Peter A. Humphrey

Intraglandular tumor extent and prognosis in prostatic carcinoma: Application of a grid method to prostatectomy specimens

The extent of tumor in prostatectomy specimens was determined by a grid method in 117 patients with prostatic adenocarcinoma. A plastic strip or ruler with squares of 3.0 mm was used, and the ratio of squares overlying carcinoma to the total number of squares overlying prostate tissue was calculated. This grid ratio, which represents an estimate of the percentage of the prostate involved by tumor, was a significant prognosticator closely tied to the likelihood of tumor progression and to survival time, as assessed by logistic regression analysis and a proportional hazard model. The grid ratio was better than histologic grade in predicting tumor progression and patient survival; also, the ratio was more objective than histologic grade as judged by interobserver agreement values. Only slight improvement in prognostication was obtained with concurrent use of both extent and grade. The grid ratio method was slightly better in predicting tumor progression and patient survival than a second method of assessing the percentage of prostatic tissue involved by tumor, the pathologists percentage estimate. These results indicate that it is important to quantitate tumor extent within prostatectomy specimens; such quantitation need not require step-sectioning of the entire prostate and an expensive and time-consuming method such as computerized morphometrics but rather may be performed by a simple estimate of the percentage of the prostate involved by tumor. Reporting of histologic grade and tumor extent in the prostate gland is recommended as both appear to be important in identifying those patients at risk for a poor outcome after prostatectomy for prostatic carcinoma.

Immunoreactive prostatic specific antigen in male periurethral glands.

Prostatic specific antigen (PSA) is considered an antigen unique to benign and malignant prostatic tissue. Recent evidence in the literature has raised serious doubts about the specificity of this antigen. In this study twenty male urethral specimens were evaluated for PSA and prostatic acid phosphatase (PAP) from patients without evidence of prostatic cancer. Eight of these 20 urethral specimens exhibited strong immunostaining for both PSA and PAP, localized in the periurethral glands. Five of the 17 urethral biopsies were positive for both antigens, while all three of the whole mount autopsy specimens stained positive for PSA and PAP. Within the autopsy series, there was heterogenous staining of the periurethral glands within the same specimen. This evidence disproves the fact that PSA and PAP are organ specific as previously described. More than likely any tissue of cloacal origin has potential for staining positive for prostatic specific antigen and prostatic acid phosphatase.

Overexpression of p53 and HER-2/ neu proteins as prognostic markers in early stage breast cancer

ObjectiveOverexpression of the p53 and HER-2/neu oncogenes are the two most common genetic abnormalities associated with breast cancer. Shorter survival time has been reported in patients with tumors with p53 or HER-2/neu. This report analyzes a retrospective cohort of early stage breast cancers for both oncogenes and relates overexpression to clinicopathologic parameters and survival. MethodsImmunostaining for p53 and HER-2/neu was performed on 230 paraffin-embedded specimens of stage I and II breast cancers diagnosed and treated at Duke University Medical Center between 1984 and 1987. Positive staining for both p53 and HER-2/neu in paraffin-embedded tissues indicates an underlying genetic abnormality: point mutations in the p53 gene and amplification of the HER-2/neu gene. ResultsIn this cohort of patients, 24% were positive for p53 and 17% for HER-2/neu. Four per cent were positive for both oncogenes. Significant correlations were found between p53 immunostaining and increasing tumor size, stage, and low estrogen and progesterone receptor contents. Univariate analysis showed that p53 and HER-2/neu were indicators of overall and failure-free survival. An additive effect on survival was observed in patients with both oncogene abnormalities. Nodal status, HER-2/neu, and p53 all attained independent prognostic value in a multivariate analysis. ConclusionsThe p53 and HER-2/neu oncogenes have proven but limited prognostic value. An approach that combines several molecular genetic markers with established pathologic criteria may help physicians to make more accurate predictions of prognosis in patients with early stage breast cancer.

Deletion-mutant epidermal growth factor receptor in human gliomas: Effect of type II mutation on receptor function

Malignant human glioma D-298 MG amplifies a rearranged epidermal growth factor receptor (EGFR) gene (c-erbB proto-oncogene), resulting in an in-frame deletion of 83 amino acids in domain IV of the extracellular domain of the EGFR. EGF and transforming growth factor-a (TGF-a) bound to the mutant EGFR with high affinity and enhanced the intrinsic mutant EGFR kinase activity. The mutant EGFR was capable of transducing EGF-stimulated glioma cell proliferation and invasiveness in an in vitro three-dimensional spheroid model. The deletion-mutant EGFR in D-298 MG is capable of being activated by growth factor; this suggests that overexpression of this mutant EGFR protein rather than structural alteration may be the more significant biologic event.

