Timothy E. Kute

c-erbB-2 Expression and Response to Adjuvant Therapy in Women with Node-Positive Early Breast Cancer

BACKGROUND The role of molecular markers in predicting the response to treatment of breast cancer is poorly defined. The Cancer and Leukemia Group B (CALGB) conducted a randomized adjuvant-chemotherapy trial (CALGB 8541) comparing three doses (high, moderate, and low) of cyclophosphamide, doxorubicin, and fluorouracil in 1572 women with node-positive breast cancer. This study (CALGB 8869) was designed to determine whether the DNA index, the S-phase fraction, c-erbB-2 expression, or p53 accumulation could be used as a marker to identify a subgroup of patients more likely than others to benefit from high doses of chemotherapy. METHODS Tissue blocks were obtained from 442 patients randomly selected from the larger CALGB trial. Paraffin sections from the primary lesions were analyzed for DNA content, S-phase fraction, c-erbB-2 expression, and p53 accumulation. RESULTS Patients randomly assigned to the high-dose regimen of adjuvant chemotherapy had significantly longer disease-free and overall survival if their tumors had c-erbB-2 overexpression. No further information was gained by adding the data on S-phase fraction or p53 accumulation to the analysis. There was no clear evidence of a dose-response effect in patients with minimal or no c-erbB-2 expression. CONCLUSIONS There is a significant dose-response effect of adjuvant chemotherapy with cyclophosphamide, doxorubicin, and fluorouracil in patients with overexpression of c-erbB-2 but not in patients with no c-erbB-2 expression or minimal c-erbB-2 expression. Overexpression of c-erbB-2 may be a useful marker to identify the patients who are most likely to benefit from high doses of adjuvant chemotherapy.

Development of Herceptin resistance in breast cancer cells

Herceptin, a humanized antibody to HER‐2, is now utilized in the clinic for metastatic breast cancer treatment. The response rate for HER‐2+ patients is only 30% and little is known as to mechanisms of resistance. The mechanism of Herceptin action is also unknown but has been related to cell cycle inhibition.

The relation of flow cytometry to clinical and biologic characteristics in women with node negative primary breast cancer

Flow cytometry (FC) analysis including DNA index (ploidy status) and cell kinetics (%S and %S + G2/M) was done on frozen tissue of the primary lesions of 101 women with node negative (N−) breast cancer who were studied prospectively. Currently, 19% (19/101) of the patients have recurred. No significant relations have been found between recurrence or survival and age, estrogen/progesterone receptor status, tumor size, and tumor type. The DNA index (ploidy) was not related to any clinical variable, time to recurrence, or survival. Aneuploid tumors did, however, have significantly higher %S phase activity. Patients with %S activity less than or equal to the median value were significantly different from those patients with %S above the median. They were older and had a higher frequency of ER/PR positive and well‐ or moderately differentiated tumors. Patients with %S + G2/M greater than the median value showed shorter time to recurrence (P = .055) and shorter survival (P = .006), whereas %S alone was significantly associated only with survival. Multivariate analysis showed that neither DNA index nor cell kinetics was significantly associated with time to relapse. DNA index was not significantly associated with survival; %S was of borderline significance whereas %S + %G2/M was a significant independent predictor of survival. Although FC data may provide independent information related to survival in N–women, additional research in a larger number of patients is needed to define its precise role in patient management.

Malignant papillary cystic tumor of the pancreas

An example of the rare papillary cystic tumor of the pancreas was diagnosed cytologically by aspiration of the primary neoplasm. Subsequently, it metastasized, proving its low‐grade malignant behavior. Diagnostic cytomorphologic features included abundant straight and branched papillary tissue fragments, and uniform, pale nuclei with folds or grooves. Although the primary tumor had a typical histologic appearance, metastases demonstrated increased nuclear pleomorphism and hyperchromasia, bizarre tumor giant cells, and an increased mitotic rate. Vimentin was diffusely positive, whereas neuron‐specific enolase and somatostatin were focally and weakly reactive. Neurosecretory and zymogen granules were absent ultrastructurally. By flow cytometric study, the tumor was aneuploid (DNA Index = 1.3).

