Received: 13 April 2020 | Revised: 25 July 2020 | Accepted: 14 August 2020

DOI: 10.1111/pai.13352

ORIGINAL ARTICLE

A randomized trial of oral immunotherapy for pediatric cow's

milk-induced anaphylaxis: Heated vs unheated milk

Ken-ichi Nagakura1,2 | Sakura Sato3,4 | Yoko Miura1 | Makoto Nishino1,4 |

Kyohei Takahashi1,4 | Tomoyuki Asaumi1 | Kiyotake Ogura1,4 |
Motohiro Ebisawa2,3 | Noriyuki Yanagida1

1
Department of Pediatrics, National Hospital
Organization Sagamihara National Hospital, Abstract
Kanagawa, Japan Background: Severe reactions may develop during cow's milk (CM) oral immunother-
2
Department of Pediatrics, Jikei University
apy (OIT). We investigated the safety and efficacy of low-dose OIT with heated milk
School of Medicine, Tokyo, Japan
3
Department of Allergy, Clinical Research
(HM) or unheated milk (UM) in children with anaphylaxis.
Center for Allergy and Rheumatology, Methods: Children with symptom onset after ingestion of 3-mL HM on a double-
National Hospital Organization Sagamihara
National Hospital, Kanagawa, Japan
blind, placebo-controlled food challenge were randomly assigned to the HM (n = 17)
4
Course of Allergy and Clinical Immunology, or UM (n = 16) group. HM group ingested milk powder heated at 125°C for 30 sec-
Juntendo University Graduate School of onds, whereas the UM group used UM. Patients were hospitalized for 5 days; the HM
Medicine, Tokyo, Japan
or UM was gradually increased to 3 mL/day; 3-mL/day ingestion was continued at
Correspondence home. One year later, the patients underwent 2-day consecutive 3- and 25-mL HM-
Noriyuki Yanagida, Department of
Pediatrics, National Hospital Organization oral food challenges (OFCs) after 2-week avoidance.
Sagamihara National Hospital, 18-1, Results: At baseline, milk- and casein-specific immunoglobulin E (IgE) levels were
Sakuradai, Minami-ku, Sagamihara,
Kanagawa 252-0392, Japan. 56.0 and 51.4 kUA/L in the HM group, and 55.2 and 65.6 kUA/L in the UM group,
Email: [email protected] respectively. One year later, 35% and 18% in the HM group and 50% and 31% in
Funding information UM group passed the 3 and 25 mL OFCs, respectively. Rates of moderate or severe
This work was supported by the Japan symptoms and respiratory symptoms per home dose were significantly lower in the
Dairy Association (J-milk) and the Practical
Research Project for Allergic Disease and HM than in the UM group (0.7% and 1.2% vs 1.4% and 2.6%, respectively, P < .001).
Immunology from the Japan Agency for β-lactoglobulin-specific IgG4 levels significantly increased from baseline only in the
Medical Research and Development (grant
no. 17ek0410019h0003). The funders UM group, whereas casein-specific IgG4 levels significantly increased from baseline
played no role in the study design, data in both groups.
collection and analysis, decision to publish,
or preparation of the manuscript. Conclusions: HM-OIT induced immunological changes more safely than the UM-OIT.
The possibility of lower treatment efficacy with HM-OIT needs to be evaluated in
larger studies.
Editor: Alexandra Santos

KEYWORDS

anaphylaxis, casein, cow's milk allergy, desensitization, heated, milk, oral immunotherapy,
randomized controlled trial, unheated, β-lactoglobulin

This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction
in any medium, provided the original work is properly cited and is not used for commercial purposes.
© 2020 The Authors. Pediatric Allergy and Immunology published by John Wiley & Sons Ltd

Pediatr Allergy Immunol. 2020;00:1–9.  wileyonlinelibrary.com/journal/pai | 1

2 | NAGAKURA et al.

