Effects on the immune system and toxicity of carboplatin/paclitaxel combination chemotherapy in patients with stage III-IV ovarian and non small cell lung cancer and its role in survival and toxicity.

PURPOSE: To examine the impact of paclitaxel and carboplatin combination chemotherapy on the parameters of the immune system in patients with non small cell lung cancer (NSCLC) and with ovarian cancer before, during and after chemotherapy, and the effect of this combination on the overall patient survival. METHODS: 24 patients with NSCLC and 20 with ovarian cancer (all in stage IIIb-IV) treated with 6 courses of paclitaxel and carboplatin combination chemotherapy were separated into two groups according to their survival group A: long survival (> 12 months for NSCLC; > 30 months for ovarian cancer) group B: short survival (<12 months for NSCLC; <30 months for ovarian cancer). At the same time we studied some immunological parameters (CD3, CD4, CD8, CD56, CD34, IL-3, IFN-γ) in relation with the induced toxicity during chemotherapy. The results were analysed using the ANOVA method. RESULTS: We observed a statistically significant difference of CD4 and CD4/CD8 after chemotherapy between groups A and B (p<0.001 and p< 0.006 respectively), implying that the further increase of T-helper cells after chemotherapy had a positive impact on survival. In addition, statistically interesting was the difference in values of IFN-γ between patients of groups A and B before and after chemotherapy (p< 0.039 and p< 0.027, respectively). Patients with high IL-3 had little chance of toxicity. CONCLUSION: Our findings support that with carboplatin/ paclitaxel combination chemotherapy, important parameters of the immune system (IFN-γ, CD4, CD4/CD8) can be used as prognostic factors for survival, while others (IL-3) as indicators of toxicity.

Comparison of new nitrosoureas esters with modified steroidal nucleus for cytogenetic and antineoplastic activity.

Nitrosourea is decomposed under physiological conditions to react with biological macromolecules by two mechanisms: alkylation (with proteins and nucleic acids) and carbamoylation (with proteins but not nucleic acids). It has been suggested that the alkylating action is responsible for the therapeutic effects of nitrosoureas, and that the carbamoylation activity leads to toxicity effects. In order to reduce systemic toxicity and improve specificity and distribution for cancer therapy, 2-haloethyl nitrosourea has been esterified with modified steroids, which are used as biological platforms for transporting the alkylating agent to the tumor site in a specific manner. The cytogenetic and antineoplastic effect were studied of seven newly synthesized esters of N,N-bis(2-chloroethyl)alanyl carboxyl derivatives with a modified steroidal nucleus (compounds 1-7). As a very sensitive indicator of genotoxicity the Sister Chromatid Exchange (SCE) assay was used and as a valuable marker of cytostatic activity the cell Proliferation Rate Index (PRI) in cultures of normal human lymphocytes was used. The order of magnitude of the cytogenetic activity on a molar basis (15, 30, 120 microM) of the compounds was 7>>6>3>5>2>4>1. The most active compound 7 has an enlarged (seven carbon atoms) A ring modified with a lactam group (-NHCO-) with the nitrosourea moiety esterified at position 17 In the group of seven substances a correlation was observed between the magnitude of SCE response and the depression in PRI (r=-O, 65, p<0.001). According to the criterion of activity of National Cancer Institute (NCI), the order of antineoplastic activity of compounds on lymphoid L1210 leukemia is 7>6>2>5>4>3>1 and on lympocytic P388 leukemia cells is 7>2>6>5>4>3>1. The present results are in agreement with previous suggestions that the effectiveness in cytogenetic activity may well be correlated with antitumor effects [T/C: 248% for the compound 7 in 250 mg/kg b.w.; T/C: mean survival time of drug-treated animals (T) (excluding long term survivals) vs. corn-oil-treated controls (C)].

A preclinical survey on the efficacy of lactandrate in the treatment of colon carcinoma.

