Clinical significance of Src expression and activity in human neoplasia.

Src, a 60 kDa non-receptor tyrosine kinase, is the product of normal c-src of the human genome and member of the Src protein tyrosine kinases family (SFK). As described by Martin and Rous, a genetic recombination between c-src and the RSV oncogene of Rous sarcoma virus results in a modified Src protein, with increased intrinsic activity and transforming potential in animal and human tissues. Several in vitro and in vivo studies supported this theory providing insight in the signalling pathways involved. Accumulating evidence from studies on clinical samples supported the role of Src in the process of carcinogenesis and disease progression in several human malignancies. Some studies have further reinforced the significance of the kinase in malignacy by correlating its expression and/or activity with important clinicopathological parameters, such as tumour stage, histopathological grade, proliferative capacity and most importantly patient's survival. This review is a comprehensive report of the published evidence on the expression and clinical significance of Src in human malignancy, which constitutes the background of the current studies and clinical trials on the use of Src inhibitors as novel potent antineoplastic strategy.

Metallothionein expression in human neoplasia.

The metallothionein family is a class of low-molecular-weight, cysteine-rich proteins with high affinity for metal ions. Four major isoforms (metallothionein-1, -2, -3, and -4) have been identified in mammals, involved in many pathophysiological processes, including metal ion homeostasis and detoxification, protection against oxidative damage, cell proliferation and apoptosis, drug and radiotherapy resistance and several aspects of the carcinogenic process. In the present review we examine the expression of metallothionein in different human tumours and its correlation with histopathological variables, tumour cell proliferation or apoptosis, resistance to radiation or chemotherapy, patient survival and prognosis. A variable profile of metallothionein and its isoforms' expression has been observed in different cancer types. Although metallothionein expression has been implicated in carcinogenic evolution, its use as a marker of tumour differentiation, cell proliferation and prognosis predictor remains unclear. Detailed studies focused on the expression of metallothionein isoforms and isotypes in different tumour types could elucidate the role of this group of proteins in the carcinogenic process, delineating its possible clinical significance for the management of patients.

Focal adhesion kinase expression is not a prognostic predictor in colon adenocarcinoma patients.

AIM: Focal adhesion kinase (FAK) is an enzyme of the tyrosine kinase group linked to signaling pathways between cells and their extracellular matrix. FAK expression in tumor cells in vitro may correlate with their ability for invasion and metastasis. METHODS: FAK protein expression was examined immunohistochemically in 80 cases of colon adenocarcinoma, and correlated with clinicopathological parameters; tumor proliferative capacity, reflected by Ki-67 antigen expression; and survival. RESULTS: All tumor samples were FAK positive compared to normal colonic mucosa. FAK protein overexpression was seen in 32 out of 80 cases. FAK protein overexpression did not correlated with tumor histological grade, stage, Ki-67 positivity or survival. CONCLUSIONS: Raised FAK protein expression was noted by immunohistochemistry in human colon carcinoma cases. The implication are discussed.

Patterns of expression of Rb and p16 in astrocytic gliomas, and correlation with survival.

The retinoblastoma pathway is a key cell cycle regulatory complex that controls the passage of cells through the G1 checkpoint and is a frequent target of genetic alterations in gliomas. In this study, we examined the expression of Rb and p16 in 170 primary astrocytic gliomas by immunohistochemical techniques, and correlated the expression with overall survival to determine their prognostic value as immunomarkers. There were 130 patients with glioblastoma multiforme (GBM) and 40 with anaplastic astrocytoma (AA). Alterations in the levels of Rb or p16 expression were seen in the majority (>90%) of the gliomas studied. The expression of Rb was completely absent or low in 47.5% of the GBM and 67.5% of the AA. The remainder of the tumors was immunopositive for Rb to varying degrees. Immunoreactivity for p16 was absent in 56% of the GBM and 77.5% of the AA. Kaplan-Meier survival plots (log-rank test) and Cox proportional hazards regression analysis, adjusted for age and histology, showed that neither Rb nor p16 expression independently predicted survival. The results of our study suggest that although genetic alterations of Rb and p16 are common in gliomas, immunohistochemical analysis of these markers correlates poorly with prognosis.

Fenretinide activates caspases and induces apoptosis in gliomas.

