Purging of contaminating breast cancer cells from hematopoietic stem cell grafts by adenoviral GAL-TEK gone therapy and magnetic antibody cell separation

The presence of contaminating tumor cells in auto]ogous bone marrow or peripheral blood stem cell (PB-SC) preparations increase the likelihood of relapse in women receiving transplants for metastatic breast cancer. We describe a new technique for purging breast cancer cells CBCCs) that combines two independent strategies: (a) the specific enrichment of CD34⁺ progenitor stem cells by magnetic antibody cell separation (MACS), and then (b) infection of the contaminating BCCs with a recombinant adGALTEK marker/suicide gene adenovirus (ad-v), followed by the addition of ganciclovir (GCV). Infection with this ad-v results in three to four times greater expression of ad-v-delivered reporter gene in BCCs than in CD34⁺ cells. In addition -2 h, -low multiplicity of infection (50:1) adGALTEK infections of BCC lines (MCF-7 and BT474) eradicated >99% of BCCs after 72 h of exposure to 20 μM GCV. However, exposure to both adenovirus and GCV at the MOIs and doses used had little effect on hematopoietic stem cells to form colonies in colony-forming unit assays. adGALTEK infection in our model system (10³-10⁵ BCCs added into 10⁷ HSCs) also resulted in the 3 to 5 log eradication of clonogenic BCCs after the addition of GCV. MACS enrichment/purification of CD34⁺ cells from PB-SC contaminated with 2 x 10⁶ to 5 x 10⁷ BCCs followed by adGAL-TEK infection and GCV addition resulted in 5-7-log depletion of clonogenic BCCs as well as enrichment of CD34⁺ progenitor cells to >98%, with the recovery of >70% of hematopoietic stem cells. This adenoviral purging system is so robust that poor MACS purification, resulting in 1.5-log depletion of BCCs, still permits excellent ad-v infection and BCC killing.