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Blood, Vol. 108, Issue 1, 319-327, July 1, 2006
BCR/ABL kinase induces self-mutagenesis via reactive oxygen species to encode imatinib resistance
Blood Koptyra et al.
108: 319
Supplemental materials for: Koptyra et al
Files in this Data Supplement:
- Figure S1. BCR/ABL expression and tyrosine kinase activity in cells used for the experiments (JPG, 51.1 KB)
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Parental 32Dcl3 cells and freshly transformed BCR/ABL counterparts were cultured continuously for 2 weeks (P-E and B/A-E, respectively) or 10 weeks (P-L and B/A-L, respectively) in the presence of IL-3. B/A-L cells were eventually pre-incubated for 48h with IM or PDTC. ABL proteins, phosphotyrosine proteins, and  -Actin were detected in total cell lysates by SDS-PAGE followed by Western analysis.
- Figure S2. ROS-induced, time-dependent accumulation of mutations causing ouabain resistance in BCR/ABL-transformed cells (JPG, 42.9 KB)
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Parental 32Dcl3 cells (P) and BCR/ABL-early cells were cultured in the absence (B/A) or presence of PDTC (B/A+PDTC). Cells with mutated Na+/K+ ATPase were detected in clonogenic assay using ouabain as a selective compound.
- Figure S3. Primary CML cells contain elevated levels of ROS (JPG, 56.2 KB)
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Mononuclear bone marrow cells were obtained from 4 healthy volunteers (N), 4 CML-CP, and 4 CML-BC patients. ROS levels were analyzed by fluorescence. Each diagram represents an individual sample.
- Figure S4. VE diet reduced ROS levels in BMC and SPL (JPG, 42.8 KB)
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SCID mice have been fed with control chow (transparent peaks) or VE diet (gray peaks) starting from one week before and continuously after i.v. injection of 5×104 freshly established 32Dcl3-BCR/ABL cells. ROS levels were measured 4 weeks after leukemia injection in the mononuclear fractions of BMC and SPL.
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