G1 checkpoint protein and p53 abnormalities occur in most invasive transitional cell carcinomas of the urinary bladder

The G1 cell cycle checkpoint regulates entry into S phase for normal cells. Components of the G1 checkpoint, including retinoblastoma (Rb) protein, cyclin D1 and p16INK4a, are commonly altered in human malignancies, abrogating cell cycle control. Using immunohistochemistry, we examined 79 invasive transitional cell carcinomas of the urinary bladder treated by cystectomy, for loss of Rb or p16INK4a protein and for cyclin D1 overexpression. As p53 is also involved in cell cycle control, its expression was studied as well. Rb protein loss occurred in 23/79 cases (29%); it was inversely correlated with loss of p16INK4a, which occurred in 15/79 cases (19%). One biphenotypic case, with Rb+p16– and Rb-p16+ areas, was identified as well. Cyclin D1 was overexpressed in 21/79 carcinomas (27%), all of which retained Rb protein. Fifty of 79 tumours (63%) showed aberrant accumulation of p53 protein; p53 staining did not correlate with Rb, p16INK4a, or cyclin D1 status. Overall, 70% of bladder carcinomas showed abnormalities in one or more of the intrinsic proteins of the G1 checkpoint (Rb, p16INK4a and cyclin D1). Only 15% of all bladder carcinomas (12/79) showed a normal phenotype for all four proteins. In a multivariate survival analysis, cyclin D1 overexpression was linked to less aggressive disease and relatively favourable outcome. In our series, Rb, p16INK4a and p53 status did not reach statistical significance as prognostic factors. In conclusion, G1 restriction point defects can be identified in the majority of bladder carcinomas. Our findings support the hypothesis that cyclin D1 and p16INK4a can cooperate to dysregulate the cell cycle, but that loss of Rb protein abolishes the G1 checkpoint completely, removing any selective advantage for cells that alter additional cell cycle proteins. © 1999 Cancer Research Campaig

Protein p16 as a marker of dysplastic and neoplastic alterations in cervical epithelial cells

BACKGROUND: Cervical carcinomas are second most frequent type of women cancer. Success in diagnostics of this disease is due to the use of Pap-test (cytological smear analysis). However Pap-test gives significant portion of both false-positive and false-negative conclusions. Amendments of the diagnostic procedure are desirable. Aetiological role of papillomaviruses in cervical cancer is established while the role of cellular gene alterations in the course of tumor progression is less clear. Several research groups including us have recently named the protein p16(INK4a )as a possible diagnostic marker of cervical cancer. To evaluate whether the specificity of p16(INK4a )expression in dysplastic and neoplastic cervical epithelium is sufficient for such application we undertook a broader immunochistochemical registration of this protein with a highly p16(INK4a)-specific monoclonal antibody. METHODS: Paraffin-embedded samples of diagnostic biopsies and surgical materials were used. Control group included vaginal smears of healthy women and biopsy samples from patients with cervical ectopia. We examined 197 samples in total. Monoclonal antibody E6H4 (MTM Laboratories, Germany) was used. RESULTS: In control samples we did not find any p16(INK4a)-positive cells. Overexpression of p16(INK4a )was detected in samples of cervical dysplasia (CINs) and carcinomas. The portion of p16(INK4a)-positive samples increased in the row: CIN I – CIN II – CIN III – invasive carcinoma. For all stages the samples were found to be heterogeneous with respect to p16(INK4a)-expression. Every third of CINs III and one invasive squamous cell carcinoma (out of 21 analyzed) were negative. CONCLUSIONS: Overexpression of the protein p16(INK4a )is typical for dysplastic and neoplastic epithelium of cervix uteri. However p16(INK4a)-negative CINs and carcinomas do exist. All stages of CINs and carcinomas analyzed are heterogeneous with respect to p16(INK4a )expression. So p16(INK4a)-negativity is not a sufficient reason to exclude a patient from the high risk group. As far as normal cervical epithelium is p16(INK4a)-negative and the ratio p16(INK4a)-positive/ p16(INK4a)-negative samples increases at the advanced stages application of immunohisto-/cytochemical test for p16(INK4a )may be regarded as a supplementary test for early diagnostics of cervical cancer

