In vivo and in vitro evaluation of combretastatin A-4 and its sodium phosphate prodrug

Abstract

SummaryThe anti-tumour effects and mechanism of action of combretastatin A-4 and its prodrug, combretastatin A-4 disodium phosphate, were examined in subcutaneous and orthotopically transplanted experimental colon tumour models. Additionally, the ability of these compounds to directly interfere with endothelial cell behaviour was also examined in HUVEC cultures. Combretastatin A-4 (150 mg kg -1 , intraperitoneally (i.p.)) and its water-soluble prodrug (100 mg kg -1 , i.p.) caused almost complete vascular shutdown (at 4 h), extensive haemorrhagic necrosis which started at 1 h after treatment and significant tumour growth delay in MAC 15A subcutaneous (s.c.) colon tumours. Similar vascular effects were obtained in MAC 15 orthotopic tumours and SW620 human colon tumour xenografts treated with the prodrug. More importantly, in the orthotopic models, necrosis was seen in vascularized metastatic deposits but not in avascular secondary deposits. The possible mechanism giving rise to these effects was examined in HUVEC cells. Here cellular networks formed in type I calf-skin collagen layers and these networks were completely disrupted when incubated with a non-cytotoxic concentration of combretastatin A-4 or its prodrug. This effect started at 4 h and was complete by 24 h. The same non-cytotoxic concentrations resulted in disorganization of F-actin and β-tubulin at 1 h after treatment. In conclusion, combretastatin A-4 and its prodrug caused extensive necrosis in MAC 15A s.c. and orthotopic colon cancer and metastases, resulting in anti-tumour effects. Necrosis was not seen in avascular tumour nodules, suggesting a vascular mechanism of action.

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“…In the first of these studies, a single dose of combretastatin A-4 phosphate (100 mg/kg) induced extensive haemorrhagic necrosis, with a rim of viable tumour at the periphery of the primary colorectal tumour 24 h after drug treatment. This effect was also observed in vascular metastatic deposits in the kidney, lymph nodes and abdominal wall, but no effect was observed in avascular deposits in the lung (Grosios et al, 1999), supporting the hypothesis that the tumour necrosis induced by the drug is secondary to vascular effects in the orthotopic setting. In the follow-up study, a single dose of combretastatin A-4 phosphate (150 mg/kg) also induced haemorrhagic necrosis in the colorectal tumours within the liver 24 h after drug treatment (Holwell et al, 2002).…”

“…The activity of combretastatin A-4 phosphate has been reported in several physiologically relevant mouse models, including orthotopic models of mouse (MAC 15) or human (SW620) colorectal cancer (Grosios et al, 1999), models of human colon cancer (DLD-1 and HT29) metastasis to the liver (Holwell et al, 2002) and orthotopic models of human NSCLC (KNS-62, Colo-699) (Boehle et al, 2001). In the first of these studies, a single dose of combretastatin A-4 phosphate (100 mg/kg) induced extensive haemorrhagic necrosis, with a rim of viable tumour at the periphery of the primary colorectal tumour 24 h after drug treatment.…”

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“…CA4P is a relatively simple stilbene that has been shown to interact with tubulin at or near the colchicine-binding site, resulting in the inhibition of tubulin assembly and therefore disruption of microtubular function (McGown and Fox, 1989). Treatment with combretastatin A4, and its more soluble prodrug CA4P (Pettit et al, 1995), results in vascular shutdown within the tumour causing massive haemorrhagic necrosis (Dark et al, 1997;Grosios et al, 1999). In animals, CA4P treatment results in significant antitumour effects at less than one-tenth of the MTD (Dark et al, 1997).…”

Fibrinogen E fragment selectively disrupts the vasculature and inhibits the growth of tumours in a syngeneic murine model

“…similar to the concentration used in our in vitro studies). Tumour volumes were calculated daily using calipers (Grosios et al, 1999). After 8 days, mice were killed and tumours and normal tissues excised, divided into two halves and fixed in either: (i) 10% neutral buffered formalin or (ii) zinc-based fixative (Beckstead, 1994), the processed into paraffin wax.…”