Molecular-based allergy
diagnostics
Suparat Sirivimonpan, MD.
18/10/2013
A WAO - ARIA - GA2LEN consensus
document on molecular-based allergy
diagnostics

World Allergy Organization Journal 2013, ...
Topic outline
Introduction
 Definitions and concepts
 Clinical application








Increase accuracy and resolve cr...
Diagnosis
•

Clinical : History, Physical examination

•

Allergic test
•

•

Skin prick test
Blood test

crude extracts

...
Currently available tests detect only sensitization,
not clinical allergy, cannot predict prognosis and severity

Nature
A...
Introduction
Molecular allergy (MA) diagnostics is increasingly entering
routine care
•

More than 130 allergenic molecule...
Introduction
WAO - ARIA - GA2LEN consensus document :
practical guide for the indication, determination, and
interpretatio...
Definitions and concepts
Definitions and concepts
•

Allergen extract
•
•

A crude, unfractionated mixture of allergenic and nonallergenic proteins...
Allergen component
Almost anything containing proteins can be an allergen
source
- each allergen source contains many diff...
Definitions and concepts
Stability of allergens
•Acid
•

pH :

susceptible to acid pH (peptic digestion)  not able to cro...
Allergen component

Protein stability
- allergen that stable to heat and digestion are more
likely to cause severe reactio...
Definitions and concepts
•

Primary allergen :
•

original sensitizing molecule (i.e., driving trigger)

•Secondary
•

sen...
Definitions and concepts
Cross-reactivity
•

phenomenon of an IgE antibody recognizing, binding,
and inducing an immune re...
Definitions and concepts
Cross-reactivity
•

IgE cross-reactivity often occurs between the following:
a) Allergenic molecu...
Definitions and concepts
Co-sensitization
Genuine sensitization to more than one allergen
source (e.g., timothy grass and ...
Definitions and concepts
Co-sensitization
Genuine sensitization to more than one allergen
source (e.g., timothy grass and ...
Definitions and concepts
sIgE sensitization
presence of allergen-specific IgE (sIgE) antibodies in the blood
• presence or...
Definitions and concepts
sIgE detection based on allergen extracts
Singleplex or multiplex platforms
• such as CAP, radioa...
Systematic allergen nomenclature
•World

Health Organization and International Union of
Immunological Species (WHO/IUIS)
•...
Clinical application
Clinical application
increased accuracy in allergy diagnosis and prognosis
• plays an important role in three key aspects ...
Increase accuracy and resolve cross-reactivity
Allergic individuals may produce IgE antibodies to allergens that
are eithe...
•

World Allergy Organization Journal 2013, 6:17
World Allergy Organization Journal 2013, 6:17
S ta b le

World Allergy Organization Journal 2013, 6:17
S ta b le

World Allergy Organization Journal 2013, 6:17
L a b ile

World Allergy Organization Journal 2013, 6:17
L a b ile

World Allergy Organization Journal 2013, 6:17
Increase accuracy and resolve cross-reactivity
While, MA studies have not fully elucidated the
underlying mechanism govern...
Food

World Allergy Organization Journal 2013, 6:17
Food

World Allergy Organization Journal 2013, 6:17
Food

World Allergy Organization Journal 2013, 6:17
A e ro a lle rg e n s

World Allergy Organization Journal 2013, 6:17
A e ro a lle rg e n s

World Allergy Organization Journal 2013, 6:17
A e ro a lle rg e n s

World Allergy Organization Journal 2013, 6:17
O th e r alle rg e n s

World Allergy Organization Journal 2013, 6:17
Increase accuracy and resolve cross-reactivity
Clinical uses of MA :
• identify the offending allergenic molecule
• distin...
Increase accuracy and resolve cross-reactivity
In the field of pollen-related food allergy : increase accuracy of
diagnosi...
Increase accuracy and resolve cross-reactivity
•

In patients sensitized to different pollen species
• detecting new relev...
Increase accuracy and resolve cross-reactivity
In conventional SPTs, some allergens can be poorly represented
in extracts ...
Increase accuracy and resolve cross-reactivity
All allergy diagnostics, including MA, should be evaluated within
framework...
Assess the risk and type of reaction
Ability to improve risk assessment, particularly for food allergies
• Different foods...
Assess the risk and type of reaction
Information may only be applicable to the specific population which
has been studied
...
Food allergens
Allergens resistant to heat and digestion often trigger more severe
allergic reactions

Storage protein, LT...
Food allergens
In addition, the amount of a molecule present in a food
source is also a parameter to take into considerati...
Food allergens

Peanut
Ara h1,2,3 (storage proteins) genuine peanut reaction
• Ara h 8 (PR-10,Bet v 1-homologue)  milder...
Food allergens

