Effect of five antineoplastic agents on tumor xenografts with different growth rates.

The effects of cyclophosphamide (Cy), doxorubicin (Dx), cisplatin (DDP), melphalan (L-PAM), and vincristine (VCR) on various human and animal tumor lines with different growth rates, growing as xenografts in NMRI (nu/nu) mice, were studied. Two types of response were observed: For Cy and Dx, the response of the xenografts was negatively correlated with tumor volume doubling time (TD), indicating that rapidly growing tumors were more sensitive to these drugs than were slowly growing tumors. For DDP, L-PAM, and VCR, the effects were positively correlated with the TD, indicating that slowly growing tumors were more sensitive to these drugs than rapidly growing tumors. The data are discussed in relation to the effects of the drugs on proliferating and nonproliferating cells obtained with other cell lines.

Polysomal polyadenylated RNA and albumin messenger RNA in Mastomys liver and in a chemically induced hepatocellular carcinoma.

A hepatocellular carcinoma line (H78) was established from a primary liver tumor induced in Mastomys natalensis by a single administration of dimethylnitrosamine. Six to 8 months after transplantation (passages 5 to 7), well-differentiated tumors, still containing glucogen-storing cells, were isolated and used for the preparation of RNA. Polysomal polyadenylated RNAs from Mastomys liver and from H78 tumor line were then compared by hybridization kinetics. Total kinetic complexities were 6.6 X 10(9) and 6.3 X 10(9) daltons for liver and tumor, respectively. Complexities of the high and middle abundant class were reduced in the hepatoma. Heterologous hybridization reactions revealed that, in terms of RNA mass, all or most of the polysomal polyadenylated RNA present in the liver was also present in the tumor and vice versa. However, shifts in the relative abundance of messenger RNA sequences were detected. In contrast to most other transplanted hepatomas, H78 has approximately the same content of albumin messenger RNA on its polysomes as has untreated liver.

Differential expression of intermediate-filament proteins in murine sarcoma 180 ascites or solid tumor.

The intermediate-filament proteins in Sarcoma 180 ascites cells and solid tumors generated by s.c. injection of ascites cells in NMRI or nude mice were analyzed by one- and two-dimensional gel electrophoresis and identified by immunological methods. The ascites form of Sarcoma 180 coexpresses keratin and vimentin, whereas the solid tumor ceases to synthesize keratins but continues to express vimentin. These reversible changes in the expression of intermediate-filament proteins may be due to a change in the differentiation program induced by environmental conditions like growth with or without cell contact.

Five-year follow-up study of independent clinical and flow cytometric prognostic factors for the survival of patients with non-small cell lung carcinoma.

Fresh surgical specimens of tumors of 187 patients with previously untreated non-small cell lung carcinomas were investigated by means of flow cytometry. The aim of the study was to look for cellular prognostic indicators for survival times of these patients in addition to the well-known clinical prognostic factors. All patients had a minimum of 5 years follow-up. Patients with aneuploid tumors had significantly shorter survival times than did those with diploid tumors (P less than or equal to 0.001). Identical results are obtained when the analysis is restricted to just those patients with T3 tumors or to patients with metastatic tumors at time of surgery or who were classified as Stage III (P less than or equal to 0.01). These data indicate that DNA ploidy is a strong and independent prognostic factor in patients with non-small cell lung carcinoma. Patients having tumors with a high proliferative activity died significantly (P less than 0.05) earlier than patients having tumors with lower proliferative activity. As with tumor ploidy, survival time in patients with high or low proliferative tumor activities was independent of whether the patients had T3-tumors, metastases, or were in Stage III. Univariate and multivariate analyses of the data in this study demonstrate two groups of independent prognostic factors for the survival of patients with non-small cell lung carcinoma: a group of clinical factors and a group of flow cytometric factors.

Prognostic relevance of ploidy, proliferation, and resistance-predictive tests in ovarian carcinoma.

In a cooperative study specimens of 37 patients with stage III and IV ovarian carcinomas who had been treated with chemotherapy were investigated utilizing flow cytometry and an in vitro short-term test for predicting resistance. Patients with aneuploid tumors had significantly shorter survival rates than did those with diploid tumors. Patients whose tumors showed a low G0/G1 cell proportion or a high proliferation pool (S- and G2/M cell-proportion) seemed to die earlier. There was also a tendency for patients with in vitro resistant tumors to die earlier under chemotherapy than those with sensitive tumors.

