Peter W. Nichols

ACCUMULATION OF NUCLEAR P53 AND TUMOR PROGRESSION IN BLADDER CANCER

BACKGROUND We have previously demonstrated a strong association between nuclear accumulation of p53 protein, as determined by immunohistochemical analysis, and mutations in the p53 gene. The purpose of this study was to determine the relation between nuclear accumulation of p53 and tumor progression in transitional-cell carcinoma of the bladder. METHODS Histologic specimens of transitional-cell carcinoma of the bladder (stages Pa, noninvasive disease, to P4, disease with direct extension into adjacent organs or structures) from 243 patients who were treated by radical cystectomy were examined for the immunohistochemical detection of p53 protein. Nuclear p53 reactivity was then analyzed in relation to time to recurrence and overall survival. RESULTS The detection of nuclear p53 was significantly associated with an increased risk of recurrence of bladder cancer (P < 0.001) and with decreased overall survival (P < 0.001). In patients with cancer confined to the bladder, the rates of recurrence for stage P1, P2, and P3a tumors that had no detectable nuclear p53 reactivity at five years were 7, 12, and 11 percent, respectively, as compared with 62, 56, and 80 percent, respectively, for tumors that had p53 immunoreactivity. Similar results were obtained when the presence or absence of p53 in the nuclei of the tumor cells was studied in relation to overall survival. In a multivariable analysis stratified according to grade, pathological stage, and lymph-node status, nuclear p53 status was an independent predictor (and in cancer confined to the bladder, the only independent predictor) of recurrence and overall survival (P < 0.001). CONCLUSIONS In patients with transitional-cell carcinoma confined to the bladder, an accumulation of p53 in the tumor-cell nuclei detected by immunohistochemical methods predicts a significantly increased risk of recurrence and death, independently of tumor grade, stage, and lymph-node status. Patients with transitional-cell carcinoma confirmed to the bladder that demonstrates nuclear p53 reactivity should be considered for protocols of adjuvant treatment.

The Role of Adjuvant Chemotherapy Following Cystectomy for Invasive Bladder Cancer: A Prospective Comparative Trial

We assigned 91 patients with deeply invasive, pathological stage P3, P4 or N+ and Mo transitional cell carcinoma of the bladder (with or without squamous or glandular differentiation) to adjuvant chemotherapy or to observation after radical cystectomy and pelvic lymph node dissection. For most patients chemotherapy was planned as 4 courses at 28-day intervals of 100 mg./M.2 cisplatin, 60 mg./M.2 doxorubicin and 600 mg./M.2 cyclophosphamide. A significant delay was shown in the time to progression (p = 0.0010) with 70% of the patients assigned to chemotherapy free of disease at 3 years compared to 46% in the observation group. Median survival time for patients in the chemotherapy group was 4.3 years compared to 2.4 years in the observation group (p = 0.0062). In addition to treatment groups, important prognostic factors included age, gender and lymph node status. The number of involved lymph nodes was the single most important variable. We recommend adjuvant chemotherapy for patients with invasive transitional cell carcinoma after definitive surgical resection.

The Rationale for EN Bloc Pelvic Lymph Node Dissection for Bladder Cancer Patients with Nodal Metastases: Long-Term Results

From August 1971 through June 1989, 591 consecutive patients underwent curative pelvic lymphadenectomy with en bloc radical cystectomy for bladder cancer. Of these patients 132 (22%) had pathologically proved nodal metastases. The incidence of positive nodes increased with increasing pathological stage of the primary tumor: stage PIS (0.75%), stage P1 (13%), stage P2 (20%), stage P3a (24%), stage P3b (42%) and stage P4 (45%). The median followup for the 31 patients still alive was 5.5 years (range 2.6 to 18.8). Recurrent bladder cancer was documented in 89 patients (67%) with a median interval to progression of 1.5 years. Pelvic recurrence as the first site of progression was uncommon, occurring in 15 patients (11%). The actuarial 2, 3, 5 and 10-year survival rates were 55%, 38%, 29% and 20%, respectively. Increased risk of progression and death was associated with advanced pathological tumor stage (stage P3b or greater, p < 0.001 and p < 0.001, respectively) and 6 or more positive nodes (p < 0.001 and p = 0.012, respectively). There was no significant difference in survival and interval to progression among patients who received preoperative irradiation or adjuvant chemotherapy compared to those treated with surgery alone. This retrospective analysis further substantiates the philosophy that single stage pelvic lymphadenectomy with en bloc radical cystectomy can provide long-term progression-free survival, particularly for patients with localized primary tumors and minimal metastatic nodal disease.

