Figure 2.
Interrogating prosurvival protein dependency in vivo in Ph+, Ph-like, and Ph− B-ALL PDX using BH3-mimetic drugs alone and in combination. (A) Irradiated NSG mice were transplanted with 1 × 106 primary B-ALL cells (Ph− 01-160-2015 or Ph+ 01-228-2012). Engraftment was confirmed at 6 weeks by detection of hCD45 in peripheral blood. Cohorts of 4 mice per group were then treated with (i) vehicle, (ii) S55746 (BCL-2 inhibitor), (iii) S63845 (MCL-1 inhibitor) or (iv) S55746 combined with S63845, and (v) dasatinib or (vi) dasatinib + S55746 (for 01-228-2012). Mice were euthanized on day 8, and immunohistological analysis for hCD45+ was performed on sternal bone marrow and spleen for infiltration by B-ALL cells captured at 100× magnification using the Aperio ScanScope. Representative cases from each cohort are shown. Flow cytometric analysis of flushed femurs showing the percentage of human CD45+ cells from B-ALL 01-160-2015 or 01-228-2012 after indicated treatments (mean ± 1 standard deviation and individual mouse values shown). B-ALL 01-160-2015 vehicle (n = 4), S63845 (n = 4), S55746 (n = 5), S55746+S63845 (n = 5). B-ALL 01-228-2012 vehicle (n = 4), S63845 (n = 3), S55746 (n = 3), S55746+S63845 (n = 4), dasatinib (n = 3), dasatinib + S63845 (n = 4). (B-C) NSG mice were transplanted with 1 × 106 Ph-like ALL cells (P2RY8-CRLF2 + JAK2 IR682RG). Leukemia engraftment and progression were assessed in groups of 6 female mice per treatment arm as shown with dosage regimen described in the “Methods.” (B) Treatment began when the mice engrafted 1% to 5% and human CD45+ cells and CD19+ proportion in peripheral blood (% hCD45+CD19+) were monitored weekly. *P = 0.0024 for Venetoclax + S63845 compared to vehicle by Mantel-Cox test. (C) Representative recipient sternal sections stained for hCD19 and hCD45 after treatment with vehicle (top) or Venetoclax + S63845 (middle and bottom, 2 independent recipients). Images taken at 20× magnification using the Panoramic Scan II by 3D Histech. (D) When venetoclax and S63845 were combined, fatal murine ATLS was observed with multiorgan involvement consisting of widely disseminated microemboli composed of nuclear and cytoplasmic debris derived from lysed leukemia cells, seen as deeply basophilic homogenous material as indicated by the arrows in spleen, lung, and kidney glomerulus.17,18 Images captured using the Olympus Model  BX41 and Olympus U-CMAD 3 camera at 200× magnification (spleen and lung) and 400× magnification (kidney). H&E, hematoxylin and eosin.

Interrogating prosurvival protein dependency in vivo in Ph+, Ph-like, and Ph− B-ALL PDX using BH3-mimetic drugs alone and in combination. (A) Irradiated NSG mice were transplanted with 1 × 106 primary B-ALL cells (Ph− 01-160-2015 or Ph+ 01-228-2012). Engraftment was confirmed at 6 weeks by detection of hCD45 in peripheral blood. Cohorts of 4 mice per group were then treated with (i) vehicle, (ii) S55746 (BCL-2 inhibitor), (iii) S63845 (MCL-1 inhibitor) or (iv) S55746 combined with S63845, and (v) dasatinib or (vi) dasatinib + S55746 (for 01-228-2012). Mice were euthanized on day 8, and immunohistological analysis for hCD45+ was performed on sternal bone marrow and spleen for infiltration by B-ALL cells captured at 100× magnification using the Aperio ScanScope. Representative cases from each cohort are shown. Flow cytometric analysis of flushed femurs showing the percentage of human CD45+ cells from B-ALL 01-160-2015 or 01-228-2012 after indicated treatments (mean ± 1 standard deviation and individual mouse values shown). B-ALL 01-160-2015 vehicle (n = 4), S63845 (n = 4), S55746 (n = 5), S55746+S63845 (n = 5). B-ALL 01-228-2012 vehicle (n = 4), S63845 (n = 3), S55746 (n = 3), S55746+S63845 (n = 4), dasatinib (n = 3), dasatinib + S63845 (n = 4). (B-C) NSG mice were transplanted with 1 × 106 Ph-like ALL cells (P2RY8-CRLF2 + JAK2 IR682RG). Leukemia engraftment and progression were assessed in groups of 6 female mice per treatment arm as shown with dosage regimen described in the “Methods.” (B) Treatment began when the mice engrafted 1% to 5% and human CD45+ cells and CD19+ proportion in peripheral blood (% hCD45+CD19+) were monitored weekly. *P = 0.0024 for Venetoclax + S63845 compared to vehicle by Mantel-Cox test. (C) Representative recipient sternal sections stained for hCD19 and hCD45 after treatment with vehicle (top) or Venetoclax + S63845 (middle and bottom, 2 independent recipients). Images taken at 20× magnification using the Panoramic Scan II by 3D Histech. (D) When venetoclax and S63845 were combined, fatal murine ATLS was observed with multiorgan involvement consisting of widely disseminated microemboli composed of nuclear and cytoplasmic debris derived from lysed leukemia cells, seen as deeply basophilic homogenous material as indicated by the arrows in spleen, lung, and kidney glomerulus.17,18  Images captured using the Olympus Model  BX41 and Olympus U-CMAD 3 camera at 200× magnification (spleen and lung) and 400× magnification (kidney). H&E, hematoxylin and eosin.

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