Laser-facilitated powder-allergen delivery for epicutaneous immunotherapy

描述（由申请人提供）：本提案的总体目标是开发一种基于激光的微递送粉末状过敏原技术，用于更安全、更快速的表皮免疫治疗，称为¿EPIT。迄今为止，过敏原特异性免疫疗法仍然是对ige介导的过敏有长期临床益处的唯一治疗方法，但由于过敏反应的风险和不便，只有不到5%的患者选择这种治疗。EPIT有可能缩短治疗时间，降低过敏风险，因为表皮不仅富含抗原呈递细胞（APCs），而且是一个非血管化组织。不幸的是，通过完整的皮肤将过敏原表皮传递面临许多挑战，因为皮肤对大分子是不可渗透的，而打破皮肤屏障通常会引发不必要的Th2免疫反应。我们和其他人一起，使用烧蚀微分数激光（A¿FL）在皮肤中产生一系列可自我再生的微通道（MCs），用于疫苗递送，并引发强烈的Th1免疫反应。这些MCs为EPIT提供了独特的优势，因为它们不仅作为粉末状过敏原进入表皮的“自由”途径，而且还维持表皮中的过敏原不断刺激免疫系统，有利于免疫耐受。此外，由于过敏原在很大程度上限制在MCs内，因此可以很好地预防过敏反应。我们已经创造了一种简单的方法来制造基于粉末的微阵列贴片（PMP），当应用到激光治疗的皮肤上时，它可以产生有效的表皮递送。我们假设激光促进的粉末状过敏原/佐剂的表皮递送可以大大增强EPIT的功效，而不会产生过敏反应的风险。我们将使用花生蛋白提取物（PPE）以及Th1或免疫抑制佐剂来制造PMP。在C3H/HeJ致敏小鼠以及致敏新生猪中，通过激光穿孔皮肤对PPE/佐剂进行表皮递送将得到优化（Aim 1.1）。局部先天免疫将在接种部位被测量，体液免疫对花生过敏原的反应将在血液样本中被检测。我们还将研究Th1、Th2和T调节性（Treg）细胞，以证实Th1/Treg对过敏原的免疫（Aim 1.2）。最后，我们将在小鼠花生过敏模型中寻求三剂量的EPIT在治疗花生过敏方面是否优于传统的18剂量皮下免疫疗法（目的2）。在工业化国家，ige介导的过敏发生率越来越高。这种EPIT如果成功，将导致更安全、更快速地治疗花生过敏，并可很容易地扩展到其他ige介导的过敏，这将对公共卫生系统产生深远影响。

DESCRIPTION (provided by applicant): The overall objective of this proposal is to develop a laser-based technology for micro-delivering powdered allergens for safer and faster epicutaneous immunotherapy, named ¿EPIT. To date, allergen specific immunotherapy remains the only treatment for long-lasting clinical benefit to IgE-mediated allergies, but fewer than 5% o patients choose the treatment because of a risk of anaphylaxis and inconvenience. EPIT can potentially shorten the duration of the treatment and reduce anaphylactic risk because the epidermis is not only rich in antigen-presenting cells (APCs), but also a non-vascularized tissue. Unfortunately, epidermic delivery of allergen via intact skin faces numerous challenges as the skin is impermeable for macromolecules, whereas breaking skin barrier often triggers unwanted Th2 immune responses. We, along with others, have used ablative micro-fractional laser (A¿FL) to generate an array of self-renewable microchannels (MCs) in the skin for vaccine delivery and to elicit strong Th1 immune responses. These MCs offer unique advantages for EPIT as they not only serve as "free" paths for powdered allergens to enter the epidermis but also sustain the allergen in the epidermis constantly stimulating immune system that favors immune tolerance. Moreover, because the allergen is largely restricted inside the MCs anaphylaxis can be well prevented. We have created a simple way to fabricate a powder-based microarray patch (PMP), which gave rise to efficient epidermic delivery when applied onto laser-treated skin. We hypothesize that the laser-facilitated epidermic ¿-delivery of powdered allergens/adjuvants can greatly augment efficacy of EPIT without incurring a risk of anaphylaxis. We will fabricate PMP with peanut protein extract (PPE), along with Th1 or immunosuppressant adjuvants. Epidermic delivery of PPE/adjuvants via laser-perforated skin will be optimized in naive and sensitized C3H/HeJ mice as well as in sensitized newborn pigs (Aim 1.1). Local innate immunity will be measured at the inoculation site and humoral immune responses against peanut allergens will be assayed in blood samples. We will also investigate Th1, Th2 and T regulatory (Treg) cells to corroborate Th1/Treg-predominant immunity against the allergen (Aim 1.2). Finally, we will seek in a murine peanut allergy model whether three doses of ¿EPIT are superior to conventional 18 doses of subcutaneous immunotherapy in treatment of peanut allergy (Aim 2). IgE-mediated allergy occurs at an increasing rate in industrialized countries. This ¿EPIT, if successful, would result in safer and faster treatments of peanut allergy and can be readily extended to other IgE-mediated allergies, which will have profound impact on public health systems.