No abstract

MelphalanMelphalan (Alkeran, 50 mg per vial, Wellcome, Beckenham, UK) was diluted in 10 ml solvent. Further dilutions were made in 0.9% sodium chloride to give a concentration of 1 µg µl -1 . A volume of 40 µl (= 40 µg) was added to the perfusion circuit.Prerequisites for effective isolated limb perfusion using tumour necrosis factor alpha and melphalan in rats Summary An isolated limb perfusion (ILP) model using soft tissue sarcoma-bearing rats was used to study prerequisites for an effective ILP, such as oxygenation of the perfusate, temperature of the limb, duration of the perfusion and concentration of tumour necrosis factor (TNF). Combination of 50 µg TNF and 40 µg melphalan demonstrated synergistic activity leading to a partial and complete response rate of 71%. In comparison to oxygenated ILP, hypoxia was shown to enhance anti-tumour activity of melphalan alone and TNF alone but not of their combined use. Shorter perfusion times decreased anti-tumour responses. At a temperature of 24-26°C, anti-tumour effects were lost, whereas temperatures of 38-39°C or 42-43°C resulted in higher response rates. However, at 42-43°C, local toxicity impaired limb function dramatically. Synergy between TNF and melphalan was lost at a dose of TNF below 10 µg in 5 ml perfusate. We conclude that the combination of TNF and melphalan has strong synergistic anti-tumour effects in our model, just as in the clinical setting. Hypoxia enhanced activity of melphalan and TNF alone but not the efficacy of their combined use. For an optimal ILP, minimal perfusion time of 30 min and minimal temperature of 38°C was mandatory. Moreover, the dose of TNF could be lowered to 10 µg per 5 ml perfusate, which might allow the use of TNF in less leakage-free or less inert perfusion settings.

We report a rare case of a primary leiomyosarcoma of the adrenal gland. A 73-year-old woman presented with an inferior vena cava syndrome. MR imaging was suggestive of a large tumour originating from the right adrenal gland. Angiography revealed a tumour vascularised by the right adrenal artery. At explorative laparotomy a tumour of 27 cm in diameter was found which was completely fixed to the liver; the tumour was therefore considered unresectable. As a consequence of the mechanical problems caused by this large tumour, the patient died 3 weeks after the operation. Autopsy revealed no distant metastases or other primary tumour site.

Isolated hepatic perfusion (IHP) with melphalan with or without tumour necrosis factor alpha (TNF-a) is currently performed in clinical trials in patients with hepatic metastases. Previous studies led to the hypothesis that the use of TNF-a in isolated limb perfusion causes specific destruction of tumour endothelial cells and thereby induces an increased permeability of tumour vasculature. However, whether TNF-a contributes to the therapeutic efficacy in IHP still remains unclear. In an in vivo rat liver metastases model we studied three different tumours: colon carcinoma CC531, ROS-1 osteosarcoma and BN-175 soft-tissue sarcoma which exhibit different degrees of vascularisation. IHP was performed with melphalan with or without the addition of TNF-a. IHP with melphalan alone resulted, in all tumour types, in a decreased growth rate. However in the BN-175 tumour addition of TNF-a resulted in a strong synergistic effect. In the majority of the BN-175 tumour-bearing rats, a complete response was achieved. In vitro cytoxicity studies showed no sensitivity (CC531 and BN-175) or only minor sensitivity (ROS-1) to TNF-a, ruling out a direct interaction of TNF-a with tumour cells. The response rate in BN-175 tumour-bearing rats when TNF-a was coadministrated with melphalan was strongly correlated with drug accumulation in tumour tissue, as only in these rats a five-fold increased melphalan concentration was observed. Secondly, immunohistochemical analysis of microvascular density (MVD) of the tumour showed a significantly higher MVD for BN-175 tumour compared to CC531 and ROS-1. These results indicate a direct relation between vascularity of the tumour and TNF-a mediated effects. Assessment of the tumour vasculature of liver metastases would be a way of establishing an indication for the utility of TNF-a in this setting.

An isolated limb perfusion (ILP) model using soft tissue sarcoma-bearing rats was used to study prerequisites for an effective ILP, such as oxygenation of the perfusate, temperature of the limb, duration of the perfusion and concentration of tumour necrosis factor (TNF). Combination of 50 microg TNF and 40 microg melphalan demonstrated synergistic activity leading to a partial and complete response rate of 71%. In comparison to oxygenated ILP, hypoxia was shown to enhance anti-tumour activity of melphalan alone and TNF alone but not of their combined use. Shorter perfusion times decreased anti-tumour responses. At a temperature of 24-26 degrees C, anti-tumour effects were lost, whereas temperatures of 38-39 degrees C or 42-43 degrees C resulted in higher response rates. However, at 42-43 degrees C, local toxicity impaired limb function dramatically. Synergy between TNF and melphalan was lost at a dose of TNF below 10 microg in 5 ml perfusate. We conclude that the combination of TNF and melphalan has strong synergistic anti-tumour effects in our model, just as in the clinical setting. Hypoxia enhanced activity of melphalan and TNF alone but not the efficacy of their combined use. For an optimal ILP, minimal perfusion time of 30 min and minimal temperature of 38 degrees C was mandatory. Moreover, the dose of TNF could be lowered to 10 microg per 5 ml perfusate, which might allow the use of TNF in less leakage-free or less inert perfusion settings.