Soy
•

Gly m 5 and/or Gly m 6  severe reactions

•

Gly m 4 (PR-10)  OAS
•

•

In birch pollen-allergic ...
Food allergens

Hazelnut
•

Cor a 1 (PR-10)  local reactions like OAS

•

Cor a 8 (LTP) , storage proteins (Cor a 9,Cor a...
Food allergens

Walnut
•

Storage protein (Jug r 1, Jug r 2) or LTP (Jug r 3)
 severe reactions

Walnut allergens have no...
Food allergens

Wheat
•

ω-5-gliadin (Tri a 19)
 immediate allergic reactions in children
 systemic exercise-induced rea...
Food allergens

Rosaceae fruits
Apple, peach, Appricot, Plum
• PR-10 proteins (Mal d 1, Pru p 1) or profilins (Pru p 4)
•
...
Food allergens

Egg
High levels of sIgE antibodies to ovomucoid (Gal d 1)
 risk factor for persistent egg allergy, includ...
Food allergens

Milk
Casein (Bos d 8) and beta-lactoglobulin (Bos d 5) sIgE
antibodies
 persistent allergy to milk, inclu...
Food allergens

Fish
•

Parvalbumins (e.g., Gad c1 and Cyp c 1)
• major allergens in fish
• typically stable to heat and d...
Food allergens

Shellfish
Tropomyosin
- high degree of cross-reactivity across a wide
variety of species, including mites
...
Food allergens

Meat allergy
Galactose-α-1,3-galactose (α-Gal)
sugar structure found on glycoproteins and glycolipids of n...
Food allergens

Meat allergy
α-Gal : present on the chimeric antibody cetuximab
• Patients sensitized to α-Gal may react w...
Inhalants

Pet dander
Fel d 1 : asthma in cat-allergic individuals
• Lipocalins (Mus m 1, Equ c 1, Fel d 4, Can f 1, 2), k...
Inhalants

Pollen
Research in pollen allergy has focused on distinguishing genuine
allergens from those that are cross-rea...
Inhalants

Pollen
•

Profilin sensitization
•
•
•

common among pollen allergic patients
associated with mild or no clinic...
Inhalants

Mites
No specific sensitization profile has been described as a
risk factor for lower airway disease or disease...
Inhalants

Molds
Aspergillus fumigatus
Asp f 2, 4, and 6 may suggest ABPA
• Asp f 1 and/or Asp f 3 may be more indicative ...
Inhalants

Cockroach
Per a 2 correlates with severity of airway allergy in cockroachallergic patients
• Per a 2 is current...
Other allergens

Latex
•

Sensitization to Hev b 8 (profilin)
•

seems to be clinically irrelevant

•

not related to clin...
Other allergens

Hymenoptera venoms
Most hymenoptera venom allergens possess CCDs
 responsible for a portion of clinicall...
Specific immunotherapy (SIT)
Correct diagnosis, selection of truly eligible patients, and
identification of primary sensit...
Specific immunotherapy
Mixing numerous allergens appears to achieve good clinical
efficacy
•

•

there may be an inability...
Grass, Pollen

In the example mentioned above a patient with sIgE antibodies against Phl p 1 and Phl p 5 but no sIgE to Be...
HDM

• if a patient is sensitized to a traditional house dust mite extract

but their IgE antibodies against Der p 10 (tro...
Hymenoptera venom

Sensitization to the major allergens Api m 1 of honeybee and Ves v 5 and/or
Ves v 1 of yellow jackets m...
Specific immunotherapy
Most commercial allergen extracts used in SIT are well
standardized for major allergens
• contain o...
Specific immunotherapy
Theoretically, identification of molecules  tailored SIT??
• In practice, this does not seem feasi...
Micro-array technology
Micro-array technology
MA diagnostics has been available on singleplex
platforms
• ImmunoCAP, ImmuLite, and HyTech platfor...
ImmunoCAP Immuno-Solid Phase Allergen Chip
(ISAC) (Phadia AB)

Fluorescence is measured with a laser scanner and results a...
ISAC
Using a standard calibration curve, results are reported within a
range of 0.3 - 100 ISAC Standardized Units (ISU-E)
...
ISAC
•

ImmunoCAP
• results : kU/L
• not interchangeable
• correlate well

ImmunoCAP technology measures IgE binding under...
When comparing ISAC with other sIgE measuring assays, concordance of results vary
between allergens tested
World Allergy O...
ISAC
Results of the ISAC assay are reproducible at a level that is
generally accepted and agreed upon
•

•

Special attent...
ISAC
ISAC is generally not recommended for monitoring quantitative
IgE levels over time in clinical routine
•

•Because

t...
Micro-array technology
ISAC results for some allergen sources such as cashew nut,
sesame, dog, mugwort, and ragweed can be...
•