Albumin messenger RNA after partial hepatectomy and sham operation.

Albumin mRNA was quantified in nuclear RNA and in polysomal and post-polysomal poly(A)-containing RNA from untreated, hepatectomized and laparotomized rat livers. A prominent reduction of albumin-specific RNA sequences was observed in all subcellular fractions 50 h after partial hepatectomy, compared to both untreated and sham-operated livers. Surprisingly, laparotomy itself also induced a dilution of albumin-specific sequences at earlier times after operation in nuclei and in post-polysomal supernatant, but not in polysomes.

Effect of highly active antiretroviral therapy on survival in patients with AIDS-associated pulmonary Kaposi's sarcoma treated with chemotherapy.

PURPOSE: Kaposi's sarcoma (KS) is the most common AIDS-related malignancy. Pulmonary involvement by KS (PKS) has carried a poor prognosis with median reported survival ranging from 3 to 10 months. We studied whether the introduction of highly active antiretroviral therapy (HAART; triple antiretroviral therapy including a protease inhibitor and two reverse transcriptase inhibitors) has been associated with improved survival for AIDS patients with PKS. PATIENTS AND METHODS: A retrospective study was performed of 37 consecutive patients with PKS and human immunodeficiency virus infection in the tumor registry at a large municipal hospital in New York City between 1994 to 1997. There were 16 patients from 1994 to 1995 (pre-HAART period) and 21 patients from 1996 to 1997 (post-HAART period). The primary end point was survival, which was defined as time from start of chemotherapy until death from any cause. RESULTS: Patients were analyzed by the date of diagnosis (pre- v post-HAART period) and whether or not they received HAART. Kaplan-Meier analysis showed significantly better survival in patients diagnosed in the post-HAART period (P =.0025). Additional Kaplan-Meier analysis indicated that patients on HAART had substantially better survival (P <.0001). Cox multivariate analyses showed that HAART therapy was associated with a reduced risk of death (hazard ratio = 0.09; 95% confidence interval, 0.03 to 0.69). CONCLUSION: In patients with AIDS-associated PKS and undergoing chemotherapy, administration of HAART was associated with increased survival.

Biological characterization of subgroups of squamous cell lung carcinomas.

Recently, Pezzella et al. (Am. J. Pathol., 1997, 151: 1417-1423, 1997) reported on a subgroup of non-small cell lung carcinomas that had no morphological evidence of neoangiogenesis but appeared to grow and were highly aggressive. In this investigation, we subdivided 87 squamous cell lung carcinomas into four subgroups according to angiogenesis (low and high vessel density) and tumor growth (low and high tumor cell proliferation). The aim was to find differences, if any, in the angiogenic status and clinical behavior between these subgroups. We identified a group of tumors with low angiogenesis and high tumor cell proliferation that was characterized by high expression of vascular endothelial growth factor, low expression of basic fibroblast growth factor, reduced apoptosis, increased incidence of metastases, and short survival times. These data show that even squamous cell lung carcinomas are a heterogeneous group of tumors that can be subdivided in tumors with different biological properties and different clinical behaviors.

Angiostatin expression in non-small cell lung cancer.

Angiostatin, a potent inhibitor of angiogenesis, tumor growth, and metastasis, was examined in a panel of human lung cancer cell lines with Western blot analysis and in 143 primary non-small cell lung carcinomas with immunohistochemistry. Thirty-four of 143 cases (24%) stained positively. Patients with angiostatin-positive tumors survived longer (146 weeks) than patients with angiostatin-negative tumors (77 weeks; log-rank test: P = 0.07; rank-sum test: P = 0.02). To determine whether combining stimulating and inhibiting factors might improve the prognostic capability, both angiostatin and vascular endothelial growth factor (VEGF) were analyzed together with respect to patient survival. The median survival time of patients with angiostatin-positive/VEGF-negative carcinomas was 184 weeks, whereas the median survival time of patients with angiostatin-negative/VEGF-positive tumors was only 52 weeks. The angiostatin-positive tumors exhibited an increased incidence of apoptosis and a reduced capability to be transplanted into nude mice, but these differences did not reach or were only of borderline statistical significance.