Detection of methylated apoptosis-associated genes in urine sediments of bladder cancer patients.

Purpose: There is increasing evidence for a fundamental role for epigenetic silencing of apoptotic pathways in cancer. Changes in DNA methylation can be detected with a high degree of sensitivity, so we used the MethyLight assay to determine how methylation patterns of apoptosis-associated genes change during bladder carcinogenesis and whether DNA methylation could be detected in urine sediments. Experimental Design: We analyzed the methylation status of the 5′ regions of 12 apoptosis-associated genes (ARF, FADD, TNFRSF21, BAX, LITAF, DAPK, TMS-1, BCL2, RASSF1A, TERT, TNFRSF25, and EDNRB) in 18 bladder cancer cell lines, 127 bladder cancer samples, and 37 samples of adjacent normal bladder mucosa using the quantitative MethyLight assay. We also analyzed the methylation status in urine sediments of 20 cancer-free volunteers and 37 bladder cancer patients. Results: The 5′ regions of DAPK, BCL2, TERT, RASSFIA, and TNFRSF25 showed significant increases in methylation levels when compared with nonmalignant adjacent tissue (P ≤ 0.01). Methylation levels of BCL2 were significantly associated with tumor staging and grading (P ≤ 0.01), whereas methylation levels of RASSF1A and ARF were only associated with tumor stage (P ≤ 0.04), and TERT methylation and EDNRB methylation were predictors of tumor grade (P ≤ 0.02). To investigate clinical usefulness for noninvasive bladder cancer detection, we further analyzed the methylation status of the markers in urine samples of patients with bladder cancer. Methylation of DAPK, BCL2, and TERT in urine sediment DNA from bladder cancer patients was detected in the majority of samples (78%), whereas they were unmethylated in the urine sediment DNA from age-matched cancer-free individuals. Conclusions: Our results indicate that methylation of the 5′ region of apoptosis-associated genes is a common finding in patients with bladder carcinoma. The ability to detect methylation not only in bladder tissue, but also in urine sediments, suggests that methylation markers are promising tools for noninvasive detection of bladder cancers. Our results also indicate that some methylation markers, such as those in regions of RASSF1A and TNFRSF25, might be of limited use for detection because they are also methylated in normal bladder tissues.

Hypomethylation of a LINE-1 Promoter Activates an Alternate Transcript of the MET Oncogene in Bladders with Cancer

It was recently shown that a large portion of the human transcriptome can originate from within repetitive elements, leading to ectopic expression of protein-coding genes. However the mechanism of transcriptional activation of repetitive elements has not been definitively elucidated. For the first time, we directly demonstrate that hypomethylation of retrotransposons can cause altered gene expression in humans. We also reveal that active LINE-1s switch from a tetranucleosome to dinucleosome structure, acquiring H2A.Z- and nucleosome-free regions upstream of TSSs, previously shown only at active single-copy genes. Hypomethylation of a specific LINE-1 promoter was also found to induce an alternate transcript of the MET oncogene in bladder tumors and across the entire urothelium of tumor-bearing bladders. These data show that, in addition to contributing to chromosomal instability, hypomethylation of LINE-1s can alter the functional transcriptome and plays a role not only in human disease but also in disease predisposition.

The Role of Radical Cystectomy in the Management of High Grade Superficial Bladder Cancer (PA, P1PIS and P2)

Between January 1979 and 1987, 411 consecutive patients were considered candidates for bilateral pelvic iliac lymph node dissection and radical cystectomy for the management of bladder cancer. From this group 160 were identified as having pathological stage P2 or less disease, including 11 who also had positive nodes. The 5-year actuarial survival rate for the respective stages at 95% confidence limits was 100% for stage P0/A, 80% for stage P1, 78% for stage P1 with stage PIS, 85% for pure stage PIS, 76% for stage P2 and 87% for stage P2 with stage PIS. Additionally, we identified a group of patients with stage P2 transitional cell carcinoma who were at significant risk for development of metastatic disease. Of 46 patients with stage P2 transitional cell carcinoma 18 had vascular space invasion resulting in 6 of 18 cancer-related deaths (33%). Our study demonstrates that radical cystectomy has been highly effective in curing patients with high grade superficial disease, including those with superficially invasive disease associated with nodal metastases.