World Allergy Organization Journal 2013, 6:17
Patients most likely to benefit from MA diagnostics
Patients who are poly-sensitized, have an unclear symptom
and/or sensi...
Molecular-based allergy diagnostics
• MA diagnostics may offer additional information for
- early diagnosis of allergies
-...
Unmet needs!
1) Large-scale, population-based multicenter studies : categories of
patients
2) Large studies that include w...
Unmet needs!
4) Identification and clinical evaluation of the most relevant
allergens have to be further investigated in m...
• International

guidelines recommendation

1st Step : a thorough clinical case history

2nd Step : allergen extract-based...
Conclusions
MA diagnostics is increasingly entering routine care
and can improve management of allergic patients
•

Benefi...
Conclusions
Sensitivity compared to ImmunoCAP is still often lower
• ISAC can have high clinical relevance by detecting
se...
Conclusions

Disadvantage
Interpretation of such sensitization in clinically
unresponsive patients is difficult or even im...
Conclusions
Although MA diagnostics is a complex area, it provides
novel and relevant information for the allergist and wi...
Thank you
Molecular based allergic diagnosis
Upcoming SlideShare
Loading in...5
×

Molecular based allergic diagnosis

2,000

Published on

Molecular based allergic diagnosis

Presented by Suparat Sirivimonpan, MD.

October 18, 2013

Published in: Health & Medicine, Technology
0 Comments
1 Like
Statistics
Notes
  • Be the first to comment

No Downloads
Views
Total Views
2,000
On Slideshare
0
From Embeds
0
Number of Embeds
1
Actions
Shares
0
Downloads
176
Comments
0
Likes
1
Embeds 0
No embeds

No notes for slide

Transcript of "Molecular based allergic diagnosis"

