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技术，用于微递送粉末状过敏原，以实现更安全、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IgE介导的过敏症的持久临床益处的唯一治疗，但由于过敏反应的风险和不便，只有不到5%的患者选择该治疗。EPIT可以潜在地缩短治疗的持续时间并降低过敏性风险，因为表皮不仅富含抗原呈递细胞（APC），而且是非血管化的组织。不幸的是，经由完整皮肤的过敏原的表皮递送面临许多挑战，因为皮肤对于大分子是不可渗透的，而破坏皮肤屏障常常触发不需要的Th 2免疫应答。我们与其他人沿着，使用烧蚀微分数激光（A？FL）在皮肤中产生一系列可自我更新的微通道（MC），用于疫苗输送，并引发强烈的Th 1免疫反应。这些MC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”路径，而且还维持表皮中的过敏原不断刺激有利于免疫耐受的免疫系统。此外，由于过敏原在很大程度上被限制在MC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），当应用于激光治疗的皮肤时，该贴片产生了有效的表皮递送。我们假设，粉末状过敏原/佐剂的激光促进表皮递送可以大大提高EPIT的疗效，而不会引起过敏反应的风险。我们将用花生蛋白提取物（PPE），沿着与Th 1或免疫抑制剂佐剂一起制备PMP。将在未处理和致敏C3 H/HeJ小鼠以及致敏新生猪中优化通过激光穿孔皮肤的PPE/佐剂的表皮递送（目的1.1）。将在接种部位测量局部先天免疫，并在血液样品中测定针对花生过敏原的体液免疫应答。我们还将研究Th 1、Th 2和调节性T细胞（Treg），以证实Th 1/Treg占主导地位的抗过敏原免疫（目标1.2）。最后，我们将在鼠花生过敏模型中寻找在花生过敏的治疗中，3个剂量的上级EPIT是否优于常规的18个剂量的皮下免疫疗法（目的2）。IgE介导的变态反应在工业化国家的发生率越来越高。这……如果成功，EPIT将导致更安全，更快速的花生过敏治疗，并可以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的影响。

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.