Practical application of molecular genetic testing as an aid to the surgical pathologic diagnosis of sarcomas: a prospective study.

The strong correlation of specific reciprocal translocations with individual tumor types and the demonstration that polymerase chain reaction (PCR)-based methods can detect translocations in tissue samples have stimulated interest in the role of molecular genetic testing in diagnostic surgical pathology. To evaluate the clinical utility of PCR-based molecular analysis of soft tissue neoplasms in routine surgical pathology, 131 consecutive soft tissue tumors submitted for molecular genetic testing at a tertiary care teaching hospital were prospectively analyzed over a 36-month period. RT-PCR was used to test tumor RNA for fusion transcripts characteristic of malignant round cell tumors (including Ewing sarcoma/primitive neuroectodermal tumor, desmoplastic small round cell tumor, and alveolar rhabdomyosarcoma), spindle cell tumors (including synovial sarcoma and congenital fibrosarcoma), and fatty tumors (myxoid liposarcoma). DNA sequence analysis was used to confirm the identity of all PCR products, and the PCR results were compared with the histopathologic diagnosis. We found that sufficient RNA for RT-PCR-based testing was recovered from 96% of the 131 cases and the percentage of tumors that tested positive for the associated characteristic fusion transcript was in general agreement with those reported in the literature. DNA sequence analysis of PCR products identified both variant transcripts and spurious PCR products, underscoring the value of product confirmation steps when testing formalin-fixed, paraffin-embedded tissue. Only in rare cases did testing yield a genetic result that was discordant with the histopathologic diagnosis. We conclude that PCR-based testing is a useful adjunct for the diagnosis of malignant small round cell tumors, spindle cell tumors, and other miscellaneous neoplasms in routine surgical pathology practice.

Stratification of pathologic features in radical prostatectomy specimens that are predictive of elevated initial postoperative serum prostate‐specific antigen levels

Background. Prostate‐specific antigen (PSA) is an important marker for adenocarcinoma of the prostate and is of clinical utility in assessment of residual carcinoma after radical prostatectomy. Although elevated postoperative serum PSA levels have been linked to pathologic stage in radical prostatectomy specimens, limited data are available on the relationship of postoperative PSA levels to margin positivity, intraglandular tumor extent, and histologic grade.

Differential Immunoreactivity of Epidermal Growth Factor Receptor in Benign, Dysplastic and Malignant Prostatic Tissues

To investigate epidermal growth factor receptor (EGFr) presence in the prostate, monoclonal antibody (clone EGFR1) immunohistochemical examination of radical prostatectomy specimens was performed (n = 37). All prostatic specimens contained benign prostatic hyperplasia (BPH) and/or dysplasia (prostatic intraepithelial neoplasia or PIN), as well as prostatic carcinoma (CaP). Areas of dysplasia were further categorized as to the basal cell layer and the luminal cell area. BPH, PIN, and CaP tissues in each specimen were analyzed by a single observer and graded on a scale from 0-4+. Fifteen samples were also analyzed for EGFr content utilizing a Cell Analysis Systems (CAS 200) image cytometer. EGFr immunoreactivity of BPH basal cells was significantly higher than EGFr immunoreactivity in areas of CaP (p < 0.001). EGFr staining of BPH basal cells was also significantly higher than that seen in PIN luminal cells (p < 0.001). Immunoreactivity of EGFr in PIN basal cells was significantly higher than in PIN luminal cells (p < 0.001). EGFr staining of basal cells in BPH tissues was higher than that seen in the PIN basal cell layer but the difference was not statistically significant (p = 0.06). The amount of staining present in PIN luminal cells was also significantly greater than in CaP tissues (p = 0.002). Quantitative image analysis utilizing the CAS 200 image cytometer was performed on BPH and CaP areas exclusively. EGFr immunoreactivity in basal cells of the BPH tissues was significantly greater than that seen in CaP tissues (p < 0.001). The decreased EGFr immunoreactivity in CaP may reflect a differentiating role for EGFr in normal tissues. Loss of EGFr influence may be associated with an increased proliferative state in PIN and CaP. Destruction or alteration of the epidermal grwoth factor receptor by a protease, such as prostatic specific antigen, may also explain our findings. At the present time the meaning of the different amounts of EGFr in the various types of prostate tissues is unknown.

