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Exploring the Diagnosis and Profile of Cannabis Allergy

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Background: Cannabis allergy (CA) has mainly been attributed to Can s 3, the nsLTP (non-specific lipid transfer proten) of Cannabis sativa. Nevertheless, standardized diagnostic tests are lacking and research on CA is scarce. Objective: To explore the performance of five cannabis diagnostic tests and the phenotypic profile of CA. Methods: 120 CA patients were included and stratified according to the nature of their cannabis-related symptoms, 62 healthy and 189 atopic controls were included. Specific (s)IgE hemp, sIgE and BAT rCan s 3, BAT with a crude cannabis extract and a skin prick test (SPT) with a nCan s 3-rich cannabis extract were performed. Clinical information was based on patient-history and a standardized questionnaire. Results: Firstly, up to 72% of CA reporting likely-anaphylaxis (CA-A) are Can s 3 sensitized. Actually, the Can s 3-based diagnostic tests show the best combination of positive and negative predictive values; 80% and 60%, respectively. sIgE hemp displays 82% sensitivity but only 32% specificity. Secondly, Can s 3+CA reported significantly more cofactor mediated reactions and displayed significantly more sensitizations to other nsLTPs than Can s 3-CA. Finally, the highest prevalence of systemic reactions to plant-derived foods was seen in CA-A, namely 72%. Conclusions: The most effective and practical tests to confirm CA are the SPT with a nCan s 3-rich extract and the sIgE rCan s 3. Can s 3 entails a risk of systemic reactions to plant-derived foods and cofactor-mediated reactions. However, as Can s 3 sensitization is not absolute, other cannabis allergens probably play a role.
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Original Article
Exploring the Diagnosis and Profile of Cannabis
Allergy
Ine Ilona Decuyper, MD
a,b
, Athina Ludovica Van Gasse, MD
a
, Margaretha A. Faber, MD, PhD
a
, Jessy Elst, MSc
a
,
Christel Mertens, MLT
a
, Hans-Peter Rihs, PhD
c
, Margo M. Hagendorens, MD, PhD
a,b
, Vito Sabato, MD, PhD
a
,
Hilde Lapeere, MD, PhD
d
, Chris H. Bridts, MLT
a
, Luc S. De Clerck, MD, PhD
a
, and Didier Gaston Ebo, MD, PhD
a
Antwerp, Ghent, Belgium; and Bochum, Germany
What is already known about this topic? Cannabis allergy, although rare, can manifest with severe and generalized
symptoms and has been linked to Can s 3, the nonspecic lipid transfer protein present in Cannabis sativa.
What does this article add to our knowledge? This article compares the performance of multiple cannabis diagnostic
methods and explores clinical and in vitro characteristics of cannabis allergy in one of the largest cannabis allergic
populations described up until now.
How does this study impact current management guidelines? There are no guidelines available on cannabis allergy
diagnosis or management. This articles perspective on diagnostic performances could aid in accurately approximating
post-test probabilities and gives insight into the prole of Western European cannabis allergic patients.
BACKGROUND: Cannabis allergy (CA) has mainly been
attributed to Can s 3, the nonspecic lipid transfer protein
(nsLTP) of Cannabis sativa. Nevertheless, standardized
diagnostic tests are lacking and research on CA is scarce.
OBJECTIVE: To explore the performance of 5 cannabis
diagnostic tests and the phenotypic prole of CA.
METHODS: A total of 120 patients with CA were included and
stratied according to the nature of their cannabis-related
symptoms; 62 healthy and 189 atopic controls were included.
Specic IgE (sIgE) hemp, sIgE and basophil activation test
(BAT) with a recombinant Can s 3 protein from Cannabis sativa
(rCan s 3), BAT with a crude cannabis extract, and a skin prick
test (SPT) with an nCan s 3-rich cannabis extract were
performed. Clinical information was based on patient history
and a standardized questionnaire.
RESULTS: First, up to 72% of CA reporting likely-anaphylaxis
(CA-A) are Can s 3 sensitized. Actually, the Can s 3-based
diagnostic tests show the best combination of positive and
negative predictive values, 80% and 60%, respectively. sIgE
hemp displays 82% sensitivity but only 32% specicity.
Secondly, Can s 3DCA reported signicantly more cofactor-
mediated reactions and displayed signicantly more sensitiza-
tions to other nsLTPs than Can s 3-CA. Finally, the highest
prevalence of systemic reactions to plant-derived foods was seen
in CA-A, namely 72%.
CONCLUSIONS: The most effective and practical tests to
conrm CA are the SPT with an nCan s 3-rich extract and the
sIgE rCan s 3. Can s 3 sensitization entails a risk of systemic
reactions to plant-derived foods and cofactor-mediated reactions.
However, as Can s 3 sensitization is not absolute, other cannabis
allergens probably play a role. Ó2018 Published by Elsevier
Inc. on behalf of the American Academy of Allergy, Asthma &
Immunology (J Allergy Clin Immunol Pract 2018;-:---)
Key words: Cannabis allergy; Diagnosis; BAT; Specic IgE; Skin
prick test; Can s 3; nsLTP; Cofactor; Basophil; Anaphylaxis;
Hemp
Cannabis is one of the most consumed drugs worldwide.
1
Despite its widespread use, reports on cannabis allergy (CA)
remain rare and generally deal with relatively small numbers of
cases.
2-6
Nevertheless, from these reports evidence is accumu-
lating that CA can manifest with severe and generalized
a
Department of Immunology-Allergology-Rheumatology, University Hospital of
Antwerp, University of Antwerp, Antwerp, Belgium
b
Department of Pediatrics, University Hospital of Antwerp, University of Antwerp,
Antwerp, Belgium
c
IPAInstitute for Prevention and Occupational Medicine, German Social Accident
Insurance, Ruhr-University Bochum, Bochum, Germany
d
Ghent University Hospital, University of Ghent, Ghent, Belgium
This work was supported by the Agency for Innovation by Science and Technology
(grant number 140185). Furthermore, V. Sabato is a senior clinical researcher of
the Research Foundation Flanders/Fonds Wetenschappelijk Onderzoek (FWO:
1804518N). D. G. Ebo is a senior clinical researcher of the Research Foundation
Flanders/Fonds Wetenschappelijk Onderzoek (FWO: 1800614N). A. L. Van
Gasse is a fellow of the Research Foundation Flanders/Fonds Wetenschappelijk
Onderzoek (FWO: 1113617N).
Conicts of interest: The authors declare that they have no relevant conicts of
interest.
Received for publication July 17, 2018; revised August 17, 2018; accepted for
publication September 15, 2018.
Available online --
Correspondence author: Didier Gaston Ebo, MD, PhD, Department of Immunology,
Allergology, Rheumatology, Faculty of Medicine and Health Science, University
of Antwerp, Campus Drie Eiken T5.95, Universiteitsplein 1, 2610 Antwerp,
Belgium. E-mail: immuno@uantwerpen.be.
2213-2198
Ó2018 Published by Elsevier Inc. on behalf of the American Academy of Allergy,
Asthma & Immunology
https://doi.org/10.1016/j.jaip.2018.09.017
1
Abbreviations used
BAT- Basophil activation test
CA- Cannabis allergy
CA-A- Likely-anaphylaxis to cannabis
CA-C- Cutaneous symptoms to cannabis
CA-R- Respiratory symptoms to cannabis
CA-RC- Localized respiratory and cutaneous symptoms to
cannabis
CBA- Cytometric bead array
CS- Cannabis sativa
HC- Healthy controls
NPV- Negative predictive value
nsLTP- Nonspecic lipid transfer protein
PþLTP- Atopic pollen-sensitized participants without an nsLTP
sensitization
PþLTPþ- Atopic pollen and nsLTP-sensitized participants
PPV- Positive predictive value
rCan s 3- Recombinant Can s 3 protein from CS
sIgE- Specic IgE
SPT- Skin prick test
symptoms and a variety of cross-reactive plant-derived food
allergies, mainly attributed to a Can s 3 sensitization, the
nonspecic lipid transfer protein (nsLTP) from Cannabis sativa
(CS). As a matter of fact, in some European surveys, Can s 3 has
been demonstrated to be a major allergen.
7-9
NsLTPs are heat
stable allergens widely distributed throughout the plant kingdom
and showing extensive in vitro and in vivo cross-reactivity.
10
Both
the severe phenotype and the extensive cross-reactivity associated
with CA can be attributed to the physiochemical properties of
Can s 3. Other putative cannabis allergens are ribulose-1,5-
bisphosphate carboxylase/oxygenase, oxygen-evolving enhancer
protein 2, and a thaumatin-like protein.
2,4
However, unlike Can
s3,
3
these allergens have not yet been successfully isolated nor
expressed as a recombinant protein and are currently unavailable
for diagnosis.
So far, in the majority of studies on CA, diagnosis is docu-
mented by prick-prick tests with buds or leaves
4-6,9
and therefore
are difcult to standardize, because of the heterogeneous
composition of the different source materials. The clinical
severity and cross-reactivity of CA together with the unpredict-
ability of the source materials used for skin testing constitute
strong incentives for more reliable cannabis diagnostic tests,
in vitro or in vivo.
