Figure 4.

A Fas-IFP enhances antitumor efficacy in vivo (A) Overall survival of ID8_VEGF tumor-bearing mice treated with repeated doses of P14 Thy1.1⁺ CD8 T cells transduced with TCR₁₀₄₅ or TCR₁₀₄₅ and Fas-4-1BB_tm IFP. Tumor-bearing mice received a single dose of cyclophosphamide (180 mg/kg) before the first T cell infusion followed by injections of 1×10⁷ TCR₁₀₄₅ or TCR₁₀₄₅/Fas-4-1BB_tm T cells and peptide-pulsed irradiated splenocytes (5:1 APC:T cell ratio) i.p. every 14 days with 1×10⁴ U IL-2 s.c. for 10 days after each infusion. Treatment was initiated 45–52 days after tumor injection, when tumors were detectable by US. Arrows above survival curve indicate the timing of T cell infusions. Survival data are aggregated from three independent experiments, n=9–13 total per group. Log-rank (Mantel-Cox) test. (B) IHC quantification of CD8 +T cells in lungs of untreated (white), TCR₁₀₄₅ (gray) or TCR₁₀₄₅/Fas-4-1BB_tm IFP (red) treated mice at necropsy. (C) Histology scoring of hematoxylin and eosin IHC stained lungs from untreated (white), TCR₁₀₄₅ (gray) or TCR₁₀₄₅/Fas-4-1BB_tm IFP (red) treated mice at necropsy. All lungs contained metastatic tumor. Scoring ranged from 0 (tissue within normal limits), to 4 (neutrophilic interstitial pneumonia with or without lymphoplasmacytic and neutrophilic perivasculitis) and was performed by a trained pathologist. (B, C) Cumulative data from three independent experiments, n=9–10 per group, except the untreated mice in (C), for which only 2 mice had metastatic tumors in the lungs. One-way ANOVA with multiple comparisons. All error bars represent SD (D)Immunohistochemistry (IHC) staining for MSLN in tumors of untreated, TCR₁₀₄₅ or TCR₁₀₄₅/Fas-4-1BB_tm IFP treated mice at necropsy. Scale bar=100 µm. Images are representative of 9–10 mice per group from three independent experiments. ANOVA, analysis of variance; IFP, immunomodulatory fusion protein; MSLN, Mesothelin; n.s, not significant.