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

Virginia Chapter ACP Annual Meeting2013

Scott P. ComminsDivision of Allergy and Clinical Immunology

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

I. Background

How oncologists helped uncover a novel food allergy

High incidence of hypersensitivity reactions to cetuximab infusions in mid-Missouri: Association with prior history of atopyR. 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

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.

Regarding cetuximab HSR:

Measurement of IgE ab to Purified Allergens Using Streptavidin Solid Phase

Streptavidin Solid Phase + Biotinylated “Allergen”

“Allergen” of interest (cetuximab) + Biotin

Solid Phase*

*Routine assay technique in parallel with Phadia standard curve gives results in IU/ml.

Then add serum; wash; add labeled anti-IgE; detect chemilum.

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

Cetuximab

* * Site not glycosylated

N-88

N-43

N-299

Qian J, Liu T,…Zhou Q. Analytical Biochem, 2007.

Typical Fc region oligos: core fucosylated

biantennary oligos; no alpha-gal or NGNA

Specificity of the IgE Antibodies that Cross-React with Cetuximab

Chung CH et al. N Engl J Med 2008;358:1109-1117

II. Defining a “new” food allergy

Specificity of the IgE Antibodies That Cross-React with Cetuximab

Chung CH et al. N Engl J Med 2008;358:1109-1117

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

Skin Testing Results: Often <4mm SPT

IntradermalPrick test

Prick test performed using lancette and intradermal testing with 25 gauge needle in the same patient on a single clinic visit.

Patient data, presentation, symptom delay, and laboratory evaluation

Patient Age /Sex Race Symptoms ANA = Anaphylaxis AE = Angioedema U = Urticaria

Time to Reaction (hours)

Alpha-Gal IgE Titer (IU/mL)

Beef IgE Titer (IU/mL)

Total IgE Titer (IU/mL)

E001 44 / M W ANA 6  67.0 3.48 157

E008 80 / F W AE, U 6  0.52 0.35 45.9

E018 26 / M W ANA 5 59.3 16.6 274

E022 74 / F W AE, U 3  9.03 8.90 66.6

E026 47 / M W ANA 6  >100 32.9 1081

E030 56 / M W U 4-6  80.6 15.8  709

E031 55 / F W U 4  68.9 26.2 243

E036 66 / M W U 4-6 21.1  19.2 262

E054 45 / M W ANA 3 2.42 0.35 244

E076 50 / M W U 4  9.25 3.24 247

E115 58 / F W ANA 2-4  >100 47.8 1622

E128 37 / M W U 4-6 26.8 5.65 886

E162 39 / F W AE, U 6 11.1 1.51 30.5

Alph

a-Ga

l

Beef

Pork

Lam

bCh

icke

n

Turk

ey

Fish Ca

t

Fel d

1

Dog

Dust

Mite

Tim

othy

Gra

ss

0.1

1

10

100

Limit ofdetection

▼x1 ♦x1 x3 x12 x11 x12 x1 x7 x1 x9 ♦x4

Foods InhalantsS

peci

fic Ig

E A

bs (I

U/m

L)

r =0.81 r =0.98

CAP-RAST of Specific IgE Abs in Patients with anti-Gal IgE

Asthma(N = 68)

Clinic controls(N = 59)

No cat in home(N = 112)

0.35

0.1

1

10

100

1000

0.35

Alph

a-ga

l spe

cific

IgE

(IU/m

L)

Limit ofdetection

• x 8 • x 13 • x 58 • x 55Number <0.35

Cat in home(N = 96)

Anaphylaxis/ urticaria subjects

Alpha-gal specific IgE is associated with anaphylaxis but not asthma (Figure adapted from Commins, Kelly…Platts-Mills, AJRCCM, 2012)

= Reactions include anaphylaxis

r = 0.60, p<0.001

30% 10%

Sex (% male) 69%

Mean age at presentation(Range)

12 (4-17)

Geometric Mean of Total IgE (95% CI) 147 IU/mL (105-206 IU/mL)

Symptoms at presentationAnaphylaxis* 44%

Gastrointestinal/Oral

64%

Urticaria 92%Angioedema 31%

Average time to symptoms (Range) 4.68 hrs (10 mins to 24 hrs)

Tick exposure 100%

Redness and Itching at site of

Tick bite

87%

Tick borne illness# 10%

Emergency Department (ED) visits 46%**

Medications administered in ED Epinephrine (19%)Antihistamines (35%)Oral Steroids (19%)

Parenteral Steroids (17%)IV Fluids (17%)

Hospital Admissions 8%

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

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

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.