In 2 preliminary studies, we have standardized and presented
initial performance results using 4 different cannabis diagnostic
tests, namely, a basophil activation test (BAT) with recombinant
Can s 3 protein from CS (rCan s 3), a BAT with a crude CS
extract, a skin prick test (SPT) with an nCan s 3-rich extract, and
nally, a specic IgE (sIgE) rCan s 3 assay using a cytometric
bead array (CBA) technique. These diagnostic tests were
compared with sIgE industrial hemp by uorescence enzyme
immunoassay (FEIA) ImmunoCAP. All 4 of our diagnostic tests
have been found reliable in diagnosing CA
7,8
and revealed Can s
3 sensitization in up to 75% of patients with CA with an
anaphylaxis-like phenotype. Alternatively, the sIgE hemp assay,
albeit displaying an excellent sensitivity, was shown to be poorly
reliable because of an important proportion of clinically irrele-
vant positive results in cannabis-tolerant individuals sensitized to
pollen and/or nsLTPs.
Importantly, for robust validation purposes, our recent study
8
was restricted to patients with an anaphylaxis-like phenotype on
cannabis exposure. However, in general practice, physicians
might frequently encounter patients with less compelling
histories such as isolated respiratory symptoms and in whom Can
s 3 sensitization seems less predominant.
3
Therefore, this study
investigates the diagnostic test performances and intertest
differences between these 5 diagnostic tests in a larger study
population expressing distinct clinical phenotypes on cannabis
exposure. Secondly, this study explores the clinical and molecular
characteristics of CA; the sensitization proles, the severity of
cross-reactivities with other plant-derived foods, and the signi-
cance of cofactors, as patients presenting with nsLTP-related
allergies have frequently been reported to necessitate a cofactor
to become symptomatic.
11,12
METHODS
Inclusion
Patients and controls were included through the outpatients
clinic of Allergology at the Antwerp University Hospital and the
Dermatology department of the Ghent University Hospital,
Belgium. The local ethics committees of both hospitals approved this
study (B300201524055), and patients or their representatives signed
an informed consent in accordance with the Declaration of Helsinki.
Patients with respiratory, gastrointestinal, cardiovascular, and/or
cutaneous symptoms on exposure to cannabis were included.
Exposure to cannabis was dened as active smoking, ingestion, and/
or direct cutaneous contact with cannabis. Patients with generalized
symptoms in 2 or more organ systems were categorized as likely-
anaphylactic according to the criteria dened by Sampson et al.
13
Furthermore, 2 distinct control groups were included: rst,
healthy controls (HC) without pollen or nsLTP sensitization;
secondly, a so-called atopic control group comprising patients with a
documented pollen allergy with (PþLTPþ) or without nsLTP
(PþLTP) sensitization. Controls were further stratied according
to exposure and tolerance to cannabis, that is, uneventful exposure.
Denitions of pollen and nsLTP sensitizations are shown in this
articles Online Repository at www.jaci-inpractice.org.
Information on CA, cofactor-associated reactions (reported plant-
derived food allergies with a history of overt or more severe/gener-
alized reactions in the presence of nonsteroidal anti-inammatory
drugs, alcohol, or physical exercise than when the reaction
occurred in the absence thereof), and severity of plant-derived food-
associated reactions was gathered by history taking and a standard-
ized questionnaire. Three cofactors were dened in this study: the
use of alcoholic beverages, nonsteroidal anti-inammatory drugs,
and/or the performance of exercise within 3 hours preceding
occurrence of an allergic reaction. A systemic reaction was dened as
grade 1 or higher as dened by the World Allergy Organization
criteria of systemic allergic reactions.
14
Patients with chronic
spontaneous urticaria, uncontrolled asthma, eosinophilic esophagi-
tis/colitis, or systemic mastocytosis were excluded.
Diagnostic tests
Basophil activation test. BAT with rCan s 3 and a crude CS
extract were performed, as detailed in this articles Online Repository
at www.jaci-inpractice.org and previously validated as described in
detail elsewhere.
8
Results were expressed as net percentages of
CD63
þ
basophils, calculated by subtraction of the spontaneous
expression from the allergen-induced CD63 expression. A result
J ALLERGY CLIN IMMUNOL PRACT
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2DECUYPER ET AL
>5% CD63
þ
basophils was considered positive as dened by
previous validation.
8
Total and specific IgE. Total and sIgE to industrial hemp,
rBet v 1 and rBet v 2 from birch (Betula verrucosa), rPhl p 1 and rPhl
p 5b from timothy grass (Phleum pratense), nArt v 1 and nArt v 3
from mugwort (Artemisia vulgaris), rAra h 9 from peanut (Arachis
hypogeae), rCor a 8 from hazelnut (Corylus avellana), rMal d 3 from
apple (Malus domesticus), rJug r 3 from walnut (Juglans regia), rPru
p 3 from peach (Prunus persica), rPar j 2 from wall pellitory (Pari-
etaria judaica), and nAna 2 c from bromelain (Ananas comosus), as a
marker for sensitization to cross-reactive carbohydrate determinants,
were quantied by the FEIA ImmunoCAP technique (Thermo-
Fisher Scientic, Uppsala, Sweden) according to the manufacturers
instructions. All sIgE assays are readily available, except for industrial
hemp, which is available for research use only and was kindly pro-
vided by ThermoFisher Scientic. Specic IgE to rCan s 3 was
quantied using a ow CBA technique (BD Biosciences, Franklin
Lakes, NJ). The method was validated as previously described.
8
Results 0.10 kU
A
/L were considered positive.
Skin prick tests. SPT implied an nCan s 3-rich CS extract that
was prepared as described elsewhere.
7
SPT responses were read after
15 minutes and considered positive when the wheal exceeded 3 mm
(largest diameter). A positive control with histamine (10 mg/mL)
and a negative saline control without allergen (ALK-Abello Ltd,
Berkshire, United Kingdom) were performed to rule out non-
responsiveness or dermographism of the skin, respectively.
Statistical analysis
IBM SPSS version 24.0 (IBM, Chicago, Ill) software was used for
data analysis. Data are expressed as medians and interquartile ranges.
Nonparametric tests and
c
2
analysis were used where appropriate.
Test performances were compared by using McNemars test. Where
needed, missing values were imputed by using a multiple-imputation
model with 5 imputations based on all available information that
were subsequently pooled in SPSS. Signicance levels for the pooled
imputed data were calculated according to the method described by
Schafer et al.
15
APvalue of <.05 was regarded as statistically
signicant.
RESULTS
Demographics
As shown in Figure 1, a total of 371 individuals were
included; 120 patients with symptoms on cannabis exposure
(CA) of which 21% (n ¼25) were classied as likely-
anaphylactic (CA-A), 19% (n ¼23) presented with mild and
localized respiratory and cutaneous symptoms (CA-RC), 51%
reported isolated respiratory symptoms (CA-R), and 9% report
isolated cutaneous symptoms (CA-C). The remaining 251 par-
ticipants were control individuals, either HC or atopics with a
pollen sensitization (PþLTPþ), with or without nsLTP sensi-
tizations (PþLTP). As displayed in Figure 1, 50% to 60% of
each control group reported the regular use of cannabis in the
past 12 months without any symptoms apart from the known
psychoactive effects; the other half reported no previous contact
with cannabis. All patients with CA displayed symptoms during
active smoking, except for 3 patients denying any previous direct
contact with cannabis (no active smoking, ingestion, or cuta-
neous contact) but who had experienced symptoms on passive
exposure to cannabis smoke. Furthermore, in total 34 patients
with CA reported respiratory and/or cutaneous symptoms on
n = 371
CA
n = 120
likely-
anaphylaxis
n = 25
respiratory
symptoms
n = 61
cutaneous
symptoms
n = 11
localized
respiratory and
cutaneous symptoms
n = 23
HC
n = 62
CS exposed
n = 37
not CS exposed
n = 25
P+
n = 90
CS exposed
n = 47
not CS exposed
n = 43
P+LTP+
n = 99
CS exposed
n = 49
not CS exposed
n = 50
FIGURE 1. Inclusion overview. CA, Cannabis allergic patients; CS,Cannabis sativa;HC, healthy controls; PþLTP, pollen-sensitized
controls without an nsLTP sensitization; PþLTPþ, pollen and nsLTP-sensitized controls.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 3
isolated passive exposure to cannabis smoke apart from symp-
toms on active smoking. Finally, 4 patients also reported
symptoms on ingestion of cannabis processed as space cake,
cannabis seeds or oil, resulting in anaphylaxis in 2 of the cases.
The individual symptoms reported by CA-A are shown in
Table E1 in this articles Online Repository at www.jaci-
inpractice.org. In summary, 23 of 25 reported respiratory
symptoms and/or cutaneous symptoms, 4 patients also
mentioned cardiovascular symptoms comprising palpitations
and/or hypotension, and nally, 5 patients additionally reported
gastrointestinal symptoms comprising abdominal pain, nausea,
and vomiting.
Table I displays demographic data of the different study
groups revealing similar age, sex ratios, and asthma prevalence in
all groups. In contrast, atopic dermatitis and elevated total IgE
values were signicantly more prevalent in the PþLTPþgroup
than in the CA group and in the PþLTPgroup. Total IgE was
also signicantly higher in the PþLTPgroup compared with
the CA group. Finally, importantly, 84% of patients with CA
showed a pollen sensitization and 72% an nsLTP sensitization.
It is important to note that pollen sensitization was predomi-
nated by Bet v 1; 72% of CA sensitized) and 79% of PþLTPþ
exhibited a Bet v 1 sensitization.
