Tissue inhibitor of metalloprotease (TIMP)‐1 and proliferative behaviour of clonal breast cancer cells

In the present paper, we have examined whether human tissue inhibitor of metalloprotease1 (hTIMP1) is able to exert a growth factorlike effect on two clonal cell lines (BC3A and BC61), isolated from a parental line of human breast carcinoma cells (8701BC), and endowed with different growth and invasive behaviour in vitro and in nude mouse. The data obtained indicate that only the more tumorigenic clonal cell line (BC61) is responsive to hTIMP1 treatment by increasing its proliferative rate in a dosedependent manner. It was also found that BC61 cells selectively express a transmembrane protein of about 80kDa able to bind hTIMP1 in vitro and in vivo with high affinity (K_d of 0.07 ± 0.004 nM), and that treatment of BC61 cells with a proliferationpromoting concentration of hTIMP1 is able to stimulate tyrosinetargeted phosphorylation. The cumulative results obtained strongly support the hypothesis that hTIMP1, classically regarded as a collagenase inhibitor, may be a crucial element of the extracellular signalling network during breast cancer development by controlling cell growth phenotype in autocrine and paracrine manner, and that intratumoural heterogeneity for the biological response to TIMP1 may exist within the composite cell population of the primary tumour site.     

Intracavitary VEGF,bFGF, IL-8, IL-12 levels in primary and recurrent malignant glioma

Intracavitary levels of VEGF, bFGF, IL-8 and IL-12 were evaluated by ELISA in 45 patients, 7 with recurrent anaplastic astrocytoma (rAA), 12 with glioblastoma (GBM) and 26 with recurrent glioblastoma (rGBM). In 25 patients plasma levels of the molecules were also quantitated. Twenty-three healthy controls were also studied for plasma concentrations of the same molecules.Plasma levels of VEGF (mean 33.89 ± 6.71pg/ml) and bFGF (mean 11.1 ± 3.24pg/ml) were higher in patients than in controls (mean 16.78 ± 3.7pg/ml for VEGF, mean 0.21 ± 0.09pg/ml for bFGF) (p = 0.04 and p = 0.001, respectively) while plasma IL-12 levels were lower (mean 45.6 ± 1.5pg/ml in patients, mean 79.7 ± 1.3pg/ml in controls) (p = 0.009).Intracavitary VEGF levels were 5–53.307 fold higher (mean 90,900 ± 24,789pg/ml) than in the corresponding plasma. Also IL-8 concentrations were higher in intracavitary fluid (mean 6,349.76 ± 1,460.93pg/ml) than in plasma (mean 43.44 ± 24.82pg/ml). Maximum VEGF levels were found in tumor fluid of recurrent glioblastoma patients (mean 147,678 ± 39,903pg/ml), intermediate levels in glioblastoma patients (mean 20,322 ± 11,892pg/ml) and lower levels in rAA patients (mean 9,111 ± 5,789pg/ml). The data also suggest that higher intracavitary levels of VEGF and IL-8, and lower IL-12 levels, may be correlated with shorter adjunctive survival times, but more data will need to be collected to establish this correlation clearly.     

Increased risk of second cancers following breast cancer: Role of the initial treatment

Objectives and methods.The risk of second primary malignancies (SMN) was studied in a cohort of 4,416 one-year survivors of a breast cancer. The role of the menopausal status and of the initial treatment modalities (surgery, radiotherapy, and chemotherapy) was investigated. Results.Excluding second primary breast cancer and non-melanoma skin cancer, a total of 193 (4.4%) patients developed a SMN between 1973 and 1992, compared with 136 expected (Standardised Incidence Ratio, SIR=1.4, 95% CI (1.2–1.6)). No trend towards either an increase or a decrease was noted in the SIR with time after treatment (p=0.2). The greatest increase in the relative risk concerned soft tissue cancers (SIR=13.0, 95% CI: 6.8–22.3), followed by leukaemia (SIR=3.1, 95% CI: 1.7–5.0), melanoma (SIR = 2.7, 95% CI: 1.4–4.8), kidney (SIR=2.5, 95% CI: 1.2–4.5), ovary (SIR=2.0, 95% CI: 1.2–3.1) and uterine tumours (SIR=1.9, 95% CI: 1.4–2.5). The SIR was 3.0 (95% CI 1.8–4.7) in women under 40 at the time of the breast cancer, 1.9 (95% CI : 1.4 – 2.4) in those aged 40–49 and 1.2 (95% CI 1.0–1.4) in those aged 50 or more. In the 2,514 women who had received radiotherapy as initial treatment without chemotherapy, the SIR for all SMN was 1.6 (95% CI: 1.1–2.3) fold higher than in those who had not received radiotherapy as initial treatment. Conclusion.In conclusion, this study confirms the increased risk of second malignancies in women treated for a breast cancer, and particularly in those who were younger at the time of treatment for breast cancer. Our results also suggest that radiotherapy may play a role in the onset of these second lesions.     

