It’s Nothing to Sneeze At – An Update in Allergy

1. It’s Nothing to Sneeze At – An Update in Allergy Virginia Chapter ACP Annual Meeting 2013 Scott P. Commins Division of Allergy and Clinical Immunology

2. Disclosures We are grateful for generous support from the Oakey Food Allergy Fund These studies were carried out with generous support from the NIH-NIAID (K08 AI085190 & R21 AI087985) Disclosure: Up-to-Date card

3. I. Background How oncologists helped uncover a novel food allergy

4. High incidence of hypersensitivity reactions to cetuximab infusions in mid-Missouri: Association with prior history of atopy R. Owera, A. Gill, S. Haddadin, R. Khozouz and M. C. Perry University of Missouri Columbia, Columbia, MO Abstract, 2008 ASCO Annual Meeting Proceedings, Vol 26, No 15S, 2008:20747

5. Regarding cetuximab HSR: The story got stranger as O’Neil talked to more oncologists. He heard that a colleague in Nashville, Tennessee, was finding the same problem. But when O’Neil spoke to oncologists from other areas of the country, they didn’t know what he was talking about. A prominent colorectal oncologist in New York “thought we were lying or crazy,” O’Neil recalls. From “A Mysterious Allergy Afflicts the South” by Sheila Read in Endeavors, 24:2, 2-3, 2008.

6. Solid Phase* Measurement of IgE ab to Purified Allergens Using Streptavidin Solid Phase “Allergen” of interest (cetuximab) + Biotin Streptavidin Solid Phase +Biotinylated“Allergen” Then add serum; wash; add labeled anti-IgE; detect chemilum. *Routine assay technique in parallel with Phadiastandard curve gives results in IU/ml.

7. IgE Antibodies Binding to Cetuximab in Sera from 76 Case Subjects and 462 Controls

8. Cetuximab N-88 * * Site not glycosylated N-43 N-299 Typical Fc region oligos: core fucosylated biantennary oligos; no alpha-gal or NGNA Qian J, Liu T,…Zhou Q. Analytical Biochem, 2007.

9. Specificity of the IgE Antibodies that Cross-React with Cetuximab Chung CH et al. N Engl J Med 2008;358:1109-1117

10. II. Defining a “new” food allergy

11. Specificity of the IgE Antibodies That Cross-React with Cetuximab Chung CH et al. N Engl J Med 2008;358:1109-1117

12. JACI, February 2009 • 24 patients • Virginia & Missouri • Symptoms delayed 3-6 hours after eating mammalian meat • Prick skin test often less than 4mm • Intradermal skin test positive

13. Skin Testing Results: Often <4mm SPT Prick test Intradermal Prick test performed using lancette and intradermal testing with 25 gauge needle in the same patient on a single clinic visit.

14. Patient data, presentation, symptom delay, and laboratory evaluation

15. Foods Inhalants CAP-RAST of Specific IgE Abs in Patients with anti-Gal IgE r =0.81 r =0.98 Specific IgE Abs (IU/mL) Limit of detection ♦x4 ♦x1 ▼x1 x3 x12 x11 x12 x1 x7 x1 x9

16. 1000 100 10 1 0.35 0.35 0.1 Alpha-gal specific IgE is associated with anaphylaxis but not asthma (Figure adapted from Commins, Kelly…Platts-Mills, AJRCCM, 2012) = Reactions include anaphylaxis Alpha-gal specific IgE (IU/mL) Limit of detection • x 8 • x 13 • x 58 • x 55 Number <0.35 Cat in home (N = 96) No cat in home (N = 112) Asthma (N = 68) Clinic controls (N = 59) Anaphylaxis/ urticaria subjects

17. 30% 10% r = 0.60, p<0.001

18. IgE Ab to alpha-gal in kids (n=45)

19. IgE Ab to alpha-gal in kids (n=45)

20. Delayed symptoms induced by oral exposure to mammalian proteins in patients with IgE ab to alpha-gal. • Despite high titer IgE antibodies that bind epitopes on a range of mammalian proteins including beef, pork and lamb: • No awareness of immediate response in the mouth (i.e. no lip swelling or tingling). • Anaphylaxis occurring after eating beef starts with skin itching 3-6 hours later. • Skin prick tests generally <4mm in diameter while intradermals often ≥8mm.

