Over-expression of Bcl-2 protects against apoptosis induced by the bioreductive cytotoxic drug SR4233 (Tirapazamine).

The human B-cell lymphoma cell line PW undergoes radiation-induced programmed cell death (PCD). Bcl-2 transfected PW cells, that overexpressed Bcl-2, were significantly more radioresistant than parental or neomycin control transfected PW cells. The viability of Bcl-2 transfected cells was significantly greater than that of parental PW cells treated with the bioreductive cytotoxin SR4233 under aerobic conditions. Bcl-2 transfectants were also significantly more resistant to hypoxia-induced PCD. However, there was no significant difference in the viability of parental and Bcl-2 transfected cells exposed to SR4233 under hypoxic conditions (pO(2)<100 ppm). Incubation of parental PW cells with N-acetyl cysteine decreased the cytotoxicity of SR4233 under aerobic but not anaerobic conditions. Depletion of cellular glutathione with buthionine sulphoxamine killed nearly 100% of control PW cells, but none of the Bcl-2 transfectants under the same conditions. The TBARS assay for lipid peroxidation showed that Bcl-2 transfectants had a significantly lower level of lipid peroxidation than parental PW cells following a 24 hour constant exposure to SR4233 under aerobic conditions. These results suggest that Bcl-2 overexpression inhibits PCD induced by the bioreductive cytotoxin SR4233 under aerobic conditions as well as PCD induced by hypoxia, and that there are other pathways leading to PCD that are unaffected by Bcl-2 overexpression.

Treatment of hormone-refractory prostate cancer with 90Y-CYT-356 monoclonal antibody.

A Phase I dose-escalation study using 90Y-CYT-356 monoclonal antibody was performed in 12 patients with hormone-refractory prostate carcinoma. Biodistribution studies using 111In-CYT-356 were performed 1 week before 90Y-CYT-356 administration. Of the 12 patients, 58% had at least one site of disease imaged after administration of 111In-CYT-356. The dose of 90Y ranged from 1.83-12 mCi/m2. Both 111In and 90Y-CYT-356 were tolerated well, without significant nonhematological toxicity. Myelosuppression was the dose-limiting toxicity and occurred at dose levels of 4.5-12 mCi/m2. Of the patients receiving

Yttrium-90-labeled anti-CD20 monoclonal antibody therapy of recurrent B-cell lymphoma.

A Phase I/II dose escalation study of 90Y-murine anti-CD20 monoclonal antibody (mAb) in patients with recurrent B-cell lymphoma was performed. The primary objectives of the study were: (a) to determine the effect of the preinfusion of unlabeled anti-CD20 mAb on the biodistribution of 111In-anti-CD20 mAb; (b) to determine the maximal tolerated dose of 90Y-anti-CD20 mAb that does not require bone marrow transplantation; and (c) to evaluate the safety and antitumor effect of 90Y-anti-CD20 mAb in patients with recurrent B-cell lymphoma. Eighteen patients with relapsed low- or intermediate-grade non-Hodgkin's lymphoma were treated. Biodistribution studies with 111In-anti-CD20 mAb were performed prior to therapy. Groups of three or four patients were treated at dose levels of approximately 13.5, 20, 30, 40, and 50 mCi 90Y-anti-CD20 mAb. Three patients were retreated at the 40-mCi dose level. The use of unlabeled antibody affected the biodistribution favorably. Nonhematological toxicity was minimal. The only significant toxicity was myelosuppression. The overall response rate following a single dose of 90Y-anti-CD20 mAb therapy was 72%, with six complete responses and seven partial responses and freedom from progression of 3-29+ months following treatment. Radioimmunotherapy with

Intratumoral radioimmunotherapy of a human colon cancer xenograft using a sustained-release gel.

