Immunotherapy with biochemically dissociated fractions of Propionibacterium acnes in a murine ovarian cancer model.

The antitumor effect of two strains of Propionibacterium acnes (PAI and PAII) and chemically derived fractions from the whole bacterial cell were studied using a murine ovarian teratocarcinoma (MOT) model. When injected i.p. in high doses (700 to 1400 micrograms/mouse), both strains produce survival of a significant proportion of tumor-bearing mice (30 to 90%). On a weight to weight basis, however, PAI was significantly more effective than PAII. PAI and PAII were extracted using pyridine, which yielded four fractions, i.e., pyridine-extracted strains PAI and PAII (PA-PEI and PA-PEII, respectively) which are composed of the cell wall material extracted by pyridine, and the residues of PA-PEI and PA-PEII (PA-RI and PA-RII, respectively) which are composed of the residue material following the chemical extraction. The chemical composition of PA-PEI was different from that of PA-PEII (the latter had proportionately three times as many carbohydrates and one-third of the protein content of the former) and so were their antitumor properties in the MOT model. PA-PEI had markedly reduced antitumor effect when compared to the untreated cell on a per weight basis. Furthermore, curability was only seen when using a high dose (1400 micrograms/mouse). By contrast, the cell wall components extracted by pyridine from PAII (PA-PEII) had powerful antitumor effects, i.e., greater than 50% of mice given 1400-micrograms injections survived. The material contained in PA-PEII was further fractionated on the basis of its organic solubility in chloroform:methanol solvent. The water-soluble and solvent-insoluble fractions retained most of the antitumor effects of PA-PEII, while the water-insoluble and solvent-soluble fractions were only moderately effective, suggesting that the active moiety(ies) was associated with the nonlipid components of this fraction. Both residue fractions (PA-RI and PA-RII) were as effective on a per weight basis in controlling the growth of 10(5) tumor inoculum as were whole untreated cells. However, periodate oxidation of PA-RI resulted in complete loss of its antitumor effects. When surviving mice that had no evidence of tumor persistence following a tumor challenge (10(5) MOT cells) and i.p. treatment with PA were subsequently rechallenged with 10(4) tumor cells, survival was significantly prolonged, as compared to tumor-challenged (10(4) MOT) naive mice. In addition, 10 to 20% of these rechallenged mice had complete eradication of the tumor inoculum (no evidence of disease for greater than 120 days).(ABSTRACT TRUNCATED AT 400 WORDS)

Single cell multiple biomarker analysis in archival breast fine-needle aspiration specimens: quantitative fluorescence image analysis of DNA content, p53, and G-actin as breast cancer biomarkers.

Fine-needle aspiration (FNA) is a sensitive and cost-effective method for evaluating breast lesions. However, the diagnosis of early premalignant lesions is less reliable by FNA because of a lack of distinctive cytological features. Accurately defining the risk of such lesions at the individual level may have significant impact in breast cancer prevention and management. The main objective of this preliminary study was to develop a method to study multiple biomarkers on archival FNA slides using quantitative fluorescence image analysis (QFIA). Biomarkers p53, G-actin, and DNA content were labeled with an immunofluorescence technique and measured by QFIA simultaneously on a single cell basis. QFIA allows the labeling and measurement procedures to be carried out in situ, without the need to remove cells from the slide while preserving the morphology of the cells. FNA slides from 72 incident patients were obtained for this study. Fifty-six cases had an adequate number of cells for the actual analysis (25 benign breast lesions, 14 proliferative breast diseases with nuclear atypia, and 17 malignant lesions). The DNA content (> or = 5c) and G-actin (average gray mean, > 90) were positive in 81% and 88% of malignant lesions, respectively. These were significantly higher than the corresponding positive rates in benign lesions (7% and 15%, respectively; P <0.01 for both). None of the benign cases were positive for G-actin and DNA simultaneously, and none of the malignant cases were negative for G-actin and DNA together. p53 was positive in 33% of malignant lesions and 8% of benign lesions (P >0.05). Our study demonstrates the feasibility of evaluating multiple biomarkers by QFIA on archival FNA-fixed specimens. The G-actin and DNA content assayed by QFIA may be potential intermediate end point markers for breast cancer individual risk assessment.

