Mo1127 Mast Cells in Addition to Eosinophils Are Markedly Elevated in Patients with Eosinophilic Gastritis and/or Gastroenteritis Evan S. Dellon1, Kathryn A. Peterson2, Robert M. Genta3, Joseph A. Murray4, Nirmala Gonsalves5, Mirna Chehade6, Marc E. Rothenberg7, Paneez Khoury8, Adam C. Bledsoe4, Camilla Shaw9, Henrik S. Rasmussen9, Bhupinder Singh9, Alan T. Chang9, Bradford A. Youngblood9, Ikuo Hirano5 1University of North Carolina, Chapel Hill, NC; 2University of Utah, Salt Lake City, UT; 3Baylor College of Medicine, Houston, TX; 4Mayo Clinic Rochester, Rochester, MN; 5Northwestern University, Chicago, IL; 6Icahn School of Medicine at Mount Sinai, New York City, NY; 7Cincinnati Children's Hospital Medical Center, Cincinnati, OH; 8NIH/NIAID, Bethesda, MD; 9Allakos, Inc., Redwood City, CA

BACKGROUND OBJECTIVE Figure 6. Study Diagnostic Criteria Figure 8. Eosinophils and Mast Cells Are Consistently • Pathologic accumulation and over-activation of eosinophils are • To quantify the extent of eosinophil and mast cell infiltration in Elevated in Gastric and Duodenal Biopsies implicated in multiple chronic inflammatory diseases in the GI tract gastric and duodenal biopsies from symptomatic patients including eosinophilic esophagitis (EoE), gastritis (EG), diagnosed with EG/EGE Eosinophilic Esophagitis Stomach Eosinophils (EoE) ESOPHAGUS Mast Cells gastroenteritis (EGE), and colitis - collectively termed eosinophilic ≥30 eos/hpf (n=40) ≥15 eos/hpf in 1 hpf 350 gastrointestinal disorders (EGIDs) METHODS 300 hpf 200 • Patients with EGIDs have decreased quality of life due to Eosinophilic Gastritis Figure 3. Screening Protocol (EG) STOMACH debilitating symptoms such as dysphagia, abdominal pain, 150 nausea, vomiting, and diarrhea Symptom PRO EGD with Biopsy Histologic Criteria ≥30 eos/hpf in 5 hpfs 100 • While the pathogenesis of EGIDs has historically been thought to Eosinophilic Gastroenteritis be driven by eosinophils, mast cells have also been shown to be (EGE) SMALL elevated in EoE.1,2 ≥30 eos/hpf in 3 hpfs INTESTINE 50 • Subjects with prior • Multiple biopsies were • Single pathologist Normal Mean Peak Cell Count / / CountCell Peak Mean MC Level4-8 • The role of mast cells in other EGIDs, has yet to be established diagnosis or suspected taken from each evaluated stained biopsy Eosinophilic Gastritis and 0 EG/EGE entered symptomatic subject samples and counted Gastroenteritis Duodenum Figure 1. Pathogenesis of EGIDs screening according to a eosinophils (Fig. 5) (EG+EGE) Antigen ≥30 eos/hpf (n=55) • Subjects with an standardized protocol: • Entry criteria Stomach: ≥30 eos/hpf in 5 hpfs Eosinophil 200 ⁃ 8-10 gastric biopsies ⁃ ≥30 eos/hpf in 5 hpfs Duodenum: ≥30 eos/hpf in 3 hpfs Mast Cell average weekly score of ⁃ 4-6 duodenal biopsies (stomach) and/or ≥30 hpf HEALTHY EGIDs ≥3 for abdominal pain, ⁃ 4-6 esophageal biopsies eos/hpf in 3 hpfs 150 diarrhea and/or nausea (only if subject had a for ≥2 weeks on a PRO (duodenum) Lumen history of EoE or if EoE ⁃ No other known cause RESULTS questionnaire (Fig. 4) features were observed for GI symptoms or 100 Mucosa qualified for an upper during EGD) tissue eosinophilia Table 1. Baseline Characteristics endoscopy (EGD) with biopsy 50 Sub- EG EGE EG+EGE Enrolled mucosa (n=10) (n=25) (n=30) (n=65) Figure 4. Daily Symptom Questionnaire Normal 4-8 Mean Peak Cell Count / / CountCell Peak Mean MC Level Age, Median Years (Range) 39 (18-56) 40 (19-74) 41 (18-68) 40 (18-74) 0 Blood Instructions: Think of the last 24 hours and choose the number that best describes the Vessel worst experience of your EG/EGE symptoms during that time. Figure 8. Data are plotted as box and whisker plots of peak mean cell count/HPF. The box plot represents the Female, n (%) 7 (70%) 16 (64%) 17 (57%) 40 (62%) 25th and 75th percentiles, the internal horizontal line indicates the median and the T-bars indicate the minimum and maximum counts. The shaded gray bar indicates normal mast cell counts in the stomach & small intestine.4-8 • EG and EGE affect 45,000 - 50,000 patients in the US, though Over the past 24 hours, please rate 0 1 2 3 4 5 6 7 8 9 10 Race - White, n (%) 10 (100%) 23 (92%) 27 (90%) 60 (92%) 3 the intensity of your this number may be significantly underestimated History of Atopic/Allergic Diseasea, n (%) 10 (100%) 17 (68%) 23 (77%) 50 (77%) 1. … Abdominal pain • Current treatment options such as diet restriction and No Worst Peripheral Blood Eos (cells/μL), Mean 318 492 978 690 2. … Nausea Table 2. Tissue Eosinophil and Mast Cell Counts corticosteroids have limited efficacy and/or are inappropriate for Symptom Possible 3. … Diarrhea chronic use 4. … Vomiting Stomacha Duodenumb Esophagusc To qualify for a screening endoscopy and biopsy a Medical history at screening of asthma, rhinitis, food allergy, atopic dermatitis, seasonal allergy, environmental allergy, or pollen allergy 5. … Fullness before finishing meal (n=40) (n=55) (n=25) • There is a significant unmet need for novel therapies for this study, patients must have reported ≥3 6. … Loss of appetite average score for at least 2 weeks for abdominal Eos MC Eos MC Eos MC 7. … Abdominal cramping pain and/or nausea and/or diarrhea Figure 2. AK002 Mechanism of Action 8. … Bloating Figure 7. Distribution of Patients by Diagnosis Mean 90 65 64 56 82 48 AK002 9. … Difficulty swallowing (EoE only) Median 77 63 57 53 87 45 Activating Siglec-8 Receptors Figure 5. Counting Eosinophils and Mast Cells SD 56 27 27 20 39 26 Screened Mast Cells Eosinophils n=113 Eosinophils and mast cell counts quantified in (a) 5 peak hpfs (b) 3 peak hpfs (c) 1 peak hpf