TGFβ Signaling Pathways and Human Diseases

Recent progress in deciphering the TGF pathway has uncovered a new signaling molecule, the Smads, and with this finding now gives us insights into how TGF-like signals are transmitted from outside the cell to the nucleus. As we learn more about how TGF regulates normal development, we also are gaining insights into diseases that are caused by mis-regulation or mutation of various components of the signaling pathways.     

START: A European state-of-the-art on-line instrument for clinical oncologists

START (State-of-the-Art Oncology in Europe), a freely available resource on the Internet, is a European information base of current clinical approaches to human tumours. Its aim is to help clinical oncologists make appropriate clinical decisions by providing them with updated information reflecting state-of-the-art cancer treatment as perceived by the European oncology community. It is based upon contributions from authors and internal reviewers from all over Europe as selected by START's editorial board under the supervision of an advisory board and a scientific committee. Close collaborations with the main European cancer societies are ongoing. An external feedback process augments the mechanisms for rendering START a truly European instrument. START is concerned with evidence-based cancer medicine, and the main clinical options are thus codified and their bases indicated in accord with a scale worked out from the perspective of clinical decision-making. Therapeutic options may be standard, investigational, or suitable for individual clinical use (within the context of a decision made jointly by the patient and the physician). The goal of instruments such as START is to improve the quality of patient care. In addition, START hopes to make contributions to the methodology by which medical research is transformed into clinical decisions.     

Phenotypic analysis of 1-B-d-arabinofuranosylcytosine deamination in patients treated with high doses and correlation with response

Summary Two phenotypes for 1-B-d-arabinofuranosylcytosine (ara-C) deamination corresponding to a ratio of distribution for slow (ratio, 14) vs fast (ratio, >14) deaminators of 70%30%, have been determined on the basis of studies on plasma ratios of 1-B-d-arabinofuranosyluracil/ara-C (ara-U/ara-C) in 56 subjects treated with high-dose ara-C (3 g/m² infused i.v. over 3 h). A positive correlation of age with the concentration of ara-U was observed. In a subgroup of 36 patients with leukemia, the ara-U/ara-C pattern was similar to that observed for all 56 subjects. In these leukemic patients, who were treated with combinations of ara-C plus other conventional agents, a tendency toward a positive response (complete response +partial response) was found for those showing low ara-U/ara-C ratios (slow deaminators). The phenotypic effect of deamination in acute leukemia needs to be evaluated prospectively.This work was supported by the Don Monti Memorial Research Foundation     

Expression of TGFα in Meningiomas

The objective of this study was to examine the expression of transforming growth factor (TGF), a mitogen for many cell types, and its receptor in basic subtypes of meningiomas as well as in meningiomas of varying grade. Formalin-fixed tissues from 26 meningiomas including 15 benign (5 meningothelial, 5 transitional, and 5 fibrous variants), 6 atypical, and 5 malignant examples were immunohistochemically examined for both TGF protein and EGF/TGF receptor protein. In addition, in situ hybridization (ISH) was used to detect TGF mRNA expression. Immunostaining for TGF was strongest in fibrous and atypical meningiomas, followed closely by transitional and malignant tumors. Only weak reactivity was observed in the meningothelial variant. In all but 4 tumors (2 fibrous, 2 atypical), ISH showed TGF mRNA to be present, the signal being stronger in malignant than in conventional or atypical tumors. Lastly, immunostaining for EGF/TGF receptor was positive in all tumors studied. Strong TGF protein expression in meningiomas is commonly associated with fibrous morphology. Although the frequent detection of both TGF protein and its mRNA, as well as of EGF/TGF receptor within tumors of all type and grades, suggests that TGF serves to promote tumor growth, its possible role in tumorigenesis or malignant progression is uncertain.In summary, demonstration of these substances is of no utility in the classification or grading of this common tumor because the differences in their expression among the various meningioma subtypes were not statistically significant.     

