Jeff T. Hutchins, PhD Vice President, Pre-Clinical Research Peregrine Pharmaceuticals, Inc. March 25, 2015

Transcription

1 Expansion and Activation of T-cells via the Targeting of the Immunosuppressive Ligand Phosphatidylserine: Combination Strategy with Other Checkpoint Inhibitors Jeff T. Hutchins, PhD Vice President, Pre-Clinical Research Peregrine Pharmaceuticals, Inc. March 25, 2015 Immune Checkpoint Inhibitors: Validate Novel Pathways, Discover Predictive Biomarkers, Optimize Clinical Strategy

2 Combination Therapy is the Future More Patients Responding to Therapy Combination Treatment * * Hypothetical for illustrative purposes only 1. Why do only a sub-set of patients respond to IO Therapy? Role of the immunosuppressive tumor microenvironment Lack of T-cell activation and myeloid cell differentiation Checkpoint Blockade Yervoy and Opdivo are registered trademarks of Bristol Meyers Squibb. Keytruda is a registered trademark of Merck. Targeted Therapy Conventional Therapy 2. How can this patient pool be expanded? Ways a PS blockade changes this environment Consequences of PS targeting -- activation and expansion of T-cells

3 Immune Activity in the Tumor Environment Tumor-Promoting Phenotype: Patients unable to benefit from checkpoint inhibitors Tumor-Rejecting Phenotype: Patients more likely to respond to checkpoint inhibitors MDSC Myeloid-Derived Suppressor Cell Highly immunosuppressive Secrete anti-inflammatory cytokines MDSC Myeloid-Derived Suppressor Cell Reduction is current goal of immunotherapy M2 Mac M2 Macrophage Secrete anti-inflammatory cytokines and growth factors M1 Mac M1 Macrophage Secrete inflammatory cytokines Kill tumor cells through ADCC DC Immature Dendritic Cell Incapable of antigen presentation to T-cells DC Mature Dendritic Cell Present tumor antigens to T-cells Co-stimulatory signaling for T-cells T-reg T-Regulatory Cell Inhibits cytotoxic T-cells CD8 T-cell Cytotoxic (CD-8+) T-cell Receptor specific tumor cell killing Increased PD-1 expression TGF- IL-10 Cytokines Inhibit anti-tumor immune responses Keep Immune cells in quiet state TNF- IL-12 Cytokines Stimulate anti-tumor immunity

4 Immune Suppression in the Tumor Environment Creates a High Hurdle for Immunotherapies MDSCs M2 Macrophages TGF- IL-10 Immature DC Naïve T-cells Maintain an immunosuppressive tumor environment Lack of antigen presentation T-cells do not activate Low levels of PD-1 expression Ineffective response to tumors MDSC M2 TGF-!" IL-10 M1 Macrophage Immature DC Poor Response to Tumor Naïve T cell

5 Bavituximab: Immune System Activation with Broad Applicability Immune Signaling Target Monoclonal antibody targeting PS Broad Applicability Activity in multiple cancer types and infectious disease Significant Clinical Experience 19 studies, 500+ patients treated with bavituximab, currently in Phase III Good safety profile alone and in combination with other therapies (no IR safety events) Immune Activation MOA and

6 PS Ligand Engages Multiple Receptors PS Expressing Cells Tumor cell Tumor microvesicle Tumor blood vessel PS PS PS PS PS PS PS MFG-E8 Gas6, Protein S TIM-1 TIM-3 TIM-4 BAI1 Stabilin-2 Immune Cells 3 5 MDSC M2 Macrophage Immature DC T-Cells Tyro-3, Axl, Mer (TAM) Wu, Y., Tibrewal, N., and Birge, R.B. (2006). Phosphatidylserine recognition by phagocytes: a view to a kill. Trends Cell Biol. 16, Park et al, (2007). BAI1 is an engulfment receptor for apoptotic cells upstream of the ELMO/Dock180/Rac module. Nature 450, Hanayama, R., Tanaka, M., Miwa, K., and Nagata, S. (2004). Expression of developmental endothelial locus-1 in a subset of macrophages for engulfment of apoptotic cells. J. Immunol. 172, Park et al, (2007). Rapid cell corpse clearance by stabilin-2, a membrane phosphatidylserine receptor. Cell Death Differ 15, Ishimoto Y, Ohashi K, Mizuno K, Nakano T. Promotion of the uptake of PS liposomes and apoptotic cells by a product of growth arrest-specific gene, gas6. J Biochem (Tokyo) 2000;127: Scott RS, McMahon EJ, Pop SM, et al. Phagocytosis and clearance of apoptotic cells is mediated by MER. Nature 2001;411: Miyanishi M, Tada K, Koike M, Uchiyama Y, Kitamura T, Nagata S. Identification of Tim4 as a phosphatidylserine receptor. Nature 2007;450: Hanayama R, Miyasaka K, Nakaya M, Nagata S. MFG-E8-dependent clearance of apoptotic cells, and autoimmunity caused by its failure. Curr Dir Autoim- mun 2006;9: Jinushi M, Nakazaki Y, Dougan M, Carrasco DR, Mihm M, Dranoff G. MFG-E8 mediated uptake of ap- optotic cells by APCs links the pro- and antiinflammatory activities of GM-CSF. J Clin Invest 2007;117: Donna L. Bratton and Peter M. Henson, Apoptotic Cell Recognition: Will the Real Phosphatidylserine Receptor(s) Please Stand up? Current Biology Vol 18 No 2