Activation Inhibition 1 hpf 1 hpf CONCLUSIONS/DISCUSSION AK002 Mast Cell Inflammatory Mast Cell Inhibition Symptomatic • 65 patients with active EG, EGE, or EG+EGE were enrolled in this Eosinophil Response Eosinophil ADCC/Apoptosis n=88 Phase 2 randomized controlled trial • Siglec-8 is an inhibitory receptor selectively expressed on human • In addition to elevated tissue eosinophils, 64 of 65 (98%) EG/EGE eosinophils and mast cells, and therefore represents a novel ≥30 Eos/hpf patients also had elevated mast cell counts in gastric and/or target for the treatment of EGIDs n=65 duodenal tissue biopsies • AK002 is a novel, humanized, non-fucosylated IgG1 monoclonal • Eosinophils and mast cells were identified by H&E and immunohistochemistry (IHC), • In patients with EoE, mast cells were also elevated in esophageal antibody to Siglec-8 respectively. IHC was performed by staining with a mouse anti-human mast cell tryptase biopsies primary antibody (Clone AA1) followed by incubation with a peroxidase-labelled anti- EG EGE EG+EGE • These data suggest the potential for a pathogenic role for both • Engagement of Siglec-8 receptor by AK002 triggers: mouse polymer secondary antibody and then counterstained with hematoxylin. n=10 (15%) n=25 (38%) n=30 (46%) mast cells and eosinophils in EGIDs and that treatments for ⁃ Antibody dependent cell mediated cytotoxicity (ADCC) against eosinophils (blood) • Mast cells and eosinophils were counted in 5 separate non-overlapping high-power field ⁃ Inhibition of mast cells and apoptosis of tissue eosinophils (tissue) (hpf; area of 0.237 mm2) for each biopsy sample EGIDs may need to target both cell types for optimal effect • ENIGMA, a Phase 2 multi-center, randomized, double-blind, • The patients qualified for study if the average of the peak counts in 5 hpfs in the EoE EoE EoE placebo-controlled study of AK002, represents the largest clinical stomach (or 3 hpf in the duodenum) was ≥ 30 eos/hpf Acknowledgements: • The patients were determined to have elevated mast cells if the average of the peak n=4 (40%) n=6 (24%) n=15 (50%) We thank the patients who participated in this study, investigators, and study staff. trial of patients with EG and EGE This study was funded by Allakos, Inc. counts in 5 hpfs in the stomach (or 3 hpfs in the duodenum) was ≥30 mast cells/hpf References: 1) Caldwell et al. J Allergy Clin Immunol. 2014. 5) Tison et al., J Allergy Clin Immun. 2010 Feb;125(2):AB182 2) Youngblood et al. Gastroenterology. 2019 6) Walker et al., Aliment Pharmacol Ther. 2009 Apr 1;29(7):765-73 3) Jensen ET, Martin CF, Kappelman MD, Dellon ES. J Pediatr Gastroenterol Nutr. 2016 Jan;62(1):36-42. 7) Martinez et al., Gut. 2013 Aug;62(8):1160-8 4) Hahn et al., Am J Surg Pathol. 2007 Nov;31(11):1669-76 8) Doyle et al., Am J Surg Pathol. 2014 Jun;38(6):832-43

Presented at DDW 2019