Comparative polymerase chain reaction analysis of c-myc amplification on archival breast fine-needle aspiration materials.

The oncogene c-myc is a key regulator of cell cycle progression (from G1 to S phase). The amplification of c-myc can either induce cell proliferation or apoptosis. As a part of our ongoing effort to develop methods for multiple tumor marker analysis, this study was carried out to determine whether biomarkers such as c-myc amplification could be analyzed on genetic materials collected from archival fine-needle aspiration (FNA) smears. A novel comparative PCR analysis was used to analyze c-myc amplification semiquantitatively. Genomic DNA was prepared using cells obtained from archival FNA materials that had undergone quantitative fluorescence image analysis (QFIA) for other biomarkers. Of the 72 cases selected from 1995 for this study, 53 had an adequate amount of DNA for analysis. A novel comparative PCR analysis was used to analyze c-myc amplification quantitatively. For each batch of experiments, DNA from the high c-myc expressing cells, HL-60, and DNA from the low expressing cells, K562, were served as positive and negative controls, respectively. c-myc amplification was observed in 16 (94.1%) of 17 malignant lesions, 5 (41.7%) of 12 proliferative breast diseases with nuclear atypia, and 4 (16.7%) of 24 other benign lesions (fibroadenoma or fibrocystic disease). The overall difference of c-myc expression among these groups was highly significant by chi2 analysis (P = 0.0002). We conclude that multiple phenotypic markers and genotypic markers may be combined in a risk assessment biomarker profile on small FNA samples that can be obtained on multiple occasions relatively noninvasively from the patient. The results of this study suggest that c-myc amplification may be a biomarker of breast cancer risk. However, additional large, prospective studies are needed to confirm the current observation.

Studies of esophageal balloon cytology in Linxian, China.

Esophageal cancer is the second leading cause of cancer death in China. Esophageal cancer has a very poor prognosis, principally because most tumors are asymptomatic until they are unresectable. Esophageal balloon cytology is an early detection method developed by Chinese scientists to identify resectable early cancers and precursor lesions. Previous studies have reported high sensitivities for detecting esophageal cancer in symptomatic patients. The current report describes several studies evaluating this diagnostic technique in asymptomatic individuals. A comparison of Chinese and U. S. cytological diagnoses of the same esophageal samples showed that the Chinese categories of precancerous neoplasia were more inclusive than the corresponding U. S. categories. Comparisons of both Chinese and U. S. cytological diagnoses with concurrent histological findings showed low (14-36%) sensitivities for the cytological detection of biopsy-proven cancers. Prospective follow-up studies of several screened cohorts showed a consistent progression of risk for developing esophageal cancer with increasing severity of initial cytological diagnosis. These preliminary studies suggest that esophageal balloon cytology is a useful technique that can benefit from additional research to improve its optimal performance.

Immunohistochemical evaluation of K-ras, p53, and HER-2/neu expression in hyperplastic, dysplastic, and carcinomatous lesions of the pancreas: evidence for multistep carcinogenesis.

The pathobiology of precursor lesions leading to invasive pancreatic adenocarcinoma remains a controversial area, but knowledge of the mechanisms of tumorigenesis may lead to possibly earlier detection, prevention, and treatment in the future. We hypothesize that ductal hyperplasia and dysplasia of the pancreas represent precursor lesions and are part of a continuous developmental spectrum evolving into ductal adenocarcinoma of the pancreas. To further define this sequence, we studied the immunohistochemical markers HER-2/neu, K-ras, and p53 in 15 adenocarcinomas and 15 nonmalignant specimens of the pancreas. The 15 nonmalignant specimens of the pancreas included both normal pancreas and chronic pancreatitis. Overall, HER-2/neu was positive in normal ducts, ductal hyperplasia, dysplasia, and carcinoma cells in 0 of 30, 11 of 20 (55%), 10 of 15 (67%), and 12 of 15 (80%), respectively, with progressive increase in the intensity of staining; p53 was positive in 1 of 30 (3%), 0 of 20, 3 of 15 (20%), and 13 of 15 (80%), respectively, and K-ras was positive in 1 of 30 (3%), 6 of 20 (30%), 11 of 15 (73%), and 8 of 15% (53%), respectively. These data support the hypothesis that ductal hyperplasia and dysplasia of the pancreas represent precursor lesions, and, in a fashion similar to that in colorectal tumorigenesis, pancreatic cancer seems to accumulate progressive genetic alterations.