  1. 1. Molecular-based allergy diagnostics Suparat Sirivimonpan, MD. 18/10/2013
  2. 2. A WAO - ARIA - GA2LEN consensus document on molecular-based allergy diagnostics World Allergy Organization Journal 2013, 6:17
  3. 3. Topic outline Introduction  Definitions and concepts  Clinical application      Increase accuracy and resolve cross-reactivity Assess the risk and type of reaction Specific immunotherapy Micro-array technology
  4. 4. Diagnosis • Clinical : History, Physical examination • Allergic test • • Skin prick test Blood test crude extracts • Specific IgE • Component resolve diagnosis (CRD)
  5. 5. Currently available tests detect only sensitization, not clinical allergy, cannot predict prognosis and severity Nature Allergenic source contain a mixture of allergenic and non-allergenic molecules
  6. 6. Introduction Molecular allergy (MA) diagnostics is increasingly entering routine care • More than 130 allergenic molecules are commercially available for in vitro sIgE testing • Clinicians and immunologists specifically trained in allergology must keep abreast of the new and rapidly evolving evidence available for MA diagnostics • World Allergy Organization Journal 2013, 6:17
  7. 7. Introduction WAO - ARIA - GA2LEN consensus document : practical guide for the indication, determination, and interpretation of MA diagnostics aimed for clinicians specifically trained in allergology • World Allergy Organization Journal 2013, 6:17
  8. 8. Definitions and concepts
  9. 9. Definitions and concepts • Allergen extract • • A crude, unfractionated mixture of allergenic and nonallergenic proteins, polysaccharides, and lipids extraction from an allergen source •Allergenic • • molecule (allergen component) A molecule (i.e., protein or glycoprotein) bind sIgE antibodies  “allergen” Allergens can be isolated from • natural allergen sources (native, purified allergen) • recombinant DNA technology (recombinant allergen) World Allergy Organization Journal 2013, 6:17
  10. 10. Allergen component Almost anything containing proteins can be an allergen source - each allergen source contains many different allergenic protein (allergen component) - each allergen component commonly has several different epitopes - epitope is the actual three-dimensional binding site for an antibody Clinical and Experimental Allergy;2010 ,40: 896-904
  11. 11. Definitions and concepts Stability of allergens •Acid • pH : susceptible to acid pH (peptic digestion)  not able to cross gastric barrier (except possibly in patients treated with antacid drugs) •Temperature • (cooking or boiling) : Temperature susceptibility  does not maintain its allergenicity after cooking/heating procedures World Allergy Organization Journal 2013, 6:17
  12. 12. Allergen component Protein stability - allergen that stable to heat and digestion are more likely to cause severe reaction - allergen that heat and digestion labile are more likely to be tolerated or only cause mild/local symptom Clinical and Experimental Allergy;2010 ,40: 896-904
  13. 13. Definitions and concepts • Primary allergen : • original sensitizing molecule (i.e., driving trigger) •Secondary • sensitization : cross-reactivity World Allergy Organization Journal 2013, 6:17
  14. 14. Definitions and concepts Cross-reactivity • phenomenon of an IgE antibody recognizing, binding, and inducing an immune response to similar allergenic molecules (homologues) present in different species (reacts to both Bet v 1 in birch pollen and Cor a 1 in hazelnut  PR10) • ≥ 50-70% sequence homology between primary structures of proteins World Allergy Organization Journal 2013, 6:17
  15. 15. Definitions and concepts Cross-reactivity • IgE cross-reactivity often occurs between the following: a) Allergenic molecules in closely related species b) Well preserved molecules with similar function present in widely different species that belong to the same protein family (tropomyosin : Der p 10 in house dust mite ,Pen m 1 in black tiger shrimp) World Allergy Organization Journal 2013, 6:17
  16. 16. Definitions and concepts Co-sensitization Genuine sensitization to more than one allergen source (e.g., timothy grass and birch) • sensitization is not due to cross-reactivity • World Allergy Organization Journal 2013, 6:17
  17. 17. Definitions and concepts Co-sensitization Genuine sensitization to more than one allergen source (e.g., timothy grass and birch) • where the sensitization is not due to cross-reactivity • World Allergy Organization Journal 2013, 6:17
  18. 18. Definitions and concepts sIgE sensitization presence of allergen-specific IgE (sIgE) antibodies in the blood • presence or absence of clinical symptoms • a) Mono-sensitization: - Sensitization to one allergen source (D. pteronyssinus) or to a closely related taxonomical family or group of allergen sources (i.e., mites) b) Poly- (or multi-) sensitization: - Sensitization to three or more allergen sources (e.g., mite, birch, and grass pollen). World Allergy Organization Journal 2013, 6:17
  19. 19. Definitions and concepts sIgE detection based on allergen extracts Singleplex or multiplex platforms • such as CAP, radioallergosorbent test (RAST) • interchangeably • • sIgE detection based on allergenic molecules Singleplex or multiplex platforms • identify cross-reacting molecules • World Allergy Organization Journal 2013, 6:17
  20. 20. Systematic allergen nomenclature •World Health Organization and International Union of Immunological Species (WHO/IUIS) • Using their Latin family name (genus and species) • Arachis hypogaea : Ara h (peanut) • A number : distinguish various allergens from the same species (e.g., Ara h 1, Ara h 2, etc.) • The numbers are assigned to the allergens in the order of their identification • Allergenic molecules are classified into protein families, according to their structure and biological function •www.allergen.org World Allergy Organization Journal 2013, 6:17 Clinical and Experimental Allergy;2010 ,40: 896-904