‘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

III. Ticks. Really?

Geographical distribution of cetuximab hypersensitivity reactions

As reported by O’Neil et al, JCO 2007

Distribution of known cases of delayed anaphylaxis to mammalian meat

Dots = single casesSmaller stars = 5 to 24 cases within a stateLarger stars = states with 25 or more cases

Geographical Incidence of Rocky Mountain Spotted Fever

Data from CDC website for 2009; accessed 2/2011

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

Geographical range of Amblyomma americanum population

Data from CDC website; accessed 10/2010

Geographical range of Amblyomma americanum population

Data from CDC website; accessed 9/2012

welcome to the party

Bites from the larvae of Amblyomma or Dermacentor ticks and IgE ab to alpha-Gal.

Test 10/26/06 5/21/07 10/9/07 11/6/07*

Cetuximab(alphaGal) 0.49 16.9 48.3 130

Total IgE 127 199 350 532

•Multiple bites by seed ticks are not uncommon in the Southeast.

Photograph: August 23, 2007.

*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.

*

Subject #1

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

Subject #2

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

* Subject #3

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

1 10 100 1000 100000.1

1

10

100

1000

Total IgE versus IgE to Alpha-Gal

Total IgE (IU / mL)

IgE

to A

lpha

-Gal

(IU

/ m

L)

Report tick or chigger bite(s)

No history obtained for tick bite(s)

Specifically deny tick bite(s)

R= 0.78P <0.001

R amblyommii?

IV. Delayed food allergy

-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

-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

-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

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)

Approximately 8.25 hours after eating prosciutto

Mammalian meat challengeSubject #FC-08: sIgE to alpha-gal = 30.3 IU/mL ; total IgE = 146

Before 3hrs 45min after eating meat

Subject #FC-08: Mammalian meat challenge

Diffuse urticaria appeared at4hrs 15min after eating meat

(left arm shown here)

Mammalian meat challengeSubject #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

UrticariaTime to

symptoms (hrs:min)

Treatment required

Tryptase elevation

Allergics (n=10) 7*/10 3:50 – 6:00 7/10 1**/9

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

Gating StrategyGating Strategy

HLA-

DR/l

inea

ge 1

FIT

C

CD63

APC

CD123 PerCP-Cy5.5

CD 123

% o

f max

BasophilsNegative for • Lineage Cocktail 1 (CD3, CD14, CD16, CD19,CD20, and CD56) • HLA-DR

Positive for • CD123 (IL-3Ra)• FcεRI (IgE receptor)• IgE

FACS Detection of CD63, a marker of Basophil Activation

nuclear membrane

Histamine

CD63-APC

-APC

APC-linked anti-CD63 ab

IL-3 Rα

PerCP-Cy5.5-linked anti-CD123 ab

1 hr0.72%

2 hrs0.54%

3 hrs0.87%

CD63

CD123

In vivo Basophil Activation during Mammalian Meat Challenge

#FC-02

4 hrs - 16.11% 5 hrs - 14.12% 6 hrs - 5.12%

In vivo Basophil Activation during Mammalian Meat Challenge

Baseline1.96%

2 hrs3.08%

4 hrs1.8%

5 hrs8.74%CD63

IgE

-FC07 - sIgE to alpha-gal = 9.3 IU/mL; Total IgE = 204-Whole blood was collected, fixed & stained

Delayed anaphylaxis Urticaria

Time to symptoms (hrs:min)

Treatment required

Allergics 2/10 7/10 3:50 – 6:00 7/10

Controls 0/10 0/10 n/a 0/10

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

Delayed anaphylaxis Urticaria

Time to symptoms (hrs:min)

Treatment required

Basophil activation

Allergics 2/10 6/10 3:50 – 6:00 7/10 8/10

• The answer could be fat. Recall that fat is absorbed differently from protein or carbohydrates in a process that takes 3-4 hrs to enter the blood stream

Why the delay?

Thoracic duct blood stream

The delivery of lipid to the circulation takes time (about 4 hours)

VLDL IDL LDL HDL

=Alpha-gal moiety,as a glycolipid

Further conversion

• The answer could be long-chain triglycerides associated with chylomicrons. Studies suggest LCT enhance dietary antigen absorption into the circulation via chylomicron formation.

• The answer could be the nature of a carbohydrate-based food allergy.

Why the delay?

• The report of delayed reactions occurring 3-6 hours after eating mammalian meat has been confirmed on multiple occasions and at two medical centers

• In subjects with IgE to alpha-gal, basophil activation in vivo coincides with the appearance of clinical symptoms

• Lack of basophil activation before 4 hours suggests that there is delayed appearance of the antigen into the bloodstream

Conclusions

UvaHayley JamesSusan MozzicatoLisa WorkmanAlicia ClarkJudith WoodfolkWalter OliveiraWhitney StevensShama SatinoverLibby KellyPeter HeymannAmy StallingsJulia WisniewskiJosh KennedyAnubha TripathiJonathon Posthumus

ContributorsBarrett LewisMichael LandJoey LaneHeather GütekünstJane PurserGary RakesMadeline DillonTina HatleyLuis MatosSaju Eapen Dane McBrideJoseph BelleauHarold MoessnerMarty FiserEddie Shields