Cytotoxicity of ketoconazole in malignant cell lines

Summary The cytotoxic effects of ketoconazole, an antifungal agent known to have some activity against human prostate cancer, adrenal cancer, and male metastatic breast cancer, were evaluated using colony-growth and clonogenic assays in eight malignant cell lines. The cytotoxicity of ketoconazole showed a dose-and time-dependent pattern, with the following concentrations, inhibiting 90% of the growing colonies (IC₉₀): MCF 7 (human breast cancer) 7.25 g/ml, T 47 D (human breast cancer) 9.0 g/ml, MiaPaCa (human pancreatic carcinoma) 10.0 g/ml, COLO 357 (human pancreatic carcinoma) 9.5 g/ml, HCT 8 (human colonic adenocarcinoma) 27.1 g/ml, DU 145 (human prostatic cancer) 40.0 g/ml, AR 42 J (rat pancreatic carcinoma) 9.0 g/ml, and L1210 (murine leukemia) 8.6 g/ml. Since a concentration of 10 g/ml can be achieved in humans, the use of ketoconazole in human malignancies might be worthy of clinical evaluation.This investigation was supported in part by a grant from the German Volkswagen Foundation, Hannover, Federal Republic of Germany and by a gift from Dr Virgil Gianelli, Stockton, California     

Pharmacokinetic basis for the comparative antitumour activity and toxicity of chlorambucil,phenylacetic acid mustard and β,β-difluorochlorambucil (CB 7103) in mice

Summary This report describes the relationship between the pharmacokinetics, antitumour activity and toxicity of chlorambucil (CHL), phenylacetic acid mustard (PAAM) and ,-difluorochlorambucil (-F₂CHL) in mice. Pharmacokinetics were studied by HPLC, antitumour activity by a regrowth delay assay using the KHT murine sarcoma and toxicity by acute LD₅₀. For both antitumour activity and acute toxicity the order of potency was: PAAM>CHL>-F₂CHL. CHL and PAAM exhibited identical therapeutic indices, whereas that for -F₂CHL was somewhat improved. CHL is metabolized by mitochondrial -oxidation to the 3,4-dehydro derivative (DeHCHL) and PAAM, and the latter is further metabolized to its monodechloroethylated derivative DeC-PAAM, presumably by hepatic microsomal enzymes. Administered PAAM gave only one metabolite, DeC-PAAM. Unexpectedly, despite ,-disubstitution, -F₂CHL was also -oxidized to give DeHCHL and PAAM, but at reduced rates. Further, metabolic switching was demonstrated with the appearance in large amount of 2 new, unidentified metabolites, which may be dechlorethylation products. The pharmacokinetics of administered CHL, PAAM and -F₂CHL differ in that the plasma clearance was fastest for CHL, slowest for PAAM and intermediate for -F₂CHL. For the metabolites, CHL produced peak plasma concentrations of DeHCHL and PAAM, respectively, 7-fold and 2-fold greater than those produced by -F₂CHL. However, despite these differences, exposures to total bifunctional nitrogen mustards were similar following administration of the 3 drugs and therefore cannot account for their differential activity. In contrast, there was a good correlation between potency and PAAM exposure, which is highest after treatment with PAAM, intermediate after CHL and lowest after -F₂CHL. In plasma, 3.2% of PAAM is present as nonprotein-bound free drug, compared to 1.3% for DeHCHL, 0.9% for CHL and 0.45% for -F₂CHL. We propose the amount of free bifunctional nitrogen mustard, itself partly dependent on the extent of metabolism, to be of major importance for the in vivo potency of CHL analogues.     

