Killing effects of 5-fluorouracil on human biliary tract cancer cell lines

Previous studies have established that a partially quaternized derivative of chitosan, N-trimethyl chitosan chloride (TMC), can be used as an absorption enhancer for large hydrophilic compounds across mucosal surfaces. This study evaluates and compares the effects of the degree of quaternization of TMC, in a neutral environment, on the permeability of intestinal epithelial cells in vitro, where normal chitosan salts are ineffective as absorption enhancers. The effects of TMC-H [61.2% quaternized, (0.05-1.5% w/v)], TMC-L [12.3% quaternized, (0.5-1.5% w/v)], and chitosan hydrochloride [0.5-1.5% w/v] on the transepithelial electrical resistance (TEER) and permeability, for the hydrophilic model compound [14C]mannitol, of intestinal epithelial Caco-2 cell monolayers, were investigated at pH values of 6.20 and 7.40. The viability of the monolayers was checked with the trypan blue exclusion technique. At a pH of 6.20, all the polymers caused a pronounced reduction (37-67% at 0.5% w/v concentrations) in the TEER of Caco-2 cells. On the contrary, at a pH of 7.40, only TMC-H was able to decrease the TEER values, even in a concentration as low as 0.05% w/v (35% reduction). Comparable results were obtained with the permeation of [14C]mannitol. Large increases in the transport rate (18-23-fold at 0.5% w/v concentrations) were found at pH 6.20, whereas only TMC-H was able to increase the permeation of [14C]mannitol at pH 7.40 (31-48-fold at 0.05-1.5% w/v concentrations of TMC-H). For all the polymers studied, no deleterious effects to the cells could be demonstrated with the trypan blue exclusion technique. It is concluded that highly quaternized TMC is a potent absorption enhancer and the potential use of this polymer, especially in neutral and basic environments where normal chitosan salts are not effective, is expected to be an important contribution to the development of effective delivery systems for hydrophilic compounds such as peptide drugs.     

Combination chemotherapy studies with gemcitabine and etoposide in non-small cell lung and ovarian cancer cell lines

Gemcitabine (2',2'-difluorodeoxycytidine, dFdC) and etoposide (4'-demethylepipodo-phyllo-toxin-9-4,6-O-ethylidene-beta-D-g lucopyranoside, VP-16) are antineoplastic agents with clinical activity against various types of solid tumors. Because of the low toxicity profile of dFdC and the differences in mechanisms of cytotoxicity, combinations of both drugs were studied in vitro. For this purpose, we used the human ovarian cancer cell line A2780, its cis-diammine-dichloroplatinum-resistant and VP-16 cross-resistant variant ADDP, and two non-small cell lung cancer cell lines, Lewis Lung (LL, murine) and H322 (human). The interaction between the drugs was determined with the multiple drug effect analysis (fixed molar ratio) and with a variable drug ratio. In the LL cell line, the combination of dFdC and VP-16 at a constant molar ratio (dFdC:VP-16 = 1:4 or 1:0.125 after 4- or 24-hr exposure, respectively) was synergistic (combination index [CI], calculated at 50% growth inhibition = 0.7 and 0.8, respectively; CI <1 indicating synergism). After 24- and 72-hr exposure to both drugs at a constant ratio, additivity was found in the A2780, ADDP, and H322 cell lines (dFdC:VP-16 = 1:500 for both exposure times in these cell lines). When cells were exposed to a combination of dFdC and VP-16 for 24 or 72 hr, with VP-16 at its IC25 and dFdC in a concentration range, additivity was found in both the LL and H322 cells; synergism was observed in the A2780 and ADDP cells, which are the least sensitive to VP-16. Schedule dependency was found in the LL cell line; when cells were exposed to dFdC 4 hr prior to VP-16 (constant molar ratio, total exposure 24 hr), synergism was found (CI = 0.5), whereas additivity was found when cells were exposed to VP-16 prior to dFdC (CI = 1.6). The mechanism of interaction between the drugs was studied in more detail in the LL cell line; dFdCTP accumulation was 1.2-fold enhanced by co-incubation with VP-16, and was even more pronounced (1.4-fold) when cells were exposed to VP-16 prior to dFdC. dCTP levels were decreased by VP-16 alone as well as by the combination of both compounds, which may favor phosphorylation of dFdC, thereby increasing dFdCTP accumulation. DNA strand break (DSB) formation was increased for exposure to both compounds together compared to exposure to each compound separately, this effect being most pronounced when cells were exposed to VP-16 prior to dFdC (38% and 0% DSB for dFdC and VP-16 alone, respectively and 97% DSB for the combination). The potentiation in DSB formation might be a result of the inhibition of DNA repair by dFdC. Provided the right schedule is used, VP-16 is certainly a compound eligible for combination with dFdC.     