7 Mechanisms of PS Immunosuppression Mediated by TAM and TIMs Receptor Signaling TIM Receptors Apoptotic cells and debris with exposed PS TAM Receptors Apoptotic cells polarize to M2 macrophages CD4 Th2 bias Tim-1 Th1 response T cell exhaustion CD8! " Tim-3 Innate immunosuppressive actions stimulated by ligand PS complexes CD8 Activated T cell feedback inhibits innate immunity Tolerance Tim-4 Freeman et al, Immunol Rev. 235: (2010) Graham et. al. Nature Reviews 14: (2014) TAM signaling inhibits NK antimetastatic effects

8 Strong TAM Kinase Domain Activation Requires PS Courtesy of Lew et. al., and Lemke; elife 2014;3:e03385

9 PS Signaling Pathway Placing the on the Immune System PS Signaling Immune System Signaling Immune Suppression PS is engaged By PS receptors Tumor Cell = Exposed PS PS MDSC PSR PS PSR M2 Exposed PS is a checkpoint that places brakes on the immune system PS receptor signaling causes increased anti-inflammatory cytokine expression MDSCs accumulate DCs fail to mature T-cells remain naïve M1 Macrophages fail to develop TGF- MDSC Naive 2. Immune System Signaling 3. Immune T-cell Suppression IL-10 M1 Immature DC MDSC MDSC Naive T-cell

