Activity of the vascular-disrupting agent 5,6-dimethylxanthenone-4-acetic acid against human head and neck carcinoma xenografts

Abstract

Head and neck squamous cell carcinomas (HNSCC) constitute a majority of the tumors of the upper aerodigestive tract and continue to present a significant therapeutic challenge. To explore the potential of vascular-targeted therapy in HNSCC, we investigated the antivascular, antitumor activity of the potent vascular-disrupting agent (VDA) 5,6-dimethylxanthenone-4-acetic acid (DMXAA) against two HNSCC xenografts with markedly different morphologic and vascular characteristics. Athymic nude mice bearing subcutaneous FaDu (human pharyngeal squamous cell carcinoma) and A253 (human submaxillary gland epidermoid carcinoma) tumors were administered a single dose of DMXAA (30 mg/kg, i.p). Changes in vascular function were evaluated 24 hours after treatment using contrast-enhanced magnetic resonance imaging (MRI) and immunohistochemistry (CD31). Signal enhancement (E) and change in longitudinal relaxation rates (deltaR1) were calculated to measure alterations in vascular perfusion. MRI showed a 78% and 49% reduction in vascular perfusion in FaDu and A253 xenografts, respectively. CD31-immunostaining of tumor sections revealed three-fold (FaDu) and two-fold (A253) reductions in microvessel density (MVD) 24 hours after treatment. DMXAA was equally effective against both xenografts, with significant tumor growth inhibition observed 30 days after treatment. These results indicate that DMXAA may be clinically beneficial in the management of head and neck cancers, alone or in combination with other treatments.

Figure 1

Vascular differences between FaDu and A253 xenografts. Graph shows percent enhancement (E) in MR signal intensity following contrast agent administration in FaDu and A253 human HNSCC implanted subcutaneously in nude mice (two-tailed Student's t test, *P < .05).

Figure 2

Vascular response of tumor and kidney tissues to DMXAA. Change in T₁ relaxation rates (ΔR₁) of control and DMXAA-treated FaDu and A253 tumors calculated from serial T₁-weighted MR images acquired before and 24 hours after administration of albumin-GdDTPA. The ΔR₁ values of kidneys before and 24 hours after DMXAA treatment are included. Values represent mean ± SEM (two-tailed Student's t test, ***P < .001, **P < .01, *P < .05).

Figure 3

Change in vascular volume and permeability following DMXAA. Graph shows change in T₁ relaxation rates (ΔR₁) over time of untreated control tumors (squares) and tumors treated with 30 mg/kg DMXAA (circles) for FaDu (left panel) and A253 (right panel) xenografts. Vascular volume and permeability values were calculated from ΔR₁ using linear regression analysis. Significant differences were seen between the vascular volumes (Y-intercepts) of control FaDu and control A253 xenografts (P < . 0001). Twenty-four hours after treatment, only FaDu tumors exhibited a significant reduction in vascular volume versus control (P < .0001). Analysis of the slopes of the plots also revealed a significant difference in permeability between control and DMXAA-treated FaDu tumors (P < .001).

Figure 4

Effect of DMXAA therapy on HNSCC xenografts. Photomicrographs of control and DMXAA-treated FaDu (upper two rows) and A253 (lower two rows) xenografts are shown before and 24 hours after DMXAA treatment. The left column shows H&E- stained tumor sections (original magnification, x200), and the right column shows CD31-immunostained tumor sections (original magnification, x400). Control FaDu xenografts consist of uniformly poorly differentiated regions (panel A) with increased MVD (panel B; arrows), whereas A253 tumors consist of hypoxic, avascular, well-differentiated islands (panel E) with fewer vessels (panel F; arrows). Twenty-four hours after DMXAA treatment, both FaDu (panel C) and A253 (panel G) tumors showed extensive necrosis and loss of CD31 staining (FaDu, panel D; A253, panel H) indicative of significant DMXAA-induced vascular damage (arrows).

Figure 5

Estimates of MVD in FaDu and A253 xenografts following DMXAA treatment. Bar graphs show MVD counts for control and DMXAA-treated FaDu and A253 tumors per HPF (original magnification, x400). Significant reduction in MVD was seen 24 hours after DMXAA treatment (two-tailed Student's t test, **P < .01, *P < .05).

Figure 6

Visualization of FaDu and A253 vascular response to DMXAA. T₁ relaxation maps (lower panel) of a nude mouse bearing bilateral FaDu (yellow arrows) and A253 (white arrows) xenografts. Maps (B) and (D) represent the precontrast and postcontrast images acquired before DMXAA treatment. Maps (F) and (H) represent the precontrast and postcontrast images acquired 24 hours after DMXAA treatment. Twenty-four hours after DMXAA treatment, no detectable MR signal enhancement was seen in FaDu tumors after contrast agent administration (map H) compared to precontrast images (map F). At the same time point, A253 showed enhancement, indicating the presence of functional vessels (maps F and H). Representative proton images are also shown (images A, C, E, and G).

Figure 7

Tumor growth inhibition following DMXAA. Nude mice bearing bilateral FaDu and A253 xenografts were injected with 30 mg/kg DMXAA, and tumor growth was monitored for a period of 30 days. Figure shows change in median tumor volume between DMXAA-treated tumors and untreated controls. DMXAA resulted in significant inhibition (P < .001) of the growth of both xenografts compared to untreated controls.