A phase II study of 5-fluorouracil, leucovorin and cisplatin (FLP) for metastatic gastric cancer

The modulation of 5-fluorouracil (5-FU) with folinic acid (leucovorin, LV) is more efficacious than 5-FU alone in the treatment of metastatic colorectal cancer, and the combination of 5-FU with cisplatin is currently one of the most active regimens in advanced gastric cancer. A phase II study was therefore conducted to test the efficacy and toxicity of the combination of 5-FU, LV and cisplatin (FLP) in metastatic gastric cancer. 28 patients entered the study. Metastatic sites were observed in the liver (in 21 patients), the peritoneum (in 8), the lymph nodes (in 7) or the bones (in 1) and a local recurrence was noted in 4 cases. The performance status (using World Health Organisation criteria) was 0 for 13 patients and 1 or 2 for the others. Cycles of treatment were administered every 28 days and consisted of LV 200 mg/m2/day for 5 days followed by 5-FU 400 mg/m2/day for 5 days with cisplatin 100 mg/m2 on day 2. The response rate for the 27 evaluable patients was 51.8% (95% confidence interval (CI), 33-70.6%). There were four complete responses (14.8%) and 10 partial responses (37%). Median survival was 11 months and 4 patients were alive at 2 years. Both response rate and survival were better for patients with a good performance status. The overall toxicity was very low, except for 1 patient who died of dehydration and cardiac failure. In conclusion, the FLP protocol was effective and well tolerated in patients with metastatic gastric cancer.     

Differentially potentiating effects by dipyridamole on cytotoxicity of 5-fluorouracil against three human maxillary cancer cell lines derived from a single tumor

Twenty specimens of six species of Nile fishes were examined for the presence of fungi. Of which 2 were from Alestes nurse; 3 from Bagrus docmac; 4 from Barbus bynni; 6 from Chrysichthys auratus; 4 from Lates niloticus and 1 from Malapterurus electricus. Forty-three fungal species in addition to 1 variety appertaining to fifteen genera were recovered from skin (15 genera and 34 species + 1 variety); gills, kidney (12 genera and 30 species + 1 variety, each); liver (11 genera and 30 species + 1 variety) and intestine (13 genera and 30 species + 1 variety) of all specimens, using glucose Czapek-Dox medium at 28 degrees C. The most common genera were Aspergillus, Penicillium and Trichoderma.     

Sensitization of human bladder cancer cells to Fas-mediated cytotoxicity by cis-diamminedichloroplatinum (II)

