Pre-clinical characterization of GMP grade CCL21-gene modified dendritic cells for application in a phase I trial in non-small cell lung cancer.
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 x 10(6) 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.


CONCLUSIONS
Viable and biologically active clinical grade CCL21 gene-modified DC can be generated from cryopreserved PBMC.
Baratelli
Felicita
F
UCLA Lung Cancer Research Program of the Jonsson Comprehensive Cancer Center, Division of Pulmonary and Critical Care Medicine, Department of Medicine, Los Angeles, CA 90095, USA. fbaratelli@mednet.ucla.edu
Takedatsu
Hiroko
H
Hazra
Saswati
S
Peebles
Katherine
K
Luo
Jie
J
Kurimoto
Pam S
PS
Zeng
Gang
G
Batra
Raj K
RK
Sharma
Sherven
S
Dubinett
Steven M
SM
Lee
Jay M
JM
D000256:Adenoviridae / Q000235:genetics; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000276:immunology / Q000628:therapy*; D002469:Cell Separation; D002478:Cells, Cultured; D054421:Chemokine CCL21 / Q000235:genetics* / Q000592:standards; D019742:Chemokines, CC / Q000096:biosynthesis; D002634:Chemotaxis, Leukocyte / Q000502:physiology; D017321:Clinical Trials, Phase I as Topic; D015925:Cryopreservation; D016207:Cytokines / Q000096:biosynthesis; D003713:Dendritic Cells / Q000276:immunology*; D004353:Drug Evaluation, Preclinical; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D016130:Immunophenotyping / Q000379:methods; D007963:Leukocytes, Mononuclear / Q000166:cytology; D008175:Lung Neoplasms / Q000235:genetics / Q000276:immunology / Q000628:therapy*; D013997:Time Factors; D014161:Transduction, Genetic
C515478:CCL21 protein, human; D054421:Chemokine CCL21; D019742:Chemokines, CC; D016207:Cytokines