Efficient gene transfer into human CD34(+) cells by a retargeted adenovirus vector

Area of Science:

• Gene Therapy
• Virology
• Hematopoietic Stem Cell Biology

Background:

• Adenovirus serotype 5 (Ad5) vectors require coxsackievirus-adenovirus receptors (CAR) and alpha(v) integrins for efficient gene transfer.
• Limited expression of these receptors on hematopoietic stem cells restricts Ad5-mediated gene transfer.
• Adenovirus serotype 35 (Ad35) shows higher tropism for CD34(+) cells.

Purpose of the Study:

• To screen adenovirus serotypes for efficient interaction with human CD34(+) cells.
• To develop a chimeric adenovirus vector with enhanced tropism for hematopoietic stem cells.
• To evaluate the gene transfer efficiency of the chimeric vector compared to standard Ad5 vectors.

Main Methods:

• Screening of different Ad serotypes for attachment, internalization, and replication in CD34(+) and K562 cells.
• Generation of a chimeric Ad5 vector (Ad5GFP/F35) with an Ad35 fiber.
• Transduction of CD34(+) cells with Ad5GFP/F35 and Ad5GFP vectors, followed by assessment of GFP expression and viral genome presence.

Main Results:

• Adenovirus serotype 35 demonstrated the highest tropism for CD34(+) cells.
• The chimeric Ad5GFP/F35 vector efficiently transduced CD34(+) cells via a CAR- and alpha(v) integrin-independent pathway.
• Ad5GFP/F35 achieved 54% GFP expression in CD34(+) cells, compared to 25% for Ad5GFP.
• Transduction efficiency was higher than reported due to unactivated CMV promoter in some cells.
• Ad5GFP/F35 transduced a broader spectrum of CD34(+) cells, including CD34(+) c-Kit(+) cells (55% vs. 13%).

Conclusions:

• A chimeric Ad5 vector incorporating the Ad35 fiber significantly enhances gene transfer into hematopoietic stem cells.
• This retargeted vector overcomes limitations associated with CAR and alpha(v) integrin dependency.
• The findings support the potential of Ad5GFP/F35 for stable gene transfer into hematopoietic stem cells.