Direct in situ hybridization for rapid detection of cytomegalovirus in bronchoalveolar lavage.

Rapid detection of cytomegalovirus (CMV) from pulmonary specimens in immunosuppressed persons may provide an origin for pneumonia. In situ DNA hybridization has been effective for detection of CMV in otherwise nondiagnostic histologic material. Studies comparing bronchoalveolar lavage (BAL) with open-lung biopsy have shown the former to be superior in detecting most pulmonary pathogens affecting immunocompromised patients. Fifty consecutive BAL specimens were studied to compare direct in situ DNA hybridization, routine tissue culture, and conventional cytologic examination to assess the efficacy of the hybridization technic to rapidly detect CMV. Using tissue culture as the standard, a sensitivity of 90% (28 of 31) and specificity of 63% (12 of 19) were observed with the CMV probe. Discrepant results between the probe and tissue culture were present in ten cases. There were seven probe-positive, culture-negative cases, three of which had systemic CMV infection, including two patients with inclusions noted by conventional cytologic examination. Three probe-negative, culture-positive cases were found. In the authors' laboratory, the predictive value of a positive CMV probe is 80% (28 of 35). In contrast to the probe, conventional cytologic examination revealed CMV inclusions in only 23% (7 of 31) of the culture-positive cases. An average of 21 days was required for CMV cultures to become positive; probe results were available within 24 hours. The authors conclude direct in situ DNA hybridization is a useful rapid method for the detection of CMV in BAL specimens submitted for cytologic examination.

Polymerase chain reaction-based K-ras mutation detection of pancreatic adenocarcinoma in routine cytology smears.

The cytologic diagnosis of pancreatic carcinoma is notoriously difficult, particularly in distinguishing benign atypia from well-differentiated adenocarcinoma. Mutation of codon 12 in the K-ras oncogene is frequently found with pancreatic cancers. Detection by polymerase chain reaction (PCR) followed by restriction endonuclease digestion can provide a powerful tool to improve and confirm diagnosis. The authors examined the utility of PCR-based detection in the diagnosis of pancreatic carcinoma using routinely obtained cytology smears that could be collected at most hospitals. Pancreatic cytology smears were collected retrospectively from 60 patients. DNA was extracted from the slides and amplified by PCR using mismatched primers that generated a Bst-N1 recognition site with the wild type codon 12 but not with the mutant allele. Results were compared with clinical follow-up. K-ras codon 12 mutations were observed in 44 of 46 (95.7%) cases of pancreatic cancer, but not in 12 benign cases nor in 2 cases of islet cell tumor. The amplification and digestion steps proved robust and sensitive, capable of detecting mutant K-ras alleles from cytology smears that contained only small foci of suspicious cells. Our results indicate that K-ras mutation analysis can be done reliably within 1 to 2 days from routine cytology slides without special handling, increasing the sensitivity of diagnosis in ambiguous cases while maintaining cost-effective and relatively noninvasive sampling strategy.

Malignant neoplasms arising in endometriosis.

