Needle-free Epicutaneous For t 2 DNA Vaccine is Effective for
Preventing and Treating Biting Midge (Forcipomyia taiwana)
allergy in a murine model
To the Editor,
Allergen-specific immunotherapy (ASIT) remains the only treatment
capable of inducing immune tolerance to the corresponding allergen and
potentially treating the root cause of the allergic
disease.1 As the treatment course of protein-based
vaccines for ASIT is time-consuming, an easily administered epicutaneous
anti-allergic DNA-based vaccine is an attractive method, especially in
light of the COVID-19 pandemic.2
The biting midge, Forcipomyia taiwana , is the most prevalent
cause of biting insect allergy in Taiwan. It is a tiny hematophagous
midge that attacks en masse. As many as 60% of exposed individuals
develop allergic reactions to the bites.3 The midge is
widely distributed throughout Taiwan and southern China. For t 2 is the
most predominant,with 75% of midge-allergic patients showing specific
IgE to For t 2.4 Allergic reactions to midge bites are
not limited to humans but also seen in livestock, such as horses,
cattle, sheep, and donkeys, causing significant veterinarian problems.
E.coli -expressed For t 2 recombinant protein (rFor t 2) was used
as an allergen to sensitize and challenge the mice.5For t 2-encoding fragment (GenBank accession EU678971) was amplified by
PCR. The PCR products were subcloned into pVAX1 (Life Technologies,
Carlsbad, CA) . The experiments were designed using two
approaches: therapeutic and prophylactic (Fig 1). The therapeutic
approach is to imitate ASIT in human with established allergy while the
prophylactic approach to non-allergics. Twenty-five μg For t 2 DNA was
determined as the optimal dose after dose-finding experiments
(Supplementary Fig S1). For each treatment, the hair of the abdominal
area of the mice was removed using a depilatory,tape-stripped,then
patched with 25 μg For t 2 DNA vaccine for one hour and removed.A total
of three treatments were given spaced one week apart (Fig 1 and Fig S2).
For t 2 proteins were detected in the patched skin and the immune organ
spleen at 24 hoursand had significantly increased at 48 hours after last
treatment (Fig S3). Scratch bouts after rFor t 2 challenge were used as
a clinical surrogate of itch. We measured For t 2-specific IgE, IgG1 and
IgG2a in the sera as well as mRNA and proteins of IL-13,
interferon-gamma, IL-10, and FOXP3 in the culture supernatants of
splenocytes after stimulation with various doses of rFor t 2 at 37℃ for
3-5 days by ELISA and real-time quantitative PCR. Histopathology of the
challenged skins was examined.
After epicutaneous DNA vaccination, the allergen-induced itchin both
groups significantly improved, and For t 2-specific IgE and IgG1/IgG2a
ratio decreased significantly at week 6 or week 8 (Fig 2). Levels of
mRNA and protein of IL-13 decreased significantly, but IFN-gamma and
IL-10 remained unchanged. Expression of FOXP3 mRNA increased (Fig 2,
protein data not shown). Eosinophils infiltration in the challenged skin
significantly decreased (Fig S4).
This is the first study to demonstrate an epicutaneous needless
anti-allergic DNA vaccine that effectively treats an established
allergic condition and prevents the development of an allergic disease
using biting midge allergy as a model. After epicutaneous DNA
vaccination, in addition to allergen-induced itch, the changes of
multiple biomarkers suggest that immune tolerance was induced after the
epicutaneous DNA vaccine.
Our data show that though the molecular weight of the For t 2 DNA
vaccine is as high as 4000 base pairs, it can penetrate the dermal
barrier and translate the corresponding protein in the targeted skin and
the spleen of the vaccinated mice. It is possible that the DNA vaccine
passes the epidermis via the hair follicles as the skin is tape-stripped
before epicutaneous vaccination.6
The mode of this anti-allergic epicutaneous DNA vaccine may potentially
be used in other specific immunotherapies for other allergens.
Mey- Fann Lee1
Chi-Sheng Wu2
Shyh-Jye Lin3
Yi-Hsing Chen2,4*
1Department of Medical Research, Taichung Veterans
General Hospital, Taichung, Taiwan
2Division of Allergy, Immunology and Rheumatology,
Taichung Veterans General Hospital, Taichung, Taiwan
3School of Medical Laboratory and Biotechnology, Chung
Shan Medical University, Taichung, Taiwan
4School of Medicine, National Yang Ming Chiao Tung
University, Taipei, Taiwan