PURPOSE: There has been a recent and dramatic increase in the pace of drug development for colorectal cancer which holds promise to further improve curative therapy. We tested lactandrate, an alkylating ester of D-lactam androsterone, for antineoplastic activity against colon adenocarcinoma in vitro and in vivo. MATERIALS AND METHODS: The cytostatic and cytotoxic activity of lactandrate were evaluated in vitro against 9 human colon adenocarcinoma cell lines. The in vitro testing was performed with the sulforhodamine B (SRB) colorimetric assay and the mean concentrations of each drug that generated 50% (GI50) or total (100%) growth inhibition (TGI), as well as the drug concentrations that produced cytotoxicity against 50% of the cultured cells (IC50) were calculated. The in vivo antitumour effect was determined against two rodent colon carcinomas, the Colon 26 and the relatively chemoresistant Colon 38 carcinoma, as well as against the human xenograft CX-1 colon carcinoma. RESULTS: Lactandrate displayed a satisfactory activity against the 9 human colon cancer cell lines, inducing significant growth inhibition and cytotoxicity. Lactandrate induced antiproliferative activity against colon cancer cell lines linearly correlated with the carcinoembryonic antigen (CEA) production. There was a non-linear polynomial correlation between CEA production and the cytotoxic effect of lactandrate. The more differentiated cell lines DLD-1 and HCC2998 appeared more resistant to the cytostatic effect of lactandrate. In vivo, the compound produced a significant antitumour activity against Colon 26 and Colon 38, as well as a moderate antitumour effect against CX-1 colon carcinoma. CONCLUSION: Preclinical research supports the high in vitro and in vivo antitumour potential of lactandrate against colon carcinoma. Therefore, lactandrate represents an important candidate drug for further clinical development.

Potential antitumor agent: steroidal bilactam ester of p-N, N-bis (2-chloroethyl) aminophenylacetic acid.

7 alpha-, 17 alpha-Diaza-7, 17-dioxo-B, D-dihomo-5-androsten 3 beta-p-N, N-bis (2- chloroethyl)aminophenylacetate, a modified steroidal alkylating agent, is active in the treatment of P388 and L1210 leukemias in vivo. The compound was also tested in vitro against L1210 and P388 leukemias, on DNA, RNA and protein synthesis and showed high inhibition effect. Also increases the frequency of Sister Chromatid Exchanges and reduces the replication index of human lymphocytes.

Cytostatic effect of homo-aza-steroidal esters in vivo and in vitro. Structure-activity relationships.

The p-[N,N-bis (2-chloroethyl)amino]phenylacetate esters of 3 beta-hydroxy-N-methyl-17 alpha-aza-D-homo-5 alpha-androstan-17-one and 3 beta-hydroxy-17 alpha-aza-D-homo-5 alpha-androstane have been prepared and their antitumor activity evaluated against L1210 leukemia, P388 leukemia, Ehrlich ascites tumor (EAT) and Lewis Lung Carcinoma (LLC). The results are compared with those of the p-[N,N-bis (2-chloroethyl)amino]phenylacetate of 3 beta-hydroxy-17 alpha-aza-D-homo- 5 alpha-androstan-17-one. The above compounds were also tested in vitro against L1210, P 388, EAT and BHX cell cultures. All compounds were found to be active and their structure-activity relationship is discussed.

Structure--anticancer and structure--genetic activity relationships of homo-aza-steroidal esters of N,N-bis(2-chloroethyl)aminocinnamic acid isomers.

Four steroidal lactams of the A- and D-rings were used for the esterification in the C-3 or C-17 positions, respectively, of their nuclei with the N,N-bis(2-chloroethyl)aminocinnamic acid isomers. The condensation reaction of the hydroxylic group of the steroidal lactams with each mustard was effected in dichloromethane in the presence of the catalyst p-dimethylaminopyridine and dicyclohexylcarbodiimide as dehydrating agent. The esters were obtained in pure form after column chromatography, and their structures were verified and confirmed by analytical methods (IR and UV spectra). The 12 esters were tested in vivo against P388, L1210 leukemias, Ehrlich ascites tumor, and melanoma B16. The esters 3 alpha-hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17- oic-13,17-lactam-o,m,p-N,N-bis(2-chloroethyl)aminocinnamates, in which the alkylating agents are linked to the modified steroid in the axial position, are inactive in the above experimental animal tumor systems. The effect of the homo-aza-steroidal esters of N,N-bis(2-chloroethyl)aminocinnamic acid isomers on the incorporation of radioactive precursors into DNA, RNA, and proteins of L1210, P388 leukemias, Ehrlich ascites tumor, and baby hamster kidney cells was investigated. Higher inhibitory effects on the incorporation of the radioactive precursors was obtained with the ortho-derivatives, yielding > 40% inhibition of thymidine incorporation in all tumor lines tested. The effect of four esters in which the m- N,N-bis(2-chloroethyl)aminocinnamic acid is linked to the modified steroids on sister chromatid exchanges in human lymphocyte culture was investigated.

Mutagenicity of four homo-aza-steroidal esters of m-N,N-bis(2-chloroethyl)aminocinnamic acid in the Ames test.