PURPOSE: The resistance of bladder cancer to anticancer chemotherapeutic drugs is a major problem. Several immunotherapeutic approaches have been developed to treat drug-resistant tumor cells. The Fas antigen (Fas)-Fas ligand pathway is involved in cytotoxic T lymphocyte and natural killer cell-mediated cytotoxicity. Like the Fas ligand, anti-Fas monoclonal antibody (mAb) induces apoptosis in tumor cells expressing Fas. Several anticancer drugs also mediate apoptosis and may share with Fas common intracellular pathways leading to cell killing. We reasoned that treatment of drug-resistant cancer cells with a combination of anti-Fas mAb and drugs might overcome their resistance. This study has investigated whether anticancer drugs synergize with anti-Fas mAb in cytotoxicity against bladder cancer cells. MATERIALS AND METHODS: Cytotoxicity was determined by a 1-day microculture tetrazolium dye assay. Synergy was assessed by isobolographic analysis. RESULTS: Treatment of the T24 human bladder cancer cell line with anti-Fas mAb in combination with 5-fluorouracil, mitomycin C or methotrexate did not overcome resistance to these agents. However, treatment of T24 tumor cells with a combination of anti-Fas mAb and cisdiamminedichloroplatinum (II) (CDDP) resulted in a synergistic cytotoxic effect. In addition, the CDDP-resistant T24 line (T24/CDDP) was sensitive to treatment with a combination of anti-Fas mAb and CDDP. Synergy by combination of anti-Fas mAb and CDDP was also achieved in three other bladder cancer lines and four freshly derived human bladder cancer cells. The combination of anti-Fas mAb and carboplatin also resulted in a synergistic cytotoxic effect on T24 cells; however, the combination of anti-Fas mAb and trans-diamminedichloroplatinum (II) resulted in an additive cytotoxic effect. Treatment with CDDP enhanced the expression of Fas on T24 cells. The synergy achieved in cytotoxicity with anti-Fas mAb and CDDP was also achieved in apoptosis. Incubation of T24 cells with anti-Fas mAb increased the intracellular accumulation of CDDP. Treatment of freshly isolated bladder cancer cells with CDDP enhanced their susceptibility to lysis by autologous lymphocytes. CONCLUSIONS: This study demonstrates that combination treatment of bladder cancer cells with anti-Fas mAb and CDDP overcomes their resistance. Synergy was achieved with established CDDP-resistant bladder cancer cells and freshly isolated bladder cancer cells. In addition, the sensitization required low concentrations of CDDP, thus supporting the potential in vivo application of combination of CDDP and immunotherapy in the treatment of CDDP- and/or immunotherapy-resistant bladder cancer.     

A phase II trial of weekly infusional 5-fluorouracil in combination with low-dose leucovorin in patients with advanced colorectal cancer

We examined 59 breast cancers for p53 and bcl-2 protein expression by immunohistochemistry. The results were correlated with Ki-67 immunostaining. p53-negativity was noted in 40 cases and the remaining 19 tumours were p53-positive. Thirty-six tumours showed strong expression of bcl-2 and in 23 no staining for this protein was observed. We found statistically significant reverse correlation between expression of p53 and bcl-2 in majority of carcinomas: 31 cases were bcl-2 positive and p53-negative, and 14 tumours were bcl-2-negative and p53-positive. Six carcinomas showed no nuclear staining for Ki-67 and in the remaining 53 the percent of cancer cells positive for Ki-67 ranged from 1 to 60 (mean: 14.6). In these 53 cases we found that bcl-2-positive tumours were characterized by lower proliferation than bcl-2-negative tumours, the mean value of Ki-67 immunostaining being 10.7% and 23.0%, respectively. p53-negative tumours showed lower proliferation than p53-positive tumours: mean Ki-67 index was 10.2% and 23.9%, respectively. We conclude that immunohistochemically detected p53 and bcl-2 proteins show a significant inverse relationship in majority of breast carcinomas and their expression correlates with tumour proliferation (Ki-67 immunostaining).     

Phase II trials of 5-fluorouracil and leucovorin in patients with metastatic gastric or pancreatic carcinoma

BACKGROUND: Previous trials in patients with colorectal carcinoma have indicated that enhancement of 5-fluorouracil (5-FU) by leucovorin (LV) can result in an improved response rate and increased survival. METHODS: Phase II trials were performed with patients who had either gastric or papcreatic adenocarcinoma with inetastases. Forty-one gastric carcinoma patients and 31 pancreatic carcinoma patients with measurable disease were treated with 5-FU, 425 mg/m2 intraveneosly (i.v.) on Days 1-5 plus LV, 20 mg/m2 i.v., on Days 1-5, reported at 4 and 8 weeks, and then every 5 weeks thereafter. RESULTS: The patients with metastatic gastric carcinoma had a median survival of 4.8 months. There was a 22% objective response rate, including a 4.9% complete response rate and a 17.1% partial response rate. Among the 31 patients with pancreatic carcinoma, there was a median survival of 5.7 months. No patients in this group showed a response. CONCLUSIONS: The response rate for patients with metastatic gastric adenocarcinoma was modest and this regimen may provide temporary palliation for some patients. However, 5-FU and LV treatment is ineffective against metastatic pancreatic carcinoma.     