Vascular endothelial growth factor in newly diagnosed and recurrent childhood acute lymphoblastic leukemia as measured by real-time quantitative polymerase chain reaction.

PURPOSE: Overexpression of vascular endothelial growth factor (VEGF) is associated with increased angiogenesis, growth, and metastasis in solid tumors, but to date the significance of VEGF in leukemia has received only limited attention. Therefore, this study examined the cellular VEGF levels in 31 newly diagnosed and 22 recurrent cases of childhood acute lymphoblastic leukemia (ALL). EXPERIMENTAL DESIGN: VEGF was determined with real-time quantitative PCR methods. Kaplan-Meier statistical analyses were conducted for the relapse-free intervals and the overall survival times. The groups were compared by log-rank and rank-sum tests. RESULTS: The VEGF levels were significantly higher in recurrent ALL compared with newly diagnosed ALL (28.0 versus 3.1 units; P = 0.001). Kaplan-Meier estimates were conducted to analyze the prognostic value of VEGF levels in newly diagnosed ALL with regard to the relapse-free intervals and the overall survival times. In this analysis, the median relapse-free interval of patients with low VEGF levels was more than 10 years, whereas the relapse-free interval of patients with high VEGF expression was only 1.2 years. The median overall survival time for the collective with low VEGF levels was >10 years, whereas the survival of the group of patients with high VEGF levels was 3.9 years. This difference was not statistically significant. This may be attributable to the small number of patients involved. CONCLUSION: Our data suggest that VEGF may play an important role in the pathophysiology of ALL. The expression of VEGF raises the possibility of using angiogenesis inhibitors as a novel therapeutic strategy in childhood ALL.

Development of tumor-infiltrating lymphocytes in breast cancer after neoadjuvant paclitaxel chemotherapy.

PURPOSE: Neoadjuvant chemotherapy for breast cancer creates new possibilities for the analysis of biological factors in the tumor and/or host, which may play a role in the response to treatment. In this study we analyzed whether changes in local antitumor immunity take place after neoadjuvant paclitaxel therapy and if they correlate with response to treatment. EXPERIMENTAL DESIGN: Neoadjuvant chemotherapy (paclitaxel, 200 mg/m2 q2w, 4 treatments) was followed by definitive surgical management. Histological sections from the pre- and post-treatment surgical specimens of 25 patients were analyzed for the extent of lymphocytic infiltration and presence of tumor infiltrating lymphocytes (TILs). The cumulative apoptotic response in the tumor after the first dose of paclitaxel was also studied in 10 of 25 patients. RESULTS: Pretreatment lymphocytic infiltrate in the tumor was minimal in the majority of patients and showed no relationship with clinical response. In the patients without TILs before treatment, development of TILs after treatment was noted in 0/3 (0%) patients with stable disease, 3/12 (25%) patients with clinical partial response, and 4/6 (67%) patients with clinical complete response and pathological residual disease. These correlated with the tumor cell apoptotic response to the first dose of paclitaxel. CONCLUSIONS: These results suggest that development of TILs after treatment correlates with clinical response to neoadjuvant paclitaxel therapy. The possible mechanism(s) whereby neoadjuvant chemotherapy may lead to induction of antitumor T cells is discussed. Immunological processes may influence the response of breast cancer patients to neoadjuvant treatment.

Paclitaxel-induced apoptosis and mitotic arrest assessed by serial fine-needle aspiration: implications for early prediction of breast cancer response to neoadjuvant treatment.

The extent of tumor reduction from neoadjuvant chemotherapy for breast cancer correlates with outcome. We investigated whether the initial cellular responses to paclitaxel are related to the extent of tumor reduction. Eleven women with breast cancer received paclitaxel (every 2 weeks for 4 cycles) as neoadjuvant treatment. Serial fine-needle aspirations (FNA; 25-gauge, 1 pass) were obtained before treatment and at 24, 48, 72, and 96 h after the first paclitaxel dose. Microscopic counts of apoptotic and mitotic indices were performed. The change in cancer volume from treatment was determined using radiological measurements with allowance for change in the histopathological amount of cancer. Apoptotic and mitotic responses usually subsided within 4 days. The duration of the initial apoptotic response was different for women with different treatment results. Cumulative apoptotic response for the first 4 days inversely correlated with the proportion of residual cancer after neoadjuvant treatment. FNA is a versatile clinical method to obtain breast cancer cells for therapy response studies. Apoptotic response to the first dose of paclitaxel is almost complete within 4 days, implying that more frequent (weekly) paclitaxel dosing might be beneficial. The apoptotic response to the first dose of paclitaxel appeared to predict the amount of cancer reduction from this treatment. This is a promising start toward the development of an early chemopredictive assay for paclitaxel treatment of breast cancer.