Four new chemicals, the homo-aza-steroidal esters of m-N,N-bis(2-chloroethyl)aminocinnamic acid, originaly synthesized to be used as antineoplastic agents, were tested for their mutagenic activity in the Ames test. 3 beta-Hydroxy-13 alpha-amino-13,17-seco-5-androsten-17-oic-13,17- lactam ester (ACALE3) and 3 alpha-hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17-oic-13,17-lactam ester of m-N,N-bis(2-chloroethyl)aminocinnamic acid (ACALE4) were found to induce base-pair substitutions, causing dose-dependent increases in his+ revertants in strains TA100 and TA1535, while no dose-dependent relations were established when 3 beta-hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17-oic-13,17-lactam ester (ACALE1) and 17 beta-hydroxy-3-aza-A-homo-4 alpha-androsten-4-one ester of m-N,N-bis(2-chloroethyl)aminocinnamic acid (ACALE2) were tested. The presence of metabolic activation enzymes in the test system had no effect in his+ revertants in strains TA100 and TA1535. The chemicals tested although having the same alkylating moiety and a similar chemical structure exhibited different mutagenic activities.

On the formation of steroidal amidoesters of 4-[N,N-bis(2-chloroethyl)amino]benzoic acid and their cytotoxic activity.

The 4-[N,N-bis(2-chloroethyl)amino]benzoate of 17beta-acetamido-5alpha-androstan-3beta-ol, 17beta-acetamido-5-androsten-3beta-ol, 3beta-acetamido-5alpha-androstan-17beta-ol and 3alpha-acetamido-5beta-androstan-17beta-ol have been prepared and their antineoplastic effect evaluated against MIA Pa-Ca-2 pancreatic carcinoma, T47D breast carcinoma and A431 squamus cell carcinoma. Among the compounds tested, the compound 17beta-acetamido-3beta-hydroxy-5-androsten-4-[N, N-bis(2-chloroethyl)amino]benzoate appeared to possess a significant cytotoxic effect against A431 cells.

Proliferation of human acute leukemia cells in culture on treatment with an homo-aza-steroid.

When cultures of human leukemia cells from 9 untreated patients with acute leukemia were treated with 3 beta-hydroxy-13 alpha-amino-13,17-seco-5 alpha-androstan-17-oic-13, 17-lactam, an increased proliferating activity was exhibited. The leukemia cells were cultured for 48h and the amounts of lactam (4, 12 and 24 ng/ml) were added at zero time in the continuous presence of (3H)-thymidine. Our data suggested that in high concentrations lactam acts as a toxic agent while in the lower ones it seems to stimulate proliferation of the blast cell.

Effect of an homo-aza-steroidal ester on estrogen receptor.

A new modified steroid esterified with the cytotoxic moiety, p-[N,N-bis(2-chloroethyl)amino]phenyl-acetic acid, has been synthesized and tested for interaction with estrogen receptor and cytotoxic activity on the MCF-7 cell line.

Preclinical evaluation of the homo-aza-steroid ester 13beta-hydroxy-13alpha-amino-13,17- seco-5alpha-androstan-17-oic-13,17-lactam-p-bis(2-chloroethyl)aminophenoxy acetate for the treatment of malignant melanoma.

PURPOSE: To investigate the in vitro and in vivo activity of an homo-aza-steroid alkylating ester, namely 13beta-hydroxy- 13alpha-amino-13,17-seco-5alpha-androstan-17-oic-13,17- lactam-p-bis (2-chloro ethyl) aminophenoxy acetate (HASE), in comparison with dacarbazine (DTIC) in the treatment of malignant melanoma. MATERIALS AND METHODS: Cytotoxicity was assessed in vitro by the MTT assay using a panel of 6 malignant melanoma human cell lines, with or without the presence of rat liver microsome assay. B16 melanoma-bearing mice were used to evaluate in vivo the antitumor activity of the tested compounds. RESULTS: In all cases of in vitro screening, HASE displayed significantly higher (p <0.0001) cytostatic and cytotoxic activity than DTIC. Moreover, the antitumor activity of HASE in B16 melanoma-bearing mice was satisfactory, prolonging the mice lifespan at 67%, compared to 43% achieved by DTIC. Furthermore, HASE significantly inhibited the tumor growth (tumor growth rate: <42%) as this was defined by tumor volume and weight differences, presenting higher antitumor effect than DTIC. CONCLUSION: HASE displayed superior in vitro and in vivo activity than DTIC in the treatment of melanoma. Thus, HASE may be considered as a significant candidate anticancer agent for further development.

Antitumour effect of a- and d- lactam androgen nitrogen mustards on non-small cell lung carcinoma.

