Solid Tumor-Targeted Infiltrating Cytotoxic T Lymphocytes Retained by a Superantigen Fusion Protein

Successful immune-mediated regression of solid tumors is difficult because of the small number of cytotoxic T lymphocytes (CTLs) that were traffic to the tumor site. Here, the targeting of tumor-specific infiltrating CTLs was dependent on a fusion protein consisting of human epidermal growth factor (EGF) and staphylococcal enterotoxin A (SEA) with the D227A mutation. EGF-SEA strongly restrained the growth of murine solid sarcoma 180 (S180) tumors (control versus EGF-SEA, mean tumor weight: 1.013 versus 0.197 g, difference  = 0.816 g). In mice treated with EGF-SEA, CD4+, CD8+ and SEA-reactive T lymphocytes were enriched around the EGFR expressing tumor cells. The EGF receptors were potentially phosphorylated by EGF-SEA stimulation and the fusion protein promoted T cells to release the tumoricidal cytokines interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). Intratumoral CTLs secreted cytolytic pore-forming perforins and granzyme B proteins near the surface of carcinomas, causing the death of many tumor cells. We additionally show that labeled EGF-SEA was directly targeted to the tumor tissue after intravenous (i.v.) injection. The findings demonstrate that antibody-like EGF-SEA plays an important role in arresting CTLs in the solid tumor site and has therapeutic potential as a tumor-targeting agent

Differential modulation of the TRAIL receptors and the CD95 receptor in colon carcinoma cell lines

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) and CD95 ligand (CD95L) are potent inducers of apoptosis in various tumour cell types. Death receptors DR4 and DR5 can induce and decoy receptors DcR1 and DcR2 can inhibit TRAIL-mediated apoptosis. The study aim was to investigate whether anticancer agents can modulate similarly TRAIL-receptor and CD95 membrane expression and TRAIL and CD95L sensitivity.Three colon carcinoma cell lines (Caco-2, Colo320 and SW948) were treated with 5-fluorouracil (5-FU), cisplatin or interferon-γ. TRAIL-receptor and CD95 membrane expression was determined flow cytometrically. Sensitivity to TRAIL or CD95L agonistic anti-CD95 antibody was determined with cytotoxicity and apoptosis assays. SW948 showed highest TRAIL sensitivity. The protein synthesis inhibitor cycloheximide decreased FLICE-like inhibitory protein levels in all cell lines, and the TRAIL-resistant cell lines Caco-2 and Colo320 became sensitive for TRAIL. Exposure of the cell lines to 5-FU, cisplatin and interferon-γ left TRAIL-receptor membrane expression and TRAIL sensitivity unaffected. CD95 membrane expression and anti-CD95 sensitivity was, however, modulated by the same drugs in all lines. Cisplatin and interferon-γ raised CD95 membrane levels 6–8-fold, interferon-γ also increased anti-CD95 sensitivity. These results indicate that the CD95 and TRAIL pathways use different mechanisms to respond to various anticancer agents. Induced CD95 membrane upregulation was associated with increased anti-CD95 sensitivity, whereas no upregulation of TRAIL-receptor membrane expression or TRAIL sensitisation could be established. For optimal use of TRAIL-mediated apoptosis for cancer therapy in certain tumours, downregulation of intracellular inhibiting factors may be required