Functional consequences of the first reported mutations of the proto-oncogene PTTG1IP/PBF.

Pituitary tumor-transforming gene 1-binding factor (PTTG1IP; PBF) is a multifunctional glycoprotein, which is overexpressed in a wide range of tumours, and significantly associated with poorer oncological outcomes, such as early tumour recurrence, distant metastasis, extramural vascular invasion and decreased disease-specific survival. PBF transforms NIH 3T3 fibroblasts and induces tumours in nude mice, while mice harbouring transgenic thyroidal PBF expression show hyperplasia and macrofollicular lesions. Our assumption that PBF becomes an oncogene purely through increased expression has been challenged by the recent report of mutations in PBF within the Catalogue of Somatic Mutations in Cancer (COSMIC) database. We therefore sought to determine whether the first 10 PBF missense substitutions in human cancer might be oncogenic. Anisomycin half-life studies revealed that most mutations were associated with reduced protein stability compared to wild-type (WT) PBF. Proliferation assays narrowed our interest to two mutational events which significantly altered cell turnover: C51R and R140W. C51R was mainly confined to the endoplasmic reticulum while R140W was apparent in the Golgi apparatus. Both C51R and R140W lost the capacity to induce cellular migration and significantly reduced cell invasion. Colony formation and soft agar assays demonstrated that, in contrast to WT PBF, both mutants were unable to elicit significant colony formation or anchorage-independent growth. However, C51R and R140W retained the ability to repress radioiodide uptake, a functional hallmark of PBF. Our data reveal new insight into PBF function and confirm that, rather than being oncogenic, mutations in PBF are likely to be passenger effects, with overexpression of PBF the more important aetiological event in human cancer.

Manipulation of PBF/PTTG1IP phosphorylation status; a potential new therapeutic strategy for improving radioiodine uptake in thyroid and other tumors.

CONTEXT: The clinical effectiveness of ablative radioiodine treatment of thyroid tumors is limited by the availability of the sodium iodide symporter (NIS) at the plasma membrane (PM) for uptake of ¹³¹I. A significant proportion of well-differentiated thyroid tumors are unable to concentrate sufficient radioiodine for effective therapy, and in other tumor models such as breast tumors, where radioiodine uptake would be an attractive therapeutic option, uptake is insufficient. OBJECTIVE: Pituitary tumor-transforming gene-binding factor (PBF; PTTG1IP) is overexpressed in multiple cancers and significantly decreases NIS expression at the PM. The goal of this study was to identify a method by which PBF repression of NIS may be overcome in human tumors. RESULTS: Here, we identify PBF as a tyrosine phosphoprotein that specifically binds the proto-oncogene tyrosine protein kinase Src in mass spectrometry, glutathione S-transferase pulldown and coimmunoprecipitation assays. Src induction leads to phosphorylation at PBF residue Y174. Abrogation of this residue results in PM retention and a markedly reduced ability to bind NIS. The Src inhibitor PP1 inhibits PBF phosphorylation in multiple cell lines in vitro, including human primary thyroid cells. Of direct clinical importance to the treatment of thyroid cancer, PP1 stimulates iodide uptake by transfected NIS in TPC1 thyroid carcinoma cells and entirely overcomes PBF repression of iodide uptake in human primary thyroid cells. CONCLUSIONS: We propose that targeting PBF phosphorylation at residue Y174 via tyrosine kinase inhibitors may be a novel therapeutic strategy to enhance the efficacy of ablative radioiodine treatment in thyroid and other endocrine and endocrine-related tumors.

Comparison of pulsatile ocular blood flow in Indians and Europeans.