PURPOSE: We tested 3 alkylating esters of D-lactam androsterone, 3 alkylating esters of A-lactam testosterone and the alkylating nitrogen mustard components of these esters, for antineoplastic activity on non-small cell lung carcinoma (NSCLC) in vitro and in vivo. MATERIALS AND METHODS: Cytostatic and cytotoxic activity was evaluated in vitro against 10 human NSCLC cell lines. The in vitro testing was performed with the MTT metabolic-colorimetric assay and the mean concentrations of each drug that generated 50% or total (100%) growth inhibition (GI50 and TGI, respectively) as well as the drug concentrations that produced cytotoxicity against 50% of the cultured cells (IC50) were calculated. Furthermore, the in vivo antitumour effect was determined against the relatively chemo-resistant Lewis lung carcinoma (LLC) on mice. The acute toxicity of the tested compounds was appointed in C57BL mice and the antitumor effect on LLC was assessed from the percent increase in median lifespan of the treated animals over the untreated (control) (T/C%). RESULTS: The lactam steroidal esters presented lower toxicity and increased antineoplastic activity in vitro and in vivo compared to their respective alkylating components. An A-lactam testosterone ester namely: 17beta-hydroxy- 3-aza-A-homo-4alpha-androsten-4-one-p-N,N-bis (2chloroethyl) amino phenoxy acetate (ALT-CAPOA) performed significantly higher anticancer activity in vitro and in vivo. This compound generated 37.5% 90-day disease free survivors (cures) against LLC. CONCLUSION: These results indicate a high antitumor potential of lactam steroid alkylating esters depended on the alkylating moiety as well as on the modified steroidal carrier. Preclinical research supports that ALT-CAPOA generates well-tolerated toxicity as well as superior antitumor activity against NSCLC. These significant results call for further clinical development.

m-N-N-bis(2-chloroethyl)aminocinnamic acid and four new homo-aza-steroidal esters induce chromosomal abnormalities and affect protein synthesis in human lymphocytes in vitro.

The alkylating agent m-N,N-bis(2-chloroethyl)aminocinnamic acid (m-ACA) and four new homo-aza-steroidal esters were studied for their ability to induce chromosomal abnormalities and to affect protein synthesis in human lymphocytes in vitro. A mitotic index reduction and an increase in the total number of aberrations were observed. Analysis of chromosomal abnormalities has shown that these are mainly chromatid breaks. A decrease in protein synthesis was also observed that seems to fit with the order of activity of the above compounds reflected in the induction of chromosomal aberrations. The observation that protein synthesis and the induction of chromosomal aberrations are affected by these chemicals may reflect interactions between these molecules and DNA that result in structural chromosome changes and decreased protein synthesis.

Effect of a delta 5-homo-aza-steroidal ester in P388 and L1210 murine leukemias.

3 beta-Hydroxy-13 alpha-amino-13,17-seco-5-androsten-17-oic-13,17-lactam P-N,N-bis-(2-chloroethyl)-amino phenylacetate gives results in P388 and L1210 leukemias in mice, by the intraperitoneal route of administration.

Comparative study of sister chromatid exchange induction and antitumor effects by homo-aza-steroidal esters of [p-[bis(2-chloroethyl)amino]phenyl]butyric acid.

The present work was undertaken in order to test the hypothesis that the Sister Chromatid Exchange (SCE) assay in vitro can be used for the prediction of in vivo tumor response to newly synthesized potential chemotherapeutics. The effect of three homo-aza-steroidal esters containing the -CONH- in the steroidal nucleus, 1, 2, and 3 on SCE rates and on cell kinetics in cultured human lymphocytes was studied. The antitumor activity of these compounds was tested on leukemia P388- and leukemia L1210-bearing mice. The three substances induced statistically significant enhancement of SCEs and of cell division delays. Compounds 1 and 3 were identified, on a molar basis, as more effective inducers of SCEs and of cell division delays compared with compound 2. Compounds 1 and 3 had upon both experimental tumors better therapeutic effects compared with compound 2 at equitoxic doses. Therefore, the order of the antitumor effectiveness of the three compounds coincided with the order of the cytogenetic effects they induced.

A comparative study on cytogenetic and antineoplastic effects induced by two modified steroidal alkylating agents.

We investigated the effects of two newly synthesized steroidal derivatives of nitrogen mustard on sister chromatid exchange rates and on human lymphocyte proliferation kinetics. The compound 33-hydroxy-5alpha,22alpha-spirostan- 12-one-p-(N,N-bis(2-chloroethyl)amino)phenylacetate(1) was, on a molar basis, less effective in inducing sister chromatid exchange and suppressing cell proliferation rate indices than compound 3beta-hydroxy-12alpha-aza-C-homo-5alpha,22alpha-spirostan-12-one-p-(N,N-bis(2-chloroethyl)amino)phenylacetate(2). A correlation was observed between the magnitude of the sister chromatid exchange response and the depression of cell proliferation index. We also studied the effects of the aforementioned compounds on Lewis lung carcinoma. The order of the percent inhibition of tumor growth achieved by the compounds coincides with the order of the cytogenetic effects they induce.