Cytokines induce thymidine phosphorylase expression in tumor cells and make them more susceptible to 5′-deoxy-5-fluorouridine

The present study shows that various cytokines such as tumor necrosis factor (TNF), interleukin-1 (IL-1), and interferon- (IFN) make tumor cells much more susceptible to the cytostatic 5-deoxy-5-fluorouridine (5-dFUrd) than to 5-fluorouracil (5-FUra) and other cytostaties. These three cytokines increased the susceptibility of human cancer cell lines (COLO201, MKN45 and WiDr) but did not affect that of normal fibroblast WI38 cells. The cytokine mixture induced a 50-fold increase in the susceptibility of COLO201 to 5-dFUrd, whereas a 12-fold increase and a less than 5-fold enhancement in the susceptibility to 5-FUra and other cytostatics, respectively, were observed. The increased susceptibility would be a result of the induction of thymidine phosphorylase (TdR Pase), which is the essential enzyme for the conversion of 5-dFUrd to 5-FUra. The cytokine mixture increased TdR Pase activity by up to 47 times and greatly induced its mRNA expression in the cancer cell lines. These results suggest that the therapeutic benefit of 5-dFUrd would be improved by its use in combination with the cytokines.     

A phase II study to evaluate the combination of fludarabine, mitoxantrone and dexamethasone (FMD) in patients with follicular lymphoma

Background:Molecular response is being investigated as atherapeutic goal in follicular lymphoma (FL). High response rates in FL withthe fludarabine combination FMD have been associated with molecularremission. A phase II study of FMD in FL was therefore conducted. Patients and methods:Fifty-four patients, ten of whom were newlydiagnosed received FMD. Forty-four percent of the previously treated patientshad chemoresistant disease. Treatment comprised: fludarabine 25mg/m² days 1–3, mitoxantrone 10 mg/m² day 1, anddexamethasone 20 mg days 1–5. Blood/bone marrow was collected forquantitation of t(14;18) by real-time PCR. Results:The overall response rate was 37 of 54 (69%),complete responses being seen in 11 patients (20%), with no differencebetween newly diagnosed and the previously treated patients. However, theresponse rate in chemosensitive relapse was 84%, compared to44% in patients in whom the last prior regimen had failed. Molecularresponses were seen in 17 of 25 and PCR negativity in 8 of 25, althoughmolecular and clinical responses did not always correlate. Toxicity wasmoderate, 19 patients required admission. However, in 6 of 12 patients,subsequent G-CSF mobilised stem cell harvests failed. Conclusions:FMD was well tolerated but with a lower than expectedresponse rate. Molecular responses were seen in the majority of respondingpatients however, molecular remission was rare.     

Urinary testosterone as a marker of risk of recurrence in operable breast cancer

Summary We investigated the role of urinary testosterone levels as a marker of risk of recurrent disease in 113 operable breast cancer patients (70 premenopausal, 43 postmenopausal). Twenty-four-hour urine collections for testosterone measurement were obtained before surgical treatment, between 20–40 days thereafter, and then every 6 months for 5 years. The cutoff values to separate high testosterone (A+) from normal testosterone (A–) were 8.0µg/24h in premenopause and 4.9µg/24h in postmenopause. Urinary testosterone levels were considered high when they exceeded the cutoff value in at least 2 of the first 3 measurements (pretreatment, post-treatment, 6 months) of each patient. According to the aforementioned criterion, 33 patients (29.2%) had high testosterone levels, which were associated to axillary node involvement in 16 patients. Thirteen of the latter relapsed during the 5-year follow-up period (5/7 in premenopause, 8/9 in postmenopause). Relapse-free survival (RFS) curves were drawn only for node-positive patients owning to the small number of recurrences observed in the node-negative group. In premenopausal node-positive patients, RFS was significantly different for patients presenting high and normal urinary testosterone levels (77% vs 28%, respectively; logrank test, p< 0.006). In postmenopausal node-positive patients, RFS was also different between the two groups (54% vs 11% in high and normal excretors, respectively) but the difference was not statistically significant. The present findings suggest that urinary testosterone is a prognostic indicator of early breast cancer recurrence in node-positive patients.     