Performance of cannabis diagnostic tests
Figure 2 shows the individual results of 5 different cannabis
diagnostic tests: the sIgE industrial hemp, sIgE rCan s 3 CBA,
SPT with an nCan s 3-rich extract, and the BAT with both rCan
TABLE I. Demographic data
HC [62 CA [120 PDLTPL[90 PDLTPD[99
Age (y), median (Q
25
-Q
75
) 28.3 (24.8-36.1) 29.2 (25.1-35.2) 28.8 (22.9-37.7) 29.9 (20.1-37.1)
Sex
(% male) 42% 48% 37% 49%
Eczema*0% 37% 37% 54%
Asthma5% 30% 28% 39%
Total IgE (kU/L), median (Q
25
-Q
75
) 16.7 (6.0-46.5) 247.4 (83.0-495.0) 126.0 (65.0-314.0) 424.5 (147.0-1054.0)
Pollen sensitizationz0% 84% 100% 100%
NsLTP sensitizationx0% 72% 0% 100%
CA, Cannabis allergy; CS,Cannabis sativa;HC, healthy controls; PþLTPþ, pollen and nsLTP-sensitized controls; PþLTP, pollen-sensitized controls without an nsLTP
sensitization; nsLTP, nonspecic lipid transfer protein.
*According to patient recollection and recent use of topical CS.
According to patient recollection.
zAt least one of the following sIgEs 0.1 kU
A
/L: rBet v 1, rBet v 2, nArt v 1, rPhl p 1, rPhl 5b.
xAt least one of the following sIgEs 0.1 kU
A
/L: rPru p 3, rMal d 3, rJug r 3, rAra h 9, rCor a 8, nArt v 3, rPar j 2, rTri a 14.
FIGURE 2. Individual test results. A, Dot plots showing healthy controls (HC), pollen and nsLTP-sensitized controls (PþLT P þ), pollen-
sensitized controls without an nsLTP sensitization (PþLT P ), and cannabis allergy (CA). B, Dot plots for the different CA groups:
CA-A, CA-RC, CA-R, and CA-C. Percentages reflect the proportion of positive results and horizontal lines represent group mean.
Dindicates patients with <15% response to anti-IgE stimulation (nonresponders). Of 371, 55 (15%) were classified as nonresponders:
15 HC, 12 PþLT P ,14PþLT P þ, and 14 CA. BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CA-C, cutaneous
symptoms to cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and cutaneous symptoms to cannabis;
sIgE, specific IgE; SPT, skin prick test.
J ALLERGY CLIN IMMUNOL PRACT
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4DECUYPER ET AL
s 3 and a crude cannabis extract. Table II compares the test
performances. For more details on the difference in test perfor-
mance for sIgE rCan s 3 and sIgE hemp (considering 0.10 or
0.35 kU
A
/L cutoff), the reader is referred to Figure E1 and
Table E2 in this articles Online Repository at www.jaci-
inpractice.org.
First of all, test performances showed important variances
between the different clinical CA groups. The 3 Can s 3-based
diagnostic methods (BAT, sIgE, and SPT) displayed a similar
sensitivity, 63% to 72% in CA-A (45% to 58% in the total CA
group), and a similar specicity (81% to 87% in the total CA
group). However, up to 37% (n ¼34) of PþLTPþshowed
clinically irrelevant Can s 3 sensitizations (measured by BAT,
sIgE, or SPT): 20 of 34 reported tolerance to active cannabis use
and 14 of 34 reported no previous cannabis contact. In com-
parison, the sIgE rCan s 3 and BAT rCan s 3 showed no clini-
cally irrelevant positive results in pollen-sensitized individuals
without nsLTP sensitizations (PþLTP).
Secondly, the sIgE industrial hemp displayed a signicantly
higher sensitivity, up to 82% (P<.01) in the total CA group
compared with the Can s 3-based diagnostic tests (45% to 58%).
However, sIgE hemp also demonstrated a signicantly higher
number of clinically irrelevant positive results in PþLTPand
PþLTPþ, that is, 51% to 82% respectively compared with 0%
to 25% for the Can s 3 diagnostic tests (all P<.01). Interest-
ingly, an increase in sensitivity as seen in the sIgE hemp was not
found in the BAT with a crude cannabis extract. The latter
reached an overall sensitivity of 49% in the total CA group that
was not superior to the Can s 3-based assays. In addition, the
BAT with the crude extract was not superior to the Can s 3
diagnostic tests in terms of specicity either, showing 19% to
38% of clinically irrelevant positive results in PþLTPand
PþLTPþ. Collectively, for all diagnostic techniques, the
majority of clinically irrelevant results were seen in the PþLTPþ
group.
In summary, when all different clinical CA groups are
considered (analysis B in Table II), it appears that the 3 Can s
3-based diagnostic tests did not signicantly differ in perfor-
mance and had the best combined positive predictive value
(PPV) and negative predictive value (NPV) of around 80% and
60%, respectively. The sIgE industrial hemp lacked specicity,
whereas the BAT crude CS extract showed no advantage over the
Can s 3-based diagnostic tests.
The clinical and molecular characteristics of
CA. Figure 3 compares different clinical and in vitro charac-
teristics for the different CA proles and the control groups. The
most prominent differences were found between CA-A and
CA-R with signicantly higher numbers of Pru p 3, Mal d 3, Cor
a 8, Jug r 3, Tri a 14, Art v 3 sensitizations (all P<.01) in CA-A
than in CA-R. Furthermore, CA-A showed a higher prevalence
of systemic reactions to plant-derived foods (72% compared with
40%, P¼.02) and cofactor-mediated allergic reactions (50%
compared with 18%, P¼.01) compared with CA-R. In addi-
tion, CA-C and CA-RC showed a single difference from CA-A,
namely a considerably lower prevalence of systemic reactions to
plant-derived foods (71% in CA-A compared with 43% in
CA-RC [P<.01] and 18% in CA-C [P¼.08]). It appears that
none of the clinical nor in vitro parameters displayed signicant
differences between CA-R, CA-C, and CA-RC.
Regarding the comparison of Can s 3-sensitized and non-
sensitized CA (as demonstrated in Table E3 in this articles
Online Repository at www.jaci-inpractice.org), it became clear
that Can s 3þCA had a signicantly higher prevalence of other
nsLTP sensitizations (92%) than Can s 3-CA (39%) with higher
frequencies of all measured nsLTPs (all P<.01), except for Par j
2. Also, Can s 3þCA displayed higher frequencies of pollen
sensitizations than Can s 3-CA (92% compared with 74%) with
signicantly more Bet v 1 sensitizations in the Can s 3-sensitized
population. In addition, Can s 3þCA showed a considerably
higher prevalence of cofactor-mediated allergic reactions when
compared with Can s 3-CA (41% vs 12%; P<.01).
In a further analysis, the complete CA group was compared
with the PþLTPþgroup (as demonstrated in Table E4 in this
TABLE II. Test performance
sIgE hemp sIgE rCan s 3 BAT rCan s 3 BAT crude CS extract SPT nCan s 3-rich extract
A
Sensitivity 86% (66-97) 63% (41-81) 71% (48-89) 63% (38-84) 72% (51-89)
Specicity 32% (20-45) 87% (78-93) 85% (76-92) 67% (55-78) 81% (71-88)
PPV 33% (28-38) 56% (40-70) 54% (39-67) 35% (25-47) 51% (39-63)
NPV 86% (66-95) 90% (84-94) 93% (86-96) 87% (78-92) 91% (84-95)
LHRþ1.3 (1.0-1.6) 4.7 (2.6-8.7) 4.8 (2.7-8.6) 1.9 (1.2-3.1) 3.7 (2.3-6.0)
LHR0.4 (0.1-1.3) 0.40 (0.3-0.7) 0.3 (0.2-0.7) 0.6 (0.3-1.0) 0.4 (0.2-0.7)
B
Sensitivity 82% (74-89) 47% (38-56) 45% (35-55) 49% (37-60) 58% (49-67)
Specicity 32% (20-45) 87% (78-93) 85% (76-92) 67% (55-78) 81% (71-88)
PPV 70% (66-74) 82% (72-89) 78% (67-86) 64% (54-73) 80% (72-86)
NPV 47% (34-61) 56% (51-60) 57% (52-62) 52% (46-59) 58% (53-64)
LHRþ1.2 (1.0-1.5) 3.5 (2.0-6.2) 3.0 (1.8-5.2) 1.5 (1.0-2.2) 3.0 (1.9-4.7)
LHR0.6 (0.3-1.0) 0.60 (0.5-0.7) 0.7 (0.5-0.8) 0.8 (0.6-1.0) 0.5 (0.4-0.7)
A: calculations based on CA-A group vs cannabis-tolerant PþLTPand PþLTPþ.
B: calculations based on the whole CA group (respiratory and/or cutaneous symptoms) vs cannabis-tolerant PþLTPand PþLTPþ. Test performance for both BATs was
calculated by considering both responders and nonresponders to anti-IgE.
BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CS,Cannabis sativa;NPV, negative predictive value; PPV, positive predictive value; LHRþ, positive
likelihood ratio; LHR, negative likelihood ratio; PþLTPþ, pollen and nsLTP-sensitized controls; PþLTP, pollen-sensitized controls without an nsLTP sensitization; sIgE,
specic IgE; SPT, skin prick test.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 5
articles Online Repository at www.jaci-inpractice.org). This
exploration revealed a signicantly (P<.01) higher prevalence of
Can s 3 sensitizations in CA (63%) compared with PþLTPþ
(35%). Furthermore, a signicantly lower prevalence of Pru p 3,
Mal d 3, Jug r 3, Par j 2 (all P<.01) but also bromelain (P¼
.02) and Phl p 1 (P<.01) sensitizations was seen in the CA
group compared with PþLTPþ. Finally, as already mentioned in
the demographic paragraph, signicantly (P<.01) more eczema
was reported in the PþLTPþgroup than the CA group and
subsequently total IgE values were also signicantly higher in
PþLTPþthan in the CA group (P<.01). Although there was
no signicant difference between CA and PþLTPþconcerning
the frequency of systemic reactions to plant-derived foods (P¼
.11), CA-A did show double the frequency of systemic reactions
to plant-derived foods than PþLTPþ(71% vs 35%, P<.01).
DISCUSSION
To the best of our knowledge, this is the largest survey
exploring diagnostic performances in different clinical pheno-
types of CS allergy. Along with the observation that the diag-
nostic utilities of our tests depend on the clinical presentation, it
appears that the CA prole in this study population has the
following peculiarities.
Primarily, in terms of practicality, efciency, and standardi-
zation, the SPT with an nCans 3-rich extract and the sIgE rCan s
3 are the easiest and fastest tests to conrm a clinical suspicion of
CA, both equally reliable. However, because of unavailability, in
clinical practice, physicians will need to rely on other tests to
screen patients with a convincing history. As a matter of fact,
according to our data, it seems that the sIgE hemp assay (avail-
able on request from ThermoFisher) could serve as a suitable
diagnostic in central Europe to exclude CA, because a negative
test result reduces the risk of CA considerably (only 18% of CA
have negative sIgE hemp results). Alternatively, patients with a
convincing history together with a positive sIgE hemp should
undergo additional testing to elucidate the clinical signicance of
the hemp solid phase assay. In addition, exploration of different
cutoffs for the sIgE rCan s 3 and hemp shows that, when
considering the 0.35 kUa/L cutoff, sensitivity of both tests de-
creases by around 10% in the total CA population. Nevertheless,
sensitivity to detect CA-A remains the same for both. Even
though the specicity of sIgE hemp almost doubles, it still only
reaches a maximum of 60%, which is not ideal.
However, none of our diagnostic tests appear absolutely pre-
dictive for the clinical outcome. Nevertheless, for the time being,
on the basis of our ndings, we propose to perform the SPT with
an nCan s 3-rich extract or quantify sIgE rCan s 3 keeping in
mind that Can s 3 does not cover the entire IgE sensitization
prole, particularly in patients with a less severe/pronounced
phenotype. In addition, it could be questioned whether Can s 3-
negative patients, especially if reporting only milder symptoms to
cannabis, should effectively be categorized as CA, because their
symptoms could result from nonspecic skin or airway irritation.
Furthermore, because of ethical and legal limitations, it is
impossible to conrm CA by an oral or respiratory challenge.
Considering this hypothesis, it follows that the actual test per-
formances are possibly underestimated in this study and that Can s
3 might even play a more prominent role than already suspected.
Furthermore, it is likely that performances of a Can s 3 assay
display regional differences due to geographic differences in IgE
reactivity proles. The reason(s) why Can s 3 negative CA pa-
tients go undetected in the BAT with the full CS extract re-
main(s) elusive but could relate to a sensitization to allergens that
are poorly present in our crude extract or do not resist our
current extraction procedure. Moreover, the low presence and
the physicochemical properties of the constituent allergens might
also explain the different sensitization proles in the distinct
phenotypes, namely, the lower prevalence of nsLTP sensitiza-
tions in CA-R compared with CA-A.
Eczema
Asthma
Cofactor
SR to plant-foods
Pollen sensitization
Bet v 1
Bet v 2
Phl p 1
Phl p 5b
Art v 1
LTP sensitization
Pru p 3
Mal d 3
Cor a 8
Ara h 9
Jug r 3
Tri a 14
Art v 3
Par j 2
Can s 3*
Bromelain
P+LTP-
P+LTP+
CA total
CA-A
CA-R
CA-RC
CA-C
SCALE
0% 0%
10% 10%
20% 20%
30% 30%
40% 40%
50% 50%
60% 60%
70% 70%
80% 80%
90% 90%
100% 100%
FIGURE 3. Overview of clinical and in vitro parameters. Color variations represent increasing frequencies of positive results for the shown
variable. For example, frequency of asthmatics (specific IgE [sIgE] measurements are shown as percentage sensitized/not sensitized).
*Measured by BAT or sIgE rCan s 3.BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CA-C, cutaneous symptoms to
cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and cutaneous symptoms to cannabis; LTP, lipid transfer
protein; OAS, oral allergy syndrome defined as localized and mild oropharyngeal symptoms without generalization; PþLTP, pollen-
sensitized controls without an nsLTP sensitization; PþLTPþ, pollen and nsLTP-sensitized controls; SR, systemic reaction defined by
generalized and severe symptoms in at least 1 organ system
14
;TOL, tolerant.
J ALLERGY CLIN IMMUNOL PRACT
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6DECUYPER ET AL
Secondly, although historically sensitization to nsLTP has
mainly been recognized to occur in the Mediterranean region,
characterized by severe reactions and governed by peach,
10,16
more
recent data have accumulated showing that sensitization to nsLTP
might also occur in other European regions and frequently remain
asymptomatic with uncertainties about the route(s) of sensitiza-
tion.
17-19
In this survey, we conrm that nsLTP sensitization
occurs frequently in CA and Can s 3 is a major allergen in patients
with CA-A, but CA also implies a risk of systemic reactions to
plant-derived foods and cofactor-mediated reactions. Furthermore,
Can s 3 sensitization can occur as a result of in vitro cross-reactivity
to nsLTPs from taxonomically related or more distant sources
such as pollen and/or plant-derived foods as suggested by the Can
s 3 positive PþLTPþpatients without any previous cannabis
contact. On the other hand, it seems that a Can s 3 sensitization in
patients with CA might also mirror a primary sensitization instead
of only in vitro cross-reactivity as indicated by the signicantly
higher prevalence of Can 3 and lower prevalence of Pru p 3, Mal
d 3, Jug r 3, and Par j 2 sensitizations in CA compared with
PþLTPþ.
Another important fact to highlight is that, because of the lack
of data on the true prevalence of CA, it is likely that the number
of patients per study group in this survey do not necessarily
reect the true prevalence of CA. Therefore, the test perfor-
mances would differ depending on the characteristics of the
tested population and the geographic prevalence of CA itself.
Finally, this study was not designed to explore the different
individual types of plant-derived food allergies, as symptoms to
different plant-derived foods were only assessed by a standardized
questionnaire complemented with a history taking without sys-
tematic conrmatory testing. However, it would be interesting to
further explore the actual differences in individual plant-derived
food allergies within CA such as the differences in symptom
severity with and without peel, the types of plant-derived foods
eliciting allergic symptoms but also the comparison of these
factors between CA and other nsLTP-sensitized individuals.
In conclusion, this study is the largest study exploring diagnostic
test performance, clinical phenotypes, and biological proles of
CA. It shows that the most effective and practical tests to conrm a
clinical suspicion of CA are the SPT with an nCan s 3-rich extract
and sIgE rCan s 3. Both tests display a PPV and NPV of approx-
imately 80% and 60%, respectively. However, because of current
unavailability, screening with sIgE hemp could be a suitable tool in
symptomatic cannabis users, because a negative result considerably
reduces the likelihood of CA. Alternatively, we dissuade the general
use of sIgE hemp to diagnose CA, mainly because of its limited
PPV. Furthermore, we show that Can s 3 is a major allergen in
patients with a history of likely-anaphylaxis on cannabis exposure
and, like other nsLTP-associated allergies, CA might indicate a risk
of systemic reactions to plant-derived foods and cofactor-mediated
reactions. Because around 30% of CA-A and even higher pro-
portions in other, milder CA groups are not sensitized to Can s 3, it
is likely that other cannabis allergens might play a role in CA.
Further studies are thus warranted to identify and express other CA
allergens that could then be applied to spike natural extracts or to
compose mixtures of allergens. Lastly, additional research should
further explore the nature of plant-derived food allergies in CA as
this study was not designed to evaluate specic plant-derived food
allergies in CA.
Acknowledgments
We thank B. Van Camp, head of the Unit drug production,
Central Drug Department, Directorate of organized crime,
Belgian Federal Judicial police for his help with providing the
necessary plant materials. We would also like to thank Mrs. K.
Vandebos and N. Maes, our study nurses, for their help in the
performance of the skin prick tests and blood sampling. Finally,
our gratitude goes out to Mrs. K Wouters, for her help and
statistical advice.
REFERENCES
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from: www.unodc.org/wdr2017. Accessed June 7, 2018.
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acterization of Cannabis sativa allergens. Ann Allergy Asthma Immunol 2013;
111:32-7.
3. Rihs HP, Armentia A, Sander I, Bruning T, Raulf M, Varga R. IgE-binding
properties of a recombinant lipid transfer protein from Cannabis sativa. Ann
Allergy Asthma Immunol 2014;113:233-4.
4. Larramendi CH, López-Matas M, Ferrer A, Huertas AJ, Pagán JA, Navarro LÁ,
et al. Prevalence of sensitization to Cannabis sativa. Lipid-transfer and
thaumatin-like proteins are relevant allergens. Int Arch Allergy Immunol 2013;
162:115-22.
