New Stereoregular Functionalized Fullerenes as Nanomedicines for PDT

描述（由申请人提供）：本申请的目的是（I）设计和合成光响应性[60]富勒烯基对映体和相关纳米结构作为PDT应用的光敏剂，以及（ii）对两亲性多功能[60]富勒烯基对映体和发色团纳米结构（如1PA-PDT和2PA-PDT）进行体外和体内生物研究（2-3）吸收性）纳米药物。我们建议设计和合成新型阳离子立构规整1-吡咯啉并[60]富勒烯基对映体PF-PhOEGn和阳离子C60(>CPAF-EG6)x，用作PDT药物，具有以下特点：（1）高光稳定性，只需单剂量药物即可进行多次PDT治疗； (2) 使用 C60 天线结构增强光捕获能力，以产生细胞毒性 ROS； (3) C60-天线的分子自组装能力形成稳定的囊泡以用于大容量的PDT药物递送； (4) 双光子吸收截面的大幅增强[C60-（天线）2>4500 GM]，用于有效的 2PA-PDT 治疗。我们已经确定了富勒烯的一些分子结构特征，这些特征可以介导微生物细胞和癌细胞的广谱光杀伤作用。这种对所需结构基序的洞察已应用于富勒烯纳米药物的分子设计。我们将通过体外和体内实验研究 PDT 效率：（1）我们组装了一组病原微生物，旨在涵盖大多数引起局部感染的微生物，例如细菌。在伤口和烧伤中，并表现出内在的和获得性的多种抗生素耐药性。聚阳离子富勒烯将作为针对这些病原体的 PDT 制剂进行测试。 (2)对于富勒烯介导的针对局部感染的PDT治疗，我们将使用基因工程生物发光细菌结合光学成像来研究伤口感染的小鼠模型，以非侵入性地实时跟踪感染的进展。 (3) 我们将对一组小鼠癌细胞系进行体外 PDT，这些细胞系预计对富勒烯具有不同的富勒烯摄取，并对 PDT 产生的活性氧产生不同的细胞反应。将针对 1 光子和 2 光子激发以及研究的细胞死亡机制进行药物和光的剂量响应研究。 (4) 对于抗癌测试，我们将使用皮下小鼠肿瘤并比较瘤内注射和静脉注射富勒烯。我们将比较使用 KTP 激光器的 1 光子激发和使用飞秒 Ti:sapph 激光器的 2 光子激发。 公共健康相关性：本申请中描述的富勒烯纳米药物有可能介导当今时代两种主要致命疾病的光动力疗法。通过将富勒烯的适当分子设计与连续波或飞秒脉冲光照明相结合，可以消除多重耐药细菌感染和癌性肿瘤。本文提出的这些化合物的特定化学和光物理特征将改善这些疾病的PDT，超出目前可能的水平。

DESCRIPTION (provided by applicant): The objective of this application is to (I) design and synthesize photoresponsive [60]fullerenyl enantiomers and related nanostructures as photosensitizers for PDT applications and (ii) carry out in vitro and in vivo biostudies of amphiphilic multifunctional [60]fullerenyl enantiomers and chromophore nanostructures as 1PA-PDT and 2PA-PDT (2-3 absorptive) nanomedicines. We propose to design and synthesize novel cationic stereoregular 1-pyrrolino[60]fullerenyl enantiomers PF-PhOEGn and cationic C60(>CPAF-EG6)x for the use as PDT agents with the following characteristics: (1) high photo stability for multiple PDT treatments with only a single-dose of the drug; (2) enhancement of light-harvesting capability using the C60-antenna construction, for cytotoxic ROS production; (3) capability of molecular self-assembly of C60-antenna forming stable vesicles for high volume delivery of PDT drug; (4) large enhancement of two-photon absorption cross-sections [>4500 GM for C60- (antenna)2] for effective 2PA-PDT treatments. We have established some of the molecular structural features of fullerenes that mediate broad-spectrum photokilling of microbial cells and cancer cells. This insight into required structural motifs has been applied to the molecular design of fullerene nanomedicines. We will study PDT efficiency by both in vitro and in vivo experiments: (1) we have assembled a panel of pathogenic microorganisms designed to encompass most of the microbes that cause localized infections e.g. in wounds and burns, and which demonstrate both intrinsic and acquired multi-antibiotic resistance. Polycationic fullerenes will be tested as PDT agents against these pathogens. (2) For fullerene-mediated PDT treatment against localized infections we will study a mouse model of wound infection using genetically engineered bioluminescent bacteria combined with optical imaging to follow the progress of the infection non-invasively in real time. (3) We will carry out in vitro PDT against a panel of mouse cancer cell lines that are expected to have different uptakes of fullerenes and different cellular responses to reactive oxygen species produced by PDT. Dose response studies of both drug and light will be carried out for both 1-photon and 2-photon excitation and mechanisms of cell death studied. (4) For anti-cancer testing we will use subcutaneous mouse tumors and compare intratumoral injection with IV delivery of the fullerene. We will compare both 1-photon excitation using a KTP laser and 2-photon excitation using a femtosecond Ti:sapph laser. PUBLIC HEALTH RELEVANCE: The fullerene nanomedicines described in this application have the potential to mediate photodynamic therapy of two of the major killer diseases of the present age. Multi-drug resistant bacterial infections and cancerous tumors could be eliminated by a combination of the appropriate molecular design of fullerene and illumination with CW or femtosecond pulsed light. Particular chemical and photophysical features of these compounds proposed herein will improve PDT of these diseases beyond what is presently possible.