Adenosine and deoxyadenosine toxicity in colony assay systems for human T-lymphocytes,B-lymphocytes,and granulocytes

Summary Adenosine and deoxyadenosine toxicity was examined in colony assay systems for human T lymphocytes, B lymphocytes, and granulocytes. In the absence of deoxycoformycin, an adenosine deaminase inhibitor, no growth inhibition was observed in the three systems with concentrations of adenosine or deoxyadenosine of at least 200 M. Deoxycoformycin itself had no growth-inhibitory effect at concentrations of at least 10 g/ml. Combinations of deoxycoformycin (l g/ml) and either adenosine or deoxyadenosine gave growth inhibition in all three systems. Deoxyadenosine was the most toxic in all the systems, the LD₅₀ values being 20–25 M. The LD₅₀ values for adenosine were 45–55 M. There was no evidence of selective toxicity by adenosine or deoxyadenosine with these three colony assay systems. In the T-lymphocyte colony system deoxyadenosine appeared to be toxic to both the inducer/helper and the suppressor/cytotoxic T-lymphocyte subpopulations.     

Fas ligand expression in BRCA1-associated hereditary breast carcinoma clearly differs from that in sporadic breast carcinoma

BRCA1associated hereditary breast carcinomas (HBCs) are diagnosed at a younger age and are known to show biological aggressiveness such as a high histological grade, frequent aneuploidy, compared to sporadic breast carcinomas. However, results of studies on their prognosis have been controversial. We hypothesized that some factors such as a high incidence of cell death could offset the aggressiveness of BRCA1associated HBCs, and therefore investigated Fas and Fas ligand (Fas L) expression in 19 BRCA1associated HBCs and 56 ageadjusted control cases. Glandepithelial and myoepithelial cells in the mammary glands expressed Fas in high incidence, but all were negative for Fas L. Fas was expressed in 89.5% of BRCA1associated HBCs and 94.4% of the controls and no significant differences could be established between the two groups. However, in 73.7% of BRCA1associated HBCs, Fas L was clearly expressed in the infiltrating mononuclear cells, whereas this was observed in only 14.3% of the control cases and statistical significance was established between the two groups (p<0.0001). These results strongly suggest that carcinoma cells in BRCA1associated HBCs are more likely to be attacked by mononuclear cells via Fas L, and this may explain, at least in part, the discrepancy with respect to the prognosis. On the other hand, carcinoma cells also expressed Fas L significantly more often (p<0.0001) in BRCA1associated HBCs (52.6%) than in sporadic cases (3.6%). This can be considered a kind of counterattack against the mononuclear cells or, alternatively, may enhance the Fas–Fas L pathway in an autocrine/paracrine fashion. The clinical significance of Fas L expressing carcinoma cells remains to be elucidated.     

In vitro antitumor activity of mitomycin C derivative (RM-49) and new anticancer antibiotics (FK973) against lung cancer cell lines determined by tetrazolium dye (MTT) assay

Summary Seven small- (SCLC) and four non-small-cell (NSCLC) lung cancer cell lines were used to examine the in vitro cytotoxicity of cytotoxic drugs such as (1aS-(1a,8,8a,8b))-8-((aminocarbonyl)oxy)methyl-4,8a-dimethoxy-1,1a,2,8,8a,8b-hexahydro-7-hydroxy-5-methyl-6-nitrosoazirino(2,3: 3,4)-pyrrolo-(1,2-a)indole (RM-49) and 11-acetyl-8-carboxymethyl-4-formyl-14oxa-1, 11-diazetetracyclo(7.4.1.0^2,7,0^10,12)tetradeca-2-4-6-trien-6,9-diyl-diacetate (FK973). In vitro cytotoxicities of RM-49 and FK973 were compared with those of mitomycin C (MMC), cisplatin (CDDP), carboplatin (CBDCA), etoposide (VP16), adriamycin (ADM) and vindesin (VDS). Drug sensitivity was determined using a tetrazolium (MTT)-based assay. Average IC₅₀ values of these two drugs were not statistically different compared with that of MMC, although FK973 showed strong antitumor activity against SCLC cell lines such as LT3, N857, and H69 at the same concentration. The predicted peak plasma concentration (predicted PPC) calculated by the formula proposed by Scheithauer, log (predicted PPC) =-0.788+(0.755×log(LD₅₀)), and relative antitumor activity, RAA (PPC/IC₅₀), of RM-49 were higher than those of other drugs such as MMC, CDDP, CBDCA, and ADM against SCLC cell lines (P0.05), and those of FK973 were also higher than those of other drugs such as MMC, CDDP, CBDCA, and ADM against SCLC cell lines (P0.05). Based on these promising in vitro studies, the clinical trials of RM-49 and FK973 were warranted.     

