Figure 6. E3330 and SHP099 treatment represses clonal expansion of Tet2-KO HSPCs and prevents aberrant early signs of MPN in both young and aged Tet2 mutant mice.

(A) A schematic showing the experimental procedure used for generating chimeric mice mimicking clonal expansion of Tet2-KO cells. Tet2-KO bone marrow cells (CD45.2) were mixed with BoyJ bone marrow cells (CD45.1) at a ratio 1 to 5 (100K: 500K). Two months post-transplant, the chimeric mice were subjected to continuous treatment with LPS (0.8 mg/kg, i.p. injection every other day for 1 month) or a daily injection of E3330 or SHP099 (oral gavage, 20 mg/kg for E3330, 50 mg/kg for SHP099 for 3 months). Chimerism in peripheral blood was analyzed monthly at indicated time points.

(B) Clonal expansion of Tet2-KO HSPCs is heightened by continuous administration of LPS.

(C) Clonal expansion of Tet2-KO HSPCs is repressed by daily injection of E3330 or SHP099.

(D) Long-term treatment of juvenile Tet2-KO mice with E3330 or SHP099 results in the rescue of multiple phenotypic defects associated with Tet2 deficiency including frequency of CMPs, LSKs, HSCs, expression of Morrbid, Bim, IL-6 and pStat3 as well as spleen weight.

(E) Short-term treatment of aged Tet2-KO mice with E3330 or SHP099 prevents early hematological signs of CMML in peripheral blood. E3330, 20 mg/kg and SHP099, 50 mg/kg, oral gavage.

Transplant data in (B) and (C) are derived from using 5 recipient mice per group, mean ± s.e.m. 15 doses of LPS were given at 0.8 mg/kg over a period of one month (i.p. injection, every other day). E3330 (20 mg/kg) and SHP099 (50 mg/kg) were administered by oral gavage daily for 12 weeks in (D) or for 2 weeks in (E). Drug treatment studies in (D) and (E) are pooled data from two independent experiments (n=4 to 5 mice per group, mean ± s.e.m.).

P value: * P < 0.05, ** P < 0.01. n.s., not significant. Statistical analysis performed by unpaired, two-tailed Student’s t-test. See also Figure S6 for additional supporting data.