Concurrent paclitaxel and radiation therapy for breast cancer.

Few studies have evaluated the role of concurrent chemoradiation therapy in the management of locally advanced breast cancer. The availability of radiosensitizing chemotherapeutic agents that are effective in breast cancer and the encouraging results achieved by concurrent chemoradiation in other malignancies have prompted us to investigate this approach. Paclitaxel is a promising agent for use with concurrent radiotherapy because of its single-agent efficacy profile and its radiosensitizing effects. A clinical protocol of preoperative paclitaxel and radiation in locally advanced breast cancer is ongoing at our institution to test feasibility, measure pathologic response at mastectomy, and explore association of pathologic response with molecular tumor markers. Initially, the study was designed to test weekly paclitaxel at a dose of 60 mg/m2 during radiation therapy, delivered 5 days a week at 200 cGy fractions to a total dose of 50 Gy over 5 weeks. Due to severe skin toxicity in the first two patients, the protocol was amended to change the scheduling of paclitaxel to 30 mg/m2 twice weekly and to reduce the radiation to 180 cGy fractions to a total dose of 45 Gy, delivered 5 days a week over 5 weeks. Presently, 13 patients have been accrued; preliminary data indicate good tolerance to twice-weekly paclitaxel, and four of eight evaluable patients have achieved pathologic response (one patient who received the weekly regimen and three who received the twice-weekly regimen). In addition, sequential fine-needle aspirations of palpable breast cancers were obtained in patients enrolled in a parallel study of preoperative single-agent paclitaxel (200 mg/m2 every 2 weeks, for a total of four cycles before breast surgery). Preliminary results suggest that a steep increase in the mitotic index occurs during the first day after paclitaxel administration and plateaus between the second and the third day, then decreases to pretreatment values. The peak apoptotic index occurs at approximately 72 hours after paclitaxel administration and decreases at approximately 98 hours. These initial findings suggest that twice-weekly dosing of paclitaxel may optimize recruitment of cells into the G2/M phase of the cell cycle, the most radiosensitive phase.

Association between nm23-H1 expression, proliferation and apoptosis in non-small cell lung carcinomas.

Twelve non-small cell lung carcinomas and adjacent normal lung tissues were examined for mutations of the nm23-H1 gene by using SSCP analysis and for an expression of the nm23-H1 protein by immunohistochemistry. No mutations could be found in either the carcinomas or in the adjacent normal tissues. In contrast, six of 12 carcinomas showed protein expression while only one adjacent normal lung tissue yielded a positive staining result. Therefore, the expression of nm23-H1 protein was analysed in a larger group of non-small cell lung carcinomas (n = 185) to determine whether or not the expression of nm23 protein may be of prognostic relevance. Only a weak relationship between nm23-H1 expression and lymph node involvement was observed. However, a significant correlation between proliferation and nm23-H1 expression was detected. Additionally, a direct correlation between apoptosis and nm23-H1 expression or between myc and nm23-H1 expression was found. Finally, non-small cell lung carcinomas that expressed nm23-H1 protein were more frequently sensitive to doxorubicin than carcinomas that did not express this protein.

Analysis of c-fos, c-jun, c-erbB1, c-erbB2 and c-myc in primary lung carcinomas and their lymph node metastases.

The purpose of this study was to identify possible alterations in proto-oncogenes (c-fos, c-jun, c-erbB1, c-erbB2 and c-myc) at the protein level in primary lung carcinomas and simultaneous metastatic lymph nodes of 21 patients. The analysis showed that proteins of c-jun and c-myc were expressed in a significantly higher frequency in metastases than in primary lung tumors. Gross differences were not found between primary tumors and metastatic tumors with regard to the expression of c-erbB1, c-erbB2 and c-fos. The finding of cases with a higher expression of c-jun and c-myc in lymph nodes suggests that metastatic capability may be higher in certain cell populations.