1 | I NTRO D U C TI O N
Key Message
Cow's milk (CM) allergy is one of the main causes of food allergy, with
Although oral immunotherapy using heated milk appeared
an estimated prevalence of 2%-3%.1,2 In Japan, CM allergy is the sec-
to be safer than unheated milk, treatment efficacy might
ond commonest cause of anaphylaxis.3 Approximately 80% of children
be weaker with heated milk.
with CM allergy acquire tolerance, but school-aged children with a high
level of specific IgE (sIgE) and a history of anaphylaxis face difficulty in
acquiring tolerance.4,5 In a Spanish survey, 40% of children diagnosed
with CM allergy reacted on accidental exposure to CM during a 12- were randomized 1:1 to the HM-OIT or UM-OIT group, by using a ran-
6
month period. Therefore, patients with severe CM allergy are at risk of dom number generator. Randomization was stratified by a threshold
reactions following accidental exposure. Oral immunotherapy (OIT) has of double-blind, placebo-controlled food challenge (DBPCFC; ≤0.75 or
been reported for food allergy, but adverse reactions, including anaphy- >0.75 mL).
laxis, may occur during OIT.7,8 It is more challenging to acquire tolerance
with CM-OIT, and there is a greater likelihood of inducing symptoms
with CM-OIT than with other food antigens.8,9 Moreover, there are only 2.2 | Eligibility criteria
a few studies on OIT among patients with CM anaphylaxis.7,10
The heating process induces conformational changes of the CM epi- Participants were 5 years old or older and had a history of milk
tope—in particular, whey proteins such as β-lactoglobulin, which trigger anaphylaxis.15,16 The inclusion criterion was the development of
11
reactions in some patients with CM allergy. To improve safety of OIT, symptoms during DBPCFC with 3-mL HM. The exclusion criteria
some reports have assessed OIT by using baked or heated milk (HM).12- were negative DBPCFCs, uncontrolled atopic dermatitis, bronchial
14
In 2017, a randomized controlled trial (RCT) showed that safety did asthma, or another ongoing immunotherapy.
not differ with the use of baked milk, although the protocol and base-
line characteristics were different for the baked and raw milk groups.14
Furthermore, a comparison of the safety and efficacy of HM and un- 2.3 | Materials
heated milk (UM) with the same protocol has not been reported thus far.
We hypothesized that OIT with HM would progress more safely than For the HM group of children, we used milk powder, prepared by heat-
with UM. We conducted this study to investigate the safety and efficacy ing CM at 125°C for 30 seconds and spray-drying for 3 seconds. For
of OIT by using HM vs UM in children with anaphylactic-type CM allergy. the UM group of children, we used unheated CM, which is sterilized
at 125°C for 2 seconds according to food safety regulations, which is
considered to be ultra-high-temperature instantaneous sterilization.
2 | M E TH O DS

2.1 | Study design 2.4 | DBPCFC

This RCT was conducted at the Sagamihara National Hospital The DBPCFC was undertaken during two separate days of hospi-
(Kanagawa, Japan) between July 2016 and March 2018. Participants talization. We used cocoa cake containing 3-mL CM (102 mg CM

F I G U R E 1 Oral immunotherapy
protocol. Children received 3- and 25-mL
oral food challenge after 2-wk avoidance
at 12 mo. DBPCFC, double-blind, placebo-
controlled food challenge; HM, heated
milk; OFC, oral food challenge; UM,
unheated milk
NAGAKURA et al. | 3

F I G U R E 2 Patient inclusion flowchart. One patient in the UM group discontinued OIT because of eosinophilic esophagitis at 1 mo.
One patient in HM group and three patients in UM group did not reach desensitization because of adverse reactions (one due to mucosal
symptom in HM group, two due to respiratory symptom, and one due to mucosal symptoms in the UM group) at home. For patients who
passed the 3-mL OFC, we allowed ingestion of 10 g butter. DBPCFC, double-blind, placebo-controlled food challenge; HM, heated milk;
OFC, oral food challenge; OIT, oral immunotherapy; UM, unheated milk