Ten cases of malignant tumors arising in foci of gonadal and extragonadal endometriosis are reported and added to 195 previously reported cases from the English literature. The ovary was the primary site in 165 (78.7%) of the cases, whereas extragonadal sites represented 44 (21.3%). Endometrioid adenocarcinomas accounted for 69% of the lesions, clear-cell carcinomas 13.5%, sarcomas 11.6%, and rare cell types 6%. Extragonadal lesions were mostly endometrioid tumors (66%) and sarcomas (25%). Tumors arising in endometriosis were predominantly low grade and confined to the site of origin. Radiation therapy was often able to control completely tumors limited to the pelvis, but was not beneficial in metastatic disease. Only one patient had a response to chemotherapy. Fourteen patients received postoperative progestin therapy, with a 77% 5-year survival. Follow-up has been reported in 86 patients. The tumor was either confined to the ovary (57), confined to the extragonadal site of origin (11), or spread throughout the peritoneal cavity (18). With each of these situations, the 5-year survival was 65, 100, and 10%, respectively. Fourteen patients had malignant transformation in endometriosis associated with presumed estrogenic stimulation; most lesions (69%) were well differentiated and the 5-year survival was 82%. After surgical resection, we recommend that progestin therapy be included in the treatment of cancer arising in endometriosis. The actual frequency of malignancy arising in endometriosis may be higher than reported.

Malignancy arising in endometriosis associated with unopposed estrogen replacement.

Malignant transformation of endometriosis is a well documented phenomenon. Although it occurs most commonly in the ovaries, there have been approximately 50 reported cases of extraovarian malignant transformation of endometriosis. This paper presents two cases of malignancy arising from a dormant focus of endometriosis after total abdominal hysterectomy, bilateral salpingo-oophorectomy, and exogenous estrogen replacement therapy. These malignancies are often well differentiated and may behave similarly to estrogen-induced endometrial carcinomas. After surgical castration of a premenopausal woman with endometriosis, the use of progestins in replacement therapy may reduce the risk of malignancy arising in endometriosis.

Primary cytoreductive surgery for epithelial ovarian cancer.

Forty-seven patients with stage III or IV invasive epithelial carcinoma of the ovary underwent primary cytoreductive surgery at UCLA during the five-year period 1974 to 1979. Optimal cytoreduction (defined as largest residual tumor mass 1.5 cm or less in diameter) was achieved in 31 patients (66%), including ten of 14 (71%) who underwent laparotomy and biopsy before referral. Median survival for the suboptimal group was six months, compared with 18 months for patients whose largest residual disease was 0.5 to 1.5 cm, and 40 months if residual nodules were less than 0.5 cm (P less than .001). All patients in the suboptimal group died of disease from 22 months to seven years and four months postoperatively. Given the limited ability of chemotherapy to cure ovarian cancer, and the acceptable morbidity of extended operation, the availability of ideal initial surgical effort for patients with advanced stage disease may be the most important variable in current ovarian cancer care. Optimal cytoreduction is most effective in prolonging survival in patients first seen without clinical ascites or large metastatic disease.

Psammocarcinoma with an aggressive course.

BACKGROUND: Psammocarcinoma is an unusual variant of serous cystadenocarcinoma characterized by heavy deposits of psammoma bodies. This disease has been suggested to be similar to carcinomas of low malignant potential in its indolent clinical course. We present this case report of an aggressive course of this disease to alert others that psammocarcinoma may not always follow a benign course. CASE: A 66-year-old woman underwent staging laparotomy for bilateral ovarian cystadenofibromata with rare foci of borderline serous tumors and several small bowel peritoneal surface nodules showing infiltrating psammocarcinoma. She was not recommended for adjuvant therapy because of the previously reported indolent course of this disease. Eighteen months later she represented with small bowel obstruction and underwent an exploratory laparotomy that demonstrated diffuse recurrence of the psammocarcinoma. CONCLUSION: Psammocarcinoma may have a more aggressive course than has been suggested. Patients with this disease should have optimal tumor debulking. There may be a role for adjuvant therapy in its treatment.

Second-look laparotomy in stage III epithelial ovarian cancer: clinical variables associated with disease status.

