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Abstract Background Our previous studies have demonstrated that transduction of human dendritic cells (DC) with adenovirus encoding secondary lymphoid chemokine, CCL21, led to secretion of biologically active CCL21 without altering DC phenotype or viability. In addition, intratumoral injections of CCL21-transduced DC into established murine lung tumors resulted in complete regression and protective anti-tumor immunity. These results have provided the rationale to generate a clinical grade adenoviral vector encoding CCL-21 for ex vivo transduction of human DC in order to assess intratumoral administration in late stage human lung cancer. Methods In the current study, human monocyte-derived DC were differentiated by exposure to GM-CSF and IL-4 from cryopreserved mononuclear cells obtained from healthy volunteers. Transduction with clinical grade adenoviral vector encoding CCL21 (1167 viral particles per cell) resulted in secretion of CCL21 protein. Results CCL21 protein production from transduced DC was detected in supernatants (24–72 hours, 3.5–6.7 ng/4–5 × 106 cells). DC transduced with the clinical grade adenoviral vector were > 88% viable (n = 16), conserved their phenotype and maintained integral biological activities including dextran uptake, production of immunostimulatory cytokines/chemokines and antigen presentation. Furthermore, supernatant from CCL21-DC induced the chemotaxis of T2 cells in vitro. Conclusion Viable and biologically active clinical grade CCL21 gene-modified DC can be generated from cryopreserved PBMC.

Abstract Background Ex vivo generated dendritic cells (DC) genetically modified to express secondary lymphoid tissue chemokine (CCL-21/SLC) have been shown to stimulate potent antitumor responses in murine models. When injected intratumorally, CCL-21 colocalizes DC and lymphocyte effector cells at the tumor site. This may improve tumor antigen presentation and T cell activation by utilizing the tumor as an in vivo source of antigen for DC. In order to develop DC-based cancer therapies for intratumoral injection that could promote tumor antigen uptake and presentation in situ, we constructed and characterized an adenoviral vector that expresses human CCL-21 (AdCCL-21). Results Human monocyte derived DC were cultured in GM-CSF and IL-4 for 6 days. Following AdCCL-21 transduction, CCL-21 protein production was assessed by ELISA on day 8. DC transduced with AdCCL-21 at multiplicities of infection (MOIs) of 50:1 or 100:1 produced up to 210 ± 9 ng/ml and 278 ± 6.5 ng/ml /106 cells/48 hours, respectively. Following transduction, an immature DC phenotype was maintained and an upregulation of the costimulatory molecule, CD86 was noted. In addition, supernatant from AdCCL-21-DC caused significant chemotaxis of peripheral blood lymphocytes and mature DC. Conclusions These studies demonstrate that AdCCL-21-DC generate functional levels of CCL-21 without adversely altering DC phenotype. These findings strengthen the rationale for further investigation of AdCCL-21-DC as a DC-based therapy in cancer treatment.