Phase II study of liarozole in advanced non-small cell lung cancer

The aim of this phase II study was to investigate the efficacy and tolerability of liarozole, a novel benzimidazole derivative, in non-small cell lung cancer (NSCLC). Liarozole 300 mg twice daily orally was evaluated in 14 patients with stage IIIB and IV NSCLC. 8 patients had received prior treatment with chemotherapy and/or radiotherapy. WHO toxicity grading and response criteria were used. Liarozole was well tolerated. Grade 2 toxicities included alopecia (1 patient), dermatological toxicity (5 patients), dry mouth (2 patients) and nausea and vomiting (2 patients). Leukocytosis was seen in 5 patients, including 2 cases with an elevated white cell count pretreatment. Liarozole was discontinued in 1 patient who developed intolerable progressive pruritus associated with an erythematous rash. No objective tumour response was seen, all 14 patients developing progressive disease within 4 months of commencing treatment. Liarozole was well tolerated but was ineffective as single agent therapy in the management of NSCLC. The side-effect profile was compatible with inhibition of retinoic acid degradation.     

Treatment of advanced gastric cancer with oral etoposide, leucovorin and tegafur: experience with an oral modification of the etoposide, leucovorin and 5-fluorouracil (ELF) regimen

Recent data have suggested enhanced therapeutic activity with prolonged administration of both etoposide as well as fluoropyrimidines in the treatment of gastrointestinal malignancies. Based on this rationale, we investigated the clinical effectiveness and tolerance of an oral modification of the widely applied etoposide, leucovorin and 5-fluorouracil (ELF) regimen in patients with advanced gastric cancer. 32 patients with advanced gastric cancer were treated with oral etoposide (100 mg), leucovorin (3 x 100 mg), and tegafur (3 x 200 mg) over 14-21 days for a maximum of six cycles. Objective response was seen in only 5 patients (16%), stable disease was documented in 7 (22%), while the remaining patients progressed during therapy. The median time to progression was 2.8 months (range 0.7-12 months) and median overall survival was 6 months (range 1-18+ months). Due to grade 3 nausea/emesis, 8 patients discontinued treatment prematurely, while 12 patients experienced anorexia and progressive weight loss. Haematological toxicity was modest, with 4 patients developing asymptomatic grade 3-4 granulocytopenia. We conclude that this oral combination regimen cannot be recommended for the treatment of advanced gastric cancer.     

Phase II trial of intravenous endotoxin in patients with colorectal and non-small cell lung cancer

We report the immunological and clinical results of a phase II trial with intravenously administered highly purified endotoxin (Salmonella abortus equi) in patients with advanced cancer. 15 patients with non-small cell lung cancer and 27 with colorectal cancer were entered into the study. 37 evaluable patients received at least four injections of endotoxin (4 ng/kg body weight) and 1600 mg ibuprofen orally in 2-week intervals. Transient renal (WHO grade 0-1) and hepatic (WHO grade 0-4) toxicities occurred in several patients. Constitutional side-effects such as fever, chills and hypotension could not be prevented completely by pretreatment with ibuprofen. 3 patients in the colorectal cancer group demonstrated objective responses (1 complete remission (CR), 2 partial remission (PR)). The complete remission has been maintained for more than 3 years, while the partial remissions were stable for 7 and 8 months, respectively. Only marginal antitumour effects were seen in the lung cancer group. Tolerance of the macrophage system to the stimulatory effect of endotoxin, as measured by human necrosis factor alpha (TNF-alpha) release into serum, built up after the first administration and remained at a steady-state level after each subsequent injection. In constrast, rising CD4:CD8 ratio and release of tumour necrosis factor beta (TNF-beta) indicated the continuing activation of the lymphocyte system by repetitive injections of endotoxin.     

