Content of Endothelial Progenitor Cells in Autologous Stem Cell Grafts Predict Survival after Transplantation for Multiple Myeloma Egil S. Blix, Anders.

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Content of Endothelial Progenitor Cells in Autologous Stem Cell Grafts Predict Survival after Transplantation for Multiple Myeloma Egil S. Blix, Anders B. Kildal, Eirin Bertelsen, Anders Waage, June H. Myklebust, Arne Kolstad, Anne Husebekk Biology of Blood and Marrow Transplantation Volume 21, Issue 5, Pages 840-847 (May 2015) DOI: 10.1016/j.bbmt.2014.12.027 Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 1 CD34+CD133+VEGFR2+ EPC can be identified within ALDHhi population in stem cell grafts from NHL and MM patients. (A) Flow cytometry analysis of EPC in representative stem cell grafts from NHL and MM patients (ID 10, 39, 12, and 22). FSC/SSC gating was used to identify lymphocytes and monocytes, followed by gating on ALDHhi cells in order to define cells with stem or progenitor characteristics. EPC were subsequent defined as triple positive CD34+VEGFR2+CD133+ cells (red arrow). (B) Back-gating analysis of VEGFR2+CD133+cells shows that the majority of double positive VEGFR2+CD133+ fall within CD34 and ALDH gates. Representative sample from a MM patient (MM ID 07). (C) Bar chart illustrating distribution of EPC as percentage of CD34+ cells in stem cell grafts from patients treated with ASCT. NHL (white bars) and MM (grey bars). Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 2 The absolute number of EPC in stem cell grafts stratifies OS in MM patients after high-dose chemotherapy with ASCT. (A) Bar chart illustrating the estimated number of EPC infused to MM patients together with autologous CD34+ stem cells during ASCT (n = 24). EPC cells per kilogram was defined as a ratio of measured percentage EPC of CD34+ cells as determined by flow cytometry analysis, divided by the total number of stem cells reinfused (CD34+ cells × 106/kg). MM patients were divided into two groups depending on whether the number of EPC cells per kilogram was above or below the median value for the cohort. (B) Progression-free survival after ASCT in MM patients with higher or lower than cohort median EPC cells per kilogram was compared with Kaplan-Meyer plot with log-rank test and found to be significantly lower in the group with EPC cells per kilogram higher than cohort median. (C) MM patient OS after ASCT was compared with Kaplan-Meyer plot with log-rank test and found to be significant lower in MM patients with EPC cells per kilogram higher than cohort median. Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 3 Positive correlation between s-β2-microglobulin and the percentage of EPC in stem cell grafts from MM patients. Scatter plot of EPC as percentage of CD34+ cells in stem cell grafts as determined by flow cytometry versus s-β2-microglobulin (mg/L) in MM patients at onset of treatment (n = 23). Association between variables was evaluated by Pearson R2. Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 4 ISS stratifies OS in MM patients after ASCT. OS (A) and PFS (B) for patients MM patients after ASCT according to ISS I (n = 12) compared with ISS II or III (n = 11). Kaplan-Meyer plot with log-rank test. Significance level of P < .05. Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Supplemental Figure 1 Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Supplemental Figure 2 Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Supplemental Figure 3 Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions

Supplemental Figure 4 Biology of Blood and Marrow Transplantation 2015 21, 840-847DOI: (10.1016/j.bbmt.2014.12.027) Copyright © 2015 American Society for Blood and Marrow Transplantation Terms and Conditions