protein) or a placebo with a cocoa cake without CM. The cocoa premedication of 10 mg loratadine, and children consumed HM or
cake was heated to 90°C for 90 seconds in a 1000-W microwave. UM at half the threshold of the baseline DBPCFC (Figure 1). The OIT
One quarter of OFC food was provided initially, and the remain- comprised eight steps, from 0.1 to 3 mL (Table S2). If symptoms were
ing three quarters were provided 60 minutes later, as previously absent or mild, the OIT dose for the next day remained the same.
17
reported. Treatment was provided appropriately based on the If moderate or severe symptoms developed, the dose for the next
severity of reaction, according to the Japanese anaphylaxis guide- day was reduced by 1 or 2 steps, respectively. After discharge, the
lines (Table S1). 3 The threshold dose was defined as the accumu- starting dose was continued at home every day for 1 month. After
lated dose which patients had ingested at the time of symptom 1 month, if patients were able to asymptomatically ingest HM or UM
onset. for five consecutive days, the dose was increased at the patients'
home by 1 step up to 3 mL per day. We offered direct telephone
support for OIT patients 24 hours a day, 365 days a year. If chil-
2.5 | OIT protocol dren were able to asymptomatically ingest 3 mL of the material for
1 month, loratadine treatment was terminated; this was defined as
Patients were hospitalized for 5 days, and DBPCFC was con- “desensitization to 3 mL.” After 12 months from the start of OIT, a
ducted on the first and second day; if positive, patients were ran- 2-day consecutive 3- and 25-mL HM-OFC was conducted for both
domized, and OIT was started from the third day, together with a groups after 2 weeks of OIT cessation.
4 | NAGAKURA et al.

TA B L E 1 Patient characteristics in the

HM group (n = 17) UM group (n = 16) P-value
HM and UM groups
Age (y) 7.6 (5.2-11.2) 6.1 (5.3-10.8) .052
Male, n (%) 14 (82.4%) 11 (68.8%) .43
Current complications
BA, n (%) 9 (53%) 8 (50%) >.99
AD, n (%) 10 (59%) 10 (63%) .82
AR, n (%) 4 (24%) 4 (25%) >.99
a
History of anaphylaxis to milk 2 (1-3) 2 (1-5) .17
Most recent anaphylaxis to milk
Period until entry (mo) 13 (3-60) 9.5 (4-62) .51
HM caused anaphylaxis, n (%) 16 (94%) 14 (88%) .87
DBPCFC
Threshold to induce ≤0.75 mL: 10 (59%) ≤0.75 mL: 10 (63%) >.99
symptoms, n (%) >0.75 mL: 7 (41%) >0.75 mL: 6 (38%)
Severity of symptoms Mild 4, Moderate 13 Mild 2, Moderate 13, .65
Severe 1
Total IgE (IU/mL) 1140 (146-11 000) 648 (69-6770) .20
Specific IgE (kUA /L)
Milk 56.0 (4.3-2630) 55.2 (12.5-745) >.99
Casein 51.4 (2.9-2950) 65.6 (8.5-645) .91
α-lactalbumin 8.8 (0.05-46.3) 9.7 (0.11-49.7) .62
β-lactoglobulin 1.9 (0.05-298) 7.5 (0.05-68.2) .27
Specific IgG (mgA/L)
Casein 7.4 (3.4-29.1) 7.7 (2.7-30.9) .99
β-lactoglobulin 3.0 (1.0-8.2) 2.9 (1.0-10.8) .73
Specific IgG4 (mgA/L)
Casein 0.52 (0.14-7.09) 0.84 (0.13-7.52) .61
β-lactoglobulin 0.08 (0.03-0.38) 0.11 (0.03-1.03) .42

Note: All patients' BA, AD, and AR were well controlled during OIT protocol.
If symptoms occurred during DBPCFC, intake was discontinued, and the accumulated dose was
calculated.
Abbreviations: DBPCFC, double-blind, placebo-controlled food challenge; HM, heated milk; IgE,
immunoglobulin E; UM, unheated milk.
a
Anaphylaxis was defined by the World Allergy Organization guidelines.15,16