  21. 21. Clinical application
  22. 22. Clinical application increased accuracy in allergy diagnosis and prognosis • plays an important role in three key aspects of allergy diagnosis: • (1) Increase accuracy and resolve cross-reactivity (2) Assess the risk and type of reaction (3) Specific immunotherapy (patients and triggering allergens) World Allergy Organization Journal 2013, 6:17
  23. 23. Increase accuracy and resolve cross-reactivity Allergic individuals may produce IgE antibodies to allergens that are either unique to a single species or common to many • Proteins within the same family share common epitopes  same IgE antibody can bind to similar structures present in allergens from different allergen sources • These cross-reactive allergens give valuable information on potential sensitization and clinical reactions to several different sources • World Allergy Organization Journal 2013, 6:17
  24. 24. • World Allergy Organization Journal 2013, 6:17
  25. 25. World Allergy Organization Journal 2013, 6:17
  26. 26. S ta b le World Allergy Organization Journal 2013, 6:17
  27. 27. S ta b le World Allergy Organization Journal 2013, 6:17
  28. 28. L a b ile World Allergy Organization Journal 2013, 6:17
  29. 29. L a b ile World Allergy Organization Journal 2013, 6:17
  30. 30. Increase accuracy and resolve cross-reactivity While, MA studies have not fully elucidated the underlying mechanism governing cross-reactivity and symptom presentation  analysis of the epitopes of key allergens will provide insight into this issue • Currently, a large number of purified or recombinant allergens are commercially available • World Allergy Organization Journal 2013, 6:17
  31. 31. Food World Allergy Organization Journal 2013, 6:17
  32. 32. Food World Allergy Organization Journal 2013, 6:17
  33. 33. Food World Allergy Organization Journal 2013, 6:17
  34. 34. A e ro a lle rg e n s World Allergy Organization Journal 2013, 6:17
  35. 35. A e ro a lle rg e n s World Allergy Organization Journal 2013, 6:17
  36. 36. A e ro a lle rg e n s World Allergy Organization Journal 2013, 6:17
  37. 37. O th e r alle rg e n s World Allergy Organization Journal 2013, 6:17
  38. 38. Increase accuracy and resolve cross-reactivity Clinical uses of MA : • identify the offending allergenic molecule • distinguish specific molecules from markers of crossreactivity World Allergy Organization Journal 2013, 6:17
  39. 39. Increase accuracy and resolve cross-reactivity In the field of pollen-related food allergy : increase accuracy of diagnosis • * peanut allergic patients, - Ara h 2  genuine marker,systemic reactions -Ara h 8  cross-reactivity among food allergens ,Fagales tree pollen  mild, oral reactions •measuring IgE responses to certain food allergens may reduce the need for food challenges • World Allergy Organization Journal 2013, 6:17
  40. 40. Increase accuracy and resolve cross-reactivity • In patients sensitized to different pollen species • detecting new relevant sensitizations • by ruling out clinically irrelevant sensitizations caused by nonsymptomatic cross-reactive allergens • For example, MA diagnostics can help to distinguish baker’s asthma from pollen or wheat allergy World Allergy Organization Journal 2013, 6:17
  41. 41. Increase accuracy and resolve cross-reactivity In conventional SPTs, some allergens can be poorly represented in extracts because of the biological variability of allergen source • * Dog allergic patients • • • Can f 5  prostate-derived allergen produced by male dogs  38% of dog allergic patients Allergen extracts used in skin tests : dog hair  skin tests routinely fail to identify patients’ sensitivity to Can f 5 (low concentration in dog hair) IgE response to Can f 5 using MA diagnostics may enhance accuracy of dog allergy diagnosis World Allergy Organization Journal 2013, 6:17
  42. 42. Increase accuracy and resolve cross-reactivity All allergy diagnostics, including MA, should be evaluated within framework of a patient’s clinical history • • IgE sensitization  not necessarily imply clinical responsiveness This is of particular importance, since allergic patients respond in individualized manner to exposure to allergens from various sources •i.e., every individual produces their own unique IgE antibody profile at molecular level • World Allergy Organization Journal 2013, 6:17
  43. 43. Assess the risk and type of reaction Ability to improve risk assessment, particularly for food allergies • Different foods contain unique allergenic molecules that are stable or labile to heat & digestion • stability of a molecule & patient’s clinical history : risk of systemic versus local reactions • Labile  local reactions (typically oral symptoms) and cooked food is often tolerated • Stable allergens  systemic reactions • MA diagnostics may decrease the need for provocation testing and improve recommendations for allergen avoidance • World Allergy Organization Journal 2013, 6:17
  44. 44. Assess the risk and type of reaction Information may only be applicable to the specific population which has been studied • Disease expression differ according to local exposures patterns characteristic of the geographical region • ex. Peanut (USA, Spain, Sweden) • World Allergy Organization Journal 2013, 6:17
  45. 45. Food allergens Allergens resistant to heat and digestion often trigger more severe allergic reactions Storage protein, LTP World Allergy Organization Journal 2013, 6:17
  46. 46. Food allergens In addition, the amount of a molecule present in a food source is also a parameter to take into consideration • World Allergy Organization Journal 2013, 6:17
  47. 47. Food allergens Peanut Ara h1,2,3 (storage proteins) genuine peanut reaction • Ara h 8 (PR-10,Bet v 1-homologue)  milder or local symptoms • • LTP (Ara h 9)  southern Europe, prevalent sensitizing allergen  systemic and more severe reactions • • Further studies in other geographical regions Profilin or CCD sensitization  no or local oral symptoms  heated peanuts may be tolerated World Allergy Organization Journal 2013, 6:17
