DNA Immunization in the Female Genital Tract Using A Controlled Delivery Matrix

简介（由申请人提供）：每年估计有239,000名妇女死于子宫颈癌。其中99%以上的死亡是由生殖器感染人乳头瘤病毒（HPV）造成的。因此，迫切需要开发一种治疗性疫苗来解决这种疾病引起的癌变。本提案的总体目标是开发一种设备，该设备将解决由HPV引起的癌前病变和癌性宫颈病变。这项工作的一个目标是制造DNA疫苗递送装置，用于局部施用于女性生殖道。此外，将进行体外和体内临床前研究，以评估该装置在施用有效疫苗方面的功效。为此，提出了以下具体目标：(1)通过控制DNA的释放来表征阴道内疫苗接种的分子和免疫学事件；(2)研究DNA传递装置的剂量、持续时间和转染剂对免疫事件的影响；(3)探讨阴道内DNA传递对肿瘤消退的效果。将研究一种以可控方式释放DNA的装置。具体来说，质粒DNA将被整合到一个由生物相容性材料制成的圆盘形状的基质中。这种质粒DNA将编码人乳头瘤病毒的致癌基因，它将作为治疗性疫苗刺激免疫相关反应。临床前实验将在模型小鼠中进行，该装置将被插入阴道。DNA和mRNA在生殖道中的持续表达将通过实时PCR来评估该装置的释放。此外，将评估对该疗法的体液和细胞免疫反应。一旦这些事件被表征，该装置将被进一步修改，以探索剂量、释放持续时间和转染剂的添加对免疫反应持续时间和强度的影响。最后，将探索在小鼠模型中解决HPV特异性肿瘤时引发最大程度和持续时间反应的装置。在这些研究的结论中，对阴道内DNA免疫的更好理解将得到阐明，并将制定开发一种解决HPV特异性病变的设备的过程。一种解决宫颈病变的非侵入性装置提供了一种不受技术人员管理限制的治疗方法，对全世界妇女的健康有直接的好处。此外，这种疫苗接种策略可以应用于开发其他性传播疾病的新疗法。

DESCRIPTION (provided by applicant): Each year an estimated 239,000 women die of cervical cancer. Over 99% of these deaths result from genital infection with human papillomavirus (HPV). As a result there is a pressing need to develop a therapeutic vaccine to resolve cancerous lesions caused by this disease. The broad goal of this proposal is to develop a device which will resolve pre-cancerous and cancerous cervical lesions caused by HPV. One objective of this work is to fabricate DNA vaccine delivery devices to be administered topically in the female genital tract. Furthermore, in vitro and in vivo preclinical studies will be performed to assess the efficacy of this device in administering an effective vaccine. To this end the following specific aims have been proposed: (1) characterize molecular and immunological events of intravaginal vaccination by controlled release of DNA; (2) investigate the influence of dosage, duration, and transfection agents from DNA delivery devices on immunological events; and (3) investigate the efficacy of tumor resolution by intravaginal DNA delivery. A device will be investigated which releases DNA in a controlled fashion. Specifically, plasmid DNA will be incorporated into a matrix comprised of a biocompatible material fabricated in the shape of a disk. This plasmid DNA will encode an oncogene of HPV which will act as a therapeutic vaccine to stimulate immune relevant responses. Pre-clinical experiments will be implemented in a model mouse, where the device will be inserted in the vagina. Persistence of DNA and mRNA expression in the genital tract will be evaluated by real-time PCR to assess the release from the device. Additionally, both humoral and cellular immune responses to the therapy will be assessed. Once these events have been characterized the device will be further modified to explore the influence of dose, duration of release, and addition of transfection agents on duration and magnitude of immune response. Finally, the device that elicits a response that is largest in magnitude and duration will be explored for resolving HPV specific tumor in a mouse model. At the conclusion of these studies a better understanding of intravaginal DNA immunization will have been elucidated and processes will be made for developing a device to resolve HPV specific lesions. A non-invasive device to resolve cervical lesions has immediate benefit to women's health worldwide by providing a therapy that is not limited by skilled personal to administer. Additionally, this vaccination strategy could be applied to develop new therapies for other sexually transmitted diseases.