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Comparison of media conditions to expand TIL-derived T-cell lines from patients with peritoneal carcinomatosis including serum free, rIFN-g and polyenzyme preparation

Kim Y.M, Zavadova E., Loercher A., Freedman R.S.

Dept of Obstet. & Gynecol, College of Medicine, Univ. of Ulsan, Asan Medical Center, Seoul, Korea & Dept. Of Gyn/Oncol. M.D. Anderson Cancer Center, Houston, TX

Proceedings of the American Association for Cancer Research (1999): 40, No. 1220.

The optimal culture conditions for the expansion of TIL ex vivo have not been developed. In the interest of patient safety, serum free conditions have been promoted (J Immunol Meth 167:145-160, 1994). However, serum may contain nutritional or other components that facilitate T cell line growth. Objective: To develop improved culture conditions for ex vivo expansion of TIL. Specimens & Methods: We compared our serum free method of expansion of TIL from 19 patients under the following culture conditions: (a) + 5% FCS, (b) + 5% FCS + rIFN-g, (c) + rIFN-g, and (d) 5% FCS + rIFN-g + a polyenzyme preparation containing trypsin, chymotrypsin and papain. Proteolytic enzymes are known to neutralize TGF-b which inhibits lymphocyte proliferation. After 3 weeks in culture TIL were examined for (i) fold of proliferation, (ii) leukocyte cell surface antigens by FACS, (iii) cytokine production by RT-PCR, and (iv) cytotoxicity against allogeneic or autologous tumor cells. Results: (a) The fold of proliferation of TIL grown in FCS was significantly better than cultures without FCS. TIL with rIFN-g and polyenzyme preparation were significantly better than with rIFN-g alone (p<0.001). No differences were detected in expression of CD3⁺, TCRab⁺, CD4⁺, or CD8⁺ T cells but a significant increase in expression of CD56⁺ cells in the interferon group (p <0.001) and the interferon group with enzymes (p <0.01) as compared with medium alone. No differences were detected for in vitro cytotoxicity to autologous or allogeneic cells. TIL expanded in culture expressed mRNA for IL-2, IFN-g, GM-CSF, and b-actin, but not IL-4 and IL-10. We conclude that culture of TIL from patients with peritoneal carcinomatosis with IFN-g and polyenzyme preparation significantly stimulate the growth rate, but does not significantly change the phenotype of lymphocytes. Supported in part by Betty Ann Asch Murray fund for research in GYN/Med Oncol.


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Zánět a jeho ovlivnění SET Zvýšení využitelnosti antibiotik proteolytickými enzymy Angiologie, flebologie Gynekologie Chirurgie, traumatologie, ortopedie Imunologie Lymfologie ORL Pediatrie Revmatologie Sportovní medicína Stomatologie Urologie Literatura Časopisy s Impact faktorem Ostatní literatura SPC Wobenzym SPC Phlogenzym	Comparison of media conditions to expand TIL-derived T-cell lines from patients with peritoneal carcinomatosis including serum free, rIFN-g and polyenzyme preparation Kim Y.M, Zavadova E., Loercher A., Freedman R.S. Dept of Obstet. & Gynecol, College of Medicine, Univ. of Ulsan, Asan Medical Center, Seoul, Korea & Dept. Of Gyn/Oncol. M.D. Anderson Cancer Center, Houston, TX Proceedings of the American Association for Cancer Research (1999): 40, No. 1220. The optimal culture conditions for the expansion of TIL ex vivo have not been developed. In the interest of patient safety, serum free conditions have been promoted (J Immunol Meth 167:145-160, 1994). However, serum may contain nutritional or other components that facilitate T cell line growth. Objective: To develop improved culture conditions for ex vivo expansion of TIL. Specimens & Methods: We compared our serum free method of expansion of TIL from 19 patients under the following culture conditions: (a) + 5% FCS, (b) + 5% FCS + rIFN-g, (c) + rIFN-g, and (d) 5% FCS + rIFN-g + a polyenzyme preparation containing trypsin, chymotrypsin and papain. Proteolytic enzymes are known to neutralize TGF-b which inhibits lymphocyte proliferation. After 3 weeks in culture TIL were examined for (i) fold of proliferation, (ii) leukocyte cell surface antigens by FACS, (iii) cytokine production by RT-PCR, and (iv) cytotoxicity against allogeneic or autologous tumor cells. Results: (a) The fold of proliferation of TIL grown in FCS was significantly better than cultures without FCS. TIL with rIFN-g and polyenzyme preparation were significantly better than with rIFN-g alone (p<0.001). No differences were detected in expression of CD3⁺, TCRab⁺, CD4⁺, or CD8⁺ T cells but a significant increase in expression of CD56⁺ cells in the interferon group (p <0.001) and the interferon group with enzymes (p <0.01) as compared with medium alone. No differences were detected for in vitro cytotoxicity to autologous or allogeneic cells. TIL expanded in culture expressed mRNA for IL-2, IFN-g, GM-CSF, and b-actin, but not IL-4 and IL-10. We conclude that culture of TIL from patients with peritoneal carcinomatosis with IFN-g and polyenzyme preparation significantly stimulate the growth rate, but does not significantly change the phenotype of lymphocytes. Supported in part by Betty Ann Asch Murray fund for research in GYN/Med Oncol.
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