Low tumor uptake and normal tissue toxicity limit the efficacy of RIT for the treatment of solid tumors. In this study, an intratumoral injectable gel drug delivery system for local administration of RIT was evaluated using the LS174T human colon cancer xenograft model in SCID mice. The injectable gel is a collagen-based drug delivery system designed for intratumoral (i.t.) administration, which has previously been shown to enhance drug retention at the injection site and reduce systemic drug exposure. We compared the local (tumor) retention and biodistribution of 111In-labeled NR-LU-10 monoclonal antibody given i.t. in the injectable gel versus simple aqueous solution. 111In gel given i.t. and 111In-NR-LU-10 given intraperitoneally (i.p.) were used as controls. The results showed that tumors treated with 111In-NR-LU-10 gel maintained the highest levels of radioactivity for up to 96 h. At 48 h after the administration of 111In-NR-LU-10 gel i.t., 111In-NR-LU-10 solution i.t., 111In gel i.t., or 111In-NR-LU-10 i.p., the level of radioactivity remaining in each gram of tumor was 98, 49, 45, and 16% of the injected dose, respectively. It was estimated that if 100 microCi of 90Y-NR-LU-10 were administered similarly, tumor treated with 90Y-NR-LU-10 gel i.t. would receive a dose of 90.0 Gy, whereas normal tissues in the same animal would receive a dose of approximately 2.43 Gy. In contrast, if 90Y-NR-LU-10 were delivered i.p., a comparable tumor would receive a dose of 16.8 Gy and corresponding normal tissues would receive 3.36 Gy. Consistent with these estimates, enhanced antitumor efficacy was observed when 90Y-NR-LU-10 gel was administered i.t. Tumor growth delay time was 6.9-fold (P < 0.01) longer in these animals (14.4 days) than in animals treated with 90Y-NR-LU-10 i.p. (2.1 days). Systemic toxicity was also significantly reduced in gel-treated animals as monitored by loss of body weight. This study demonstrated that intratumoral delivery of 90Y-NR-LU-10 gel markedly increased the retention of the radioisotope in tumors, enhanced the antitumor efficacy, and reduced systemic toxicity compared to systemic administration of the radiolabeled antibody. This injectable gel drug delivery system may allow for improvement in the therapeutic index for RIT.

Delta-9-tetrahydrocannabinol treatment results in a suppression of interleukin-2-induced cellular activities in human and murine lymphocytes.

Delta-9-tetrahydrocannabinol (THC), the major psychoactive component in marijuana, has been shown to suppress a variety of interleukin-2-(IL-2)-dependent cellular functions in both murine and human lymphocytes. These effects were examined in both human peripheral blood lymphocytes (hPBL) and the IL-2-dependent murine cytotoxic T-cell line CTLL-2. Interleukin-2-induced thymidine uptake and uridine uptake were suppressed in a dose related manner when cells were co-incubated for 48 h with 100 U rhIL-2/ml and 1-10 micrograms THC/ml. Interleukin-2-induced protein synthesis was also suppressed in a dose related manner over this THC concentration range, with the hPBL being more susceptible to the suppressive effect of THC than the CTLL-2 cells. Autoradiographic analysis of the synthesized proteins from hPBL cell lysates reveals a generalized suppression of all nascent proteins in THC-treated cultures. Human natural killer cell activity is only affected at the highest concentration tested (10 micrograms THC/ml) while lymphokine-(IL-2)-activated natural killer cell activity is affected throughout the range of 1-10 micrograms THC/ml. Together these results suggest that THC interferes with the IL-2:IL-2 receptor signaling cascade at one or possibly many points causing a decrease in IL-2-induced metabolic activity and cytolytic function.

Radiobiologic studies of radioimmunotherapy and external beam radiotherapy in vitro and in vivo in human renal cell carcinoma xenografts.

