High-efficiency transduction of human monocyte-derived dendritic cells by capsid-modified recombinant AAV2 vectors

George V. Aslanidi; Angela E. Rivers; Luis Ortiz; Lakshmanan Govindasamy; Chen Ling; Giridhara R. Jayandharan; Sergei Zolotukhin; Mavis Agbandje-McKenna; Arun Srivastava

doi:10.1016/j.vaccine.2012.03.079

High-efficiency transduction of human monocyte-derived dendritic cells by capsid-modified recombinant AAV2 vectors

George V. Aslanidi, Angela E. Rivers, Luis Ortiz, Lakshmanan Govindasamy, Chen Ling, Giridhara R. Jayandharan, Sergei Zolotukhin, Mavis Agbandje-McKenna, Arun Srivastava

Research output: Contribution to journal › Article › peer-review

39 Scopus citations

Abstract

Phosphorylation of surface-exposed tyrosine residues negatively impacts the transduction efficiency of recombinant AAV2 vectors. Pre-treatment of cells with specific cellular serine/threonine kinase inhibitors also significantly increased the transduction efficiency of AAV2 vectors. We reasoned that site-directed mutagenesis of surface-exposed serine residues might allow the vectors to evade phosphorylation and thus lead to higher transduction efficiency. Each of the 15 surface-exposed serine (S) residues was substituted with valine (V) residues, and the transduction efficiency of three of these mutants, S458V, S492V and S662V, was increased by up to ∼20-fold in different cell types. The S662V mutant was efficient in transducing human monocyte-derived dendritic cells (moDCs), a cell type not readily amenable to transduction by the conventional AAV vectors, and did not induce any phenotypic changes in these cells. Recombinant S662V-AAV2 vectors encoding a truncated human telomerase (hTERT) gene were generated and used to stimulate cytotoxic T cells (CTLs) against target cells. S662V-AAV2-hTERT vector-transduced DCs resulted in rapid, specific T-cell clone proliferation and generation of robust CTLs, which led to specific cell lysis of K562 cells. These studies suggest that high-efficiency transduction of moDCs by serine-modified AAV2 vectors is feasible, which supports the potential utility of these vectors for future human DCs vaccine studies.

Original language	English (US)
Pages (from-to)	3908-3917
Number of pages	10
Journal	Vaccine
Volume	30
Issue number	26
DOIs	https://doi.org/10.1016/j.vaccine.2012.03.079
State	Published - Jun 6 2012
Externally published	Yes

Bibliographical note

Funding Information:
We thank Drs. Kenneth I. Berns and Nicholas Muzyczka for a critical reading of this manuscript. We thank Robert Ng for assistance in the preparation of Fig. 3 . This research was supported in part by a Junior Investigator Award from the University of Florida Shands Cancer Center , American Cancer Society Chris DiMarco Institutional Research Grant (to GVA), and Public Health Service grants P01 DK-058327 – Project 1 (to AS), R01 HL-097088 (to AS and SZ), and R01 GM-082946 (to SZ and MA-M) from the National Institutes of Health. GRJ was supported in part by an ‘Overseas Associate Fellowship-2006’ from the Department of Biotechnology, Government of India.

Keywords

Adeno-associated virus vectors
Capsid proteins
Dendritic cells
Gene expression
Serine-phosphorylation
Serine/threonine kinase

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10.1016/j.vaccine.2012.03.079

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title = "High-efficiency transduction of human monocyte-derived dendritic cells by capsid-modified recombinant AAV2 vectors",

abstract = "Phosphorylation of surface-exposed tyrosine residues negatively impacts the transduction efficiency of recombinant AAV2 vectors. Pre-treatment of cells with specific cellular serine/threonine kinase inhibitors also significantly increased the transduction efficiency of AAV2 vectors. We reasoned that site-directed mutagenesis of surface-exposed serine residues might allow the vectors to evade phosphorylation and thus lead to higher transduction efficiency. Each of the 15 surface-exposed serine (S) residues was substituted with valine (V) residues, and the transduction efficiency of three of these mutants, S458V, S492V and S662V, was increased by up to ∼20-fold in different cell types. The S662V mutant was efficient in transducing human monocyte-derived dendritic cells (moDCs), a cell type not readily amenable to transduction by the conventional AAV vectors, and did not induce any phenotypic changes in these cells. Recombinant S662V-AAV2 vectors encoding a truncated human telomerase (hTERT) gene were generated and used to stimulate cytotoxic T cells (CTLs) against target cells. S662V-AAV2-hTERT vector-transduced DCs resulted in rapid, specific T-cell clone proliferation and generation of robust CTLs, which led to specific cell lysis of K562 cells. These studies suggest that high-efficiency transduction of moDCs by serine-modified AAV2 vectors is feasible, which supports the potential utility of these vectors for future human DCs vaccine studies.",

keywords = "Adeno-associated virus vectors, Capsid proteins, Dendritic cells, Gene expression, Serine-phosphorylation, Serine/threonine kinase",

author = "Aslanidi, {George V.} and Rivers, {Angela E.} and Luis Ortiz and Lakshmanan Govindasamy and Chen Ling and Jayandharan, {Giridhara R.} and Sergei Zolotukhin and Mavis Agbandje-McKenna and Arun Srivastava",

