Targeted retroviral transduction of c-kit+ hematopoietic cells using novel ligand display technology

Abstract

Gene therapy for a wide variety of disorders would be greatly enhanced by the development of vectors that could be targeted for gene delivery to specific populations of cells. We describe here high-efficiency targeted transduction based on a novel targeting strategy that exploits the ability of retroviruses to incorporate host cell proteins into the surface of the viral particle as they bud through the plasma membrane. Ecotropic retroviral particles produced in cells engineered to express the membrane-bound form of stem cell factor ( IntroductionThe development of retroviral vectors that are targeted to transduce only specific cell types has been the goal of many investigators working in the field of gene therapy. [1][2][3][4][5] The availability of such vectors would revolutionize the field, opening up new avenues for gene therapy in acquired and inherited disorders by enabling in vivo delivery of therapeutic molecules to specific populations of target cells. Novel approaches to drug delivery and for use in vaccines also become possible through display of molecules on the retroviral surface. Over a period of around a decade, however, this goal has proven highly elusive. Virtually all attempts to target specific cell types have logically focused on modification of the retroviral envelope protein, as this complex glycoprotein is established to determine viral host range and facilitate virus entry though membrane fusion. 2,4,5 These modifications can be the simple substitution of the envelope protein for one from another retrovirus or other enveloped virus, such as vesicular stomatitis virus (VSV) 6 , a process referred to as pseudotyping. 2 More recently, attempts have been made to deliberately engineer envelope proteins to redirect their binding to new cell surface molecules. 4 This has involved making N-terminal extensions, insertions, or replacements to regions of the envelope protein. Commonly used modifications have been incorporation of growth factor-binding regions [7][8][9][10][11][12] or the addition of single-chain antibodies. [13][14][15][16][17] While many of these modifications have successfully redirected viral binding, this has invariably been achieved at the expense of infectivity to the extent that targeted binding actually inhibited viral entry. This property has actually been used to advantage in an approach referred to as "inverse targeting." 10,18 A number of reports, however, have indicated that low levels of targeted transduction may be achievable by coexpression of an ecotropic envelope in conjunction with the modified envelope protein. 7,9,16,19 To circumvent the problems associated with envelope modification, we have developed an alternative strategy, exploiting the natural budding mechanism of the virus to insert new binding ligands into the viral surface. We describe here successful and efficient specific targeting of transduction to human cells expressing stem cell factor (SCF) receptor (c-kit) by ecotropic retroviruses bearing surface stem cell factor. Materials and methods Cytokine...

Lentiviral vectors for cancer immunotherapy: transforming infectious particles into therapeutics

“…If a specific receptor is overexpressed on the cell membrane of a producer cell line, then this will also be displayed on the viral envelope. When overexpressing stem cell factor (SCF) on the cell membrane of an ecotropic producer cell line, Chandrashekran et al 135,136 showed that retrovirus thus produced was able to preferentially transduce c-kit expressing human stem cells. Recently, Yang et al 137 demonstrated that this approach can also be applied for lentiviruses.…”

Conjugation of Lentivirus to Paramagnetic Particles via Nonviral Proteins Allows Efficient Concentration and Infection of Primary Acute Myeloid Leukemia Cells

“…Moreover, vectors from different sources (2) and with alternative or reengineered targeting envelopes (3,21,33,40) may be particularly sensitive to centrifugation (35). Thus, magnetic concentration not only is a useful purification technology but also allows the use of additional factors for capture and/or targeting strategies that are not dependent on the modification of viral envelope proteins (7,8).…”

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“…To overcome these limitations, retroviral vectors have been extensively modified to generate self-inactivating vectors [70]. Further, packaging cell lines have been modified to attenu-future science group Modification of viral vectors by nanomaterials Review ate vector inactivation by serum complement [71] or to endow target-specificity to RVs [72], thus leading to enhanced safety and therapeutic profiles.…”