Research Article: An Efficient Method for the Delivery of the Interleukin-2 Gene to Human Hematopoietic Cells using the 
Fiber-Modified Recombinant Adenovirus

Date Published: , 2011

Publisher: A.I. Gordeyev

Author(s): V.N. Rogozhin, D.Yu. Logunov, D.V. Shchebliakov, M.M. Shmarov, E.E. Khodunova, 
I.V. Galtseva, R.V. Belousova, B.S. Naroditsky, A.L. Gintsburg.

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Abstract

Recombinant human adenovirus serotype 5 (Ad5/35F-IL2) with
modified fibres containing the C-terminal domain fiber-knob of human adenovirus
serotype 35, carrying the gene of recombinant human IL-2, has been designed. As a
result of the fiber modification, the adenovirus can efficiently deliver the genetic
information to bone marrow leukocytes and the tumor blood cells KG-1A (human
myeloblastic leukemia cells) and U937 (human histiocytic lymphoma cells), which are
normally resistant to Ad5 infection. The flow cytometry data reveal that the
modified Ad5/35F penetrates into a population of monocytes, granulocytes, and blast
cells of human bone marrow. The expression of interleukin-2 in CAR-negative bone
marrow leukocytes (3682.52 ± 134.21 pg/ml) and the cell lines KG-1A (748.3 ± 32.8
pg/ml) and U937 (421.5 ± 59.4 pg/ml) transduced with adenovirus Ad5/35F-IL2 is
demonstrated. The fiber-modified adenovirus can be used as a vector for the
efficient gene delivery of interleukin-2 to human normal and tumor hematopoietic
cells.

Partial Text

Among the vectors most commonly used to deliver genes to human and mammal cells are
vectors based on the human adenovirus serotype 5 (Ad5). The advantages of these
vectors are numerous; they are capable of transducing both dividing and non-dividing
cells [1, 2]; the adenoviral DNA is not incorporated into the host cell genome and
retains its extrachromosomal form; adenoviruses can be produced at a titer of over
10 10 pfu/ml, which enables them to be used as living recombinant
vaccines; and they ensure a high expression level of the target gene in a target
cell. One of the drawbacks of Ad5-based vectors is their low transducing activity
with respect to CAR-deficient and CAR-negative cells. Among those, hematopoeitic
cells occupy a significant place. This problem derives from the fact that binding of
the adenovirus capsid protein (fiber) and the membrane cell receptor CAR
(coxsakievirus-adenovirus receptor) is necessary for the primary interaction between
Ad5 and a cell. Therefore, a deficiency or the total absence of these receptors on
the cell surface is a factor that limits efficient gene delivery using Ad-5 based
adenovectors.

Plasmid vectors

Obtaining fiber-modified human recombinant adenoviruses serotype
5

A specific modification consisting in the substitution of the C-terminal knob domain
of the Ad5 fiber for the analogous domain of the Ad35 fiber was used to obtain
recombinant Ad5 containing the modified fiber and carrying the human
IL2 gene. A manifold increase in efficiency in the penetration
of this vector into cells, as compared with the unmodified vector, was demonstrated
on KG-1A and U937 line cultures of human tumor blood cells. A 30-fold increase in
efficiency in the penetration of the modified vector in comparison with that for the
unmodified vector was first demonstrated in experiments on the transduction of the
primary leukocyte culture from RBM taken from a healthy donor, which ensures the
expression of the human interleukin-2 gene in them. Along with the efficient
transduction of RBM monocytes [16], the
modified Ad5/35F was first shown to efficiently transduce granulocytes and blast
cells of human RBM, while this vector does not penetrate into T- and B-lymphocyte
subpopulations.

 

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