10 Multiple Pre- 2aG4 treatment of PC3 tumor-bearing mice increases production of immunostimulatory IL-12 and TNF- microenvironment and decreases the production of immunosuppressive TGF #$%&'(%)'"*+"',(*$-.%&$/)0"(/1%" /)1$%&2%2"(&',$%"342 5" 2aG4 treatment induces expression of T-cell co-stimulatory molecules on CD11b+ cells in PC3 tumors 1 Prolonged survival time and cures in rats bearing orthotopic F98 tumors treated with irradiation and 2aG4. Rats bearing orthotopic F98 gliomas were treated with 10 Gy radiation therapy (RT), 2aG4, RT + 2aG4, or RT + C44 5 d after injection of tumor cells as in Fig. 1. The survival in the RT + 2aG4 group was significantly longer than in all other groups (P < 0.001). Thirteen percent of rats treated with RT + 2aG4 were rendered disease free. After 230 d, the disease-free rats were rechallenged by intracerebral injection of into the contralateral hemisphere. None of the rats developed disease, indicating that their prior treatment had caused them to develop immunity. 3 Tumor-specific cytotoxic T-cells 3 ()*+,-,+"./0,0"" 1" Cure of castration resistant prostate cancer in TRAMP mice treated with mchn11 and androgen deprivation therapy. Survival time (until sacrifice) was analyzed by the Log-rank method. Mice treated with Cx+ mch1n11 had longer survival times than mice treated with Sx+ mch1n11, Cx+C44, or Sx+C44. * Significant for mch1n11 or castration alone versus C44 control; ** Significant for Cx+ mch1n11 versus mch1n11 or Cx or alone. Cx, castration. Sx, sham-operated. N = #$%&"""""""""""" '&&" 2aG4 directly induces repolarization of TAMs by binding to PS on their cell surface and activating them in an Fc-dependent manner. (a) FACS analyses showing that F4/80+ TAMs from disaggregated PC3 tumors acquire an M1-like phenotype when cultured for 4 days with 2aG4. The M2-like (F4/80+, Arg-1+) population and the M1-like (F4/80+, inos+) were quantified. (b) Treatment of F4/80+ TAMs with 2aG4 induces NO production in an Fc-dependent manner. (c) RT-PCR showing that F4/80+ TAMs lose M2 markers and acquire M1 markers when cultured with 2aG4. 1 2aG4 directly induces repolarization of TAMs by binding to PS on their cell surface and activating them in an Fc-dependent manner. (a) FACS analyses showing that F4/80+ TAMs from disaggregated PC3 tumors acquire an M1-like phenotype when cultured for 4 days with 2aG4. The M2-like (F4/80+, Arg-1+) population and the M1-like (F4/80+, inos+) were quantified. (b) Treatment of F4/80+ TAMs with 2aG4 induces NO production in an Fc-dependent manner. (c) RT-PCR showing that F4/80+ TAMs lose M2 markers and acquire M1 markers when cultured with 2aG4. 1 Vascular damage is caused by M1-like TAMs generated by 2aG4 treatment. a) Representative frozen sections showing TAM-mediated disintegration of vascular endothelium in PC3 tumors from mice treated for 2 weeks with 2aG4, alone or in combination with docetaxel. b) TAMs congregating around damaged vessels in 2aG4-treated mice costained for inos (blue) and F4/80 (green), indicating they were of M1-like phenotype. c) Representative sections showing that 2aG4 treatment causes a shift in the predominant polarity of TAMs from M2-like (F4/80+, green; Arg-1+, red) in C44 mice (upper) to M1-like (F4/80+, green; inos+, blue) in 2aG4-treated mice (lower). d) Histograms show the area of PC3 tumor sections occupied by M1 (F4/80+, inos+) or M2 (F4/80+, Arg-1+) TAMs. (e) RT-PCR of RNA from PC3 tumor homogenates reveals a general increase in M1 markers and a decrease in M2 markers in tumors from 2aG4-treated mice relative to C44-treated mice. (f) TAMs from 2aG4-treated mice manufacture NO. (g). TAMs from 2aG4-treated mice are tumoricidal. 1 Damage to tumor vasculature induced by treatment with 2aG4. a)denudation of vascular endothelium in PC3 tumors in mice treated with 2aG4. b) Reduction of tumor vascularity and perfusion in PC3 tumor-bearing mice treated with 2aG4 alone or in combination with docetaxel. c) 3-D power Doppler ultrasound showing a reduction in blood flow in PC3 tumors in mice treated with 2aG4 alone or in combination with docetaxel AACR 2013 Annual Meeting: Phosphatidylserine-Targeting Antibody Induces Differentiation of Myeloid-Derived Suppressor Cells into M1-like Macrophages Yi Yin, Xianming Huang, Dan Ye, Yi Yin, Philip E. Thorpe Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, TX. 2. AACR 2012 Annual Meeting: Yin et al, Cure of castration-resistant prostate cancer in TRAMP mice by reactivating tumor immunity with a phosphatidylserine-targeting antibody. Department of Pharmacology, The University of Texas Southwestern Medical Center at Dallas, Dallas, TX 3. He et al, Antiphosphatidylserine Antibody Combined with Irradiation Damages Tumor Blood Vessels and Induces Tumor Immunity in a Rat Model of Glioblastoma. Clin Cancer Res 2009;15:

11 Bavituximab Facilitates Induction of Tumor-Specific Cytotoxic T-Cells MHCII CD40 Mature DC CD80 CD86 Antigen presentation T T Tumor blood vessel cell / Tumor cell / Microvesicle Cytotoxic T-cells T Tumor-specific cytotoxicicity Generation of Antitumor T-cell Immunity in Syngeneic F98 Glioma in Rat 2 Long-term Immunity Cure of TRAMP Mice by Anti-PS Combination 1 Cytotoxic T-cell Generation 2 1. AACR 2012 Annual Meeting: Yin et al, Cure of castration-resistant prostate cancer in TRAMP mice by reactivating tumor immunity with a phosphatidylserine-targeting antibody. Department of Pharmacology, The University of Texas Southwestern Medical Center at Dallas, Dallas, TX 2. He et al, Antiphosphatidylserine Antibody Combined with Irradiation Damages Tumor Blood Vessels and Induces Tumor Immunity in a Rat Model of Glioblastoma. Clin Cancer Res 2009;15:

12 Tumor Response to Bavi Equivalent Involves Increased CD8 T cells and Decreased MDSCs (PS Targeting) EMT-6 breast tumor model in Balb/c mice (N=10/group; mean sem) % C D 8 T IL s G r o u p M e a n C c h 1 N p v a lu e = % M D S C /C D G r o u p M e a n C c h 1 N p v a lu e = T u m o r V o lu m e (m m 3 ) T u m o r V o lu m e (m m 3 )

13 Bavituximab Blocks Immunosuppression Activates Immune Response (Hits Accelerator) (Removes Brakes) Blocks PS Signaling Blocks PS receptor immunosuppressive signal IL-10 and TGF- levels decrease Activates Immune Response Fc / receptor signaling MDSC differentiation M2 to M1 macrophage polarization DCs mature, present tumor antigens to T-cells Sustained Immune Response Increased Inflammatory cytokines M1 macrophages kill tumor cells via ADCC Activated T-cells kill tumor cells Yi Yin, Xianming Huang, Kristi D. Lynn, and Philip E. Thorpe, Phosphatidylserine-Targeting Antibody Induces M1 Macrophage Polarization and Promotes Myeloid-Derived Suppressor Cell Differentiation. Cancer Immunology Research October :

14 Promising and Consistent Pre-Clinical Data Support Phase III Development in Combination with Docetaxel Rationalebased Combination Docetaxel has been shown to increase PS exposure (AACR 2013 Poster #2850) Docetaxel has immunostimulatory effects (Kodumudi et al, Clin Cancer Res 2010;16: ) Compelling Pre-clinical Data Tumor models show greater than 90% reduction in tumor growth with bavi equivalent plus docetaxel (Cancer Research 2005; 65: (10) )

15 Final Phase II 2 nd Line NSCLC Overall Survival Data with Docetaxel 4*(./)%6"4*)'$*7" 4*(./)%6"4*)'$*7"

16 Global Phase III Registration Trial N = 582 Randomized 1:1 Placebo-Controlled Bavituximab (3 mg/kg) Weekly Docetaxel Day 1, of 21-day cycles up to 6 cycles Placebo Weekly Docetaxel Day 1 of 21-day cycles for up to 6 cycles Primary Endpoint: OS Secondary Endpoints: PFS, ORR, Safety Positive EOP2 discussion with FDA and Ex-US Regulatory Agencies Single Phase III global registration trial > 150 sites (U.S., Europe and Asia Pacific) Est. 24 month enrollment and 12 month followup Powered to show approx. 2 month improvement in median overall survival Enrollment initiated January 2014 Patient Criteria: Stage IIIb/IV Non squamous NSCLC Only one prior systemic therapy for advanced disease Unselected for genetic mutations Granted Fast Track Designation January 2014

17 Promising ORR, PFS and Survival Data in Breast Cancer Single-Arm Trials n= ORR PFS OS Advanced/MBC All Types (1 Prior Chemo) Bavituximab + Docetaxel 46 61% Docetaxel 29 41% Advanced/MBC All Types (Front-Line) Bavituximab + Carbo/Paclitaxel 46 74% Carboplatin/Paclitaxel % MBC (Her2 neg) Bavituximab + Paclitaxel 14 85% Breast Docetaxel: ORR: Burstein, H., et al (2000). JCO. PFS/OS: Nabholtz, J., et al (1999). JCO. Peregrine ASCO (2010), 8/11. Breast Paclitaxel/Carboplatin: Loesch D., et al (2002). JCO. Peregrine ASCO (2010). Bavi IST: Alison Stopeck

18 A HCC Responsive Patient Treated with Bavituximab in Combination with Sorafenib Pt ID #P2-23 (7.98) Pre Post Pre Post H&E CD68 IHC score: 3 IHC score: 3 CD4 FoXP3 IHC score: 2 IHC score: 3 IHC score: 1 IHC score: 0 CD8 GranzymeB IHC score: 2 IHC score: 3 IHC score: 1 Increase in tumor immune activity: increase CD8+ & CD4+ T cells IHC score: 2

19 Hypothesis: Bavituximab Can Potentiate Additional anti-pd-1 Responders Phase II HCC Data: Increased T-cell tumor infiltration Best immune responses observed in patients with greatest % increase in PD-1 and CD8+ cells = poor immune responders