note = "Funding Information: We thank Drs. Kenneth I. Berns and Nicholas Muzyczka for a critical reading of this manuscript. We thank Robert Ng for assistance in the preparation of Fig. 3 . This research was supported in part by a Junior Investigator Award from the University of Florida Shands Cancer Center , American Cancer Society Chris DiMarco Institutional Research Grant (to GVA), and Public Health Service grants P01 DK-058327 – Project 1 (to AS), R01 HL-097088 (to AS and SZ), and R01 GM-082946 (to SZ and MA-M) from the National Institutes of Health. GRJ was supported in part by an {\textquoteleft}Overseas Associate Fellowship-2006{\textquoteright} from the Department of Biotechnology, Government of India.",

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doi = "10.1016/j.vaccine.2012.03.079",

language = "English (US)",

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T1 - High-efficiency transduction of human monocyte-derived dendritic cells by capsid-modified recombinant AAV2 vectors

AU - Aslanidi, George V.

AU - Rivers, Angela E.

AU - Ortiz, Luis

AU - Govindasamy, Lakshmanan

AU - Ling, Chen

AU - Jayandharan, Giridhara R.

AU - Zolotukhin, Sergei

AU - Agbandje-McKenna, Mavis

AU - Srivastava, Arun

N1 - Funding Information: We thank Drs. Kenneth I. Berns and Nicholas Muzyczka for a critical reading of this manuscript. We thank Robert Ng for assistance in the preparation of Fig. 3 . This research was supported in part by a Junior Investigator Award from the University of Florida Shands Cancer Center , American Cancer Society Chris DiMarco Institutional Research Grant (to GVA), and Public Health Service grants P01 DK-058327 – Project 1 (to AS), R01 HL-097088 (to AS and SZ), and R01 GM-082946 (to SZ and MA-M) from the National Institutes of Health. GRJ was supported in part by an ‘Overseas Associate Fellowship-2006’ from the Department of Biotechnology, Government of India.

PY - 2012/6/6

Y1 - 2012/6/6

N2 - Phosphorylation of surface-exposed tyrosine residues negatively impacts the transduction efficiency of recombinant AAV2 vectors. Pre-treatment of cells with specific cellular serine/threonine kinase inhibitors also significantly increased the transduction efficiency of AAV2 vectors. We reasoned that site-directed mutagenesis of surface-exposed serine residues might allow the vectors to evade phosphorylation and thus lead to higher transduction efficiency. Each of the 15 surface-exposed serine (S) residues was substituted with valine (V) residues, and the transduction efficiency of three of these mutants, S458V, S492V and S662V, was increased by up to ∼20-fold in different cell types. The S662V mutant was efficient in transducing human monocyte-derived dendritic cells (moDCs), a cell type not readily amenable to transduction by the conventional AAV vectors, and did not induce any phenotypic changes in these cells. Recombinant S662V-AAV2 vectors encoding a truncated human telomerase (hTERT) gene were generated and used to stimulate cytotoxic T cells (CTLs) against target cells. S662V-AAV2-hTERT vector-transduced DCs resulted in rapid, specific T-cell clone proliferation and generation of robust CTLs, which led to specific cell lysis of K562 cells. These studies suggest that high-efficiency transduction of moDCs by serine-modified AAV2 vectors is feasible, which supports the potential utility of these vectors for future human DCs vaccine studies.

AB - Phosphorylation of surface-exposed tyrosine residues negatively impacts the transduction efficiency of recombinant AAV2 vectors. Pre-treatment of cells with specific cellular serine/threonine kinase inhibitors also significantly increased the transduction efficiency of AAV2 vectors. We reasoned that site-directed mutagenesis of surface-exposed serine residues might allow the vectors to evade phosphorylation and thus lead to higher transduction efficiency. Each of the 15 surface-exposed serine (S) residues was substituted with valine (V) residues, and the transduction efficiency of three of these mutants, S458V, S492V and S662V, was increased by up to ∼20-fold in different cell types. The S662V mutant was efficient in transducing human monocyte-derived dendritic cells (moDCs), a cell type not readily amenable to transduction by the conventional AAV vectors, and did not induce any phenotypic changes in these cells. Recombinant S662V-AAV2 vectors encoding a truncated human telomerase (hTERT) gene were generated and used to stimulate cytotoxic T cells (CTLs) against target cells. S662V-AAV2-hTERT vector-transduced DCs resulted in rapid, specific T-cell clone proliferation and generation of robust CTLs, which led to specific cell lysis of K562 cells. These studies suggest that high-efficiency transduction of moDCs by serine-modified AAV2 vectors is feasible, which supports the potential utility of these vectors for future human DCs vaccine studies.

KW - Adeno-associated virus vectors

KW - Capsid proteins

KW - Dendritic cells

KW - Gene expression

KW - Serine-phosphorylation

KW - Serine/threonine kinase

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High-efficiency transduction of human monocyte-derived dendritic cells by capsid-modified recombinant AAV2 vectors

Abstract

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Keywords

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