2.6 | Outcomes Scientific/Phadia AB, Uppsala, Sweden) at baseline and after 1, 3, 6,

and 12 months in both groups.
The primary endpoint was the rate of total number of symptoms per
total number of ingestions at home during the 12-month study. The
secondary endpoints were symptom severity, symptoms by organ, 2.8 | Statistical analysis
the proportion of desensitization to 3 mL, passing the 3- and 25-mL
OFC, and immunological changes. We hypothesized that the primary outcome, which was that the rate
of symptoms would be reduced by 15% with HM, would be 17%
and 20% in the HM and UM groups, respectively, with a power of
2.7 | Immunological parameters 80%. Therefore, we estimated that 30 participants with a ratio of 1:1
would be sufficient to detect this difference. Values are expressed
The sIgE to milk, casein, α-lactalbumin and β-lactoglobulin; specific as the median and range. Differences in categorical data were evalu-
IgG (sIgG); and specific IgG 4 (sIgG 4) to casein and β-lactoglobulin ated with Fisher's exact test. Continuous variables were evaluated
were measured using the ImmunoCAP assay system (Thermo Fisher with Wilcoxon rank-sum tests. A P-value of <.05 was considered
NAGAKURA et al. | 5

statistically significant. Statistical analyses were conducted in SPSS TA B L E 2 Adverse symptoms and treatment at home
version 24.0 software (IBM Corp., Armonk, NY, USA).
HM group UM group
(n = 17) (n = 16) P-value

Number of intakes 4916 4383

2.9 | Ethics of OIT
Number of adverse 396 (8.1%) 419 (9.6%) .01
This study was conducted in accordance with the principles of the symptoms, n (%)
Declaration of Helsinki and was approved by the Sagamihara National Severity of symptoms
Hospital Ethics Committee (no.: 2016-003). This trial was registered Mild 363 (7.4%) 357 (8.1%) .17
at the University Hospital Medical Information Network Clinical Trials Moderate 32 (0.7%) 62 (1.4%) .0002
Registry (no: UMIN000011202). Written informed consent was ob-
Severe 1 (0.02%) 0 (0.0%) -
tained from the guardians following an explanation of the study de-
Organ system of symptoms
sign and risk of symptoms. We anonymized all data before analysis.
Skin 134 (2.7%) 127 (2.4%) .61
Mucosal 285 (5.8%) 215 (4.9%) .06
Respiratory 59 (1.2%) 105 (2.6%) <.0001
3 | R E S U LT S
Gastrointestinal 111 (2.3%) 56 (1.3%) .0003

3.1 | Study population Cardiovascular 0 (0.0%) 1 (0.02%) -

Anaphylaxis 1 (0.02%) 2 (0.05%) -
During screening, 35 children underwent the baseline DBPCFC, but Total number of 107 (2.2%) 103 (2.3%) .57
two passed the test and were excluded. Thirty-three children with symptoms requiring
any treatments
CM anaphylaxis were randomized to the HM or UM group (n = 17
Antihistamines 86 (1.7%) 90 (2.1%) .28
or 16, respectively; Figure 2). Median milk- and casein-specific IgE
levels were 56.0 and 51.4 kUA/L vs 55.2 and 65.6 kUA/L in the HM Corticosteroids 7 (0.1%) 28 (0.4%) <.0001

and UM groups, respectively (Table 1). β2-stimulant 30 (0.6%) 42 (1.0%) .06

inhalation
Adrenaline 1 (0.02%) 1 (0.02%) -

3.2 | Efficacy outcomes Note: The patients' guardians kept a daily record of ingestion,
symptoms, and treatment requirements. Patients visited the hospital
at 1, 3, 6, 9, and 12 mo from the initiation of OIT. At the hospital visit,
One patient in the UM group discontinued OIT because of eosino-
we checked the diary and recorded adverse symptoms. If moderate or
philic esophagitis at 1 month. All other patients completed the pro- severe symptoms developed, the patients' guardians reported these to
tocol in both groups and were included in the final analysis dataset. the investigators via telephone.
Although the sample size is insufficient to compare efficacy, Abbreviations: HM, heated milk; OIT, oral immunotherapy; UM,
one year later, 94% and 75% of patients achieved desensitization unheated milk.

