Page 139 - 2018_12-Haematologica-web

CDK9 as a potential molecular target in NK cell leukemia/lymphoma
same manner, were 17.0, 26.0, 26.0, and 16.8%; and 0.2, 0.3, 0.3, 0.2, and 0.2%, respectively. Thus, BAY1143572 treatment resulted in lowered percentages of ANKL cells infiltrating into the bone marrow (P=0.009; Figure 5B).
BAY 1143572 treatment reduces primary ANKL cells in the liver
The percentage of leukemia cells infiltrating into the liver of control NOG mouse No.1 was 32.4% (i.e., 36.9% [CD45-positive lymphocyte population] x 87.8% [CD16/56-positive, CD3-negative] = 32.4%) (Figure 5C, the two upper left panels), 22 days after tumor inocula- tion. In control NOG mice Nos. 2, 3, 4, and 5, and in BAY 1143572 treated NOG mice Nos. 1, 2, 3, 4, and 5, these fig- ures were 27.6, 29.7, 15.8, and 21.4%; and 0.3, 0.6, 1.8, 0.5, and 0.1%, respectively. Thus, the percentage of pri- mary ANKL cells present in the liver was also decreased by BAY1143572 treatment in these animals (P=0.009; Figure 5C).
BAY 1143572 treatment reduces primary ANKL cells in the spleen
Using the same experimental protocol as described above, 22 days after inoculation of primary ANKL tumor cells, the percentage of leukemia cells in the spleen of con- trol NOG mouse No.1 was 11.7% (Figure 5D, the upper
left two panels) whereas in control NOG mice Nos. 2, 3, 4, and 5, and in BAY1143572-treated NOG mice Nos.1, 2, 3, 4, and 5, these percentages were 17.6, 21.0, 21.1, and 19.3%; and 0.3, 0.5, 1.0, 0.9, and 2.5%, respectively. Thus, BAY1143572 treatment also decreases primary ANKL cell infiltration into the spleens of these animals (P=0.009; Figure 5D).
BAY1143572 prolongs the survival of mice inoculated with primary ANKL cells
Two of the 6 BAY 1143572-treated mice survived up to 34 days after inoculation of the primary ANKL tumor cells, whereas none of the vehicle-treated control mice were alive at that time (P=0.020; Figure 6). No toxicity attributable to BAY 1143572 was observed over this time period in any of the mice. Thus, we conclude that BAY 1143572-treatment of mice inoculated with primary ANKL cells significantly extended their survival relative to the untreated controls.
Discussion
NK-cell leukemia/lymphoma have a very poor progno- sis necessitating improved treatment regimens for these patients.1-9 Here, we begin to assess whether a selective
A
B
C
Figure 3. BAY 1143572 inhibits proliferation and affects CDK9 activity in primary ANKL cells. (A) Viability of human CD56-positive cells in control donor PBMC (n=5) assessed in the presence of recombinant human IL-2 (rIL-2, Miltenyi Biotec) at a final concentra- tion of 100 IU/mL together with different concentrations of BAY 1143572 for 24 hours. The IC50 value is indicated in each graph. (B) Viability of primary aggressive NK-cell leukemia (ANKL) cells from two separate patients (patient A, left panel, and patient B, right panel) assessed in the presence of rIL-2 at 100 IU/mL together with different concentrations of BAY 1143572 for 24 hours. The IC50 value is indicated in each graph. IC50 was defined as the concentra- tion of an inhibitor that reduced cell survival to 50% of the untreated control value, with the highest viability (no inhibitor) defined as 100%, and the lowest viability defined as 0%. (C) Primary ANKL cells from patient A were treated with the indicated concentrations of BAY 1143572 for 12 hours in the presence of 100 IU/mL rIL-2, followed by Western blotting probed with antibodies to phospho-RNAPII (Ser2), phospho-RNAPII (Ser5), and Mcl-1. Actin was the loading con- trol.
haematologica | 2018; 103(12)
2063