Fig. 6. EZH2 inhibition as a promising therapeutic strategy for TP63-rearranged TCLs.

(A) Tumor growth curve of NSG mice engrafted with SMZ1 cells injected subcutaneously (n=5/arm). Mice were treated with vehicle (Veh) or Valemetostat (VAL) when tumor size reached to 200 mm³ and sacrificed when subcutaneous tumors reached 2 cm in the longest dimension. (B) Kaplan-Meyer survival curve for mice in A. (C) Mean fluorescence intensity (MFI) of the H3K27me3 abundance in tumor samples from B determined by FACS. (D) Schematic outline of transplantation of Cre/tgTT lymphoma-infiltrated spleen cells (Tumor 1) and treating C57BL/6 recipients with Veh or VAL. C57BL/6 mice are irradiated with 5.5 Gy before receiving 0.5 million cells intravenously and treatments are initiated two days after transplantation. A cohort of mice are sacrificed after 10 days for pharmacodynamic (PD) analysis, and another cohort are observed for survival analysis. (E, F, G) Photo of spleens (E), spleen weight versus body weight ratio (F) and western blot analysis of H3K27me3 in spleen cells (G) from mice treated with VAL or Veh (n=3 mice/arm) for 10 days. (H) Kaplan-Meyer survival curve of C57BL/6 mice transplanted with Cre/tgTT lymphoma-infiltrated spleen cells and treated with Veh or VAL (n=6 mice/arm). (I) Western blot analysis for indicated proteins in spleen cells harvested from H. (J) Schematic of subcutaneous engraftment of ALK- ALCL PDX (COH1) harboring a TBL1XR1-TP63 rearrangement in NSG mice and treating recipients with VAL or Veh when tumor size reached to 200 mm³. (K) FACS analysis of H3K27me3 in tumor cells harvested from mice treated with VAL or Veh (n=3 mice/arm) for 10 days. (L, M) Tumor growth curve (L) and Kaplan-Meyer survival curve (M) of the COH1 PDX model. Mice were treated with Veh or VAL (n=6 mice/arm) by p.o. and sacrificed when the subcutaneous tumor reached 2 cm in the longest dimension. (N, O) Western blot analysis for indicated proteins in tumor cells harvested from M (N) and quantification of relative H3K27me3 abundance normalized to H3 (O). (P) Fluorescence in situ hybridization using a 5’ TP63 probe (green, 314 kb centromeric to TP63) and a 3’ TP63 probe (red, 310 kb telomeric to TP63). The translocated chromosomes are indicated with arrows. (Q, R) FACS showing abnormal T cells (purple) (Q) and associated quantifications (R) from pre-treatment and day 28 of VAL treatment in whole blood cells of Patient 2. Other cells shown are NK cells (blue), CD8+ T cells (green), and CD4+ T cells (red).

Data are presented as mean ± SEM. ** P<0.01 or **** P<0.0001 as compared between indicated groups (Student’s t-test). P value for survival by Log-rank test.