to 3-mL CM in the HM and UM groups (P = .17), respectively,

whereas 35% and 18% in the HM group, and 50% and 31% in The rate of gastrointestinal symptoms, mostly mild, in the HM
the UM group, passed the 3- and 25-mL OFC (P = .34, P = .43; group was significantly higher than in the UM group (Table 2). In
Figure 2), respectively. the UM group, one patient developed diarrhea and hematochezia;
esophagogastroduodenoscopy revealed eosinophilic gastroenteritis.
There were no significant between-group differences in the
3.3 | Safety per-patient rates of adverse symptoms (Table S4). Table S5 showed
the adverse symptom rate over the treatment period, and rates
The only adverse symptoms recorded were mild adverse symptoms from OIT initiation to 3 months; rates from 3 to 6 months in the HM
during hospitalization; rates of total adverse symptoms were 20.6% group were significantly lower than those in the UM group (P = .002,
and 32.4% in the HM and UM groups, respectively, without any sig- P < .0001, respectively).
nificant intergroup difference (P = .20; Table S3).
In the home-dosing phase, the total adverse symptom rate per
dose was 8.1% and 9.6% in the HM and UM groups, respectively 3.4 | Laboratory data
(P = .01). Rates of moderate/severe symptoms and respiratory
symptoms were 0.7% and 1.2% vs 1.4% and 2.6% in the HM and Median milk- and casein-specific IgE levels significantly decreased
UM groups (P = .0002, P < .0001), respectively. In the HM and UM from baseline after 3 months and further decreased to 25.8 and 23.7
groups, respectively, 0.1% and 0.4% symptoms necessitated cortico- vs 24.1 and 22.7 kUA/L in the HM and UM groups at 12 months,
steroid therapy (P < .0001). respectively (Figure 3); α-lactalbumin- and β-lactoglobulin-sIgE
6 | NAGAKURA et al.

HM group UM group
p < 0.0001 (kUA/L)
(kUA/L) p < 0.0001
p = 0.005
10
4
10
4

p = 0.007 p < 0.001

10
3
103

Milk-sIgE
Milk-sIgE

102 102

10
1
10
1

10
0
100
Pre 1m 3m 6m 12 m Pre 1m 3m 6m 12 m

p < 0.0001 (kUA/L)

(kUA/L) p < 0.0001
p = 0.02
10
4
104
p = 0.002 p < 0.001

103 103
Casein-sIgE
Casein-sIgE

10
2
10
2

10
1
101

100 100
Pre 1m 3m 6m 12 m Pre 1m 3m 6m 12 m

F I G U R E 3 Milk- and casein-specific immunoglobulin E (IgE) changes in the HM and UM groups; the x-axis represents milk- and casein-
specific IgE, and the y-axis represents time from start of oral immunotherapy. Milk- and casein-specific IgE levels significantly decreased
from baseline to 12 mo in both groups. The rates of reduction of milk-, casein-, α-lactalbumin-, and β-lactoglobulin-sIgE levels from baseline
to 12 months did not significantly differ between the HM and UM groups. HM, heated milk; sIgE, specific IgE; UM, unheated milk

significantly decreased from the baseline after 12 months (6.0 and or UM-OIT in children with CM anaphylaxis. We found that the
1.1 vs 6.7 and 4.6 kUA/L in the HM and UM groups, respectively; rates of total adverse symptoms, moderate or severe symptoms,
Figure S1). and respiratory symptoms in the HM group were significantly
Furthermore, the casein-sIgG and casein-sIgG 4 levels increased lower than those in the UM group. Despite a sample size was
significantly from baseline after 1 month in both groups (11.4 and insufficient to assess efficacy, HM-OIT induced immunological
0.7 vs 12.8 and 1.5 mgA/L in the HM and UM groups, respectively). changes and desensitization with 3 mL in the majority of patients,
In addition, β-lactoglobulin-sIgG and β-lactoglobulin-sIgG 4 levels and some participants even passed the 25-mL OFC. However, in
increased significantly from the baseline after 1 month only in the the HM-OIT, the proportion of patients who passed the 3- and/
UM group (2.8 and 0.1 vs 3.5 and 0.2 mgA/L in the HM and UM or 25-mL OFC tended to be lower than that of participants with
groups, respectively). In the HM group, the β-lactoglobulin-sIgG UM-OIT; moreover, β-lactoglobulin-sIgG and β-lactoglobulin-
and β-lactoglobulin-sIgG 4 levels did not significantly change during sIgG 4 levels did not significantly change, whereas β-lactoglobulin-
the course of the OIT protocol (Figure 4, Figure S2). sIgG and β-lactoglobulin-sIgG 4 levels significantly increased in
UM-OIT. Heating reduces the allergenicity of conformational
epitopes of CM protein, especially whey proteins that include
4 | D I S CU S S I O N β-lactoglobulin.11 The effects of heating could have contributed to
the results of this study.
We conducted an RCT using the same protocol, except for the Regarding safety, the rates of total, moderate/severe, and
food formulation, to investigate the safety and efficacy of HM-OIT respiratory symptoms per home dose in the HM group were
NAGAKURA et al. | 7