Interleukin 15 protects against toxicity and potentiates antitumor activity of 5-fluorouracil alone and in combination with leucovorin in rats bearing colorectal cancer

5-Fluorouracil (FUra) modulated by leucovorin (LV) is active in the treatment of colorectal cancer. Diarrhea and stomatitis are the most common dose-limiting toxicities. We have developed a model system in rats bearing a transplantable colon carcinoma sensitive to FUra therapy with dose-limiting toxicity profiles similar to what is observed in patients treated with either daily or weekly schedules of FUra plus LV. Interleukin 15 (IL-15), a cytokine that shares many biological activities with IL-2, was used at different doses (25, 100, and 400 microg/kg) and schedules (three doses before a single dose of FUra, FUra/LV daily x 5, or before each week of FUra/LV weekly x 4, or three doses before a single dose of FUra or FUra/LV daily x 5, then twice daily x 5 for a total of 11 doses) to evaluate its role in the modulation of the therapeutic selectivity of FUra alone and modulated by LV. IL-15 induced a dramatic decrease in chemotherapy-induced gastrointestinal toxicities, significant potentiation of antitumor activity, and an increased therapeutic index of FUra administered on single dose, daily x 5 and weekly x 4 schedules. In contrast, IL-2 (400 microg/kg) significantly potentiated the toxicity of FUra administered as a single i.v. push, with minimal potentiation of the antitumor activity. Taken together, the results clearly demonstrated the ability of IL-15, but not IL-2, to provide significant improvement of the therapeutic index of FUra alone and in combination with LV. The clinical relevance of the results obtained in this model system needs to be confirmed.     

A phase II study of irinotecan alternated with five days bolus of 5-fluorouracil and leucovorin in first-line chemotherapy of metastatic colorectal cancer

PURPOSE: This multicenter phase II study was designed to assess the efficacy of the alternating schedule of irinotecan (CPT-11) with bolus 5-fluorouracil (5-FU) and leucovorin (LV) in first-line chemotherapy for metastatic colorectal cancer (CRC). PATIENTS AND METHODS: Patients with histologically proven metastatic colorectal cancer, and at least one bidimensionally measurable lesion, aged 18-70, with performance status < or = 2, normal baseline biological values and no prior chemotherapy (or only adjuvant chemotherapy completed > or = 6 months before study entry) were selected. Treatment was irinotecan 350 mg/m2, i.v., day 1, alternating with leucovorin 20 mg/m2 i.v. and 5-FU 425 mg/m2, i.v. daily for five consecutive days, day 22-26 (Mayo Clinic regimen). One alternating cycle was to be performed every six weeks. Patients were evaluated for efficacy every alternating cycle. Treatment was administered until five alternating cycles, disease progression, unacceptable toxicity or patient refusal. RESULTS: Thirty-three patients (28 chemotherapy-na?ve and five with prior adjuvant treatment completed > 1 year prior to accrual) were enrolled. The objective response rate (RR) was 30% (95% CI: 16-49; 10 patients/33; nine partial response and one complete response). All responses were reviewed by an independent external review committee. An additional 49% of patients had stable disease. The median survival was 16 months, the one year survival amounted to 58% and the median progression free survival was 7.2 months. Relative dose intensity was nearly 90% for both drugs. Grade 3-4 diarrhea and neutropenia were the most frequent severe toxic events, seen in 24% and 64% of patients, respectively. CONCLUSIONS: The alternating schedule of CPT-11 350 mg/m2 with five days bolus of 5-FU and low dose LV is an active and feasible regimen as front-line therapy for metastatic CRC. It is well tolerated, without evidence of overlapping toxicity. The response rate appears promising with regard to that expected with either single agent. This regimen warrants further assessment in randomized trials.     

Phase II study of all-trans-retinoic acid and alpha-interferon in patients with advanced non-small cell lung cancer

