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Objective: To extract and purify ricin from castor beans and to evaluate its anti-cancer activity., Methods: Ricin was purified from castor beans according the modified method of Nicolson and Blaustin. The lectins were extracted in 0.01 mol/L phosphate buffered saline and isolated in the 40% to 80% fraction of ammonium sulfate precipitation. The dialyzed fractionated preparation was applied with a Sepharose 4B column. The lectins were eluted with a linear lactose gradient (0.01 mol/L approximately 0.5 mol/L). Ricin was separated from the ricinus agglutinin by gel filtration on a Sephadex G-100. MTT was applied to analyze the cytotoxicity with different dosage of ricin in different cancer cell lines., Results: There was no difference between the killing effect of normal cells and that of colon cancer cells by using the high dosage of ricin (5 x 10(-8) mol/L approximately 5 x 10(-10) mol/L). However, the cytotoxicity was significant different in those cells with the low dosage of ricin (5 x 10(-11) mol/L approximately 5 x 10(-13) mol/L). Meanwhile ricin had the similar cytotoxicity to leukemia cell K562 and colon cancer cell SW480., Conclusion: Ricin is able to kill tumor cells selectively at low concentration, but the selectivity does not appear at high concentrations.

This study was aimed to investigate the clinical outcome of ricin-immunotoxin mediated T cell partially depleted HLA/MLC mismatched allogeneic hematopoietic stem cell transplantation. 13 patients with hematological malignancies were treated by ricin-immunotoxin mediated T cell partially depleted allogeneic hematopoietic stem cell transplantations from HLA/MLC mismatched donors, including 6 cases of CML in CP(1), 1 case of ALL in CR(1), 1 case of ALL in CR(2), 1 case of ALL in relapse, 2 cases of AML in CR(1), 1 case of AML in CR(2), 1 case of MDS-RAEBT-AML (M(4)) in CR(1). The results showed that 8 cases were engrafted successfully, 2 cases of them developed grade II acute GVHD and 2 cases developed grade III-IV acute GVHD. Within following-up of 8 - 90 months, 2 patients who experienced grade III-IV acute GVHD died early after transplantation; 1 patient died of late onset of infection; the other 5 patients survived free from diseases. After failure at first infusion, 4 patients were given reinfusion of peripheral blood hematopoietic stem cells from the same donor. 3 out of 4 cases failed to engraft and only one patient got engraftment but died of related complications of transplantation. One patient was performed a second transplantation from a syngeneic donor and survive free of disease until now. In conclusion, T cell partially depleted HLA/MLC mismatched allogeneic hematopoietic stem cell transplantation by ricin-immunotoxin decreases the occurrence of severe acute GVHD but with high risk of rejection, which clinical outcome still needs further evaluation.

Bispecific monoclonal antibody (bsMAb), secreted by hybrid-hybridoma, has an intact 1g molecule construction with dual antigen binding specificities. bsMAb has several advantages over conventional conjugate in tumor targeting therapy: (1) The damage to both antibody and tumoricidal agents due to chemical crosslinking can be avoided. (2) The antigen modulation can be decreased due to monovalence binding of bsMAb with target antigen. (3) The binding affinity between Fc fragment of bsMAb with heterogenous heavy chains and Fc receptor is decreased, so that nonspecific distribution and side effects can be reduced. (4) The bsMAb can pretarget to tumor site, increasing the T/NT ratio and cytotoxicity. In the present study, A bsMAb with versatile adaptor was designed. In this bsMAb, one arm could react with gastric cancer-associated antigen, and the other with a hapten TNP. The bsMAb could mediate different tumoricidal agents crosslinked to TNP. A variant hybridoma 3H11-HAT(s) secreting McAb against gastric cancer was fused with spleen cells of mice immunized with KLH-TNP. After screening and subcloning, 10 hybrid-hybridomas were obtained, which secreted antibodies against both gastric cancer target cells BGC823 and BSA-TNP. By using special bridge method, only hybrid-hybridomas 6A3 (gamma 2b, mu), and 1G7 (gamma 2b, gamma 2b) were confirmed to secrete bsMAb. Further experiments showed that the bsMAb 6A3 and 1G7 could mediate different cytotoxicities, for example, Ricin-TNP, MMC-HSA-TNP and ADM-BSA-TNP. This system is useful for evaluating different tumoricidal agents in bsMAb targeting therapy, and has potential value in clinics.(ABSTRACT TRUNCATED AT 250 WORDS)

Three anti-breast carcinoma monoclonal antibodies 317G5, 260F9 and 741F8 linked to ricin A chain (rRTA) were selected for the present investigation. Attempts were made to understand the relative efficiency of these immunotoxins in relation to: a) binding, b) internalization, and c) clonogenic suppression of breast cancer cells in vitro. The results indicated that 317G5-rRTA combined with 260F9-rRTA had synergistic antitumor activity against SKBr3 breast carcinoma cell line. Individually, 10 micrograms/ml 260F9-rRTA or 317 G5-rRTA alone could only eliminate 2-3 logs of clonogenic SKBr3 cells. When used in combination, 260F9-rRTA and 317G5-rRTA, 1 microgram/ml each, could kill 4 logs of clonogenic tumor cells. The mechanism of the synergistic action is discussed.