  48. 48. Food allergens Soy • Gly m 5 and/or Gly m 6  severe reactions • Gly m 4 (PR-10)  OAS • • In birch pollen-allergic individuals, the combination of Gly m 4 sensitivity and intake of large amounts of mildly processed soy, such as soy drinks, can induce a severe reaction Profilin or CCD  no, or local oral symptoms  heated soy may be tolerated World Allergy Organization Journal 2013, 6:17
  49. 49. Food allergens Hazelnut • Cor a 1 (PR-10)  local reactions like OAS • Cor a 8 (LTP) , storage proteins (Cor a 9,Cor a 14)  severe symptoms • Profilin (Cor a 2) or CCD alone  no or local oral symptoms  heated hazelnuts may be tolerated. World Allergy Organization Journal 2013, 6:17
  50. 50. Food allergens Walnut • Storage protein (Jug r 1, Jug r 2) or LTP (Jug r 3)  severe reactions Walnut allergens have not been available on the market until recently, as is reflected by the lack of recent clinical studies • • Profilin or CCD alone  no, or local oral, symptoms  heated walnut may be tolerated World Allergy Organization Journal 2013, 6:17
  51. 51. Food allergens Wheat • ω-5-gliadin (Tri a 19)  immediate allergic reactions in children  systemic exercise-induced reactions in adults • LTP (Tri a 14)  some degree of cross-reactivity with other food LTPs  more knowledge is needed about its prevalence and clinical implication • Profilin or CCD  no, or local oral, symptoms  heated wheat may be tolerated World Allergy Organization Journal 2013, 6:17
  52. 52. Food allergens Rosaceae fruits Apple, peach, Appricot, Plum • PR-10 proteins (Mal d 1, Pru p 1) or profilins (Pru p 4) •  local, oral reactions (sensitive to heat & digestion) • LTP (Pru p 3)  typical of the Mediterranean area  wide range of clinical expressions (from asymptomatic to anaphylaxis)  risk marker for severe reactions including cofactor (e.g., exercise, alcohol or drugs) dependent anaphylaxis World Allergy Organization Journal 2013, 6:17
  53. 53. Food allergens Egg High levels of sIgE antibodies to ovomucoid (Gal d 1)  risk factor for persistent egg allergy, including reactions to cooked/heated egg • Undetectable levels indicate tolerance to cooked egg • World Allergy Organization Journal 2013, 6:17
  54. 54. Food allergens Milk Casein (Bos d 8) and beta-lactoglobulin (Bos d 5) sIgE antibodies  persistent allergy to milk, including heated milk • undetectable levels indicate tolerance to baked milk products • World Allergy Organization Journal 2013, 6:17
  55. 55. Food allergens Fish • Parvalbumins (e.g., Gad c1 and Cyp c 1) • major allergens in fish • typically stable to heat and digestion • high degree of cross-reactivity whereby patients sensitized to one parvalbumin may also react to parvalbumins from other fish, including carp, cod, herring, plaice, mackerel, tuna, salmon, perch, and eel World Allergy Organization Journal 2013, 6:17
  56. 56. Food allergens Shellfish Tropomyosin - high degree of cross-reactivity across a wide variety of species, including mites • Shrimp and other shellfish also contain other clinically relevant allergens - sarcoplasmic calcium-binding protein - arginine kinases • World Allergy Organization Journal 2013, 6:17
  57. 57. Food allergens Meat allergy Galactose-α-1,3-galactose (α-Gal) sugar structure found on glycoproteins and glycolipids of nonprimate mammals and new world monkeys, but not on humans • It is assumed that sensitization to α-Gal can be induced by tick bites or certain parasite infections • α-Gal– sensitized patients  delayed immediate type reactions to red meat (beef, pork, goat, deer) • Anti-α-Gal-IgE  severe allergic symptoms and with delayed-type anaphylaxis • World Allergy Organization Journal 2013, 6:17
  58. 58. Food allergens Meat allergy α-Gal : present on the chimeric antibody cetuximab • Patients sensitized to α-Gal may react with anaphylactic reaction after the administration of cetuximab • Testing for α-Gal before administration of cetuximab • • Bovine serum albumin (e.g. Bos d 6) • • • heat labile allergen present both in milk and beef may cause cross-reactivity between different mammalian meat World Allergy Organization Journal 2013, 6:17
  59. 59. Inhalants Pet dander Fel d 1 : asthma in cat-allergic individuals • Lipocalins (Mus m 1, Equ c 1, Fel d 4, Can f 1, 2), kallikrein (Can f 5), and secretoglobin (Fel d 1) : severe asthma in Swedish children • • More knowledge is needed in the area of pet allergy • many of the patients are poly-sensitized to several pets • clinical history is often inconclusive • cross-reactivities between e.g. cat, dog and horse is not fully clarified at the MA level World Allergy Organization Journal 2013, 6:17
  60. 60. Inhalants Pollen Research in pollen allergy has focused on distinguishing genuine allergens from those that are cross-reactive • Little is known regarding specific markers of severe reactions • • Some sensitivities to specific allergens (ex.Ole e 9 ,LTP Ole e 7) • may be markers of more severe symptoms in pollen allergy • increasing the risk of systemic reactions during immunotherapy World Allergy Organization Journal 2013, 6:17
  61. 61. Inhalants Pollen • Profilin sensitization • • • common among pollen allergic patients associated with mild or no clinical symptoms minority of patients: risk factor for more severe reactions in olive pollen-allergic individuals and in patients allergic to certain plant foods like melon or citrus World Allergy Organization Journal 2013, 6:17
  62. 62. Inhalants Mites No specific sensitization profile has been described as a risk factor for lower airway disease or disease severity • Higher sIgE/IgG4 ratio for Der p 2 has been associated with asthma • • Der p 10 (tropomyosin) • • minor allergen in mite-allergic patients risk for allergic reactions to shellfish or snail, which can be severe World Allergy Organization Journal 2013, 6:17
