Search

Fifteen patients with advanced squamous cell cancer of the head and neck received weekly vindesine at doses of 3 mg/m2. No patient had received prior chemotherapy, though all had received either radiotherapy or radiotherapy and surgery. Fourteen patients were evaluable for response. Two patients had documented partial remissions. Dose-limiting neutropenia was the primary toxic effect observed. It was frequent and occasionally life-threatening. Vindesine has minor but real activity in this group of patients with advanced head and neck cancer who have not received prior chemotherapy.

The presence of an estrogen-regulated protein (24K) in normal, dysplastic, metaplastic, and neoplastic cervical epithelium correlates with histologic criteria for squamous cell maturation. The 24K protein, originally discovered in the MCF-7 breast cancer cell line, was studied in 51 cases by the modified immunoperoxidase Avidin-Biotin Complex method, using an anti-24K mouse monoclonal antibody. Immunostained sections were compared to hematoxylin-and-eosin-stained sections cut from the same tissue block. The 24K protein was observed to be located primarily in the parabasal or "prickle cell layer" of normal cervical tissue (6 of 7 normal cervical tissue specimens tested were positive for 24K protein. Specimens were obtained from surgery for nonneoplastic causes) and in all cases (12 of 12) of dysplasia and carcinoma in situ. Intercellular bridges of these cells showed prominent immunostaining in normal cervix and dysplasia. 24K protein was observed as a granular cytoplasmic stain in all cases of squamous metaplasia (5 of 5) and keratinizing squamous cell carcinoma (9 of 9), and in 8 of 14 cases of nonkeratinizing squamous cell carcinoma. In this latter group, immunostaining was confined to only those cells showing cytoplasmic eosinophilia on H&E sections. In no case was the presence of the 24K protein associated with areas of mature keratin. 24K immunostaining was also observed in the reserve cells of morphologically normal endocervical glands adjacent to areas of dysplasia and carcinoma. We conclude that 24K protein is associated with squamous cell maturation and may be an important marker of reserve cell hyperplasia and squamous metaplasia.

Megakaryocytes are the bone marrow cells that generate platelets. They are relatively rare cells, comprising between 0.03 and 0.06% of all nucleated marrow cells (R. L. Berkow et al., J. Lab. Clin. Med., 103: 811-818, 1984). The study of human megakaryocyte differentiation and function has been hampered by the small number of these cells available for study. Recently we have established a human cell line (EST-IU) from the marrow of a patient with an acute nonlymphocytic leukemia and a mediastinal germ cell tumor. While this cell line seems to express many of the phenotypic characteristics of human megakaryocytes, it does not appear to express any phenotypic properties associated with cells of the erythroid, lymphoid, granulocytic, or monocytic lineages. Transmission electron microscopy demonstrates frequent multinucleated cells. Staining for platelet peroxidase reactivity revealed darkening of the perinuclear envelope and the endoplasmic reticulum, a characteristic of cells of the megakaryocytic lineage. Indirect immunofluorescence assays reveal that EST-IU expresses reactivity with anti-platelet glycoprotein antisera, anti-Factor VIII-related antigen antisera, anti-Factor V antisera, anti-thrombocyte antisera, Tab (monoclonal anti-platelet glycoprotein IIb-IIIa), and anti-fibronectin antisera. Flow cytometry-derived DNA histograms demonstrate populations with multiple ploidies, ranging from approximately 4N to 32N. Based upon morphological and histochemical characteristics, antigenic expression, and nuclear characteristics, EST-IU cells appear to have a phenotype that closely resembles human megakaryocytes. This cell line should be useful in the further cell study of the molecular and cell biology of human megakaryocytopoiesis.

Several investigators have now confirmed our original report demonstrating the myelopoietic suppressive activity of lactoferrin (LF) in vitro. In order to further clarify this activity, we used the recently produced and purified neutralizing antibody (II 2C) to LF to set up an immunoradiometric assay specific for LF and to affinity purify LF from lysates of peripheral blood polymorphonuclear neutrophils (PMN) obtained from healthy donors. Iron-saturated purified PMN LF was as active as iron-saturated affinity purified milk LF as a suppressor of the release of granulocyte-macrophage colony stimulating factors (GM-CSF) from mononuclear human peripheral blood leukocytes. The activities of both the PMN LF and milk LF were inactivated by preincubation with monoclonal anti-LF antibody (II 2C). In order to evaluate the methods of iron saturation of LF in vitro as measures of their functional activities, milk LF was iron saturated by four different methods, including ferric citrate, ferric ammonium sulphate, ferric chloride with nitriloacetate, and ferric chloride alone. The functional characteristics of all four preparations of LF saturated with iron in vitro were relatively equal and were more active than native LF. Resident mouse peritoneal macrophages separated into subpopulations of GM-CSF-producing cells by velocity sedimentation were evaluated for their LF-receptor binding capacity and for sensitivity to the suppression of GM-CSF release by LF. Iron saturated LF suppressed release of GM-CSF from only those fractions containing LF-receptor bearing cells, although not all fractions containing cells bearing receptors for LF responded to the suppressive activity of LF. These studies provide further evidence for the myelopoietic regulatory activity in vitro of PMN-derived LF, which is mediated through populations of mononuclear phagocytes having receptors for LF.

