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Purging of Contaminating Breast Cancer Cells from Hematopoietic Stem Cell Grafts by Adenoviral GAL-TEK Gene Therapy and Magnetic Antibody Cell Separation¹

Department of Molecular Hematology and Therapy [F. C. M., V. S., Q. Y., X. Z., M. A.], Department of Surgical Oncology, Breast Section [S. E. S.], and Department of Bone Marrow Transplant [R. C.], University of Texas, M. D. Anderson Cancer Center, Houston, Texas 77030

The presence of contaminating tumor cells in autologous 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 (BCCs) 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 adGAL-TEK 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) adGAL-TEK 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. adGAL-TEK 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.

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