  63. 63. Inhalants Molds Aspergillus fumigatus Asp f 2, 4, and 6 may suggest ABPA • Asp f 1 and/or Asp f 3 may be more indicative of allergic asthma • These associations must still be confirmed in other patient populations • World Allergy Organization Journal 2013, 6:17
  64. 64. Inhalants Cockroach Per a 2 correlates with severity of airway allergy in cockroachallergic patients • Per a 2 is currently not commercially available for in vitro testing • Per a 2 homologue Bla g 2 is available • Cross-reactive tropomyosin (Bla g 7)  risk for allergic reactions to shellfish or snail, which can be severe • World Allergy Organization Journal 2013, 6:17
  65. 65. Other allergens Latex • Sensitization to Hev b 8 (profilin) • seems to be clinically irrelevant • not related to clinical latex reactions The other latex allergens are linked to clinical reactions • No association between allergens and severity of reactions • The cross-reactive allergen responsible for the so called latex-fruit syndrome are not fully clarified • • although data indicate that Hev b 5, 6 and 11 play a role World Allergy Organization Journal 2013, 6:17
  66. 66. Other allergens Hymenoptera venoms Most hymenoptera venom allergens possess CCDs  responsible for a portion of clinically irrelevant IgE antibody cross-reactivity between bee and wasp venom • Detection of recombinant venom allergens can discriminate between genuine venom sensitization and cross-reactivity due to CCDs in patients with double-positive IgE results from traditional venom tests that are based on allergen extract • World Allergy Organization Journal 2013, 6:17
  67. 67. Specific immunotherapy (SIT) Correct diagnosis, selection of truly eligible patients, and identification of primary sensitizing allergen(s) are important for optimal and cost-effective patient management • In some patients, a detailed clinical history and traditional extractbased IgE testing (SPT and/or in vitro sIgE) is sufficient to identify the relevant allergen(s) • Patient demonstrates polysensitization : traditional diagnostic tests based on allergen extracts and their clinical history is not sufficient to clarify the nature of the sensitization • may occur in a relatively high proportion of patients • World Allergy Organization Journal 2013, 6:17
  68. 68. Specific immunotherapy Mixing numerous allergens appears to achieve good clinical efficacy • • there may be an inability to identify the responsible allergen in the case of adverse events Certain proteins (e.g., profilins, polcalcins, LTPs, PR10, tropomyosins) are highly conserved in a wide variety of species • The use of recombinant/purified allergens would allow for the discrimination between genuine sensitizations and cross reactivities • World Allergy Organization Journal 2013, 6:17
  69. 69. Grass, Pollen In the example mentioned above a patient with sIgE antibodies against Phl p 1 and Phl p 5 but no sIgE to Bet v 1 is truly sensitized to grass If sIgE antibodies to Phl p 12 (profilin) were also detected, profilin sensitization would probably be responsible for the positive SPT result obtained with birch extract, which contains profilin as well. SIT would be prescribed for grass only World Allergy Organization Journal 2013, 6:17
  70. 70. HDM • if a patient is sensitized to a traditional house dust mite extract but their IgE antibodies against Der p 10 (tropomyosin) and not to Der p 1, 2/ Der f 1, 2 • SIT for mites should not be given, because mite extracts mainly contain Der p 1, 2/Der f 1, 2 and have variable or low amounts of Der p 10 • World Allergy Organization Journal 2013, 6:17
  71. 71. Hymenoptera venom Sensitization to the major allergens Api m 1 of honeybee and Ves v 5 and/or Ves v 1 of yellow jackets may be helpful in discriminating between true double bee and wasp sensitization and cross-reactivity due to CCDs • World Allergy Organization Journal 2013, 6:17
  72. 72. Specific immunotherapy Most commercial allergen extracts used in SIT are well standardized for major allergens • contain only minimal or variable amounts of minor allergens • Patients with sensitization to minor allergens alone will likely not receive sufficient amounts of allergen to achieve a successful outcome by SIT • World Allergy Organization Journal 2013, 6:17
  73. 73. Specific immunotherapy Theoretically, identification of molecules  tailored SIT?? • In practice, this does not seem feasible 1. number of possible combination of sensitization profiles is large when taking into consideration all allergenic sources 2. recombinant vaccines do not perform better than traditional allergen extracts, as observed in some studies 3. each single recombinant/purified allergen would need to be individually tested and registered, which carries a substantial financial burden for manufacturers • reality of patient-tailored SIT is still a distant prospect • World Allergy Organization Journal 2013, 6:17
  74. 74. Micro-array technology
  75. 75. Micro-array technology MA diagnostics has been available on singleplex platforms • ImmunoCAP, ImmuLite, and HyTech platforms • single allergens together with the corresponding allergen extract • • Multiplex technology : • measure sIgE antibodies against multiple allergens in a single assay • testing of a large number of allergens using a small amount of serum World Allergy Organization Journal 2013, 6:17
  76. 76. ImmunoCAP Immuno-Solid Phase Allergen Chip (ISAC) (Phadia AB) Fluorescence is measured with a laser scanner and results are evaluated using a Microarray Image Analysis (MIA) software, which provides an automatic readout of the results. World Allergy Organization Journal 2013, 6:17
  77. 77. ISAC Using a standard calibration curve, results are reported within a range of 0.3 - 100 ISAC Standardized Units (ISU-E) • giving a semi-quantitative indication of IgE antibody levels • World Allergy Organization Journal 2013, 6:17