Between April 1993 and June 1994, 29 patients (pts) with unresectable, locally advanced, or metastatic non-small cell lung cancer were treated with a combination of p.o. trans-retinoic acid (TRA), 150 mg/m2/day, in three divided doses and s.c. IFN-alpha, 3 x 10(6) units/day. The age range was 41-80 years (median, 63 years). The Eastern Cooperative Oncology Group performance status was 0-1 (24 pts) and 2 (5 pts). Pts had advanced disease, refractory to conventional therapy (5 stage IIIB and 24 stage IV). Twenty-one pts had adenocarcinoma, six had squamous cell carcinoma, and two had large cell carcinoma. Only 3 pts completed 8 weeks of treatment, requiring neither interruption nor dose modification. Fatigue occurred in 88% of pts. A syndrome complex consisting of dry oral and nasal mucosa, recurrent sinus infections, and epistaxis occurred in 64% of pts. Grade II/III dermatitis was seen in 52%. Severe scrotal dermatitis was seen in 7 pts (47% of 15 males). Hypertriglyceridemia was moderate/severe in 11 pts, and 3 pts required gemfibrozil for levels up to 1660 mg/dl. Hematological toxicity was not encountered, and none of the pts had leukocytosis. One pt died with complications of myocardial infarction while on TRA/IFN-alpha. Twenty-five pts had more than 2 weeks of treatment and are evaluable for response; two pts died early with complications of cancer, and two pts declined to continue after only 3 and 5 days of treatment. Final assessment of response was by accepted clinical and radiological criteria at 8 weeks. There have been four objective responses: complete response, 2 (18+ and 17 months) and partial response, 2 (7 and 14 months). Responses were observed in all histologies. Combined differentiation treatment with TRA/IFN-alpha has modest but objective activity in non-small cell lung cancer. Toxicity is considerable. Additional studies to elucidate the biological basis of TRA/IFN-alpha and to define prognostic parameters predicting response are needed.     

Differential expression of DNA topoisomerase II alpha and II beta genes between small cell and non-small cell lung cancer

The purpose of this study is to describe the appearance of bowel-related abscesses on magnetic resonance (MR) images. Sixteen consecutive patients who had bowel-related abscesses underwent MR examination at 1.5T. MR sequences included T1-weighted fat-suppressed imaging pre- and post-intravenous gadolinium chelate administration (all patients) and breathing-independent single-shot T2-weighted half Fourier turbo (fast) spin echo (6 patients). Patients with pelvic abscesses also underwent sagittal imaging with post-gadolinium T1-weighted images (9 patients) and T2-weighted turbo (fast) spin echo (8 patients). Abscesses were confirmed by open surgery or surgical drainage (6 patients), percutaneous drainage (8 patients), or combined physical examination, fluoroscopic fistulogram, and clinical follow-up (2 patients). Oval-shaped fluid collections were identified in all of the patients, which ranged in diameter from 2 cm to 18 cm, mean: 8 cm. Abscesses were low to intermediate in signal on T1-weighted images, heterogenous and moderately high signal on T2-weighted images, and low signal on post-gadolinium images. A layering effect of lower signal material in the dependent portion of the abscess was noted in abscesses in 6 of 14 patients on T2-weighted images. Post-gadolinium images demonstrated a definable 3- to 7-mm thick abscess wall, which enhanced substantially with contrast. Definition of the wall was best shown on fat-suppressed images post-gadolinium. Substantial enhancement of surrounding periabscess tissues was demonstrated in all cases and was most clearly defined on fat-suppressed images. Image acquisition in two orthogonal planes was of value to demonstrate that fluid collections were oval, and separate from bowel. Image acquisition in the sagittal plane was useful in the evaluation of pelvic abscesses. The results from this preliminary study show that bowel-related abscesses are demonstrable on MR images using gadolinium-enhanced fat-suppressed T1-weighted and turbo (fast) spin-echo T2-weighted sequences. The presence of a thickened, enhancing lesion wall and enhancement of perilesional tissues on T1-weighted fat-suppressed images were observed in all abscesses. A layering effect of low signal intensity material in the dependent portion of the abscess was an important ancillary feature.     

Steroid-hormone receptors in cell lines and tumor biopsies of human lung cancer

Female gender is a significant independent favorable prognostic factor in lung cancer. To study the possible role of sex hormones in lung cancer, the expression of sex-steroid receptors and the glucocorticoid receptor was investigated in 29 lung-cancer cell lines stemming from small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC) by means of immunocytochemistry, ligand-binding assays and RNA expression via polymerase chain reaction. In at least 2 methods of investigation, NSCLC cell lines showed a low expression of estrogen receptor in 6, progesterone receptor in 13 and androgen receptor in 12 out of 17 cases examined; sex-steroid-receptor expression was virtually absent in SCLC cell lines. The glucocorticoid receptor was expressed in all 29 cell lines studied. Additionally, 52 tumor samples from primary lung cancer were investigated for their receptor expression by means of immunohistochemistry. Among patients with primary lung-cancer sex-steroid-receptor expression in tumor biopsies was detected most frequently in female patients (in 69% of 16 cases, vs. 42% of 36 tumors from men) and in patients with adenocarcinoma. Further research will focus on these subgroups. Immunohistology is a feasible method of studying steroid-receptor expression in lung cancer.     

