BackgroundThere is a continued need to develop more effective cancer immunotherapy strategies. Exosomes, cell-derived lipid vesicles that express high levels of a narrow spectrum of cell proteins represent a novel platform for delivering high levels of antigen in conjunction with costimulatory molecules. We performed this study to test the safety, feasibility and efficacy of autologous dendritic cell (DC)-derived exosomes (DEX) loaded with the MAGE tumor antigens in patients with non-small cell lung cancer (NSCLC).MethodsThis Phase I study enrolled HLA A2+ patients with pre-treated Stage IIIb (N = 4) and IV (N = 9) NSCLC with tumor expression of MAGE-A3 or A4. Patients underwent leukapheresis to generate DC from which DEX were produced and loaded with MAGE-A3, -A4, -A10, and MAGE-3DPO4 peptides. Patients received 4 doses of DEX at weekly intervals.ResultsThirteen patients were enrolled and 9 completed therapy. Three formulations of DEX were evaluated; all were well tolerated with only grade 1–2 adverse events related to the use of DEX (injection site reactions (N = 8), flu like illness (N = 1), and peripheral arm pain (N = 1)). The time from the first dose of DEX until disease progression was 30 to 429+ days. Three patients had disease progression before the first DEX dose. Survival of patients after the first DEX dose was 52–665+ days. DTH reactivity against MAGE peptides was detected in 3/9 patients. Immune responses were detected in patients as follows: MAGE-specific T cell responses in 1/3, increased NK lytic activity in 2/4.ConclusionProduction of the DEX vaccine was feasible and DEX therapy was well tolerated in patients with advanced NSCLC. Some patients experienced long term stability of disease and activation of immune effectors

Purpose:To determine the safety and immunologic and clinical efficacy of a dendritic cell vaccine modified to hyperexpress costimulatory molecules and tumor antigen. Experimental Design: In this phase I study, we administered one or two cycles of four triweekly s.c./intradermal injections of ex vivo generated dendritic cells modified with a recombinant fowlpox vector encoding carcinoembryonic antigen (CEA) and a triad of costimulatory molecules [rF-CEA(6D)-TRICOM]. Controls consisted of immature dendritic cells loaded with tetanus toxoid and a HLA A2^restricted peptide derived from cytomegalovirus pp65 protein.Results: Fourteen patients (11 with colorectal cancer and 3 with non^small cell lung cancer) were enrolled and 12 completed at least one cycle of immunization. There were no grade 3/4 toxicities directly referable to the immunizations. One patient had a decrease in the CEA level from 46 to 6.8 and a minor regression in adenopathy that occurred several months after completion of the immunizations. Five other patients were stable through at least one cycle of immunization (3 months). Direct analysis of peripheral blood mononuclear cells using the ELISpot assay showed an increase in the frequency of CEA-specificTcells in 10 patients (range, 10-541 CEA-specific cells/10 5 peripheral blood mononuclear cells). There was a trend for a greater peak frequency of CEA-specific T cells among those with either a minor response or a stable disease following at least one cycle of therapy. A second cycle was not associated with higher T-cell frequencies. Cytokine flow cytometry showed CEA-specific immune response among both CD4 + and CD8 + T cells in all immune responders. Conclusion: This immunization strategy is safe and activates potent CEA-specific immune responses.

A novel metallonucleoside phosphoramidite is reported for the automated solid-phase synthesis of site-specifically metallolabeled oligonucleotides.

A facile procedure for incorporating a Ru(diimine)(3)(2+) complex at the nucleobase in an oligonucleotide is reported that combines the advantages of Pd(0) cross-coupling and solid-phase DNA chemistries. These ruthenium-modified oligonucleotides form stable duplexes, and the favorable photophysical properties associated with the Ru(diimine)(3)(2+) complex are retained after site-specific covalent attachment.

A facile procedure for the incorporation of Ru(bpy)3 2+ in an oligonucleotide is reported. A Ru(bpy)3 2+ phosphoramidite is synthesized, and then attached to the 5‘-terminus of DNA using a standard protocol on an automated DNA solid-phase synthesizer. Photophysical studies of the Ru(II) tris-diimine complex as well as the corresponding labeled oligonucleotides demonstrate that the excited-state electron is localized on one specific bipyridine with the dipole directed toward the linkage to DNA, and that the Ru(II) excited state is long-lived when attached to the DNA.

The synthesis and photophysical properties of a novel ruthenium-modified nucleoside are reported. The key synthetic step to 5-[Ru(bpy)2(4-m-4‘-pa-bpy)]2+-2‘-deoxyuridine involves the Pd(0)-catalyzed cross-coupling of a propargylamine-derivatized Ru(bpy)3 2+ and 3‘,5‘-dibenzoyloxy-2‘-deoxy-5-iodouridine. The long-lived 3MLCT excited state (1300 ns) of 5-[Ru(bpy)2(4-m-4‘-pa-bpy)]2+-2‘-deoxyuridine has an emission maximum centered at 640 nm. Step-scan Fourier transform infrared (S2FTIR) time-resolved spectroscopy reveals the excited-state electron to be localized on the modified bipyridine with the excited-state dipole oriented toward the 2‘-deoxyuridine.

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