Summary The tumour vasculature is an attractive target for therapy. Combretastatin A‐4 (CA‐4) and A‐1 (CA‐1) are tubulin binding agents, structurally related to colchicine, which induce vascular‐mediated tumour necrosis in animal models. CA‐1 and CA‐4 were isolated from the African bush willow, Combretum caffrum, and several synthetic analogues are also now available, such as the Aventis Pharma compound, AVE8062. More soluble, phosphated, forms of CA‐4 (CA‐4‐P) and CA‐1 (CA‐1‐P) are commonly used for in vitro and in vivo studies. These are cleaved to the natural forms by endogenous phosphatases and are taken up into cells. The lead compound, CA‐4‐P, is currently in clinical trial as a tumour vascular targeting agent. In animal models, CA‐4‐P causes a prolonged and extensive shut‐down of blood flow in established tumour blood vessels, with much less effect in normal tissues. This paper reviews the current understanding of the mechanism of action of the combretastatins and their therapeutic potential.

The vascular system plays a role of key importance during tumour growth and metastasis formation. In addition, the effectiveness of almost all therapeutic modalities, including drug therapy and radiotherapy, is influenced by the micro-architecture and the gradients of essential nutrients around each vessel. This underlines the importance of the vascular architecture, its origin and effectiveness as a nutrient delivery system. The knowledge that tumour vasculature is abnormal has led to concepts such as angiogenic attack and vascular targeting (Folkman, 1976;Denekamp, 1984;Bicknell, 1994;Folkman and D'Amore, 1996). This, in turn, has led to many more studies of tumour vasculature and primary as well as secondary angiogenesis. However, even though our knowledge of the mechanisms underlying angiogenesis has increased dramatically in the past 25 years, few quantitative data are available on the vascular network architecture and pattern formation in tumours.The fact that the tumour vascularity differs in many aspects from the vasculature of normal organs and tissues was already recognized in the last century (Virchow, 1863; Thiersch, 1865). Thomlinson and Gray (1955) indicated the importance of intervessel spacing because of the threefold increase in radioresistance that accompanies reduction of pO 2 concentrations below critical level. The important question as to whether the vascular architecture of an individual tumour is tumour type-specific has been controversial (Warren, 1979;Vaupel and Gabbert, 1986.) This is due, at least in part, to the methodologies used. Most reports confine themselves to qualitative observations and comparisons of gross vascular patterns in host and tumour, or to blood vessel density, length and diameter measurements, which in turn vary with the staining and counting techniques (Davidson et al, 1994;Endrich and Vaupel, 1998). Recently, morphometric analyses have been introduced in which the features of tumour cells (proliferation rate, oxygenation, angiogenic growth factor production) have been mapped by sequential staining of the same section, allowing the influence of the vascular assay on clinically relevant aspects of cell populations to be mapped. Vascular and metabolic profiles (VAMP) have illustrated marked differences between different types of tumour of the same general histology, for brain tumours and those from the head and neck region. Less et al (1991) introduced the first suitable approach for determining branching patterns and vessel dimensions in corrosion casts of mammary carcinomas. They established a quantitative classification scheme which takes account of the unique features of tumour microvascular network topology. However, it should be recognized that measurements were made on planar twodimensional (2D) projected images of three-dimensional (3D) specimens. All attempts to determine distances in corrosion casts geometrically after 2D projection inevitably include a considerable error. Measurements of vascular parameters such as intercapillary distance and vessel se...

SummaryThe role of nitric oxide (NO) in the response to Photofrin-based photodynamic therapy (PDT) was investigated using mouse tumour models characterized by either relatively high or low endogenous NO production (RIF and SCCVII vs EMT6 and FsaR, respectively).-nitro-L-arginine methyl ester (L-NAME), administered to mice immediately after PDT light treatment of subcutaneously growing tumours, markedly enhanced the cure rate of RIF and SCCVII models, but produced no obvious benefit with the EMT6 and FsaR models. Laser Doppler flowmetry measurement revealed that both L-NNA and L-NAME strongly inhibit blood flow in RIF and SCCVII tumours, but not in EMT6 and FsaR tumours. When injected intravenously immediately after PDT light treatment, L-NAME dramatically augmented the decrease in blood flow in SCCVII tumours induced by PDT. The pattern of blood flow alterations in tumours following PDT indicates that, even with curative doses, regular circulation may be restored in some vessels after episodes of partial or complete obstruction. Such conditions are conducive to the induction of ischaemia-reperfusion injury, which is instigated by the formation of superoxide radical. The administration of superoxide dismutase immediately after PDT resulted in a decrease in tumour cure rates, thus confirming the involvement of superoxide in the anti-tumour effect. The results of this study demonstrate that NO participates in the events associated with PDT-mediated tumour destruction, particularly in the vascular response that is of critical importance for the curative outcome of this therapy. The level of endogenous production of NO in tumours appears to be one of the determinants of sensitivity to PDT.

No abstract

Neutrophils have become recognised as important contributors to the effectiveness of tumour eradication by photodynamic therapy (PDT). In this study, we have used the mouse SCCVII squamous cell carcinoma model to investigate the activity of neutrophils in tumours treated by PDT. Tumour levels of neutrophilic myeloperoxidase (MPO) demonstrated not only a massive and sustained sequestration of these cells in PDT-treated tumours but also revealed their activated state evidenced by the presence of released MPO. Among the adhesion molecules expressed on tumour vascular endothelium, ICAM-1 appears to be of primary importance in the invasion of neutrophils into PDT-treated tumours, because its functional blocking with monoclonal antibodies reduced the tumour cure rate. A marked upregulation of its ligands CD11b/CD18 and CD11c/CD18 found on neutrophils associated with PDT-treated tumours supports this assumption. To evaluate the role of inflammatory cytokines regulating neutrophil activity, neutralising antibodies were given to mice before PDT treatment. The results suggest that IL-1beta activity is critical for the therapeutic outcome, since its neutralisation diminished the cure rates of PDT-treated tumours. No significant effect was observed with anti-IL-6 and anti-TNF-alpha treatment. Further flow cytometry-based examination of neutrophils round in PDT-treated tumours revealed that these cells express MHC class II molecules, which suggests their engagement as antigen-presenting cells and involvement in the development of antitumour immune response.