Microvessel density, expression of proto-oncogenes, resistance-related proteins and incidence of metastases in primary ovarian carcinomas.

Relationships between the incidence of metastatic spread and microvessel density, expression of proto-oncogene products, or expression of resistance-related proteins were investigated in human ovarian carcinomas by immunohistochemistry. Ovarian carcinomas with a high microvessel density showed a significantly increased formation of metastases (P = 0.005). Tumors with positive immunoreactivity of c-jun and c-myc products had a higher metastatic spread; however, these results were not statistically significant. A marginally significant correlation existed between the expression of erbB1 (EGFR) and metastatic spread (P = 0.05). No significant relationship was found between the expression of the resistance-related proteins P-glycoprotein or glutathione S-transferase-pi and the incidence of metastases. Furthermore, no correlation was detected between expression of the heat shock protein 70 and the occurrence of metastases.

Expression of oncoproteins in primary human non-small cell lung cancer and incidence of metastases.

In the current study the relationship between the incidence of metastatic spread and expression (at the protein level) of various proto-oncogenes was investigated in 217 human non-small cell lung carcinomas. Tumors with an overexpression of proteins encoded by the oncogenes c-jun and c-myc showed a significantly increased formation of metastases (c-jun: P = 0.008; c-myc: P = 0.018). No significant correlations were found between the expression of the c-fos, c-erbB1, c-neu and c-ras products and metastatic spread.

Prognostic value of basic fibroblast growth factor and its receptor (FGFR-1) in patients with non-small cell lung carcinomas.

Tumour specimens of 206 patients with untreated non-small cell lung carcinomas (NSCLC) were analysed immunohistochemically for the expression of the basic fibroblast growth factor (bFGF) and for its receptor (FGFR-1, Flg). Seventy of the tumours showed weak expression, 109 moderate and 27 high expression of bFGF. Thirty-eight tumours had low expression of FGFR-1, 116 had moderate and 52 cases high expression. Patients with high FGFR-1 expression had significantly shorter survival times than patients with weak or moderate expressions (P < 0.05), but there was no significant correlation between bFGF expression and patient survival. The results of the multivariate analysis demonstrated that FGFR-1 in the presence of stage is not an independent prognostic factor.

High rate of expression of multidrug resistance-associated P-glycoprotein in human endometrial carcinoma and normal endometrial tissue.

The overexpression of P-glycoprotein was studied in 10 normal endometrial controls (five from the proliferative and five from the secretory phase of the menstrual cycle) and in 23 endometrial carcinomas of different histological varieties, using the C219 and JSB-1 monoclonal antibodies. Three of the tumours had been previously treated with combination chemotherapy containing doxorubicin. All endometrial carcinomas, whether treated or untreated, as well as the normal endometrial controls from both the proliferative and the secretory phase of the menstrual cycle, overexpressed P-glycoprotein. This puts endometrial carcinoma into the same category as other tumours arising in organs which normally overexpress P-glycoprotein, all of which tend to be intrinsically resistant to chemotherapy.

Association of cyclin D1 expression in lung cancer and the smoking habits of patients.

This study examined whether or not cyclin D1 expression is associated with the smoking habits of patients with non-small cell lung carcinomas (NSCLC). Immunohistochemistry was used to analyze 181 NSCLC samples for the expression of cyclin D1. Expression of cyclin D1 protein was found in 130 out of 181 cases (72%). A significant relationship between cyclin D1 expression and stage or histological classification was not observed. The carcinomas of smokers expressed cyclin D1 in 77% of the cases while carcinomas of non-smokers expressed this protein only 57% of the time (P < 0.01, Fisher's exact test). The correlation between smoking and cyclin D1 expression was maintained when the analysis was limited to squamous cell lung carcinomas. However, no correlation was found between cyclin D1 expression and the smoking habits of patients with adenocarcinomas. This can be explained by the fact that the development of adenocarcinomas--in contrast to squamous cell lung carcinomas--is not closely related to tobacco smoke.