Interferon-gamma pharmacokinetics and pharmacodynamics in patients with colorectal cancer

Purpose The study objectives were to define subcutaneous (s.c.) interferon gamma (IFN-) disposition in patients with gastrointestinal malignancies receiving 5-fluorouracil (5-FU) and leucovorin (LV) and to examine the relationship between IFN- exposures and Fas upregulation in vivo and in vitro.Methods Patients received IFN- (10, 25, 50, 75, and 100 g/m²) with LV and 5-FU, and serial samples were collected after the first dose. IFN- concentrations were measured by ELISA. A linear one-compartment model with a lag was fitted to the IFN- plasma concentration-time data. To examine the relationship between IFN- systemic exposure and biological activity in vivo, cell surface Fas upregulation was assessed in peripheral blood mononuclear cell (PBMC) subcompartments.Results The median (range) apparent IFN- clearance was 46 l/m² per hour (2.6–92 l/m² per hour). With increasing IFN- dosages, the area under the concentration-time curve (AUC₀) and C_max increased; however, significant interpatient variability was observed. IFN- AUC₀ and time above 33.3 pg/ml significantly correlated with Fas upregulation in several PBMC compartments, but dosage was significantly correlated with this pharmacodynamic marker only in CD4⁺ and CD56⁺ cells. In vitro studies in HT29 cells demonstrated that clinically relevant IFN- concentrations (1 to 10 U/ml for 6.5 h) with 5-FU/LV upregulated Fas expression 3.5-fold, similar to that in PBMC in vivo.Conclusions We characterized IFN- disposition and developed a limited sampling model for use in future pharmacokinetic studies. Our results showed that IFN- upregulates Fas in PBMC in vivo and in HT29 cells in vitro at tolerable, clinically relevant exposures and that monitoring IFN- pharmacokinetics/pharmacodynamics may be warranted in IFN- clinical use.This work was supported in part by US Public Health Service awards CA23099, CA32613 and CA23944, the Wings Cancer Foundation, and American Lebanese Syrian Associated Charities (ALSAC).     

In vivo effects in mice produced by a 3′-branched homologue of α-2′-deoxythioguanosine

Summary Studies with a 3-branched chain homolog (-3-BCTGdR) of 2-deoxythioguanosine (-TGdR) showed that it did not prolong the survival of mice bearing the Mecca lymphosarcoma. Host toxicity was quite profound and resembled that seen with 6-thioguanine (6-TG). Evidence was obtained that this nucleoside derivative was not appreciably converted to 6-TG in the mouse. Mice treated with toxic doses of 6-TG or -3-BCTGdR were found to have very similar pathological changes. The granulocytes were eliminated from the peripheral blood, bone marrow was acellular, and some more limited damage was seen in the intestinal crypts. Experiments with radiosulfur-labeled drugs demonstrated that -3-BCTGdR was incorporated into the DNA of mouse bone marrow, predominantly in the chain-terminating position, with the result that shorter chains of DNA accumulated. The new homolog, unlike -TGdR, was phosphorylated in bone marrow as well as in tumor, and incorporated well into the DNA both of bone marrow and of the neoplastic cells. In devising other homologs attention must be given to the specificity of the kinases, i.e., to whether phosphorylation is superior in tumor cells or in the growing normal cells.Abbreviations used are 6-TG 6-thioguanine - ,-TGdR ,-2-deoxythioguanosine - -3-BCTGdR 6-thioguanine--2,3-dideoxy-3-(hydroxymethyl)-d-erythro-pentofuranose - ³H-TdR Tritium-labeled (methyl) thymidine     

Evaluation of blood-brain barrier permeability and the effect of interferon in mouse glioma model