20 PD-L1/PD-1 Signaling = Suppression of T-Cell Function IL-10 MDSC APC Conversion into T-reg cell PD-L1 PD-L1 T-reg PD-1 PD-1 Endothelial Cell PD-L1 PD-1 CD8+ T-cell PD-1 PD-L1 T-reg PD-L1 + = Exhausted or PD-1 Regulatory Cell PD-L1 Tumor Cell PD-1 PD-1 PD-L1 M1 Mac Conversion into Reg. Mac M2 Mac Anti-inflammatory cytokine production Adapted from Nguyen et al, Nature Reviews: Immunology 15, (2015) and Lee et al, Advances in Bioscience and Biotechnology; 2013, 4, 19-29

21 Relationship of PD-L1 Expression and Activity in Tumor Biopsies Evaluable by IHC NSCLC PD-L1 Tumor Expression N= Responses PFS at 6 mo. OS at 6 mo. High 7 4 (57%) 67% 89% Low 21 1 (5%) 11% 33% AACR 2014 Annual Meeting, abstract CT105 MK-3475 (anti-pd-1 monoclonal antibody) for non-small cell lung cancer (NSCLC): Antitumor activity and association with tumor PD-L1 expression Melanoma PD-L1 Tumor Expression N= Responses median PFS OS at 6 mo. Positive (48%) % Negative 22 1 (5%) % AACR 2014 Annual Meeting, abstract CT104 Antitumor activity of the anti-pd-1 monoclonal antibody MK-3475 in melanoma (MEL): Correlation of tumor PD-L1 expression with outcome

22 Upstream & Downstream Checkpoint Combinations = Exposed PS Tumor Cell PS PS PSR PSR MDSC M2 Macrophage TGF- IL-10 " " Immature Dendritic Cell B7-H3 B7-H3L PD-L1 PD-L1 CTLA-4 PD-1 B7-H3L KIR T TIM-3 NK PS / PSR: Upstream PS PS Receptors: Axl TIM-1 TIM-4 Hokkaido University, Japan TIM-3 TGF- IL-10 IL-21 IDO Immunosuppressive Cytokines/ Enzymes PD-1 PD-L1 CTLA-4 TIM-3 Downstream

23 PS Blockade Synergizes with Anti-PD-1 Tumor Volume (mm 3 ) anti-pd-1 Day 42 89:;:<55" anti-pd-1 + anti-ps Mice bearing K1735 melanoma tumors were treated with anti-pd-1 or mch1n11 + anti-pd-1, 2.5 mg/kg 2x per week. " T u m o r V o lu m e (m m 3 ) T u m o r V o lu m e (m m 3 ) Response in Individual Mice a n ti-p D E x p e rim e n ta l D a y s a n ti-p D -1 + a n ti-p S E x p e rim e n ta l D a y s

24 Combination Treatment Repolarizes Immune Profile in Spleen Results of PS Blockade + anti-pd-1 Versus anti-pd-1 Alone (K1735 melanoma model) CD8+ T-cell CD8+ Cells 8"9 CD8 cells 40% PD-1+ No Change IFN- 40% CD4+ T-cell CD4+ Cells 8"9 CD4 cells No Change T-reg No Change PD-1+ No Change IL-2+ 80% IL % MDSC Mac DC MDSC / Macs 8"9 CD11b+ No Change MDSC ( via CD11b) 45% PD-L1+ CD11b cells No Change PD-L1+ CD11b/Ly6G- No Change PD-L1+ CD11b/Ly6G- No Change DC (CD11c hi /F480 lo ) 30% Mac (CD11b/F480 hi ) No Change PS Blockade + Anti-PD-1shifts spleenocytes to immune active phenotype Data support potentiating additional PD-1 responders via PS blockade.

25 PS Targeting Treatment Dramatically Reduces Expression of PDL-1 on Tumor Isolated Leukocytes 4*)'$*7" &)'/-8=" 30.5% 30.5% 7.94% 7.9% &)'/-835" 31.4% 31.4% DE;<B"*+"83-C5">"4%772" &)'/-835>&)'/-8=" 5.0% 4.97% 85 to 90% of PD-L1+ cells are CD45+ leukocytes, less than 10-20% PDL-1+ cells are tumor cells in this analysis?5;?b"*+"83-c5">"4%772"?5;fb"*+"83-c5">"4%772" PD-L1