5. Tessmer A, Berlin N, Sussman G, Leader N, Chung EC, Beezhold D. Hyper-
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New food allergies in a European non-Mediterranean region: is Cannabis sativa
to blame? Int Arch Allergy Immunol 2013;161:220-8.
8. Decuyper II, Faber MA, Lapeere H, Mertens C, Rihs HP, Van Gasse AL, et al.
Cannabis allergy: a diagnostic challenge. Allergy 2018;73:1911-4.
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Quesada J. Molecular diagnosis in cannabis allergy. J Allergy Clin Immunol
Pract 2014;2:351-2.
10. Pastorello EA, Robino AM. Clinical role of lipid transfer proteins in food
allergy. Mol Nutr Food Res 2004;48:356-62.
11. Pascal M, Munoz-Cano R, Reina Z, Palacin A, Vilella R, Picado C, et al. Lipid
transfer protein syndrome: clinical pattern, cofactor effect and prole of molecular
sensitization to plant-foods and pollens. Clin Exp Allergy 2012;42:1529-39.
12. González-Mancebo E, González-de-Olano D, Trujillo MJ, Santos S, Gandolfo-
Cano M, Meléndez A, et al. Prevalence of sensitization to lipid transfer proteins
and prolins in a population of 430 patients in the south of Madrid. J Investig
Allergol Clin Immunol 2011;21:278-82.
13. Sampson HA, Munoz-Furlong A, Campbell RL, Adkinson NF Jr, Bock SA,
Branum A, et al. Second symposium on the denition and management of
anaphylaxis: summary reportsecond National Institute of Allergy and Infec-
tious Disease/Food Allergy and Anaphylaxis Network symposium. Ann Emerg
Med 2006;47:373-80.
14. Cox LS, Sanchez-Borges M, Lockey RF. World Allergy Organization systemic
allergic reaction grading system: is a modication needed? J Allergy Clin
Immunol Pract 2017;5:58-62.e5.
15. Schafer JL. Analysis of Incomplete Multivariate Data. London: Chapman &
Hall; 1997.
16. Fernandez-Rivas M. Fruit and vegetable allergy. Chemical Immunol Allergy
2015;101:162-70.
17. Pascal M, Vazquez-Ortiz M, Folque MM, Jimenez-Feijoo R, Lozano J,
Dominguez O, et al. Asymptomatic LTP sensitisation is common in plant-food
allergic children from the Northeast of Spain. Allergol Immunopathol (Madr)
2016;44:351-8.
18. Faber MA, Van Gasse AL, Decuyper II, Uyttebroek A, Sabato V,
Hagendorens MM, et al. IgE-reactivity proles to nonspecic lipid transfer
proteins in a northwestern European country. J Allergy Clin Immunol 2017;139:
679-682.e5.
19. Azofra J, Berroa F, Gastaminza G, Saiz N, Gamboa PM, Vela C, et al. Lipid
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Immunol 2016;169:181-8.
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DECUYPER ET AL 7
ONLINE REPOSITORY
METHODS
Patient stratification. Pollen sensitization was dened by at
least 1 positive specic IgE (sIgE) result to the following re-
combinant (r) or native (n) allergens: rBet v 1 and rBet v 2 from
birch (Betula verrucosa), rPhl p 1 and rPhl p 5b from timothy
grass (Phleum pratense), and/or nArt v 1 of mugwort (Artemisia
vulgaris). Nonspecic lipid transfer protein (nsLTP) sensitization
was documented by at least 1 positive sIgE result to the following
nsLTP allergens: rAra h 9 from peanut (Arachis hypogeae), rCor a
8 from hazelnut (Corylus avellana), rMal d 3 from apple (Malus
domesticus), rJug r 3 from walnut (Juglans regia), rPru p 3 from
peach (Prunus persica), nArt v 3 from mugwort (Artemisia vul-
garis), and rPar j 2 from wall pellitory (Parietaria judaica).
Diagnostic methods. Briey, prewarmed heparinized blood
samples were stimulated with rCan s 3 (expressed in Escherichia
coli
E1
) and a crude Cannabis sativa extract (extraction method as
described elsewhere,
E2
obtained by solvent precipitation in the
presence of enzyme inhibitors). Antihuman IgE served as a
positive control (10
m
g/mL, clone G7-18, BD Biosciences,
Erembodegem, Belgium) and stimulation buffer was used to
measure spontaneous CD63 expression by quiescent cells.
Basophil activation test was performed using side scatter, anti-
IgE, and anti-CD203c (clone NP4D6, BD Biosciences) to
characterize the basophils. Subsequently, within this gate, baso-
phil activation was quantied as the percentage
CD203cþþCD63þexpressing basophils using anti-CD63
(clone H5C6, BD Biosciences). Results were expressed as net
percentages of CD63
þ
basophils, calculated by subtraction of the
spontaneous expression from the allergen-induced CD63
expression.
Results
Cannabis allergyerelated symptoms
Performance of cannabis diagnostic tests. As shown
in Figure E1 and Table E2, sIgE rCan s 3s specicity does not
change when the cutoff is increased from 0.10 to 0.35 kU
A
/L.
Specicity of sIgE hemp rises almost 2-fold (32% to 60%)
when a cutoff of 0.35 kU
A
/L is considered.
When only considering CA reporting likely-anaphylaxis (CA-
A), there is no difference in sensitivity between a cutoff of 0.10 vs
0.35 kU
A
/L for either sIgE rCan s 3 or sIgE hemp. In other
words, all sensitized CA-A have an sIgE result 0.35 kU
A
/L for
both tests. When the complete CA group is considered, sensi-
tivity decreases around 10% for both tests: sIgE hemp (82% to
72%; P<.001), sIgE rCan s 3 (47% to 38%; P¼.002).
J ALLERGY CLIN IMMUNOL PRACT
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7.e1 DECUYPER ET AL
FIGURE E1. sIgE results for sIgE hemp and sIgE rCan s 3 showing both cutoff 0.10 kU
A
/L and 0.35 10 kU
A
/L. CA-A, CA reporting likely-
anaphylaxis; CA-C, cutaneous symptoms to cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and
cutaneous symptoms to cannabis; HC, healthy controls; PþLTP, pollen-sensitized controls without an nsLTP sensitization; PþLTPþ,
pollen and nsLTP-sensitized controls; sIgE, specific IgE.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 7.e2
TABLE E1. Symptoms reported on cannabis exposure by CA-A
Patient Symptoms on cannabis exposure
PT1 AE, D, RC
PT2 AE, AP, C, U, W
PT3 AE, D, P
PT4 D, P, RC, U
PT5 AE, D, RC, U
PT6 AD, D, P, RC, U
PT7 AE, D, P, U, W
PT8 AE, C, D, RC
PT9 BP, OAS, RC, U
PT10 D, RC, U
PT11 AE, D, N, V
PT12 D, RC, U
PT13 D, RC, U
PT14 D, OAS, RC, U
PT15 D, RC, U
PT16 AE, D, P, PL, U
PT17 AE, D, OAS
PT18 AE, D, U
PT19 D, RC, U
PT20 OAS, V, U
PT21 D, N, PL, W
PT22 D, P, RC, U
PT23 N, U, V
PT24 AE, D, P, PL, RC, U
PT25 D, RC, P, U
AD, Atopic dermatitis air; AE, angioedema; AP, abdominal pain; BP, blood pres-
sure drop; C, cough; CA-A, CA reporting likely-anaphylaxis; D, dyspnea; N, nausea;
OAS, oral allergy syndrome; P, pruritus; PL, palpitations; RC, rhinoconjunctivitis; U,
urticaria; V, vomiting; W, wheezing.
J ALLERGY CLIN IMMUNOL PRACT
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7.e3 DECUYPER ET AL
TABLE E2. Differences in sIgE results between 0.1 and 0.35 kU
A
/L cutoffs
Cutoff
sIgE hemp
Pvalue
sIgE rCan s 3
Pvalue0.10 kU
A
/L 0.35 kU
A
/L 0.10 kU
A
/L 0.35 kU
A
/L
Sensitivity*CA-A 86 (66-97) 77 (55-92) .5 63 (41-81) 63 (41-81) 1
Sensitivity CA82 (74-89) 72 (63-80) .001 47 (38-56) 38 (29-48) .002
Specicityz32 (20-45) 60 (49-70) <.001 87 (78-93) 89 (83-93) 1
CA, Cannabis allergy; CA-A, CA reporting likely-anaphylaxis; PþLTP, pollen-sensitized controls without an nsLTP sensitization; PþLTP þ, pollen and nsLTP-sensitized
controls; sIgE, specic IgE.
*Calculations based on the CA-A group vs cannabis-tolerant PþLTPand PþLTPþ.
Calculations based on the whole CA group (respiratory and/or cutaneous symptoms) vs cannabis-tolerant PþLTPand PþLTPþ.
zCalculations based on the PþLTPand PþLTPþgroups. Pvalues calculated by McNemar analyses.