The synthetic retinoid fenretinide (N-[4-hydroxyphenyl] retinamide or 4HPR) has been shown to not only inhibit cell growth but also to induce apoptosis in a variety of malignant cell lines. It is being tested presently for its potential as a chemopreventive agent against several cancers. A related retinoid, 13-cis-retinoic acid (cRA), has been shown to have activity against gliomas in vitro as well as in a recent clinical study. The present study aimed at assessing the activity of fenretinide against glioma cells in vitro and comparing it with that of cRA at pharmacologically relevant doses. We hypothesized that the ability of fenretinide to induce apoptosis would make it more potent against gliomas than cRA. Four glioma cell lines (D54, U251, U87MG, and EFC-2) were treated with fenretinide (1-100 microM) and showed dose- and time-dependent induction of cell death. At pharmacologically relevant doses, fenretinide was more active against glioma cells than cRA because of its ability to induce apoptosis. Flow cytometric studies using D54 cells demonstrated no significant changes in the cell cycle distribution compared with untreated control, but a sub-G1 fraction consistent with apoptosis was detected. Terminal deoxynucleotidyl transferase-mediated nick end labeling assay indicated that the apoptotic fraction was cell cycle nonspecific. Fenretinide treatment resulted in cleavage of poly ADP-ribose polymerase, indicating an activation of the caspase 3. Immunofluorescence studies using the nuclear stain 4',6-diamidine-2'-phenylindole dihydrochloride showed nuclear condensation and an apoptotic morphology. Hence, this study demonstrates that, at clinically relevant doses, fenretinide is a potent inducer of apoptosis in gliomas acting via the caspase pathway. We also show that at clinically achievable doses, fenretinide has more activity against gliomas than comparable doses of cRA. The favorable side effect profile seen in previous clinical studies and the in vitro activity against gliomas demonstrated in this study suggest that fenretinide could be a promising therapeutic agent against gliomas.

Elevated levels of urokinase-type plasminogen activator and its receptor during tumor growth in vivo.

Urokinase-type plasminogen activator (uPA) and its receptor (uPAR) are important in the regulation of tumor tissue progenesis, cell differentiation, tumor cell motility, and tumor cell invasiveness. We have recently reported that the levels of uPA and uPAR were higher in malignant astrocytomas than in low-grade gliomas. In the present study, we measured the levels of uPA and uPAR during the growth of glioblastomas in nude mice. Using fibrin zymography, densitometry, and an enzyme-linked immunosorbent assay, we found that the enzyme activity and content of uPA were increased 4- to 10-fold during tumor formation. Using a receptor assay and an enzyme linked immunosorbent assay, we found the numbers and content of uPAR were increased 5- to 15-fold during tumor formation. In addition, immunohistochemical staining for uPA and uPAR revealed strong immunoreactivity in tumor cells with the staining more intense on day 28 than on day 14. These results suggest that the upregulation of uPA and uPAR plays a major role in the formation of gliomas.

Increased invasion of neuroglioma cells transfected with urokinase plasminogen activator receptor cDNA.

The cell-surface urokinase plasminogen activator receptor (uPAR) plays a key role in regulating plasminogen cleavage during extracellular proteolysis. Our recent results demonstrated that uPAR expression is critical for the invasiveness of human gliomas and down regulation of uPAR caused by antisense cDNA transfection inhibits the invasion of these stable antisense uPAR-transfectant clones. To study the role of uPARs in glioma cell invasion, a human neuroglioma cell line (H4) that normally produces low numbers of uPARs was transfected with the expression vector containing full-length human uPAR cDNA. Stable transfectants were analyzed for uPAR mRNA expression, receptor number, in vitro invasion and secretion of uPA and MMP-2. The uPAR-overproducing clones showed a 4-fold increase in uPAR mRNA transcription and approximately 40% increase in receptor numbers. uPAR-overproducing clones also invaded through matrigel to a significantly greater extent than did parent cell line and vector clones. However, the uPAR-overexpressing clones and parent cell lines showed similar uPA and MMP-2 activities. These results suggest that the over-production of uPAR on the surface of neuroglioma cells enhances the invasiveness.

Increased estrogen receptor and epidermal growth factor receptor gene product co-expression in surgically resected gastric adenocarcinomas.

BACKGROUND: Evidence exists that estrogens influence the action of epidermal growth factor (EGF) and its receptor (EGF-R) at multiple levels. Estrogen and antiestrogen action on gastric and other gastrointestinal malignancies has been evaluated by several groups with conflicting results, and EGF-R has been implicated in the current growth factor-mediated models for gastric cancer progression. METHODS: ERs and EGF-Rs were detected immunohistochemically in a total of 53 advanced gastric carcinomas using monoclonal antibodies (mAbs) to human ERs and EGF-Rs. RESULTS: ERs were expressed in 30 (56%) and EGF-Rs in 20 (38%) of the gastric tumors. ER(+) gastric tumors were closely associated with the intestinal type (P < 0.01), whereas EGF-R(+) tumors were significantly correlated with poor differentiation status and ER(+) expression (P < 0.01). Of EGF-R(+) tumors, 85% were also ER(+). EGF-R and ER co-expression was demonstrated in 17 tumors (32% of the group). These cases were significantly corelated with poor differentiation and large tumor size upon resection (P < 0.05). CONCLUSIONS: ER and EGF-R co-expression indicates that a functional interaction between estrogens and EGF may exist in gastric cancer and that when such an interaction becomes operative, it may lead to dedifferentiation and increased tumor growth.