Summary The aim of this study was to evaluate whether interferon [IFN] can affect intracerebrally grown glioma and how alteration of the blood-brain barrier [BBB] may influence this effect. An intracerebrally implanted glioma G-26 (G-26) mouse brain-tumor model was developed and used in these studies. Histological characterization of this intracerebrally grown tumor revealed its anaplastic character. The astrocytic origin of G-26 was evidenced by glial fibrillary acidic protein staining and electron microscopic visualization of glial filaments. A study of tumor progression and animal survival showed development of a well defined tumor nodule within approximately seven days after the implantation. The median animal survival time was 27 ±3.8 days. The integrity of the blood-brain barrier [BBB] within the tumor was evaluated by the intravenous injection of horseradish peroxidase at days 3, 7,10 and 20 after brain tumor implant and compared to sham controls. The tumor-induced BBB alteration was progressive from day 3 to day 20.Glioma-26 subcutaneously passed in C57BL/6 mice was also continously cultured in vitro. Its proliferation was inhibited by homologous mouse interferon / [MuIFN / but not by human interferon a lymphoblastoid or human interferon ß. The in vivo studies of G-26 glioma treatment with MuIFN / were performed using single bolus of IFN in osmotically altered animals or slow IFN infusion through osmotic micro-pumps. The slow infusion of IFN had no effect on animal survival. However, a statistically significant increase in animal survival was observed after single bolus IFN treatment following osmotic BBB alteration.     

Studies on the biochemical mechanism of the novel antitumor agent,CI-920

Summary Biochemical studies on a new antitumor antibiotic, CI-920, have been directed toward understanding its mode of action. The most striking effect brought on by CI-920 was a marked inhibition of macromolecular synthesis. L1210 leukemia cells exposed to 10 M CI-920 exhibited a decreased rate of DNA, RNA, and protein synthesis within 45 min, and maximal inhibition occurred within 60 min. The reduction in nucleic acid synthesis was not due to precursor depletion, since ribonucleoside and deoxyribonucleoside triphosphate levels in cells exposed to 10 M CI-920 for 2 h either remained unchanged relative to control cells or were elevated, suggesting a block more directly at the level of nucleotide incorporation. Nevertheless, CI-920 (50 M) had no effect on DNA or RNA polymerase activity as assessed in permeabilized L1210 cells. However, if viable cells were exposed to 20 M CI-920 for 1 h prior to permeabilization and then the polymerases assayed in the absence of drug, there was a 60% depression in enzyme activity. The inhibition of RNA polymerase appears to result from an effect on the enzyme rather than the template, since inhibition of RNA polymerase activity in cell-free systems from drug-treated cells could not be restored by addition of excess DNA template. DNA polymerase, however, was at least partially restored by addition of template and therefore was inconclusive in this respect. The data, then, suggest that CI-920 inhibits nucleic acid synthesis directly at the level of nucleotide incorporation, either by direct inhibition of DNA or RNA polymerase or by inactivation of an essential component of these enzyme systems. Since the drug in its parent form did not inhibit nucleic acid synthesis in cell-free systems the effects may possibly be mediated through conversion of this agent to another chemical form within viable cells.     

A Randomized Phase II and Pharmacokinetic Study of Dacarbazine in Patients with Recurrent Glioma

We conducted a randomized phase II study to determine the efficacy of dacarbazine (DTIC) in recurrent gliomas. Patients were randomly assigned to receive either DTIC 750mg/m²IV day 1 every 28 days (Arm A) or DTIC 200mg/m²IV days 1–5 every 28 days (Arm B). Pharmacokinetics were studied in 6 patients on each arm using HPLC analysis. Thirty-nine patients (30 male, 9 female), ages 27–67 years (median 53) were entered on the study (20 on Arm A, 19 on Arm B). No objective responses were seen. Median time to progression was 3 months. Median survival was 8 months. Treatment was generally well tolerated. Major toxicities were grade 1–2 nausea (33%), lethargy (28%), diarrhea (15%), alopecia (15%), and grade 3 neutropenia (8%). Four patients on Arm A had mild self-limited episodes of intravascular hemolysis occurring immediately after drug infusion, the mechanism of which is unknown. Mean AUC for DTIC, HMMTIC (5-[3-hydroxymethyl-3-methyl-1-triazeno] imidazole-4-carboxamide), and MTIC (5-[3-methyl-1-triazeno] imidazole-4-carboxamide), in Arm A were 14.8, 0.17, and 1.15mMmin, respectively. Corresponding values for Arm B (on day 1 of 5) were 1.7, 0.06, and 0.29mMmin, respectively. The predicted HMMTIC and MTIC exposure over 5 days for Arm B, based on the day 1 data, is higher than with Arm A. We conclude that DTIC is well tolerated but does not have activity in patients with recurrent gliomas. The 5-day schedule appears less toxic, and pharmacokinetic studies show that it provides greater exposure to MTIC and HMMTIC compared to the one-day schedule.