Loss of functional pRB is not a ubiquitous feature of B-cell malignancies

Human cancers frequently sustain genetic mutations that alter the function of their G1 cell cycle control check point. These include changes to the retinoblastoma gene and to the genes that regulate its phosphorylation, such as the cyclin-dependent kinase inhibitor p16(INK4a). Altered expression of retinoblastoma protein (pRb) is associated with non-Hodgkin's lymphoma, particularly centroblastic and Burkitt's lymphomas. pRb is expressed in normal B-cells and its regulatory phosphorylation pathway is activated in response to a variety of stimuli. Since human B-lymphoma-derived cell lines are often used as in vitro model systems to analyse the downstream effects of signal transduction, we examined the functional status of pRb in a panel of human B-cell lines. We identified eleven cell lines which express the hyperphosphorylated forms of pRb. Furthermore, we suggest that the pRb protein appears to be functional in these cell lines

Predictive value of expression of p16INK4A, retinoblastoma and p53 proteins for the prognosis of non-small-cell lung cancers

The predictive value of expression of p16INK4A, retinoblastoma (Rb) and p53 proteins for prognosis was evaluated in 76 patients with non-small-cell lung cancers (NSCLCs) that were potentially curatively resected between 1990 and 1995, using the results of immunostaining analyses of these proteins as reported in our previous study (Kinoshita et al, 1996). Of these NSCLCs, 22 (29%) lacked p16 protein expression and eight (11%) Rb protein, while 30 (39%) showed positive (altered) p53 protein expression. Survival of patients with p16-negative tumours was not significantly different from that of patients with p16-positive tumours (5-year survival rates 67% and 72% respectively, P = 0.8), nor was survival of patients with Rb-negative tumours significantly different from that of patients with Rb-positive tumours (5-year survival rates 42% and 69% respectively, P = 0.9). Moreover, survival of patients with p16/Rb-negative (either p16- or Rb-negative) tumours was not significantly different from that of patients with p16/Rb-positive (both p16- and Rb-positive) tumours (5-year survival rates 67% and 68% respectively, P = 0.7). In contrast, survival of patients with p53-positive (altered) tumours tended to be shorter than that of patients with p53-negative (unaltered) tumours (5-year survival rates 56% and 78% respectively, P = 0.06). In univariate analysis of potential prognostic factors, p16, Rb and p16/Rb proteins were not significant prognostic factors in the present cohort of potentially curatively resected NSCLCs. Altered p53 protein status tended to be a negative prognostic factor (P = 0.06 by the univariate analysis). These results indicate that loss of p16 protein alone, or in combination with loss of Rb protein, does not predict the clinical outcome of patients with resected NSCLCs. © 1999 Cancer Research Campaig

CALGB 30601: A phase II study of dasatinib (D) in patients (pts) with previously treated malignant mesothelioma (MM)

BACKGROUND: SRC is commonly over-expressed in MM. D is a potent inhibitor of SRC family kinases, EphA2 and PDGFRβ. There are no approved therapies for MM pts who progress on pemetrexed. We therefore conducted a phase II trial of D in MM pts who had received 1 prior pemetrexed-based regimen. METHODS: Single arm phase II. Eligible pts had unresectable MM, PS 0-1, measurable disease, and no symptomatic effusions. Primary endpoint: Progression-free survival (PFS) at 24 weeks (wks). D 70 mg BID was given orally. CT scans were obtained every 2 months. Pre- and posttreatment plasma VEGF, PDGFβ, and serum CSF-1 and mesothelin-related protein were collected. Tumor was evaluated for expression of EphA2 and PDGFRβ. RESULTS: 46 pts enrolled at 12 sites 9/07-8/09, 35 are evaluable for PFS, 22 for response, and 46 for toxicity. Pt characteristics: Male 72%; median age 68 (range 35, 81); PS 0 41%; epithelial histology 72%, pleural 76%. Median cycles given 2 (range 1-8). The starting dose was reduced to 50 mg BID after the first 23 pts enrolled because 50% of the first 12 pts enrolled had AE grade 3. Grade 3/4 toxicities (% pts): anemia 2%, fatigue 11%, pleural effusion 9%, pericardial effusion 2%, pneumonitis 2%, hypoxia 2%, nausea 4%, hyponatremia 7%, hypophosphatemia 2%.There were 3 grade 5 toxicities: 1 ARDS, 1 respiratory failure, 1 unknown. Efficacy: 24-week PFS rate 15% (95% CI 6%, 29%), median PFS 8.3 wks (7.7, 10.3), median overall survival 20.7 wks (11.4, 28.3), partial response 0%; stable disease 20%. CONCLUSIONS: Dasatinib is inactive in previously-treated MM; the 70 mg dose is poorly tolerated. Further data on biologic correlates will be presented