HM group UM group

p = 0.008 p < 0.0001

(mgA/L) (mgA/L)
p = 0.001 p < 0.0001
102 102
p = 0.0001 p = 0.0001

p = 0.0002 p < 0.0001

10 10
1 1
Casein-sIgG4

Casein-sIgG4
10 10
0 0

10 10
–1 –1

10–2 10–2
Pre 1m 3m 6m 12 m Pre 1m 3m 6m 12 m

p = 0.0006
(mgA/L) (mgA/L) p = 0.0007
102 102 p = 0.0005
n.s
p = 0.0007

10 10
1 1
β-lactoglobulin-sIgG4

β-lactoglobulin-sIgG4

10 10
0 0

10–1 10–1

10–2 10–2
Pre 1m 3m 6m 12 m Pre 1m 3m 6m 12 m

F I G U R E 4 Casein- and β-lactoglobulin-specific immunoglobulin G 4 changes in the HM and UM groups; the x-axis represents casein- and
β-lactoglobulin-sIgG 4, and the y-axis represents time from start of OIT. β-lactoglobulin-sIgG 4 levels significantly increased only in the UM
group. HM, heated milk; sIgG 4, specific IgG 4; UM, unheated milk

significantly lower than those in the UM group; especially, these patients had a history of anaphylaxis); therefore, the HM group
differences were seen in the first 6 months. Reducing adverse can be considered to have more safely undergone OIT than the
symptom rate during first 6 months, including up-dosing phase, UM group.
would be important for safely conducting OIT.18 Previous reports Regarding efficacy, during low-dose OIT with a target dose of
showed that safety did not differ when using raw or baked milk, 3 mL of HM, 38% and 19% of participants passed the 3- and 25 mL-
although the OIT protocol and patient age were different be- OFC after 2 weeks of avoidance, respectively. These rates are lower
tween the baked milk and raw milk groups.14 The present study than those with high-dose OIT but similar to the results of previous
used the same protocol, with the exception of food material, and low-dose OIT with a target dose of 3-mL.8,10,21 In addition, these
showed similar baseline profiles between the HM and UM groups. rates in the HM group were not significantly different than those
Therefore, this study design appeared appropriate for establish- in the UM group. Some reports have shown that HM ingestion ac-
ing the investigational safety of OIT. Although the gastrointes- celerates tolerance acquisition in patients with CM allergy. 22,23 The
tinal symptom rate in the HM group was higher than in the UM results of the present RCT suggest that HM-OIT is an effective strat-
group, most of symptoms involved mild pruritus of the throat or egy in patients with severe CM allergy. However, when comparing
oral cavity. This was possibly because the HM group used pow- the rates of passing the 25-mL OFC, the 17% reported for the HM
der, and the actual consumed doses were higher; therefore, the group is lower than the 31% for the UM group. In a larger-sample
increased amount of contact in the mouth may have led to the study, treatment efficacy could be significantly lower in the HM
19
increased rate of oral symptoms. Furthermore, the threshold group.
dose varies with heating, particularly in patients with severe CM Some previous OITs with a target dose of 200-mL or more
allergy. 20 In the present study, allergy-related characteristics showed a decrease in sIgE levels and an elevation in sIgG 4 lev-
among study participants were severe (high sIgE levels, and all els. 24,25 In the present study, we showed that sIgE to milk, casein,
8 | NAGAKURA et al.