Growth effects of epidermal growth factor (EGF) and a monoclonal antibody against the EGF receptor on four glioma cell lines.

SummaryEpidermal growth factor (EGF) has been shown to stimulate DNA synthesis and cell division in normal glia. At least half of malignant human gliomas (MHG) express high levels of the EGF receptor (EGFR), which are above those detected in normal brain. The demonstration that antibodies against the EGFR inhibit the growth of squamous cell carcinoma line A-431, with large numbers of EGFR, in vitro and in vivo raises the possibility that these agents could be used therapeutically against malignant human gliomas either alone or conjugated to other agents. We have measured the growth effects of EGF and an anti-EGFR monoclonal antibody, 528 (Ab-528), on four well-characterized human malignant glioma cell lines, D-263 MG, D-247 MG, U-343 MGa Cl 2∶6, and D-37 MG, with 2.9×104, 1.5×105, 8.6×105 and 1.59×106 EGFRs per cell, respectively. EGF significantly increased cell number in D-263 MG and D-37 MG by 65% and 74%, respectively, had no effect on D-247 MG, and significantly decreased cell number in U-343 MGa Cl 2∶6 by 39%. U-343 MGa Cl 2∶6 growth was inhibited 19% by Ab-528, but Ab-528 had no effect on growth of the other MHG lines. Ab-528 significantly inhibited all EGF-mediated growth effects. These studies demonstrate that, although Ab-528 alone has little antiproliferative activity on MGH, it successfully competes with EGF to reduce the biological effects of EGF-EGFR binding. Therefore, this antibody could potentially be used to target radioisotopes to MHG via the EGFR for diagnosis and therapy.

Tenascin expression in prostatic hyperplasia, intraepithelial neoplasia, and carcinoma

The expression of tenascin, an extracellular matrix glycoprotein, was studied in three human prostatic carcinoma cell lines by Northern and Western blot analyses and in human prostate tissues by immunohistochemistry and Western blot analysis. All three carcinoma cell lines expressed tenascin mRNA and protein, which were found predominantly in secreted form in culture supernatant. By immunohistochemistry, fetal prostatic tissue showed strong and diffuse tenascin immunoreactivity around developing glands. Normal adult prostatic tissue revealed only focal, scant periglandular and stromal immunoreactivity around acini and ducts. Most cases of hyperplasia and intraepithelial neoplasia showed variable periglandular immunostaining. Tenascin periglandular staining with diffuse stromal extension was noted with all grades of adenocarcinoma; however, the intensity was variable and appeared unrelated to the histologic grade. Metastatic prostatic carcinoma showed strong immunoreactivity in lymph nodes and bone marrow samples, with only weak reactivity of the normal connective tissue framework in both tissues. Western blot analysis of prostatic hyperplasia and carcinoma demonstrated the large and small isoforms of tenascin. These findings suggest a prominent role for tenascin in stromal alterations associated with both benign and malignant prostatic epithelial growth processes.

Mutation of the p53 tumor-suppressor gene is not a feature of endometrial hyperplasias

OBJECTIVEnMutation and overexpression of the p53 gene occur in approximately 20% of endometrial carcinomas. To determine whether alteration of the p53 gene is an early event in endometrial carcinogenesis, we examined the p53 gene in endometrial hyperplasias.nnnSTUDY DESIGNnGenomic deoxyribonucleic acid was extracted from 117 endometrial hyperplasias (36 simple, 40 complex, 41 atypical) and 30 endometrial cancers. Exons 5 through 8 of the p53 gene were amplified by means of the polymerase chain reaction. Mutations in the p53 gene were sought with single-stranded conformation polymorphism analysis and confirmed by direct deoxyribonucleic acid sequencing.nnnRESULTSnNone of 117 endometrial hyperplasias were found to have mutations in the p53 gene, whereas mutations were seen in three of 30 (10%) endometrial cancers (p < 0.02). The p53 mutations seen in three cancers were confirmed by direct sequencing (codons 157, 180, 272).nnnCONCLUSIONnBecause it does not appear to be a feature of endometrial hyperplasias, mutation of the p53 gene may represent a relatively late event in endometrial carcinogenesis.