High incidence of leukemia in large animals after stem cell gene therapy with a HOXB4-expressing retroviral vector
J. Clin. Invest. Xiao-Bing Zhang, et al. 118:1502 doi:10.1172/JCI34371 [Go to this article.]

Figure 2
Retroviral integration induced mutagenesis is associated with leukemogenesis. (A) Southern blot analyses of marrow samples from G374, G450, and K00339 demonstrate monoclonality. Marrow DNA was digested with BglII, which cuts the transgene once, releasing a unique band for each integrant. Digestion with SacI, which cuts the transgene twice, showed a 3.9-kb band for all the integrants. (B–D) Integration sites were determined by LAM-PCR, and the schematic representations of integration sites for G374 (B), G450 (C), and K00339 (D) are shown. Exons are represented by black boxes. MSCV indicates the integration site, and the arrow indicates the orientation. ATG denotes the translation start site. Note that exons and introns are not to scale. (E–G) Dysregulated expression of genes in close vicinity of the integration sites for G374 (E), G450 (F), and K00339 (G). SYBR Green real-time RT-PCR was performed to determine the expression levels of dog MYB (E), dog ZFP36L2 and thyroid adenoma associated (THADA) (F), and macaque PRDM16, SSBP2, and SOCS1 (G). Canine PRDM16 (E) and macaque MAGOH2 (G) expression was undetectable in samples from normal control animals. (G) Expression of NSMCE1 was undetectable in K00339 and control animals. Marrow mRNA samples from normal animals were used as controls. M, DNA ladder.