Biologic and clinical effects of continuous infusion interleukin-2 in patients with non-small cell lung cancer

BACKGROUND: Interleukin-2 (IL-2) has shown antitumor activity in some neoplasms, such as melanoma and renal carcinoma, but toxicity derived from bolus administration is significant, particularly at the cardiorespiratory level. METHODS: To test feasibility, antitumor activity, pulmonary and systemic immunologic effects, and pulmonary function changes of continuous-infusion recombinant IL-2 given to patients with non-small cell lung cancer, eleven subjects with Stage III-IV disease were treated in a standard pulmonary medicine unit with a dose of 18 million IU/m2/day from day 1 to day 13 with 1-day rest on day 7. A second induction course was given after a 3-week rest. In patients with nonprogressive disease, four maintenance courses of 6 days' duration at the same dose were planned. Immunologic tests, including lymphocyte phenotype analysis and assays for the detection of tumor necrosis factor (TNF) and of anti-IL-2 antibodies, were performed before and after treatment in serum and bronchoalveolar lavage fluid (BAL). Cardiopulmonary function tests, including spirometry, arterial blood gas analysis, diffusion capacity, and echocardiography, were obtained before, during, and after treatment. RESULTS: Twenty-one cycles (15 induction courses plus 6 maintenance courses) were administered. No patient was able to complete the six planned courses, and only 3 patients entered the maintenance phase. Reasons for discontinuation included progressive disease in five cases, toxicity in three cases, and patient request in three cases. The most common side effects were fever, hypotension, oliguria, and elevated serum creatinine and liver enzyme levels. No patient required intubation or intensive care. No objective response was seen, and the median survival time was 10 months. Lymphocytosis and eosinophilia were observed in all patients. Surface marker analysis revealed a statistically significant increase in the percentage of CD3+, CD4+, CD25+ and DR+ cells in peripheral blood. Lymphoid cells derived from BAL disclosed an increased natural killer activity after IL-2 treatment, and TNF was increased in BAL fluid. Pulmonary function tests evidenced an increased alveolar-arterial difference for oxygen allied with a decrease of forced expiratory volume in 1 second, forced vital capacity, and carbon monoxide transfer coefficient consistent with a significant, albeit not clinically relevant, interstitial lung defect. CONCLUSION: Continuous-infusion IL-2 is feasible in patients with advanced lung cancer even outside an intensive care unit, but overall compliance is poor. Although clinical pulmonary toxicity is negligible, small but statistically significant alterations of the pulmonary function are evident. In addition, this regimen produces a significant activation of the immune system at the pulmonary level.     

Co-amplification of a novel cyclophilin-like gene (PPIE) with L-myc in small cell lung cancer cell lines

Specific genetic alterations affecting proto-oncogenes of the myc gene family are frequently detected in human lung cancer. Among 11 SCLC cell lines with L-myc gene amplification, four were found to have alteration of the RLF gene by Southern blot and RT-PCR analyses. One cell line, NCI-H378, contained aberrantly-sized L-myc-hybridizing bands by Southern and Northern blot hybridization but had no alteration of RLF. Some L-myc-hybridizing cDNAs from NCI-H378 contained a novel sequence with close homology to the cyclophilins joined to antisense L-myc exon 2 sequence. Full length cDNAs isolated from human skeletal muscle containing only the novel sequence identify open reading frames of 301 and 296 amino acids and differ in the C-terminal region by 22 and 17 amino acids. This gene, tentatively named PPIE (peptidyl-prolyl cis-trans isomerase E), has 83% amino acid identity with the central conserved region of cyclophilin A, is evolutionarily conserved by Southern blot, and exhibits differential tissue expression with highest levels found in muscle and brain. Co-amplification of PPIE was observed in seven of eleven L-myc amplified cell lines. Analysis of radiation hybrids suggests that the gene order is RLF-PPIE-L-myc on chromosome 1p and pulse-field gel electrophoresis localizes all three genes to an 800 megabase Mlu I fragment. The prognostic and functional consequences of PPIE gene amplification in SCLC can now be determined.     