α-lactalbumin, and β-lactoglobulin were significantly reduced, and methodology (lead); project administration (lead); resources (lead); soft-
the sIgG and sIgG 4 to casein were significantly increased during ware (equal); supervision (equal); validation (supporting); visualization
low-dose OIT with a target dose of 3-mL. These findings are (supporting); writing – original draft (lead); writing – review and editing
similar to those of previous low-dose OITs.17,26-28 Interestingly, (equal). Sakura Sato: Conceptualization (lead); data curation (support-
β-lactoglobulin-sIgG and β-lactoglobulin-sIgG 4 levels significantly ing); formal analysis (supporting); funding acquisition (equal); inves-
increased only in the UM group. β-lactoglobulin has a conforma- tigation (equal); methodology (equal); project administration (equal);
tional structure that is denatured by heating, whereas casein is resources (equal); software (equal); supervision (lead); validation (lead);
less denatured on heating. Therefore, β-lactoglobulin-sIgG and visualization (lead); writing – original draft (equal); writing – review
β-lactoglobulin-sIgG 4 would not change in the HM group. Increase and editing (lead). Yoko Miura: Conceptualization (supporting); data
in the levels of β-lactoglobulin-sIgG and β-lactoglobulin-sIgG 4 curation (supporting); formal analysis (supporting); funding acquisition
may require exposure to unheated β-lactoglobulin. We theorized (supporting); investigation (equal); methodology (supporting); project
that these immunological results may be related to differences in administration (supporting); resources (supporting); software (sup-
treatment efficacy: higher tendency of the rates of passing the porting); supervision (supporting); validation (supporting); visualization
3- and 25-mL OFCs in UM group. The reason for the unaltered (supporting); writing – original draft (supporting); writing – review and
β-lactoglobulin-sIgG 4 despite significant decreases in β-lactoglob- editing (supporting). Makoto Nishino: Conceptualization (supporting);
ulin-sIgE might be that increases in sIgG 4 seem to require a high- data curation (supporting); formal analysis (supporting); funding acqui-
dose antigen. 21 However, the sample size was insufficient to draw sition (supporting); investigation (equal); methodology (supporting);
definitive conclusions about these immunological changes, and project administration (supporting); resources (supporting); software
therefore, larger studies are needed. (supporting); supervision (supporting); validation (supporting); visualiza-
One limitation of our study was that we did not conduct raw milk tion (supporting); writing – original draft (supporting); writing – review
OFC. The participants developed symptoms against 3-mL HM and had and editing (equal). Kyohei Takahashi: Conceptualization (supporting);
a history of anaphylaxis; thus, they had severe CM allergy. Therefore, data curation (supporting); formal analysis (supporting); funding acqui-
we did not undertake raw milk OFC due to the risk of inducing severe sition (supporting); investigation (equal); methodology (supporting);
symptoms. We instructed patients who passed the 25-mL HM-OFC project administration (supporting); resources (supporting); software
to ingest processed food containing 25-mL HM at home; we subse- (lead); supervision (supporting); validation (lead); visualization (support-
quently considered raw milk OFC after the second year. ing); writing – original draft (supporting); writing – review and editing
Second limitation was that the UM group did not use completely (supporting). Tomoyuki Asaumi: Conceptualization (supporting); data
raw CM, as most of the commercially available milk in Japan is steril- curation (supporting); formal analysis (supporting); funding acquisition
ized at 125°C for 2 seconds due to food-related regulations. (supporting); investigation (equal); methodology (supporting); project
Third limitation was that adverse symptom rates per patients administration (supporting); resources (supporting); software (sup-
were not significantly lower in the HM group than those in the UM porting); supervision (supporting); validation (supporting); visualization
group, despite significantly lower rates per intake number. However, (supporting); writing – original draft (supporting); writing – review and
the efficacy data seem to suggest that UM may be advantageous editing (supporting). Kiyotake Ogura: Conceptualization (supporting);
compared with HM. Larger studies are desired to assess this. data curation (supporting); formal analysis (supporting); funding acqui-
In conclusion, HM-OIT is apparently safer than UM-OIT in pa- sition (supporting); investigation (equal); methodology (supporting);
tients with CM anaphylaxis. Despite the slightly lower treatment project administration (supporting); resources (supporting); software
efficacy in the HM group, the number of participants was insuffi- (lead); supervision (supporting); validation (supporting); visualization
cient to compare efficacy. Larger studies are necessary to confirm (supporting); writing – original draft (supporting); writing – review and
efficacy between the HM-OIT and UM-OIT protocols. editing (supporting). Motohiro Ebisawa: Conceptualization (lead); data
curation (supporting); formal analysis (supporting); funding acquisition
AC K N OW L E D G M E N T S (equal); investigation (equal); methodology (equal); project administra-
We would like to thank all pediatricians, nutritionists, and nurses tion (equal); resources (equal); software (equal); supervision (lead); vali-
who supported this trial at Sagamihara National Hospital. dation (lead); visualization (equal); writing – original draft (equal); writing
– review and editing (lead). Noriyuki Yanagida: Conceptualization (lead);
C O N FL I C T O F I N T E R E S T data curation (supporting); formal analysis (supporting); funding acqui-
Motohiro Ebisawa serves on the clinical medical advisory board of sition (lead); investigation (equal); methodology (equal); project admin-
DBV Technologies. Sato Sakura and Motohiro Ebisawa have received istration (equal); resources (equal); software (equal); supervision (lead);
speaker honoraria from Mylan EPD. The other authors declare that validation (lead); visualization (lead); writing – original draft (equal); writ-
they have no conflicts of interest. ing – review and editing (lead).