BACKGROUND: Previous studies suggest that the radiobiologic characteristics of in vitro survival curves are important determinants of the response of tumors to both conventional radiotherapy and radioimmunotherapy (RIT). The purpose of this study was to elucidate the relationship between in vitro radiation survival curve parameters and the relative sensitivity of tumor to RIT, exponentially decreasing low dose rate (ED LDR) irradiation and conventional high dose rate (HDR) fractionated external beam radiotherapy. METHODS: Two human renal cell carcinoma cell lines, Caki-1 and A498, were used in vitro and nude mouse xenograft studies. HDR external beam gamma irradiation (dose rate, 430 centigray [cGy]/minute) and ED LDR irradiation (initial dose rate, 22-25 cGy/hour) were performed with a cesium-137 (137Cs) gamma irradiator. RIT was carried out with yttrium-90 (90Y-labeled monoclonal antibody NR-LU-10, and the absorbed radiation doses were calculated by medical internal radiation dose methodology. A clonogenic assay was used to generate radiation survival curves, and a computer FIT program was used to calculate the radiobiologic parameters. The antitumor efficacy of the different treatments was compared in vivo using a tumor regrowth delay assay in these two tumor xenograft models. RESULTS: The radiation survival curves showed that the Caki-1 cell line was more sensitive to both HDR and ED LDR irradiation than A498 in vitro. The Caki-1 cell line, compared with A498, had a larger alpha (0.39 vs. 0.15 Gy following HDR and 0.32 vs. 0.21 Gy following ED LDR) and alpha-to-beta ratio (6.92 vs. 2.60 Gy for HDR and 40.0 vs. 19.2 Gy for ED LDR), a smaller n number (5.13 vs. 23 for HDR and 1.16 vs. 3.53 for ED LDR), a lower quasi-threshold dose (Dq) (1.60 vs. 3.15 Gy for HDR and 0.35 vs. 1.76 Gy for ED LDR), and a lower surviving fraction at 2 Gy (SF2) (0.37 vs. 0.60 for HDR and 0.51 vs. 0.61 for ED LDR), suggesting that Caki-1, compared with A498, had a steep initial slope and a small shoulder. The final slope represented by the beta value and D0 dose (the dose (Gy) required to reduce the fraction of surviving cells of 37% of its previous value in the exponential region of the survival curves) did not vary significantly between these two cell lines at either HDR or ED LDR irradiation. Tumor volume doubling times were 4.0 +/- 1.5 days for Caki-1 and 4.2 +/- 1.8 days for A498 tumor xenografts. One hundred microCi/50 microg of 90Y-labeled, isotype-matched irrelevant monoclonal antibody CCOO16-3 produced a tumor growth delay time (TGD) of 2.1 days in Caki-1 tumors but had no effect on A498 tumors (P < 0.05). RIT with 100 microCi of 90Y-NR-LU-10 resulted in a TGD of 4.8 days for Caki-1 tumors, whereas 100 microCi and 150 microCi of 90Y-NR-LU-10 produced a TGD of 1.9 and 2.7 days for A498 tumors, respectively. Estimated absorbed doses were 21.9 Gy in Caki-1 tumors treated with 100 microCi of 90Y-NR-LU-10 and 14.5 Gy and 21.8 Gy in A498 tumors treated with 100 microCi and 150 microCi of 90Y-NR-LU-10, respectively. The weighted normal tissue absorbed doses were 7.4 Gy for Caki-1 tumor-bearing mice and 9.0 Gy for A498 tumor-bearing mice (P > 0.05). To compare the responses of Caki-1 and A498 xenografts to RIT with external beam ED LDR and HDR irradiation, tumor-bearing mice were treated with equivalent doses (20-22 Gy) of 1) RIT with 90Y-NR-LU-10 (100 microCi for Caki-1 and 150 microCi for A498), 2) continuous ED LDR 137Cs irradiation with a initial dose rate of 22 cGy/hour, or 3) HDR X-irradiation (2 Gy x 10 fractions in 2 weeks). The TGDs produced by RIT, ED LDR, and HDR were 5.3, 9.7, and 8.3 days for Caki-1 and 2.7, 5.1, and 5.8 days for A498. The relative efficacy of RIT in these xenograft models correlated well with the radiobiologic parameters (i.e., the size of the initial slope and shoulder) of in vitro survival curves following HDR and ED LDR irradiation in these cell lines. (ABSTRACT TRUNCATED)