A phase II trial of 5-fluorouracil, leucovorin, and carboplatin in patients with unresectable biliary tree carcinoma

BACKGROUND: Unresectable adenocarcinoma of the biliary tree are associated with a very poor prognosis. 5-fluorouracil (5-FU) combination regimens have produced objective response rates in approximately 10-20% of patients. Leucovorin increases the selective cytotoxicity of 5-FU. There also are encouraging reports of carboplatin in combination with 5-FU in other gastrointestinal tract malignancies. METHODS: Fourteen consecutive eligible patients were treated with a combination of carboplatin, 300 mg/m2, intravenously (i.v.) on Day 1 only and 5-FU, 400 mg/m2, i.v. with leucovorin, 25 mg/m2, i.v. on Days 1-4. All patients were required to have a histologically confirmed diagnosis and measurable disease. Patients were evaluated for response, survival, and toxicity. RESULTS: A total of 48 cycles of therapy were delivered. The median survival was 5 months. One patient achieved complete remission and two others partial remission, for a total response rate of 21.4%. Four additional patients had stable disease for a median duration of 4 months. The therapy was well tolerated, with moderate myelosuppression as the main dose-limiting toxicity. CONCLUSIONS: The current combination regimen of leucovorin-modulated 5-FU with carboplatin is well tolerated with appropriate supportive care, produces significant objective responses in 21% of patients with biliary tree carcinoma, and should be considered for the treatment of this disease.     

Antiproliferative effect of curcumin (diferuloylmethane) against human breast tumor cell lines

Pharmacologically safe compounds that can inhibit the proliferation of tumor cells have potential as anticancer agents. Curcumin, a diferuloylmethane, is a major active component of the food flavor turmeric (Curcuma longa) that exhibits anticarcinogenic properties in vivo. In vitro, it suppressed c-jun/Ap-1 and NF-kappaB activation and type 1 human immunodeficiency virus long-terminal repeat-directed gene expression. We examined the antiproliferative effects of curcumin against several breast tumor cell lines, including hormone-dependent and -independent and multidrug-resistant (MDR) lines. Cell growth inhibition was monitored by [3H]thymidine incorporation, Trypan blue exclusion, crystal violet dye uptake and flow cytometry. All the cell lines tested, including the MDR-positive ones, were highly sensitive to curcumin. The growth inhibitory effect of curcumin was time- and dose-dependent, and correlated with its inhibition of ornithine decarboxylase activity. Curcumin preferentially arrested cells in the G2/S phase of the cell cycle. Curcumin-induced cell death was neither due to apoptosis nor to any significant change in the expression of apoptosis-related genes, including Bcl-2, p53, cyclin B and transglutaminase. Overall our results suggest that curcumin is a potent antiproliferative agent for breast tumor cells and may have potential as an anticancer agent.     

Differential effects of taxol on two human cancer cell lines

Taxol is the prototype of a new class of drugs with promise in the treatment of various cancers. Its mechanism of action is not fully understood. We have investigated the effects of taxol on two human cancer cell lines, HT-29, derived from a colon carcinoma, and SK-MEL-28, from a melanoma. Immunofluorescence staining for microtubules, cell cycle analysis by flow cytometry, cytotoxicity assays, and DNA fragmentation studies by agarose gel electrophoresis were performed. The two cell lines responded quite differently to taxol. HT-29 cells, when treated with taxol for 24 h, showed an abundance of multiple microtubule asters and the cells were arrested in mitosis. Flow cytometry also showed arrest in mitosis and development of a hypodiploid region with increasing taxol incubation times. These cells were more sensitive to taxol exposure, which caused internucleosomal DNA fragmentation, indicative of apoptosis. The SK-MEL-28 cells, on the other hand, were less sensitive to taxol. Significant cytotoxic effects were only visible after 72 h. These cells exhibited a predominance of microtubule bundles and multinuclei, and only a few cells were arrested in mitosis. Flow cytometry revealed that the SK-MEL-28 cells became polyploid, as a result of exit from mitosis without cell division. These results suggest that the mechanism involved in taxol-induced cell death is different in these two cell lines.