AU T H O R C O N T R I B U T I O N PEER REVIEW
Ken-ichi Nagakura: Conceptualization (equal); data curation (lead); The peer review history for this article is available at https://publo​
formal analysis (lead); funding acquisition (equal); investigation (lead); ns.com/publo​n/10.1111/pai.13352.
NAGAKURA et al. | 9

DATA AVA I L A B I L I T Y S TAT E M E N T 15. Simons FE. World Allergy Organization survey on global availability
of essentials for the assessment and management of anaphylaxis by
If reasonable request, the datasets used to support the findings of
allergy-immunology specialists in health care settings. Ann Allergy
this trial are available from the corresponding author. Asthma Immunol. 2010;104(5):405-412.
16. Simons FE, Ardusso LR, Bilo MB, et al. World allergy organization
ORCID guidelines for the assessment and management of anaphylaxis.
Ken-ichi Nagakura https://orcid.org/0000-0002-9381-9044 World Allergy Organ J. 2011;4(2):13-37.
17. Yanagida N, Okada Y, Sato S, Ebisawa M. New approach for food
Sakura Sato https://orcid.org/0000-0003-3674-0759
allergy management using low-dose oral food challenges and low-
Yoko Miura https://orcid.org/0000-0001-8887-6901 dose oral immunotherapies. Allergol Int. 2016;65(2):135-140.
Makoto Nishino https://orcid.org/0000-0002-5381-8848 18. Pajno GB, Fernandez-Rivas M, Arasi S, et al. EAACI Guidelines
Kyohei Takahashi https://orcid.org/0000-0003-1220-7386 on allergen immunotherapy: IgE-mediated food allergy. Allergy.
2018;73(4):799-815.
Tomoyuki Asaumi https://orcid.org/0000-0002-7464-5421
19. Fauquert JL, Michaud E, Pereira B, et al. Peanut gastrointestinal de-
Motohiro Ebisawa https://orcid.org/0000-0003-4117-558X livery oral immunotherapy in adolescents: results of the build-up
Noriyuki Yanagida https://orcid.org/0000-0001-9643-744X phase of a randomized, double-blind, placebo-controlled trial (PITA
study). Clin Exp Allergy. 2018;48(7):862-874.
20. Remington BC, Westerhout J, Campbell DE, Turner PJ. Minimal
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