ASSESSMENT OF THE ROLE OF NEUTROPHILS ON THE ANTITUMOR EFFECT OF TNF-ALPHA IN AN IN-VIVO ISOLATED LIMB PERFUSION MODEL IN SARCOMA-BEARING BROWN-NORWAY RATS

Introduction. Isolated limb perfusion (ILP) with TNF alpha in combinat ion with melphalan and IFN gamma has resulted in an immediate and dram atic tumor response in patients. Such an effect was also noted followi ng ILP in a rat sarcoma model. This model enables us to investigate se veral factors responsible for the TNF alpha-induced tumor responses. W e applied total body irradiation (TBI) to reduce white blood cell coun t, to investigate the contribution of leukocytes to the antitumor effe ct of TNF alpha. Methods. Small fragments of the nonimmunogenic BN 175 sarcoma were implanted sc in the lower hind leg. A 5 Gy TBI was perfo rmed before ILP at a tumor diameter of approximately 15 mm. The hind l imbs of 63 rats were perfused and were divided into 6 groups: group 1, sham perfusion, n = 9; group 2, TBI + sham perfusion, n = 6; group 3, TNF alpha 50 mu g, n = 9; group 4, melphalan 40 mu g, n = 9; group 5, TNF alpha 50 mu g + melphalan 40 mu g, n = 22; group 6, TBI + TNF alp ha + melphalan ILP, n = 8. In addition, 10 rats were perfused for hist ological analysis at 24 h post-ILP. Results. We observed in Group 1: 9 /9 progressive disease (PD); Group 2: 6/6 PD; Group 3: 9/9 PD; Group 4 : 9/9 no change (NC) of tumor diameter for at least 4 days; Group 5: 6 /22 NC, 16/22 complete remission (CR), 12/16 of which showed skin necr osis at the tumor site; and Group 6: 7/8 NC and 1/8 CR (without skin n ecrosis). After TBI, WBC reduction of 80-95% was observed, while the n umber of platelets was not significantly reduced and platelet aggregat ion was maintained at 72 %. Histological analysis revealed decreased h emorrhagic necrosis associated with the absence of PMN infiltration at the tumor margins in the TBI rats. Conclusion. TBI and the associated reduction in WBC count decreased the tumor response by TNF alpha and melphalan significantly and abrogated the immediate response of skin n ecrosis at the tumor site, as found in rats treated with TNF alpha and melphalan without TBI, These data strongly suggest that leukocytes pl ay an important role in the hemorrhagic effects of TNF alpha. (C) 1998 Academic Press.