Megakaryocytes are relatively rare components of human bone marrow, making the study of their maturation difficult. Phorbol esters can act as differentiating agents in a number of cell systems including murine megakaryocytes. We report the effects of phorbol esters on the previously described long-term human megakaryocytic leukemia cell culture, EST-IU. While two nontransforming phorbols fail to affect these cells, the transforming phorbol 12-O-tetradecanoylphorbol-13-acetate (TPA) induces a phenotype with characteristics of more mature megakaryocytes in a dose-related manner. This phenotype includes an increased adherence to untreated plastic or glass, polyploidization, an increase in cell size, and increased expression of both platelet glycoproteins and factor VIII-related antigen. Two-color flow cytometric analysis allowed simultaneous determinations of DNA content and the expression of surface membrane antigens or alpha-granule constituents, providing evidence that nuclear, membrane, and cytoplasmic maturation occur in parallel in this cellular system. TPA-induced maturation of EST-IU cells provides an important new cellular model for the further study of human megakaryocyte development.

Cisplatin's role in metastatic breast cancer was initially confined to heavily pretreated patients. It had relatively little activity in these patients, except when administered with high-dose intensity (greater than 33.3 mg/m2/week). More recent trials in previously untreated patients have suggested that cisplatin is an active single agent, with response rates approaching 50% when administered at a dose of 30 mg/m2/d for four days every 3 weeks. Cisplatin has been safely incorporated into combination chemotherapy regimens, and has been used in high-dose regimens in conjunction with autologous bone marrow transplantation. Future trials should examine the use of cisplatin or platinum analogues as first-line therapy in alternating non-cross-resistant regimens, and should attempt to circumvent the dose-limiting toxicities of cisplatin when used in a dose-intensive fashion.

The presence of an estrogen-regulated protein of Mr 24,000 (24K) was studied in 43 patients diagnosed as having endometrial adenocarcinoma. using an anti-24K monoclonal antibody, a modified immunoperoxidase system (avidinbiotin complex) was used to detect the presence of 24K in formalin-fixed, paraffin-embedded tissue samples. The positivity for 24K was correlated with low tumor histologic grade, few mitotic figures, few nucleoli, and a low degree of nuclear pleomorphism. These data suggest that 24K may be a potential marker of tumor differentiation.

Formalin-fixed, paraffin-embedded tissues from eight renal oncocytomas were analyzed using a Coulter Epics V cell sorter. Samples were processed using Hedley's method modified by including centrifugation through sucrose, RNAse digestion, and propidium iodide staining. The nuclei were excited using 200 mw of laser power at 488 nm. The tissues were all from surgical specimens acquired between 1972 and 1983, and the neoplasms met stringent light and electron microscopic criteria for the diagnosis. Sixteen samples were analyzed from the eight tumors; the coefficients of variation on the interpretable G1 peaks ranged from 3.95 per cent to 7.62 per cent with a mean of 5.46 per cent. One of the tumors had been frozen and yielded uninterpretably wide peaks. Control samples of non-neoplastic kidney from six of the specimens had coefficients of variation in a similar range. All of the oncocytomas with acceptable histograms had single, sharp G1 peaks with no evidence of aneuploidy. Proliferative indices were calculated and ranged from 0.02 to 0.18 for the oncocytomas and from 0.01 to 0.03 for renal cortex. The mean of the ratios of proliferative indices between the oncocytomas and the corresponding renal cortical samples was 4.5. These data support the concept that renal oncocytomas are euploid neoplasms with significant growth rates and are consistent with their benign biologic behavior and propensity to grow locally to large size.

Hydatidiform mole will progress to malignant gestational trophoblastic neoplasia (GTN) in some cases. Aneuploidy and high proliferative activity are associated with malignant tumors. Molar pregnancy tissue was considered a precursor to malignant GTN, and was studied retrospectively using paraffin-embedded tissue to determine whether aneuploidy or proliferative rates measured on molar tissue could predict a malignant course. Tissues from 51 complete hydatidiform moles were analyzed for nuclear DNA content by flow cytometric techniques. A chart review identified the clinical course after evacuation of the mole. A satisfactory DNA histogram was generated in 40 cases. Of the 40 patients, 22 (55%) had spontaneous resolution, and 18 patients (45%) required treatment for persistent GTN. The molar tissue was found to be euploid in 27 cases and aneuploid in 13 cases. Eight of the twenty-seven euploid cases (30%) required treatment after evacuation, whereas 10 of the 13 aneuploid cases (77%) required treatment after molar evacuation. Proliferative index (PI) was compared with treatment requirements. Average PI was 0.11 +/- 0.10 for the treatment group and 0.08 +/- 0.06 for the spontaneous resolution group. The correlation of clinical course with ploidy was significant (P less than 0.01). The association with proliferative index was not (P greater than 0.05). Aneuploidy, therefore, identifies a high-risk group of molar pregnancies, and may represent those that have undergone one stage of malignant transformation.