Variables associated with a negative second-look laparotomy in patients with stage III epithelial ovarian carcinoma are analyzed. Fifty-six patients were clinically free of disease after systemic chemotherapy and were subjected to second-look laparotomy to assess tumor status. Eighteen of these patients (32.1%) had no evidence of malignancy. Eight (14.3%) additional patients with no gross evidence of disease at laparotomy had microscopic persistence; five of these had disease documented in the pelvic or para-aortic lymph nodes. Significant variables associated with a histologically and cytologically negative second-look operation were low tumor grade (P less than .01), the use of cis-platinum containing combination chemotherapy (P less than .01), patient age less than or equal to 50 years (P less than .02), small residual tumor (less than 0.5 cm) before chemotherapy (P less than .05), and metastatic tumor less than or equal to 10 cm before initial cytoreduction (P less than .05). Patients treated with six to nine cycles of combination chemotherapy had the same probability of a negative second-look laparotomy as those treated with ten to 12 cycles. Multivariate discriminate analysis indicated that patients with low tumor grade, those receiving cis-platinum containing combination chemotherapy, and those with minimal residual tumors (less than 0.5 cm) after primary cytoreductive surgery correctly classify second-look status in 78.6% of patients. Until a nonsurgical method of monitoring subclinical disease is available, a through second-look laparotomy, including a pelvic and para-aortic lymphadenectomy, should be performed.

Individualization of treatment for stage I squamous cell vulvar carcinoma.

Of 177 cases of invasive squamous cell vulvar cancer seen at the University of California at Los Angeles and the City of Hope National Medical Center from 1957 to 1980, 84 (47.5%) had stage I disease. Seventy-seven patients with stage I disease (91.7%) had stromal invasion of 5 mm or less. Correlation between lymph node status and depth of invasion was as follows: 1 mm or less, none of 34 (0%); 1.1 to 2 mm, two of 19 (10.5%); 2.1 to 3 mm, two of 17 (11.8%); 3 to 5 mm, one of seven (14.3%); more than 5 mm, three of seven (42.9%). Fifty-six patients had radical vulvectomy for the primary lesion, and 28 had more conservative excision, but the incidence of local invasive recurrence (4%) was the same in each group. None of 58 patients treated with inguinal-femoral lymphadenectomy developed a groin recurrence, but three of 26 patients (11.5%) who had omission or modification of inguinal-femoral lymphadenectomy died with groin recurrence within 12 months. These data suggest that although some modification of the standard radical vulvectomy is appropriate for the primary lesion in patients with stage I disease, patients with greater than 1 mm of stromal invasion require at least an ipsilateral inguinal-femoral lymphadenectomy.

Peritoneal adenocarcinoma (serous) of Müllerian type: a subgroup of women presenting with peritoneal carcinomatosis.

Peritoneal adenocarcinoma (serous or other subtype) of Müllerian type (PAMT) is frequently misclassified as another primary tumor. Peritoneal carcinomatosis in women without evidence of a primary site may occur secondary to a number of processes. Confusion regarding the nomenclature has made it difficult to determine the incidence and natural history of this unique malignancy. Other terms used for this tumor include mesothelioma, peritoneal papillary serous carcinoma, extra-ovarian serous carcinoma, and normal-sized ovarian carcinoma syndrome. Thirty-four patients (33 serous and one endometrioid) were identified with PAMT during 1976 through 1988. One hundred and thirty-seven patients underwent primary cytoreductive surgery for a preoperative diagnosis consistent with ovarian cancer. Twenty-nine (21.2%) were classified as PAMT (5 of the 34 had their initial surgery at other institutions). The mean age was 61.4 years. The primary symptoms and signs were abdominal pain (68%) and ascites (52%). Twenty-five (73%) had a preoperative diagnosis of ovarian cancer while the postoperative diagnosis was unknown (44%), PAMT (29%), and ovarian cancer (27%). Univariate and multivariate survival analysis were performed. Survival was independent of age, residual disease, grade, ascites, type of chemotherapy, and second-look results. In patients with residual disease < 1.5 cm, extended survival was found in (hose with ascites < 1000 ml, residual disease in pelvis only, and small residual volume but statistical significance was not obtained. Twenty-eight patients received >/=4 courses of chemotherapy after primary surgery. Twelve of 21 patients (57%) who received cisplatin (CDDP) survived between 23 and 92 months, while no patient receiving other chemotherapeutic regimens survived more than 25 months. The 2 and 3 year survival rate for CDDP was 47% and 33% vs. 14% and 0% for other regimens. Optimal cytoreductive surgery was not an independent prognostic factor as found in ovarian cancer, probably secondary to unresectable peritoneal carcinomatosis. PAMT is sensitive to chemotherapy but only the use of CDDP was associated with long term survival. Based on these results, women with peritoneal carcinomatosis consistent with PAMT should receive a CDDP-based regimen after primary surgery.

