Sensitizing Protective Microenvironments to Antibodymediated Dr. Christian P. Pallasch
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1 Sensitizing Protective Microenvironments to Antibodymediated Therapy Dr. Christian P. Pallasch
2 The Microenvironment in Lymphoma and Leukemia Lymphomagenesis Therapy Progression Seite 2 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
3 Macrophages determine antibody therapy outcome in the hmb humanized Lymphoma Model Human Cord Blood derived CD34 + HSC Eµ/CD19Pro Leskov et al.oncogene 2012 GFP + cells / spleen GF P MY C BCL2 n.s. * GFP + cells / organ Control + Antibody Treatment * * Seite 3 Control Alem. Clod. Clod + Alem. Pallasch et al. CELL 2014 Alem.: Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch Peripheral Spleen Liver Bone Blood marrow
4 Development of resistence towards Alemtuzumab in the Bone Marrow Day 4 Day 9 Day 12 Day 15 Lymphoma cells in the bone marrow are susceptible to Alemtuzumab treatment until they reach a threshold of ~5% infiltration Seite 4 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
5 The tumor microenvironment determines therapeutic response Antibodymediated phagocytosis of lymphoma cells Primary resistance to therapeutic antibody??? Seite 5 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
6 An in vivo RNAi approach towards macrophage inactivating mechanisms shrna library (MLSmCHERRY) PCR amplify hairpins Control Highthroughput sequencing Control PTGES3 FCGR2B CD52 Harvested hmb cells Antibody treatment Quantify shrna sequences MIF Alemtuzumab treated Seite 6? PGE2 FCGR2B Control shcd52 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch % Alem. specific killing shfcgr2b shptges3
7 Seite 7 Combinatorial Chemoimmunotherapy overcomes sitespecific resistance Untreated Control Alemtuzumab GMCSF + Alemtuzumab Irradiation Irradiation+ Alemtuzumab Cyclophosphamide Cyclophosphamide + Alemtuzumab Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch Pallasch et al. CELL 2014
8 CCL4 pg/ml Cyclophosphamide induced Cytokine response ehances macrophage dependent lymphoma cell depletion IL8 pg/ml * VEGF pg/ml TNFα pg/ml Irradiation CTX d2 d2 d2 d2 Seite 8 % Alem. specific killing Blocking antibody Recombinant in conditioned media cytokines Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch Media Control * Cond. Media CTX Pretreated BM * antitnfα * antivegf % Alem. specific killing Media Control Cond. Media CTX Pretreated BM IL8 * TNFα * VEGF * CCL4
9 Finding a dosing regimen synergy is time dependent GFP + leukemic cells / femur * * Interval Alemtuzumab to Cyclophosphamide / days Pallasch et al. CELL 2014 Seite 9 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
10 Cyclophosphamide Treatment rewires the Tumor Microenvironment Seite 10 untreated Cyclo. GREEN GFP+ lymphoma cells RED 70kD DextraneTexasRed/ Macrophages d0 d Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch % F4/80+ macrophages / femur CD86 CXCL9 TNF CXCR4 CD16/32 Lyve1 CD80 CD14 CD206 TLR2 Arginase1 FcgR1a CCL2 CD68 IL4 Dectin1 IL10 IL12 log2 MFI * * Untreated Cyclophosphamide nonleukemic leukemic 24 h 72 h FC * Leuk vs. 72h CTX 1.5 * * 2.5 * FC NonLeuk. vs. Leuk. 2.8 * 1.4 * 1.5 * 1.4 * 1.6 * 3.2 * 1.4 * 2.1 *
11 Cytokine Release determines Cyclophosphamide/Rituximab Resistance in Patient derived BALL % Rituximab spec. cell killing * * * hcd19 + leukemia cells / femur * n.s. Seite 11 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
12 Chemotherpy in ALL patients increases macrophage frequencies in the bone marrow Seite 12 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
13 Cyclophosphamide induced acute Cytokine Release enables macrophagedependent tumor cell depletion VEGF TNF IL8 CCL4 Differentiation Migration Activation Cyclophosphamide Treatment Pallasch et al. CELL 2014 Seite 13 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
14 Tumor Microenvironment and Therapy Perspectives and further plans 1. Microenvironmentdependent resistance?? 2. Mechanism of the Acute secretory activating response? 4. Development of Microenvironmenttailored therapeutic strategies Seite 14 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch 3. Therapeutic Implications of Macrophages
15 Acknowledgements AG Therapie und Mikromilieu Maligner Lymphome Daniela Vorholt Yvonne Meyer Nadine Kutsch Janine Schwamb Andreas Roth Jana Baller Steffen Hoppe On Yu Kwon Uniklinik Koeln Lukas Frenzel Karl Kreuzer Lukas Heukamp Clemens Wendtner Michael Hallek VJM Amsterdam Nico van Rooijen David H. Koch Institute for Integrative Cancer Research at MIT Luke Gilbert Christian Braun Yadira Soto Ilya Leskov Jianzhu Chen Frank Gertler Herman Eisen Mike Hemann Seite 15 Sensitizing protective microenvironments to antibodymediated therapy Dr. Christian P. Pallasch
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