21. ‘Delayed’ food allergy? Food allergy symptoms classically occur immediately (OAS), up to 2 hours after eating No mention of ‘delayed’ symptoms that can be attributed to food in common allergy texts

22. III. Ticks. Really?

23. Geographical distribution of cetuximab hypersensitivity reactions As reported by O’Neil et al, JCO 2007

24. Distribution of known cases of delayed anaphylaxis to mammalian meat Dots = single cases Smaller stars = 5 to 24 cases within a state Larger stars = states with 25 or more cases

25. Geographical Incidence of Rocky Mountain Spotted Fever Data from CDC website for 2009; accessed 2/2011

26. Original image: CDC; modified by D.N. Gaines for Virginia Department of Health

27. Geographical range of Amblyomma americanum population Data from CDC website; accessed 10/2010

28. Geographical range of Amblyomma americanum population welcome to the party Data from CDC website; accessed 9/2012

29. Bites from the larvae of Amblyomma or Dermacentorticks and IgE ab to alpha-Gal. • Multiple bites by seed ticks are not uncommon in the Southeast. *Titers:Total 262, Beef 19.2 Pork 10.1,Cat 17.5,Dog 19.8, IU/ml. Chicken, Turkey, Roach, Dust Mite and Ragweed all <0.35 IU/ml. Photograph: August 23, 2007.

30. Time course of IgE antibodies to alpha-gal and total IgE following episodes of multiple tick bites (red arrows) Subject #1 *

31. Time course of IgE antibodies to alpha-gal and total IgE following episodes of multiple tick bites (red arrows) Subject #2

32. Time course of IgE antibodies to alpha-gal and total IgE following episodes of multiple tick bites (red arrows) * Subject #3

33. Total IgE versus IgE to Alpha-Gal Report tick or chigger bite(s) No history obtained for tick bite(s) Specifically deny tick bite(s) 1000 IgE to Alpha-Gal (IU / mL) R= 0.78 P <0.001 100 10 1 0.1 1 10 100 1000 10000 Total IgE (IU / mL)

34. R amblyommii?

35. IV. Delayed food allergy

37. -FC02 - sIgE to alpha-gal = 22.4 IU/mL; Total IgE = 109 -4:22 after consuming meat, symptoms began as itching & hive on hand; second hive appeared on arm -Given AH and observed

38. -FC02 - sIgE to alpha-gal = 17.6 IU/mL; Total IgE = 184 -Whole blood was collected, fixed & stained -3:54 after consuming meat, symptoms began as itching, flushing and progressed: ultimately requiring epinephrine

39. -Subject #FC-07: sIgE to alpha-gal = 9.3 IU/mL; Total IgE = 204 -Whole blood was collected, fixed & stained -6 hours after consuming 56g of pork prosciutto, subject released with (disappointing) mild itch and single hive

40. Subject calls from car 30 minutes after being released to report progression of itching and “warmth to skin” Initial picture upon arriving home (approximately 7.5 hours after eating pork prosciutto)

41. Approximately 8.25 hours after eating prosciutto

42. Mammalian meat challenge Subject #FC-08: sIgE to alpha-gal = 30.3 IU/mL ; total IgE = 146 Before 3hrs 45min after eating meat

43. Subject #FC-08: Mammalian meat challenge Diffuse urticaria appeared at 4hrs 15min after eating meat (left arm shown here)

44. Mammalian meat challenge Subject #FC-10: sIgE to alpha-gal = 29.1 IU/mL ; total IgE = 201 4hrs 25min after eating mammalian meat: pruritus followed by urticaria on right flank

45. Summary of Mammalian Meat Challenge Results in Subjects with IgE to alpha-gal • *Two subjects received epinephrine (i.e., these reactions are probably better defined as delayed anaphylaxis to mammalian meat) • **Tryptase: • -Subject FC-08: 4.2ng/mL (baseline) → 18.2ng/mL (4 hrs) → 20.1 ng/mL (5 hrs) • -Subject FC-10: 4.9 (baseline) → 6.2 (4 hrs) → 9.9 (5 hrs) → 10.7 (6 hrs) • -No tryptase measurements were performed in subject FC-06

46. Gating Strategy Gating Strategy HLA-DR/lineage 1 FITC CD63 APC CD123 PerCP-Cy5.5 • Basophils • Negative for • Lineage Cocktail 1 (CD3, CD14, CD16, CD19,CD20, and CD56) • HLA-DR • Positive for • CD123 (IL-3Ra) • FcεRI (IgE receptor) • IgE % of max CD 123

47. FACS Detection of CD63, a marker of Basophil Activation PerCP-Cy5.5-linked anti-CD123 ab APC-linked anti-CD63 ab -APC CD63 IL-3 Rα -APC Histamine nuclear membrane

48. In vivo Basophil Activation during Mammalian Meat Challenge 3 hrs 0.87% 1 hr 0.72% 2 hrs 0.54% CD63 CD123 4 hrs - 16.11% 6 hrs - 5.12% 5 hrs - 14.12% #FC-02

49. In vivo Basophil Activation during Mammalian Meat Challenge Baseline 1.96% 2 hrs 3.08% 4 hrs 1.8% 5 hrs 8.74% -FC07 - sIgE to alpha-gal = 9.3 IU/mL; Total IgE = 204 -Whole blood was collected, fixed & stained CD63 IgE

50. Summary of Mammalian Meat Challenge Results in Subjects with or without IgE to alpha-gal