title: Combined immune checkpoint protein blockade and low dose whole body irradiation as immunotherapy for myeloma.
abstract: BACKGROUND
  Multiple myeloma is characterized by the presence of transformed neoplastic plasma cells in the bone marrow and is generally considered to be an incurable disease. Successful treatments will likely require multi-faceted approaches incorporating conventional drug therapies, immunotherapy and other novel treatments. Our lab previously showed that a combination of transient lymphodepletion (sublethal whole body irradiation) and PD-1/PD-L1 blockade generated anti-myeloma T cell reactivity capable of eliminating established disease. We hypothesized that blocking a combination of checkpoint receptors in the context of low-dose, lymphodepleting whole body radiation would boost anti-tumor immunity.
  
  
  METHODS
  To test our central hypothesis, we utilized a 5T33 murine multiple myeloma model. Myeloma-bearing mice were treated with a low dose of whole body irradiation and combinations of blocking antibodies to PD-L1, LAG-3, TIM-3, CD48 (the ligand for 2B4) and CTLA4.
  
  
  RESULTS
  Temporal phenotypic analysis of bone marrow from myeloma-bearing mice demonstrated that elevated percentages of PD-1, 2B4, LAG-3 and TIM-3 proteins were expressed on T cells. When PD-L1 blockade was combined with blocking antibodies to LAG-3, TIM-3 or CTLA4, synergistic or additive increases in survival were observed (survival rates improved from ~30% to >80%). The increased survival rates correlated with increased frequencies of tumor-reactive CD8 and CD4 T cells. When stimulated in vitro with myeloma cells, CD8 T cells from treated mice produced elevated levels proinflammatory cytokines. Cytokines were spontaneously released from CD4 T cells isolated from mice treated with PD-L1 plus CTLA4 blocking antibodies.
  
  
  CONCLUSIONS
  These data indicate that blocking PD-1/PD-L1 interactions in conjunction with other immune checkpoint proteins provides synergistic anti-tumor efficacy following lymphodepletive doses of whole body irradiation. This strategy is a promising combination strategy for myeloma and other hematologic malignancies.
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: Weiqing
authors.initials: W
authors.lastname: Jing
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: Jill A
authors.initials: JA
authors.lastname: Gershan
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: James
authors.initials: J
authors.lastname: Weber
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: Dominique
authors.initials: D
authors.lastname: Tlomak
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: Laura
authors.initials: L
authors.lastname: McOlash
authors.affiliation: Novartis Institutes for BioMedical Research, Inc., Cambridge, MA 02139 USA.
authors.forename: Catherine
authors.initials: C
authors.lastname: Sabatos-Peyton
authors.affiliation: Division of Hematology/Oncology/Transplant, Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI 53226 USA.
authors.forename: Bryon D
authors.initials: BD
authors.lastname: Johnson
keywords: 2B4; Blockade; CTLA4; Immune checkpoint proteins; LAG-3; Low dose whole body irradiation; Myeloma; PD-L1; TIM-3