Detection of Enzymatic Activity of Free Circulating Tyrosine Kinase in Patients with Chronic Myelogenous Leukemia (CML) and the Demonstration of Lower Levels in Resistant Patients.

Abstract 4247

Tyrosine kinases (TKs) regulate a wide variety of cellular signaling events, and aberrant activation of TKs is directly associated with tumorigenesis and cancer progression. Targeted therapy using TK inhibitors (TKIs) is commonly explored for treating various types of cancers. Measuring TK activities for diagnosis and monitoring of therapeutic efficacy may be particularly important for patients treated with TKIs. In this study, we examined the clinical utility of measurement of free circulating TK (cTK) activity directly from plasma in CML patients. The study included 46 patients with newly-diagnosed CML, 24 drug (imatinib, dasatinib, or nilotinib)-resistant CML patients, and 38 apparently healthy control subjects. Plasma levels of cTK activity were measured using a chromogenic universal TK assay kit in which samples and standards were added to a 96-well microtiter plate immobilized with synthesized poly (Glu-Tyr) peptide substrates. Specific TK activity was determined by spectrophotometry. Measured cTK activity at baseline was ∼1000-fold higher in plasma than in paired serum specimens. Overall, plasma cTK activity was significantly higher in newly diagnosed CML patients (median=802 U/mL, range=18-3932 U/mL) than in healthy donors (median=83 U/mL, range=0.6-853 U/mL) or drug-resistant CML patients (median=202 U/mL, range=17-1215 U/mL) (P<0.001). In drug–resistant CML patients, plasma cTK activity was closely correlated with cellular BCR-ABL1 kinase activation, as indicated by downstream phospho-CRKL (P=0.0005) and phospho-STAT-5 (P=0.003) levels. TKI treatment of these patients also resulted in a corresponding decrease in cTK activity over time reflecting response to therapy. Nevertheless, cTK activity was not associated with BCR-ABL1 expression level and was independent of BCR-ABL1 mutation type. In conclusion, measuring circulating TK activity in plasma/serum may reflect the leukemic cells in CML patients and can be used to evaluate response to therapy. In addition, our plasma results support previous data showing that resistant cells in CML patients demonstrate lower TK activity as compared with naïve leukemic cells.