A monoclonal antibody (323/A3) with a high degree of selectivity for binding to breast cancer cells was produced by immunization of mice with MCF-7 human breast cancer cells. The antigen recognized by 323/A3 on MCF-7 appears to be surface localized, and by enzyme-linked immunosorbent assay, the antibody was found to bind strongly with four of six breast cancer cell lines examined while no binding was detectable with nonbreast cancer cell lines. In vivo distribution of the 323/A3 antigen was screened by immunoperoxidase staining of formalin-fixed paraffin sections of normal human tissues and tumors. Among breast tissues, positive staining was detected with 75% (6 of 8) of metastatic lymph nodes, 59% (76 of 128) of primary breast tumors, 20% (13 of 63) of benign breast lesions, and 0% (0 of 10) of normal breast. No immunostaining was detected with a large variety and number of other normal human tissues with the exception of staining observed with epithelium of normal colon. Antigen distribution appears not to be disease specific, since positive staining was also observed with adenocarcinomas other than breast. The antigen recognized by the 323/A3 antibody was identified by Western blot analysis as a Mr 43,000 protein. The glycoprotein nature of the antigen was demonstrated by its binding to concanavalin A, specific elution with sugar, and immunoprecipitation of a Mr 43,000 radiolabeled protein from extracts of MCF-7 cells after pulse labeling with [3H]glucosamine. The 323/A3 antigen appears to be the same Mr 43,000 protein in cell lines as in breast tumors in vivo. Based on a comparison with the molecular weights of other known tumor-associated antigens and with their immunocytochemical tissue distribution, the Mr 43,000 glycoprotein described here represents a tumor-associated antigen previously undescribed in breast cancer or in other tumors. Since the Mr 43,000 glycoprotein is present on the surface of most breast cancer cells and is either absent or expressed at very low levels in most normal tissues including normal breast, the monoclonal antibody described here may have potential applications in diagnosis and management of breast cancer.

The presence and distribution of a protein with a mol. wt of 24,000 (24K) was determined in endometrial biopsies from regularly cycling women and in women with endometrial carcinoma. This protein, of as yet unknown function and originally found in a breast cancer cell line, was detected by immunohistochemistry using a monoclonal antibody. In regularly cycling women, the 24K protein began to appear in the glandular epithelium during the late proliferative phase and decreased after ovulation. In contrast, in the superficial epithelium, the strongest immunostaining was observed during the secretory phase. Superficial epithelial cells expressed maximal 24K immunoreactivity around day 21 of the cycle and it was clearly seen in the bulbous projections of the apical cytoplasm. These results suggest that the 24K protein may be a marker for hormonal events in the endometrium during the menstrual cycle. In endometrial carcinoma, 24K was correlated with low tumor histologic grade, few mitotic figures, few nucleoli and a low degree of nuclear pleomorphism. These data suggest that 24K may be a potential marker of tumor differentiation.

Patients with advanced disseminated germ cell tumors of the testis, retroperitoneum, and mediastinum have impaired survival compared to other patients with disseminated germ cell tumors having less bulky metastatic disease. Among patients with advanced disseminated germ cell tumors, we currently lack adequate predictors of long-term survival. Flow cytometric analysis of the paraffin-embedded, formalin-fixed tumor blocks of 50 of these patients suggests that proliferative activity is significantly correlated with survival (p less than 0.001) in multivariate analysis. Log (beta-human chorionic gonadotropin) is the only other useful predictor of long-term survival in multivariate analysis of prognostic factors in this group of patients. Flow cytometric DNA analysis may be useful in predicting survival in patients with advanced disseminated germ cell tumors.

A mouse monoclonal antibody was prepared against purified and fully iron-saturated human breast milk lactoferrin (LF). This antibody was of the IgG1 subclass, and recognized LF biosynthesized in low-density normal human bone marrow cells and LF stored in normal human polymorphonuclear neutrophils. This antibody did not recognize purified and iron-saturated human transferrin. The antibody inactivated the suppressive effects of purified and iron-saturated human milk LF and LF present in crude extracts of normal polymorphonuclear neutrophils against the release of granulocyte-macrophage colony-stimulating factors from mononuclear blood leukocytes in vitro.