Characterization of a human ovarian carcinoma cell line with estrogen and progesterone receptors

The potential significant therapeutic and prognostic roles for the sex steroid receptors in ovarian cancer are recognized. The authors present in detail the biochemical, morphologic, cytogenetic, and growth characteristics of an ovarian carcinoma cell line, BG‐1, which has functional estrogen and progesterone receptors (23 and 300 fmol/mg protein, respectively) in clinically significant levels. In particular, BG‐1 has a DNA index of 1.14, a stable karyotype with specific translocations, and produces and secretes CA 125 into the media.

Endometrial adenocarcinoma: A multiparameter clinicopathologic analysis including the DNA profile and the sex steroid hormone receptors

With endometrial adenocarcinoma, several factors, such as stage and histologic grade, are well recognized as having prognostic significance. Other variables, e.g., nuclear grade, probably are also important clinically. A preliminary investigation of the prognostic value of the ploidy, cell cycle kinetics, and sex steroid receptor contents of these tumors has been conducted and compared with more conventional prognostic factors such as stage, grades, and depth of invasion. As analyzed by flow cytometry, the proliferative activities of the carcinomas were significantly associated with prognosis. Tumors with lower levels of proliferative activity were related to improved survivals. Two thirds of the carcinomas were diploid; ploidy did not carry prognostic weight in the series. Both the estrogen and progesterone receptor status were related to survival. Patients whose tumors were positive for these receptors had a better prognosis than did those in which the receptors were not present in significant quantities.

Expression of Microtubule-Associated Protein 2 in Benign and Malignant Melanocytes : Implications for Differentiation and Progression of Cutaneous Melanoma

Cutaneous melanocytic neoplasms are known to acquire variable characteristics of neural crest differentiation. Melanocytic nevus cells in the dermis and desmoplastic melanomas often display characteristics of nerve sheath differentiation. The extent and nature of neuronal differentiation characteristics displayed by primary and metastatic melanoma cells are not well understood. Here, we describe induction of a juvenile isoform of microtubule-associated protein 2 (MAP-2c) in cultured metastatic melanoma cells by the differentiation inducer hexamethylene bisacetamide. Up-regulation of this MAP-2 isoform, a marker for immature neurons, is accompanied by extended dendritic morphology and down-regulation of tyrosinase-related protein 1 (TYRP1/gp75), a melanocyte differentiation marker. In a panel of cell lines that represent melanoma tumor progression, MAP-2c mRNA and the corresponding approximately 70-kd protein could be detected predominantly in primary melanomas. Immunohistochemical analysis of 61 benign and malignant melanocytic lesions showed abundant expression of MAP-2 protein in melanocytic nevi and in the in situ and invasive components of primary melanoma, but only focal heterogeneous expression in a few metastatic melanomas. In contrast, MAP-2-positive dermal nevus cells and the invasive cells of primary melanomas were TYRP1-negative. This reciprocal staining pattern in vivo is similar to the in vitro observation that induction of the neuronal marker MAP-2 in metastatic melanoma cells is accompanied by selective extinction of the melanocytic marker TYRP1. Our data show that neoplastic melanocytes, particularly at early stages, retain the plasticity to express the neuron-specific marker MAP-2. These observations are consistent with the premise that both benign and malignant melanocytes in the dermis can express markers of neuronal differentiation.