Lymph Node Dissection Technique Is More Important Than Lymph Node Count in Identifying Nodal Metastases in Radical Cystectomy Patients: A Comparative Mapping Study

BACKGROUND The value of lymph node dissection (LND) in the treatment of bladder urothelial carcinoma is well established. However, standards for the quality of LND remain controversial. OBJECTIVE We compared the distribution of lymph node (LN) metastases in a two-institution cohort of patients undergoing radical cystectomy (RC) using a uniformly applied extended LND template. DESIGN, SETTING, AND PARTICIPANTS Patients undergoing RC at the University of Southern California (USC) Institute of Urology and at Oregon Health Sciences University (OHSU) were included if they met the following criteria: (1) no prior pelvic radiotherapy or LND; (2) lymphatic tissue submitted from all nine predesignated regions, including the paracaval and para-aortic LNs; (3) bladder primary; and (4) category M0 disease. The number and location of LN metastases were prospectively entered into corresponding databases. MEASUREMENTS LN maps were constructed and correlated with preoperative and pathologic characteristics. Kaplan-Meier curves were constructed to estimate overall survival (OS) and recurrence free survival (RFS) among LN-positive (LN+) patients. RESULTS AND LIMITATIONS Inclusion criteria were met by 646 patients (439 USC, 207 OHSU), and 23% had LN metastases at time of cystectomy. Although there was a difference in the median per-patient LN count between institutions, there were no significant interinstitutional differences in the incidence or distribution of positive LNs, which were found in 11% of patients with ≤pT2b and in 44% of patients with ≥pT3a tumors. Among LN+ patients, 41% had positive LNs above the common iliac bifurcation. Estimated 5-yr RFS and OS rates for LN+ patients were 45% and 33%, respectively, and did not differ significantly between institutions. CONCLUSIONS LN metastases in regions outside the boundaries of standard LND are common. Adherence to meticulous dissection technique within an extended template is likely more important than total LN count for achieving optimal oncologic outcomes.

Lymphomatoid granulomatosis: A clinicopathologic study of 42 patients

&NA; We studied the histological and clinicopathological findings in 42 patients who had lymphomatoid granulomatosis (LYG). In addition to small round lymphocytes, small to intermediate lymphocytes with serpentine nuclei, large immature mononuclear lymphoid cells, abundant histiocytes, and vascular invasion by the cell infiltrate were observed in all cases. Fifty percent of lesions had occasional “atypical” cells with multi‐lobed nuclei. Three of four follow‐up autopsies showed large cell lymphoma, while one other autopsy and the single repeat biopsy showed increased numbers of large immature mononuclear lymphoid cells. Patients were most frequently men 40–60 yr old who had a history of pulmonary symptoms, such as cough or chest pain, and who showed multiple bilateral lung nodules without hilar adenopathy in the chest x‐ray. Thirteen patients (38%) died of disease, 11 of them within 12 mth of initial diagnosis. The presence of neurological signs and symptoms, increased mitoses, or increased numbers of atypical multi‐nucleated cells in the initial biopsy were not statistically significant predictors of survival.

Lymphomatoid granulomatosis: A T-cell disorder?

Abstract The histogenesis of lymphomatoid granulomatosis has been controversial since it was first defined by Liebow and colleagues. We report a typical clinical and histopathologic case of lymphomatoid granulomatosis. Immunologic markers indicate the lymphocytes comprising this patients initial lesion were predominantly T lymphocytes.

A Clinical, Histopathologic, and Electron Microscopic Study of Pneumocystis carinii Choroiditis

We studied the clinical and histopathologic features of Pneumocystis carinii choroiditis in three patients with acquired immunodeficiency syndrome. In two cases, a provisional diagnosis of disseminated P. carinii infection was made by ophthalmologic examination. The characteristic fundus changes in this infection consisted of numerous slightly elevated, plaque-like, yellow-white lesions located in the choroid and unassociated with signs of intraocular inflammation. The diagnosis was confirmed by postmortem examination of the eyes and other organs. Histopathologically, the globes showed many choroidal infiltrates that were eosinophilic, acellular, vacuolated, and frothy. Several such infiltrates were noted within the choroidal vessels and choriocapillaries. Gomoris methenamine silver stain demonstrated many cystic and crescentic organisms. Electron microscopy disclosed thick-walled cystic organisms and large numbers of trophozoites.