PURPOSE: To compare pulsatile ocular blood flow (POBF) in Europeans and Indians and provide reference values for a group of healthy Indians. PATIENTS AND METHODS: Measurement with the POBF Tonograph was performed on healthy Indian subjects in India (n=252). A further 80 subjects (40 of Indian descent and 40 Europeans) underwent measurements in Cambridge, England. The instrument used for measurement was the same for both the studies. RESULTS: The mean POBF in the Indians in India was found to be 1176 microl/min. The mean POBF value in the Europeans was found to be 1033 microl/min and that for Indians in England was 1061 microl/min. The difference between the POBF within groups was significant (one-way ANOVA P<0.05) with the POBF of Indians in India being higher than Europeans and Indians in the UK. The difference between the Europeans and Indians in the UK did not reach statistical significance. CONCLUSIONS: POBF values in Indians living in India were found to be considerably higher than the previously published normal value of 650 microl/min in European studies and other studies for other racial groups. The reason for this apparent difference may be instrument-related rather than genetic because such a large difference was not observed when a comparison was performed in the UK. In addition, the results for both groups in our comparative study were still considerably higher than reported in previous studies. The POBF of Indians in India is slightly higher than the POBF of people of Indian ethnic origin in England.

Effect of corneal parameters on measurements using the pulsatile ocular blood flow tonograph and Goldmann applanation tonometer.

AIMS: To investigate the effect of central corneal thickness and corneal curvature on intraocular pressure measurements using the pulsatile ocular blood flow tonograph and the Goldmann applanation tonometer, and to assess the agreement between the pulsatile ocular blood flow tonograph and the Goldmann applanation tonometer in intraocular pressure measurement. METHODS: 479 subjects underwent intraocular pressure measurements with the Goldmann applanation tonometer and the pulsatile ocular blood flow tonograph. Of these, 334 patients underwent additional measurement of central corneal thickness with an ultrasonic pachymeter and corneal curvature measurement with a keratometer. RESULTS: The intraocular pressure measurements obtained with both the Goldmann applanation tonometer and the pulsatile ocular blood flow tonograph varied with central corneal thickness and mean keratometric reading. Intraocular pressure measured using the Goldmann applanation tonometer increased by 0.027 mm Hg per micro m increase in central corneal thickness. Intraocular pressure measured using the pulsatile ocular blood flow tonograph increased by 0.048 mm Hg per micro m increase in central corneal thickness. For an increase of 1 mm of mean corneal curvature there was rise in intraocular pressure of 1.14 mm Hg measured by the Goldmann applanation tonometer and of 2.6 mm Hg measured by the pulsatile ocular blood flow tonograph. When compared to the Goldmann applanation tonometer, the pulsatile ocular blood flow tonograph underestimated at low intraocular pressure and overestimated at higher intraocular pressure. CONCLUSION: Central corneal thickness and corneal curvature affected measurements obtained with the pulsatile ocular blood flow tonograph more than they affected measurements obtained with the Goldmann applanation tonometer.

Repeatability and reproducibility of the BVI ultrasonic Pachymeter.

PURPOSE: To evaluate the interobserver and intraobserver reliability of a commercially available, portable, ultrasonic pachymeter. METHODS: For the interobserver variability study, 42 healthy subjects underwent repeated ultrasonic pachymetry under topical anaesthesia performed by two observers. For the intraobserver study, another 30 further healthy subjects underwent repeated pachymetry by one of the observers. Agreement was analysed by means of Bland-Altman plots and by determination of the intraclass correlation coefficient. RESULTS: For the interobserver variability study, the mean measurement difference between observers was 0.7 microm (95% CI -0.8-2.2 microm) and the intraclass correlation coefficient was 0.9958 (95% CI 0.9922-0.9977). The Bland-Altman plot showed narrow limits of agreement with respect to central corneal thickness (CCT). For the intraobserver variability study, the mean difference between the repeated measurements was 0.9 microm (95% CI -3.1-1.3 microm). The intraclass correlation coefficient was 0.9934 (95% CI 0.9863-0.9969). A Bland-Altman plot again showed narrow limits of agreement with respect to CCT. The mean CCT for 72 subjects was 538 microm (95% CI 528-545 microm). CONCLUSIONS: Measurements of CCT using the BVI Pocket Pachymeter were repeatable and had excellent interobserver reliability. Measurement variation amounted to less than 0.2% assuming a mean CCT of 538 microm.