of the heme pocket structure in the dynamics, and they suggest that proximal protein relaxation is not the limiting factor in the geminate recombination process. Recombination in cobalt derivatives is somewhat more efficient on the sub-nanosecond time scales than in corresponding iron myoglobins, consistent with other results that show a greater intrinsic reactivity toward the NO of cobalt compared with the iron heme. A comparison of results using Soret band excitation with previous Q-state excitation studies demonstrates that the ligand dissociates with a similar kinetic energy in both cases, suggesting fast intramolecular energy redistribution before dissociation.