TABLE E3. Comparison of Can s 3-sensitized and nonsensitized
CA patients
Can s 3DCA Can s 3LCA Pvalue
Eczema 31% 47% .08
Asthma 30% 35% .42
Cofactor*42% 15% <.01
Systemic reaction to plant foods52% 35% .14
Pollen-sensitized individuals*92% 74% .03
Bet v 1*82% 56% <.01
Bet v 2 14% 20% .46
Phl p 1 54% 62% .45
Phl p 5b 43% 43% 1
Art v 1 17% 8% .15
nsLTP-sensitized individuals*92% 39% <.01
Pru p 3*86% 35% <.01
Mal d 3*87% 30% <.01
Cor a 8*80% 16% <.01
Ara h 9*76% 18% <.01
Jug r 3*80% 28% <.01
Tri a 14*55% 19% <.01
Art v 3*70% 10% <.01
Par j 2 9% 20% .11
Bromelain 37% 20% .07
CA, Cannabis allergy; nsLTP, nonspecic lipid transfer protein; WAO, World
Allergy Organization.
*Signicant differences as measured by P<.05.
Dened as grade 1 or higher as described by the WAO criteria of systemic allergic
reactions.
E3
TABLE E4. Comparison of CA and PþLT P þpatients
CA PDLTPDPvalue
Eczema*37% 54% .01
Asthma 30% 39% .14
Cofactor*31% 19% .05
Systemic reaction to plant foods45% 33% .11
Total IgE (mean)*247.4 424.5 <.01
Pollen-sensitized individuals*84% 100% <.01
Bet v 1 72% 79% .27
Bet v 2 16% 23% .17
Phl p 1*57% 79% <.01
Phl p 5b 43% 54% .09
Art v 1 14% 19% .32
nsLTP-sensitized individuals*72% 100% <.01
Pru p 3*66% 88% <.01
Mal d 3*65% 82% <.01
Cor a 8 56% 61% .50
Ara h 9 54% 62% .23
Jug r 3*60% 79% <.01
Tri a 14 41% 45% .54
Art v 3 47% 42% .45
Par j 2*14% 38% <.01
Can s 3*z63% 35% <.01
Bromelain*30% 45% .02
BAT, Basophil activation test; CA, cannabis allergy; nsLTP, nonspecic lipid transfer
protein; PþLTPþ, pollen and nsLTP-sensitized controls; sIgE, specic IgE; SPT,
skin prick test; WAO, World Allergy Organization.
*Signicant differences as measured by P<.05.
Dened as grade 1 or higher as described by the WAO criteria of systemic allergic
reactions.
E3
zMeasured by sIgE, BAT, or SPT.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 7.e4
REFERENCES
E1. Rihs HP, Armentia A, Sander I, Bruning T, Raulf M, Varga R. IgE-binding
properties of a recombinant lipid transfer protein from Cannabis sativa. Ann
Allergy Asthma Immunol 2014;113:233-4.
E2. Decuyper II, Faber MA, Lapeere H, Mertens C, Rihs HP, Van Gasse AL, et al.
Cannabis allergy: a diagnostic challenge. Allergy 2018;73:1911-4.
E3. González-Mancebo E, González-de-Olano D, Trujillo MJ, Santos S, Gandolfo-
Cano M, Meléndez A, et al. Prevalence of sensitization to lipid transfer proteins
and prolins in a population of 430 patients in the south of Madrid. J Investig
Allergol Clin Immunol 2011;21:278-82.
J ALLERGY CLIN IMMUNOL PRACT
MONTH 2018
7.e5 DECUYPER ET AL
... Although the drug is perceived as harmless, the increased use has rendered a greater awareness of the possible negative effect of cannabis, including the increased risk for asthma [75] and allergic reactions following cannabis consumption [76,77]. Allergic reactions to cannabis can vary from mild rhinoconjunctivitis or urticaria to angioedema or severe systemic reactions [77,78]. The majority of patients with severe and generalized symptoms are sensitized to Can s 3, a major allergen in Cannabis allergy classified as a nonspecific lipid transfer protein (nsLTP) which is found in Cannabis sativa [79]. ...
... Due to cross-reactivity, patients sensitized to Can s 3 are at risk of developing allergic reactions upon exposure to nsLTPs from other plant foods such as Pru p 3 (peach), Mal d 3 (apple), and Jug r 3 (walnut). Furthermore, the opposite may also occur, that is, patients sensitized to other nsLTPs can develop allergic symptoms upon exposure to cannabis [78], a pattern of cross-reactivity that is known as the cannabis-plant food syndrome [80]. Taken together, these findings suggest that analysis of IgE-ab to Can s 3 and other nsLTPs could be of importance in individuals with unexpected allergic reactions upon consumption of cannabis or plant foods such as peach, apple, and walnut. ...
... Taken together, these findings suggest that analysis of IgE-ab to Can s 3 and other nsLTPs could be of importance in individuals with unexpected allergic reactions upon consumption of cannabis or plant foods such as peach, apple, and walnut. However, in the majority of studies on Cannabis allergy, a diagnosis is documented by skin prick test using buds or leaves which are difficult to standardize [78]. Decuyper et al. [78] propose that a highly sensitive diagnostic test, based on a crude extract (such as sIgE to hemp), should be used when there is clinical suspicion of allergy to cannabis. ...
Article
Full-text available
The fifth class of immunoglobulin, immunoglobulin E (IgE) was discovered in 1967 and has had immense importance for the understanding, diagnosis, and treatment of allergic disease. More than 50 years have passed and efforts to characterize, standardize, and refine allergens with the aim to improve clinical diagnosis and allergen-specific immunotherapy are still ongoing. Another important breakthrough was made in 1999 with the introduction of component-resolved diagnostics (CRD), making it possible to quantify IgE antibodies against individual allergen proteins for diagnostic purposes at a molecular level. The progress and developments made in allergy diagnosis often originate from clinical observations and case studies. Observant physicians and health-care personnel have reported their findings in the medical literature, which in turn has inspired researchers to become involved in clinical research. Allergists continuously encounter new allergies and are often asked by their patients how to prevent new reactions. In the current article, we focus on recent clinical observations that can now be explained by CRD. The examples taken concern allergic reactions toward peanuts, tree nuts, lemon kernels, health drinks, meat, insects, dog dander, cannabis, and semen. We now have an improved understanding of why patients may react in a serious or unexpected way, as illustrated by these examples, yet many other clinical observations remain unexplained. The aim of this review is to highlight the importance of clinical observations among allergic patients, focusing on systemic, or unusual and unexpected allergic reactions, where component-testing has further refined the diagnosis of IgE-mediated allergy.
... Although research on the topic is still in its infancy, and the prevalence of such an allergy remains unknown, different research groups around the world have shown that a cannabis allergy can manifest severe and generalized symptoms with detrimental effects on a patient's health and quality of life. [11][12][13][14] As such, in this article, we aim to describe cannabis allergy and discuss the difficulty discriminating "allergic" symptoms from physiological effects, as well as the challenges associated with diagnosis, and finally debate possible future diagnosis and treatment options. ...
... 18,19 Multiple reports stemming from occupational cannabis exposure have suggested that cutaneous cannabis contact and/or contact with the chemical pesticides used in plantations can induce an irritating reaction 20,21 that can be confused with allergic contact urticaria and/or eczema as seen in cannabis allergic reactions. 11,[20][21][22][23][24][25] Diagnostic challenges for cannabis allergy As with most allergies, a diagnosis of cannabis allergy should first be based on a detailed history and would ideally include a challenge test. The latter is currently a big hurdle for multiple reasons. ...
... 14,28-30 Because some of these allergens belong to a superfamily of panallergens including the nsLTP (nonspecific lipid transfer protein) family, profilins, and B verrucosa 1 homologues, cannabis-allergic patients can experience cross-allergies with multiple plant-foods and Hevea latex. 11,12,29,31 This type of cannabis allergy can be diagnosed with different allergy diagnostics, such as skin prick tests (SPTs), specific IgE (sIgE) quantification, and basophil activation tests (BATs), using crude extracts or purified/recombinant proteins. 15 As can be expected, test specificity seems to be best using recombinant allergens, 11,13,32 whereas sensitivity is found to be superior when using crude cannabis extracts. ...
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Drug allergy has been a research topic within the allergy field for decades. However, many drug reactions presumed to be of allergic nature, are not and originate from different mechanisms. Drug-induced reactions can affect numerous organ systems, present with a variety of symptoms, and have more than one mechanism of action. In this rostrum article we want to give an overview of the different allergic and non-allergic reactions that can be expected with the (illicit) use of cannabis, cocaine, opioids and alcohol. In addition, this article focuses on the different methods available to diagnose allergy related to these four drug types and highlight the pitfalls of non-allergic reactions or allergy "mimickers" complicating diagnosis of true drug allergy. Finally, the impact on current medical practices is addressed and future research in support of the allergist in diagnosis and treatment of these medical problems.
... [68][69][70][71][72][73][74] Sensitization to Can s 3, the nsLTP in cannabis, is a feature of European patients with cannabis allergy. 75,76 It was first shown in 2007 that a patient sensitized to the peach nsLTP Pru p 3 reacted to Cannabis sativa; further research confirmed that Can s 3 was a relevant cause of sensitization and reactions to other nsLTP, even via passive inhalation. 64,75,[77][78][79] Thus Cannabis sativa could act as a primary sensitizing allergen, with a significant percentage of those allergic to cannabis reporting severe allergy to plant foods. ...