Peroperative pancreatic aspirations.

Peroperative fine needle pancreatic aspirations were performed on 16 patients with suspected pancreatic masses during 1976 and 1977. Seven of the 16 aspirates demonstrated benign cells. Nine of the 16 revealed malignant cells, eight of which could be classified as to tumor type. This procedure provided a definitive diagnosis resulting in the appropriate surgery. The aspiration technique is highly accurate, rapid, uncomplicated and eliminates the problems associated with biopsy of this organ.

The use of cytology to evaluate pericardial effusions.

Pericardial effusions from 27 patients were examined cytologically during the five-year period of 1973-1977. Malignant cells were found in eight cases (30 percent). In three of these patients malignancy was unsuspected clinically, and this was the first time the cancer was diagnosed. In addition, cytology often suggested the specific histological types and possible primary sites to be determined. Special stains were also found helpful. There were no false positive reports. Although the pericardial effusions from the remaining 19 patients were negative for tumor cells, metastatic carcinoma to the pericardium was discovered at autopsy in two of these cases. Thus, cytologic examination of pericardial fluid is an important tool in the diagnosis of malignancy, but false negative results may occur.

Ependymoma-like tumor of the mesovarium.

We report a case of primary extramedullary ependymoma, arising from the mesovarium in a 47-year-old woman and discovered incidentally during routine hysterectomy for uterine leiomyomas. The tumor exhibited the light microscopic, immunohistochemical, and ultrastructural features of a typical ependymoma, including a prominent pseudorosette pattern, positive staining for glial fibrillary acidic protein, and electron microscopic visualization of microvilli, intermediate filaments, basement membranes, and long cytoplasmic processes. While the occurrence of subcutaneous ependymomas in the sacrococcygeal region and metastatic central nervous system tumors at various sites are familiar, we believe this case to be a rare example of a primary ependymoma arising in a different and unusual extramedullary location.

Amyloidoma of the skull base.

Localized amyloid tumors of the central nervous system are rare. We present the case of a 59-year-old white man with a large expansile amyloidoma at the base of the skull. To our knowledge this is the second reported case of localized amyloid tumor involving the skull.

Mixed mesodermal sarcoma of the ovary in a young patient.

Mixed mesodermal tumors (MMT) of the ovary are rare and have a poor prognosis. This ovarian malignancy usually occurs in postmenopausal women. We report an unusual ovarian MMT in a young woman given treatment similar to one used for ovarian germ cell malignancies. We believe this is the youngest patient reported with an homologous MMT of the ovary.

The cytobrush for evaluating routine cervicovaginal-endocervical smears.

While the conventional wooden spatula/cotton-tipped applicator has remained the method of obtaining Papanicolaou smears for most clinicians at the University of California at Los Angeles, the Student Health Service (SHS) adopted the Zelsmyr cytobrush as its sole method of obtaining cervical samples in late 1986. A study was done to define changes in both the adequacy of sampling and the detection rate for both squamous and glandular epithelial abnormalities with the cytobrush. To accomplish this goal, 1,000 cytobrush and 244 conventionally obtained smears were analyzed prospectively, and 3,864 SHS samples obtained in 1986 prior to the change in method were reviewed retrospectively. As compared to cervical samples obtained by conventional methods, the cytobrush smears contained significantly more endocervical cells and had fewer drying artifacts. Both methods obtained equivalent squamous samples and had similar final class distributions. No case of endocervical adenocarcinoma carcinoma in situ or invasive adenocarcinoma was detected. SHS patients who had initial "no endocervical cells" samples but whose repeat sample did contain endocervical cells retained the same class in more than 75% of cases. This study confirmed that the cytobrush technique produces Papanicolaou smears with improved sampling of the squamocolumnar junction but questioned whether that results in an increased detection rate for cervical pathology.