  78. 78. ISAC • ImmunoCAP • results : kU/L • not interchangeable • correlate well ImmunoCAP technology measures IgE binding under conditions of excess of immobilized allergen • ISAC uses low amounts of immobilized allergen allowing for competition with allergen-specific isotypes other than IgE • World Allergy Organization Journal 2013, 6:17
  79. 79. When comparing ISAC with other sIgE measuring assays, concordance of results vary between allergens tested World Allergy Organization Journal 2013, 6:17
  80. 80. ISAC Results of the ISAC assay are reproducible at a level that is generally accepted and agreed upon • • Special attention : samples contain low levels of sIgE (0.3–1 ISU-E),  higher degree of variability at low sIgE levels ImmunoCAP is more sensitive than ISAC Comparative data with the ImmuLite or HyTech platforms are not available in the literature • World Allergy Organization Journal 2013, 6:17
  81. 81. ISAC ISAC is generally not recommended for monitoring quantitative IgE levels over time in clinical routine • •Because there is a higher degree of between assay variability for ISAC compared to ImmunoCAP The interpretation of the results may be challenging 1. clinical relevance of the different allergens must be considered 2. results must be evaluated in relation to traditional diagnostic tests 3. results must be evaluated with regard to the patient’s clinical history (most important) • World Allergy Organization Journal 2013, 6:17
  82. 82. Micro-array technology ISAC results for some allergen sources such as cashew nut, sesame, dog, mugwort, and ragweed can be negative, even when the extract-based test is positive  triggering allergen is not present on the chip • World Allergy Organization Journal 2013, 6:17
  83. 83. • World Allergy Organization Journal 2013, 6:17
  84. 84. Patients most likely to benefit from MA diagnostics Patients who are poly-sensitized, have an unclear symptom and/or sensitization pattern, or who do not respond to their treatment • Patients with documented poly-sensitization to one or more inhalants, but also suffering from food allergy (i.e., from less severe manifestations such as OAS to more severe, including anaphylaxis, asthma or eczema) • Mono-sensitized patients with a clear case history and symptom profile may not benefit from MA diagnostics compared to traditional diagnostic tests World Allergy Organization Journal 2013, 6:17
  85. 85. Molecular-based allergy diagnostics • MA diagnostics may offer additional information for - early diagnosis of allergies - aid in the monitoring of the evolution of the allergic disease - useful for preventive indications to the patient World Allergy Organization Journal 2013, 6:17
  86. 86. Unmet needs! 1) Large-scale, population-based multicenter studies : categories of patients 2) Large studies that include well-characterized patients and healthy, sensitized controls representative of different geographical regions 3) Cost-utility studies :  compare effectiveness of MA diagnostics with the traditional in vitro sIgE or SPT World Allergy Organization Journal 2013, 6:17
  87. 87. Unmet needs! 4) Identification and clinical evaluation of the most relevant allergens have to be further investigated in many allergen sources.(e.g., nuts, molds, tree and weed pollen) 5) Training efforts in both the clinical and research settings 6) Development of clinical decision support is needed to prevent misinterpretation and improve knowledge as the amount of information obtained from MA diagnostics may be complex, especially as the evidence for MA is rapidly progressing. World Allergy Organization Journal 2013, 6:17
  88. 88. • International guidelines recommendation 1st Step : a thorough clinical case history 2nd Step : allergen extract-based IgE tests (in vitro specific IgE or skin prick test)  identification of the allergen source  SPT and in vitro sIgE tests provide similar information associated advantages and disadvantages of both types of tests are dependent on the clinical case 3rd Step : Molecular-based allergy (MA) diagnostics  used for select patients in whom first- and second-line investigations were inconclusive  For the majority of patients, first- and second-line investigation is sufficient to define the nature of a patient’s allergy  For experienced users MA may be included in second-line testing World Allergy Organization Journal 2013, 6:17
  89. 89. Conclusions MA diagnostics is increasingly entering routine care and can improve management of allergic patients • Benefit (1) Increase accuracy and resolve cross-reactivity (2) Assess the risk and type of reaction (3) Specific immunotherapy - small serum or blood sample - broad spectrum analysis of a patient’s IgE profile World Allergy Organization Journal 2013, 6:17
  90. 90. Conclusions Sensitivity compared to ImmunoCAP is still often lower • ISAC can have high clinical relevance by detecting sensitization patterns to important allergens and crossreacting groups • World Allergy Organization Journal 2013, 6:17
  91. 91. Conclusions Disadvantage Interpretation of such sensitization in clinically unresponsive patients is difficult or even impossible • Relatively expensive compared with traditional tests  economic consideration or budget limitations • the search for more, clinically relevant molecules is needed and ongoing • World Allergy Organization Journal 2013, 6:17
  92. 92. Conclusions Although MA diagnostics is a complex area, it provides novel and relevant information for the allergist and will soon become a standard tool in the allergist’s armamentarium • Educational programs training allergists on the use and interpretation of MA are highly needed • • World Allergy Organization Journal 2013, 6:17
  93. 93. Thank you
  1. A particular slide catching your eye?

    Clipping is a handy way to collect important slides you want to go back to later.

×