Efficacy of an osmotic pump delivered, GM-CSF-based tumor vaccine in the treatment of upper aerodigestive squamous cell carcinoma in rats

Abstract

Purpose: Upper aerodigestive tract (UADT) cancer has not experienced significant overall survival improvement for over 20 years, and no successful treatments for systemic disease exist. Most patients with UADT cancer experience immune suppression, therefore immune restorative therapies may offer promise for these patients. We presently tested the efficacy of granulocyte macrophage-colony stimulating factor (GM-CSF) delivered via 28-day continuous infusion pump, in combination with irradiated tumor cells, in a flank model of UADT cancer.

Methods: Five groups of rats were inoculated with syngeneic mucosally derived squamous carcinoma cells (FAT-7). Osmotic minipumps were implanted in the contralateral flank to deliver GM-CSF at 0 (PBS), 0.1, 1, 10, or 100 ng/day (n = 6 per group) for 28 days; 10(6) irradiated FAT-7 cells (ITC) were injected at the site of the GM-CSF infusion on days 0, 3, 7, 14, and 21 immune infiltrates in tumors were analyzed.

Results: Rats that received 10 or 100 ng/day GM-CSF/ITC had a significantly slower tumor growth rate compared to those who received 0, 0.1, or 1 ng/day (ANOVA, P < 0.01). There were increased CD 4+, CD 8+, and CD 68+ cells in tumors of GM-CSF/ITC treated animals over controls.

Conclusion: GM-CSF (10 or 100 ng/day) delivered locally via osmotic pump with ITC slows the growth rate of mucosally derived squamous cell carcinoma in rats while improving immune cell infiltrates. The efficacy of locally delivered GM-CSF immunotherapy in this model may be a first step toward this immunotherapy strategy for humans.

Fig. 1

Tumor growth inhibition with GM-CSF/ITC. Six rats per group were implanted with osmotic minipumps delivering GM-CSF at the above doses. Irradiated FAT-7 cells were injected on days 0, 3, 7, 14, and 21 (a). Significance of differences in tumor growth rate between groups depicted in AUC analysis described in Sect. ”Materials and methods” (b)

Fig. 2

Tumor AUC analysis for control animals versus animals treated with GM/ITC (GM), GM/ITC + IL-12 (GM + IL-12), and GM/ITC + indomethacin (GM + Indo)

Fig. 3

Immune histochemistry staining of infiltrating inflammatory cells in rat Fat-7 flank tumors in control tumors versus those treated with GM-CSF + ITC. a–c CD 1a staining: a control lymph node, b untreated tumor, c 100 ng/day GM-CSF + ITC; d–f CD 4 staining: d control lymph node, e untreated tumor, f 100 ng/day GM-CSF; g–i CD 8 staining: g control lymph node, h untreated tumor, i 100 ng/day GM-CSF; j–l CD 68 staining: j control lymph node, k untreated tumor, l 100 ng/day GM-CSF. Note the diffuse intratumoral staining for all markers shown in tumors treated with regional GM-CSF. In b and e peritumoral infiltrate is present but no intratumoral cells are visualized. Note (for + control panels a, d, g, and j) pan-staining of entire field with CD 1a (panel a), principal staining outside of germinal centers for CD 4+, CD 8+, and CD 68+ cells (panels d, g, and j). All images ×100

Fig. 4

Quantitation of high power field immune cell infiltrate counts in control versus GM + ITC treated animals. There was variability in the cells per randomly quantified high powered field secondary to the focal nature of some of the infiltrates. Box–whisker plots for each for each of the cells stained are depicted (CD 1a, P = 0.0005; CD 4, P = 0.0005; CD 8, P = 0.1655; CD 68, P = 0.0158)