26 Combination Treatment Repolarizes Immune Profile in Tumor Results of PS Blockade + Anti-PD-1 Versus anti-pd-1 Alone (K1735 melanoma model) CD8+ T-cell CD8+ Cells 8"9 CD8 cells 30% CD8 /T-reg Ratio 100% < 0.01 PD-1+ 50% Lag-3+ 90% < 0.01 IFN- 70% TNF- 70% Granzyme-B+ 70% " " CD4+ T-cell CD4+ Cells 8"9 CD4 /T-reg Ratio 100% < 0.01 PD-1+ No change CD137+ (41BB) 60% 0.03 IFN- 30% < 0.05 TNF- 100% < 0.01 IL-2+ 60% 0.02 MDSC Mac MDSC / Macs 8"9 MDSC ( via CD11b) 40% 0.02 inos+ CD11b cells 90% PD-L1+ TIL 80% <0.01 PS Blockade + Anti-PD-1 increases TIL ; shifting TIL and cytokines towards tumorrejecting phenotype. Data support potentiating additional PD-1 responders via PS blockade.

27 Key Takeaway: PS Blockade Optimizes Tumor Environment for Additional Anti-PD-1 Responders Results of PS Blockade + anti-pd-1 versus anti-pd-1 alone (K1735 melanoma model) MDSC 40% reduction* of myeloid-derived suppressor cells CD8+ T-cell 30% increase* in CD8+ T-cells 50% increase* in PD-1 expression Positive environment for increasing anti-pd-1 Responders PD-L1 on TILs PD-1 80% decrease* PD-L1 expression on TIL (CD45+) * Statistically significant (p = < 0.02)

28 PS Targeted Combination Therapy in PD-1 Resistant Melanoma Increases the Anti-tumor Response Rate C o n t r o l I g G ( C 4 4 ) M e d ia n T u m o r V o lu m e ( m m 3 ) C o n t r o l Ig G C h 1 N 1 1 (Anti-PS) A n ti-p D -1 C h 1 N A n ti-p D -1 p = p = T u m o r V o l u m e ( m m 3 ) T u m o r V o l u m e ( m m 3 ) T u m o r V o l u m e ( m m 3 ) E x p e r i m e n t a l D a y s C h 1 N E x p e r i m e n t a l D a y s A n t i - P D E x p e r i m e n t a l D a y s E x p e r im e n t a l D a y s A n t i - P D N B16F10 Melanoma Tumor Model in C57BL Mice (10/group) T u m o r V o l u m e ( m m 3 ) E x p e r i m e n t a l D a y s 28

29 Immune Cell Changes in the Tumor Microenvironment in the Mouse B-16F10 Model Treatment Type (Day 23, after 4 treatments) CD45 (WBC) CD3* (mature T-cells) Immune Cell Type (%) CD8** (Cyto T- cells) CD4** (helper T- cells) T reg Tumor Size (mm 3 ) Control mab (2) <:;5! FG;D! AA;E! <:;D! 5G;E! FF<A! ch1n11 (anti- PS) (7) <F;G! EF;G! AF;D! FF;G! 5:;G! <5@A! Anti-CTLA-4 (5) 5@;?! F?;D! EA;5! E:;E!?;F! <<:E! Anti-PD-1 (6) <G;F! E<;<! E?;E! FF;?! 5?;<! 5G?G! ch1n11 + Anti-PD-1 (5) 2&3&! 4535! 6738! <:;A! 5G;F! 967! "H"8%$1%)'&0%"*+"43EA"I7%,1*1J'%K"L*2/'/M%"1%772" HH"8%$1%)'&0%"*+"43F"I(&',$%"#-1%77K"L*2/'/M%"1%772" (:$:,0:,+$;;/"0,<=,-,+$=:"+>$=<*"

30 Key Observations - Summary Phosphatidylserine (PS) is externalized in the tumor microenvironment and is a major immunosuppressive signal PS is a global immune checkpoint Antibody-mediated blockade of PS signaling breaks immune tolerance reactivating innate and adaptive immunity and results in durable anti-tumor responses in multiple pre-clinical models Combination pre-clinical studies in immuno-competent models with PD-1 and CTLA-4: increase CD8 T cells and M1 macrophages decrease MDSCs and M2 macrophages demonstrate statistically significant anti-tumor responses

31 =L%1/&7"#N&)O2"'*P" Powerful science. Novel therapies. Dedicated to patients. Xianming Huang, Ph.D. Rolf Brekken, Ph.D. Bruce Freimark, Ph.D. Nikoletta Kallinteris Dmitry Gabrilovich, M.D., Ph.D.