... 75,76 It was first shown in 2007 that a patient sensitized to the peach nsLTP Pru p 3 reacted to Cannabis sativa; further research confirmed that Can s 3 was a relevant cause of sensitization and reactions to other nsLTP, even via passive inhalation. 64,75,[77][78][79] Thus Cannabis sativa could act as a primary sensitizing allergen, with a significant percentage of those allergic to cannabis reporting severe allergy to plant foods. 75,8 0-83 Due to cross-reactivity between nsLTPs, patients sensitized to Can s 3 may develop sensitization and relevant clinical symptoms to a wide range of fruits, vegetables, and cereals, and also to wine, beer, Hevea latex, and tobacco ( Figure 2). ...
Article
Cannabis is the most widely used recreational drug in the world. Cannabis sativa and Cannabis indica have been selectively bred to develop their psycho‐active properties. The increasing use in many countries has been accelerated by the COVID‐19 pandemic. Cannabis can provoke both type 1 and type 4 allergic reactions. Officially recognised allergens include a pathogenesis‐related class 10 allergen, profilin and a non‐specific Lipid Transfer Protein. Other allergens may also be relevant, and recognition of allergens may vary between countries and continents. Cannabis also has the potential to provoke allergic cross‐reactions to plant foods. Since cannabis is an illegal substance in many countries, research has been hampered, leading to challenges in diagnosis since no commercial extracts are available for testing. Even in countries such as Canada, where cannabis is legalized, diagnosis may rely solely on the purchase of cannabis for prick‐to‐prick skin tests. Management consists of avoidance, with legal issues hindering the development of other treatments such as immunotherapy. Education of healthcare professionals is similarly lacking. This review aims to summarise the current status of cannabis allergy and proposes recommendations for the future management of this global issue.
... Mediterranean), such as northern Europe. [85][86][87] An important factor able to influence the clinical outcome of patients sensitized to LTP is the presence or absence of co-sensitization to PR-10 or profilins. 70,71 Monoreactive LTP patients show more frequently severe reactions than polyreactive individuals. ...
... Finally, LTP sensitization may be initiated by the inhalation/contact of Cannabis LTP, Can s 3, even via passive smoke, as demonstrated even in paediatric patients who had developed LTP-triggered anaphylaxis after a second-hand exposure to cannabis smoke.82 Cannabis LTP sensitization might precede the development of food allergy[83][84][85][86][87][88] ...
Article
Sensitization to lipid transfer protein (LTP), the most frequent cause of food allergy in southern Europe still shows several controversial, but also intriguing aspects. Some of these include the degree of cross‐reactivity between LTPs from botanically distant sources, the definition of risk factors, the role of some cofactors, clinical outcomes, geographical differences and the identification of the primary sensitizer in different areas. This review article tries to analyze and comment on these aspects point by point suggesting some explanatory hypotheses with the final scope to stimulate critical thoughts and elicit the scientific discussion about this issue in the readership.
... Since nonspecific lipid transfer proteins are widespread in vegetables and fruits, there is the possibility of a cross-allergy between marijuana and many food products such as peaches, tomatoes, bananas, and apples. This is the so-called "Cannabis-fruit/vegetable syndrome" (Nayak et al., 2013;Decuyper et al., 2015Decuyper et al., , 2019Jackson et al., 2020). ...
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The global incidence of respiratory diseases and complications is increasing. Therefore, new methods of treatment, as well as prevention, need to be investigated. A group of compounds that should be considered for use in respiratory diseases is cannabinoids. There are three groups of cannabinoids – plant-derived phytocannabinoids, synthetic cannabinoids, and endogenous endocannabinoids including the enzymes responsible for their synthesis and degradation. All cannabinoids exert their biological effects through either type 1 cannabinoid receptors (CB1) and/or type 2 cannabinoid receptors (CB2). In numerous studies (in vitro and in vivo), cannabinoids and inhibitors of endocannabinoid degradation have shown beneficial anti-inflammatory, antioxidant, anti-cancer, and anti-fibrotic properties. Although in the respiratory system, most of the studies have focused on the positive properties of cannabinoids and inhibitors of endocannabinoid degradation. There are few research reports discussing the negative impact of these compounds. This review summarizes the properties and mechanisms of action of cannabinoids and inhibitors of endocannabinoid degradation in various models of respiratory diseases. A short description of the effects selected cannabinoids have on the human respiratory system and their possible use in the fight against COVID-19 is also presented. Additionally, a brief summary is provided of cannabinoid receptors properties and their expression in the respiratory system and cells of the immune system.
... 14,72-74 Research from northern/central Europe suggests marijuana inhalation is an effective entry route to nsLTP sensitization in the absence of Pru p 3-mediated allergy.[75][76][77] Decuyper and colleagues reported that 45% of 120 patients allergic to C. sativa and sensitized to Can s 3, and the nsLTP in C. sativa had severe and generalized allergy to plant foods.78 Passive inhalation of smoke from C. sativa can also lead to sensitization to Can s 3, and other nsLTP allergens.79 ...
Article
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Background Discovered and described 40 years ago, non‐specific lipid transfer proteins (nsLTP) are present in many plant species and play an important role protecting plants from stressors such as heat or drought. In the last 20 years, sensitization to nsLTP and consequent reactions to plant foods has become an increasing concern. Aim The aim of this paper is to review the evidence for the structure and function of nsLTP allergens, and cross‐reactivity, sensitization, and epidemiology of nsLTP allergy. Materials and Methods A Task Force, supported by the European Academy of Allergy & Clinical Immunology (EAACI), reviewed current evidence and provide a signpost for future research. The search terms for this paper were “Non‐specific Lipid Transfer Proteins”, “LTP syndrome”, “Pru p 3”, “plant food allergy”, “pollen‐food syndrome”. Results Most nsLTP allergens have a highly conserved structure stabilised by 4‐disulphide bridges. Studies on the peach nsLTP, Pru p 3, demonstrate that nsLTPs are very cross‐reactive, with the four major IgE epitopes of Pru p 3 being shared by nsLTP from other botanically related fruits. These nsLTP allergens are to varying degrees resistant to heat and digestion, and sensitization may occur through the oral, inhaled or cutaneous routes. In some populations, Pru p 3 is the primary and sole sensitizing allergen, but many are poly‐sensitised both to botanically un‐related nsLTP in foods, and non‐food sources of nsLTP such as Cannabis sativa, Platanus acerifolia, (plane tree), Ambrosia artemisiifolia (ragweed) and Artemisia vulgaris (mugwort). Initially, nsLTP sensitization appeared to be limited to Mediterranean countries, however more recent studies suggest clinically relevant sensitization occurs in North Atlantic regions and also countries in Northern Europe, with nsLTP sensitisation profiles being broadly similar. Discussion These robust allergens have the potential to sensitize and provoke symptoms to a large number of plant foods, including those which are raw, cooked or processed. It is unknown why some sensitized individuals develop clinical symptoms to foods whereas others do not, or indeed what other allergens besides Pru p 3 may be primary sensitising allergens. It is clear that these allergens are also relevant in non‐Mediterranean populations and there needs to be more recognition of this. Conclusion Non‐specific LTP allergens, present in a wide variety of plant foods and pollens, are structurally robust and so may be present in both raw and cooked foods. More studies are needed to understand routes of sensitization and the world‐wide prevalence of clinical symptoms associated with sensitization to these complex allergens.
... There are currently no known allergens of Cannabis indica listed in the database. It is estimated that only 72% of patients with a reported allergy to cannabis are sensitized to Can s 3 [10]. Published data is not currently available regarding the allergenicity of Can s 4. One study looking at genotyping of various cannabis strains showed that C. sativa and C. indica have distinguishable pools of genetic diversity and there is only a moderate correlation between the genetic structure of marijuana strains and their reported C. sativa and C. indica ancestry [11]. ...
Article
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Background Cannabis use is growing domestically due to recent legalization in many jurisdictions. There are two main species of cannabis, Cannabis sativa and Cannabis indica , and thousands of different commercially available cannabis strains. Although there are multiple reports of cannabis allergy in the literature, to our knowledge, there is no prior published report of selective cannabis strain allergy. Case presentation A 31-year-old male was referred for allergy assessment due to several episodes of localized pruritus and erythema after direct contact with various strains of cannabis. He had noted that the severity of his reaction appeared to be strain dependent. He developed a severe local reaction involving bilateral periorbital edema shortly after coming into direct contact with one particular strain of cannabis. He denied any adverse symptoms after inhalation of cannabis. Fresh skin prick testing was performed to various strains of cannabis and had positive testing to the three of the five tested strains. Conclusions We believe this is the first reported case of selective cannabis strain allergy based on patient history and skin prick testing. This case report outlines the variability in different strains of cannabis and stresses the importance of further research into cannabis allergen identification. Multiple cannabis allergens should be included and incorporated into commercial extracts when they become routinely available.
Article
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Sensitization to one or more non‐specific lipid transfer proteins(nsLTP), initially thought to exist mainly in southern Europe,is becoming accepted as a cause of allergic reactions to plant foods across Europe and beyond.The peach nsLTP allergen Pru p 3 is a dominant sensitizing allergen, and peaches a common food trigger, although multiple foods can be involved. A frequent feature of reactions is the requirement for a co‐factor (exercise, alcohol, non‐steroidal anti‐inflammatory drugs, Cannabis sativa)to be present for a food to elicit a reaction. The variability in the food and co‐factor triggers make it essential to include an allergy focused diet and clinical history in the diagnostic work‐up. Testing on suspected food triggers, should also establish whether sensitization to nsLTP is present, usingpurified or recombinant nsLTP allergens such as Pru p 3. Theavoidance of known trigger foods and advice on co‐factors is currently the main management for this condition. Studies on immunotherapy are promising, but it is unknown whether such treatments will be useful in populations where Pru p 3 is not the primary sensitizing allergen. Future research should focus on the mechanisms of co‐factors, improving diagnostic accuracy and establishing the efficacy of immunotherapy.