Relationship of flow cytometry results to clinical and steroid receptor status in human breast cancer

SummaryFlow cytometry (FC), estrogen receptor (ER), and progesterone receptor (PR) analyses were performed on 226 breast cancers. The presence of steroid receptors was inversely proportional to proliferative index and percent aneuploidy. Within the two ER (+ and −) groups, the presence of PR did not add significantly to the comparison. The mean proliferative index for the diploid tumors was 17.5±6.8 compared to 27.8±9.8 for aneuploid tumors (p<.001). The degree of aneuploidy, or DNA index, was not related to cell cycle kinetics or steroid receptor status. In 163 tissues analyzed for percent tumor present, a correlation between the relative number of aneuploid cells and percent tumor in the histologic review was observed. A study of the diploid tumors indicated greater than 75% had at least 10% tumor cells by histologic review. Since with FC one can observe at least 10% aneuploid cells in a tumor sample, it is our opinion that the percent aneuploidy in this study is not artifactually low due to sampling error. There was no significant relationship between nodal status or number of positive nodes and proliferative index, aneuploidy, or steroid receptor status. Metastatic tumors had a higher mean proliferative index, but this was not statistically significant. There was a relationship between age and proliferative index but not between age and ploidy status. In a small group of patients there was a trend for proliferative index and percent aneuploidy to be higher in the poorly differentiated and larger tumors when compared to the well differentiated and smaller tumors.

Relationship between Flow Cytometric Features and Clinical Behavior of Meningiomas

Meningiomas have a wide range of biological potential and clinical behavior. Histological findings are helpful in recognizing the malignant potential of a given tumor, but often fail to correlate with gross features, liability of recurrence, and extent of associated cerebral edema. To find alternate approaches to improve the correlation between biological and clinical behavior, 20 meningiomas were studied by flow cytometry (FC), an assessment that has been applied to meningiomas previously. Such FC features as DNA index (DI) and proliferative index (PI, %G2 + %S) were correlated with size, location, brain invasion, associated edema, and recurrence. Tumors with severe edema had significantly higher PIs (19.5 +/- 4.1) than those with moderate (12.6 +/- 4.5) or minimal (8 +/- 0) edema (P less than 0.05). The PI was greater than 16 in those tumors that recurred (n = 3) or invaded the brain (n = 3). Six tumors were aneuploid (DI, 1.33 +/- 0.17; PI, 17.1 +/- 5.3). These were uniformly large when compared with the diploid tumors, which were more variable in size. All of the aneuploid tumors were associated with moderate to severe cerebral edema. Two partly psammomatous tumors with high PIs and foci of high cellularity suggesting recent growth were associated with severe edema. One of these exhibited brain invasion. These preliminary data indicate that FC may have a potential use in the clinical management of meningiomas.

Low cathepsin D and low plasminogen activator type 1 inhibitor in tumor cytosols defines a group of node negative breast cancer patients with low risk of recurrence

Prognostic factors are highly needed to divide node negative breast cancer patients into groups of low versus high risk of recurrence and death. In order to invade and spread, cancer cells must degrade extracellular matrix proteins. Accordingly, tumor levels of molecules involved in this degradation might be associated with patient outcome. Previous work has demonstrated that high levels of the aspartyl protease cathepsin D in breast cancer are associated with a poor prognosis and similar findings have been reported for molecules involved in the urokinase pathway of plasminogen activation. Interactions between different protease systems have been described and data suggest that several proteolytic enzymes may be operable at the same time in a tumor. In the present study we measured cathepsin D (n=162), uPA (n=116), uPAR (n=109) and PAI-1 (n=135) in tumor cytosols obtained from a population of node negative breast cancer patients. A significant correlation was found between levels of uPA, uPAR, and PAI-1. Levels of cathepsin D were directly related to levels of uPA and uPAR. With a median observation time of 4.81 years, univariate survival analyses showed that high levels of uPA and cathepsin D significantly predicted a shorter disease free survival, while only high levels of cathepsin D were able to significantly predict a shorter overall survival. Tumor levels of uPAR and PAI-1 gave mixed results depending on the cut-off point choosen. Interestingly, multivariate analysis demonstrated that PAI-1 and cathepsin D were independent significant prognostic indicators for disease-free survival while only cathepsin D was helpful in overall survival. The five year relapse rate of patients with low PAI-1 and low cathepsin D was 13% while patients who had greater than the median value for both of these molecules had a 5 year relapse rate of 40%. These data would indicate that at least two different protease systems are active in spread of node negative breast cancer and that measurement of these molecules may aid in the decisions to be made when offering adjuvant treatment to these patients.