Article
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Background: The sensitisation profile at molecular level in plant-food allergy is complex. Several allergens may be involved, with different potential for severe reactions. lipid transfer proteins (LTP) are considered the most relevant plant-food allergens in adults in Mediterranean countries, but less is known in children. Aim: To describe the clinical pattern and sensitisation profile of children with plant-food allergy and LTP sensitisation from Northeast Spain. Methods: Children with history of immediate reaction to plant-food(s), positive skin-prick-test to the culprit plant-food(s) and specific-IgE to plant-food LTPs were analysed. Results: 130 children were included. 69.2% (90/130) had reacted to ≥2 taxonomically unrelated plant-foods. Peach, walnut, hazelnut and peanut were most frequently involved. Reactions severity ranged from anaphylaxis (45.4%, 59/130) to oral symptoms only. Sensitisation to a particular plant-food LTP not always caused clinical symptoms with that plant-food; 69% (40/58) and 63% (17/27) of peach- and walnut-tolerant subjects had positive rPru p 3 and nJug r 3 specific IgE, respectively. 65.4% (85/130) of children were also sensitised to storage proteins, which was associated to anaphylaxis and nut allergy. However, 60% of patients without nuts/seeds allergy were sensitised to storage proteins. Specific-IgE levels to LTPs and/or storage proteins were not useful to predict allergy (vs. tolerance) to peach, walnut, peanut or hazelnut. Conclusions: Sensitisation to LTP and/or storage proteins without clear clinical significance is relatively common. Prospective longitudinal studies are required to evaluate the relevance of these silent sensitisations over time. Caution is required when interpreting the results of molecular-based diagnostic tools in clinical practice.
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Background Due to the increasing social, medical and occupational exposure to Cannabis sativa, allergic reactions grow in frequency but little is known about the IgE-reactivity of single Cannabis allergens. Methods To identify the mature peptide sequence of the lipid transfer protein (LTP) and to study the IgE-binding reactivity of a recombinant Can s 3 (rCan s 3), a cDNA was synthesized from total RNA of Cannabis sativa L. ssp. sativa cv. Kompolti leaves obtained from the botanical garden of the University Bonn. The LTP gene was amplified with a primer mix deduced from published amino acid sequences. The LTP variant was identified in five independent clones by sequencing in the pDrive vector system followed by the expression of the mature LTP in the pMAL-vector. After isolation of a soluble recombinant maltose-binding protein (MBP)-Can s 3 fusion protein, aliquots were biotinylated and coupled to Streptavidin-ImmunoCAPs. Results Sera of 16 (6 Spanish and 10 German) subjects with allergic symptoms to Cannabis were tested. Specific IgE (sIgE) values >0.35 kUA/L were regarded as positive. Twelve out of 16 sera (75%) showed sIgE to Cannabis (range: 0.42-31.80 kUA/L). Five of them (31%) displayed sIgE to rCan s 3 (range: 0.40-14.10 kUA/L) but no sIgE to MBP. Specific IgE-reactivity to Pru p 3 was detected in all Cannabis-positive sera from Spain but only in 3 out of 6 German sera. All sera with sIgE to rCan s 3 showed also sIgE to Pru p 3. Conclusions These results show for the first time IgE-binding of a recombinant Cannabis allergen (rCan s 3). Due to the 60% amino acid identity between Can s 3 and Pru p 3 a cross-reactivity is possible, but also to the LTPs of other plants. Since rCan s 3 is now available for sIgE diagnostics, implementation in larger studies may help to further elucidate the impact of this allergen.
Article
There is no universally accepted grading system to classify the severity of systemic allergic reactions (SARs), including anaphylaxis. Although a consensus definition for anaphylaxis was established in 2005, the signs and symptoms required to define a reaction as anaphylaxis are inconsistently applied in research and clinical practice. As a result, it is difficult to compare and evaluate safety outcomes in surveys, clinical practice and trials, and pharmacovigilance data. In 2010, the World Allergy Organization (WAO) proposed a uniform grading system to classify allergen immunotherapy SARs. The basis of the grading system is the organ system(s) involved and reaction severity. The final grade is determined by the physician/health care professional after the event is over. Although the 2010 WAO grading system was developed to classify allergen immunotherapy SARs, with appropriate modifications, it can be used to classify SARs from any cause. The purpose of this Rostrum is to present a proposed modification of the 2010 WAO SAR grading system that will make it applicable to all SARs due to any cause. The modified grading system allows for classification of less severe SARs, which may be underreported or overreported in clinical trials and surveillance studies, depending on the criteria specified for adverse event reporting. The universal use of the proposed modified SAR grading system will allow for better safety comparisons across different venues and treatment protocols.
Article
Background: Plant food allergies associated with lipid transfer protein (LTP) have been widely described in the Mediterranean Basin. Objective: The aim of this work was to describe the clinical profile and pollen sensitization of plant food- allergic patients sensitized to LTP in a non-Mediterranean area. Methods: Patients with clear IgE-mediated symptoms associated with plant foods and a positive skin prick test (SPT) to Pru p 3 were included in a prospective study in the north of Spain. Reported symptoms were analyzed together with a battery of food and pollen SPTs and specific IgE components by ISAC microarray. Cross-inhibition studies were performed by ImmunoCAP with plane tree, mugwort and rPru p 3. Results: Among the 72 patients included, the most frequent food allergy reported was to peaches (69%) followed by nuts (walnuts 55%, peanuts 54% and hazelnuts 43%). Most patients suffered from symptoms with multiple plant foods (a median of 6 foods per patient). Regarding the patients' pollen sensitization, 36% were sensitized to mugwort pollen (72% showing sIgE to Art v 3), 33% to grass pollen and 24% to plane tree pollen (94% with sIgE to Pla a 3). Inhibition studies showed that specific IgEs against mugwort and plane tree pollen are inhibited by Pru p 3 in a strong manner, whereas Pru p 3 was less inhibited by pollen extracts. Conclusions: LTP syndrome occurs in a non-Mediterranean area and is related to multiple sensitizations to foods and pollens such as plane tree and mugwort. In these pollen sensitizations, Pru p 3 seems to be the primary sensitizer.
Article
Fruit and vegetable allergies are the most prevalent food allergies in adolescents and adults. The identification of the allergens involved and the elucidation of their intrinsic properties and cross-reactivity patterns has helped in the understanding of the mechanisms of sensitisation and how the allergen profiles determine the different phenotypes. The most frequent yet contrasting fruit and vegetable allergies are pollen-food syndrome (PFS) and lipid transfer protein (LTP) syndrome. In PFS, fruit and vegetable allergies result from a primary sensitisation to labile pollen allergens, such as Bet v 1 or profilin, and the resulting phenotype is mainly mild, consisting of local oropharyngeal reactions. In contrast, LTP syndrome results from a primary sensitisation to LTPs, which are stable plant food allergens, inducing frequent systemic reactions and even anaphylaxis. Although much less prevalent, severe fruit allergies may be associated with latex (latex-fruit syndrome). Molecular diagnosis is essential in guiding the management and risk assessment of these patients. Current management strategies comprise avoidance and rescue medication, including adrenaline, for severe LTP allergies. Specific immunotherapy with pollen is not indicated to treat pollen-food syndrome, but sublingual immunotherapy with LTPs seems to be a promising therapy for LTP syndrome. © 2015 S. Karger AG, Basel.
Article
Background: Although allergy to Cannabis sativa was first reported over 40 years ago, the allergenicity has scarcely been studied. The objectives of this study were to investigate the frequency of sensitization to this plant, to analyze the clinical characteristics and allergenic profile of sensitized individuals and to identify the allergens involved. Methods: Five hundred and forty-five individuals in Spain attending allergy clinics with respiratory or cutaneous symptoms underwent a skin-prick test (SPT) with C. sativa leaf extract. The extract was characterized by SDS-PAGE and 2-dimensional electrophoresis. Specific IgE to C. sativa was measured in positive SPT individuals. The clinical and allergenic profiles of sensitized individuals were investigated and the most-recognized allergens sequenced and characterized by liquid chromatography-mass spectrometry/mass spectrometry. Results: Of this preselected population, 44 individuals had positive SPT to C. sativa (prevalence 8.1%). Prevalence was higher in individuals who were C. sativa smokers (14.6%). Two individuals reported mild symptoms with C. sativa. Twenty-one individuals from 32 available sera (65.6%) had positive specific IgE to C. sativa. Twelve sera recognized at least 6 different bands in a molecular-weight range of between 10 and 60 kDa. Six of them recognized a 10-kDa band, identified as a lipid transfer protein (LTP) and 8 recognized a 38-kDa band, identified as a thaumatin-like protein. Conclusions: There is a high prevalence of sensitization to C. sativa leaves. The clinical symptoms directly attributed to C. sativa were uncommon and mild. The sensitization profile observed suggests that C. sativa sensitization may be mediated by two mechanisms, i.e. cross-reactivity, mainly with LTP and thaumatin-like protein, and exposure-related 'de novo' sensitization.