Aptamer mediated targeting of Fanconi Anemia oral cancer initiating cells

描述(申请人提供)：我们寻求使用一种新的细胞内化SELEX(通过指数富集法的配体的系统进化)来分离硫代适配子结合的脂质体纳米粒(TA-NP)，该纳米粒内化并将其siRNA传递到口腔鳞状细胞癌的肿瘤起始细胞(TIC)的胞浆中，最大限度地减少非靶标传递。在体内，siRNA从内体到胞浆的传递和释放仍然是将抗癌siRNA药物转移到临床的两大障碍。口腔鳞状细胞癌，俗称口腔癌，是头颈部最常见的恶性肿瘤，对全球公众健康的影响很大。范可尼贫血(FA)是一种易患OC的遗传性癌症综合征。有证据表明，TIC是野战癌变、耐药和疾病复发的关键驱动因素。一种选择性靶向TIC并将抗癌药物输送到TIC的治疗工具为OC治疗提供了巨大的希望。我们的数据和已发表的研究表明：(1)与散发性OC相比，TIC在FA-OC中更为丰富；(2)Chk1和CD147促进了TIC的生存和化疗耐药，其过度表达导致OC预后不良。本应用的主要目的是以FA-OC-TIC为靶细胞制备TA-NP，用于胞质递送siRNA。我们推测，分离的TA-NP将促进siRNA靶向传递到OC-TIC胞浆，并沉默OC-TIC中的CD-147/Chk1，扰乱它们的生态位，使它们对化疗敏感，容易被吉西他滨化疗消除。目的：(1)：阐明RNAi介导的CD147/Chk1沉默在FA-OC-TIC中的作用；(2A)：筛选针对FA-OC细胞TIC部分的TA-脂质体NP(TA-NP)，同时利用基于阳性和阴性细胞摄取的SELEX来避免肝细胞摄取；(2B)：确定所选TA-NP在体外和小鼠OC异种移植瘤中的靶向性和治疗效果。我们将在FA-OC细胞系中分离ALDH+TIC，并检测siRNA介导的CD147和Chk1沉默的效果。(2)我们将使用带有正负选择的改良共轭SELEX来鉴定OC-TIC特异性内化TA-NP。(3)我们将验证分离的TA-NP-SRNA在体外和在小鼠原位FA-OC异种移植中的靶向性和沉默效果。RNAi介导的CD147和Chk1在OC-TIC中的抑制作用将有助于深入了解它们在OC-TIC的肿瘤生长和耐药特性中的潜在作用。所提出的TA-NP在体内靶向OC-TIC的治疗药物和显像剂方面将具有重要的临床潜力。

DESCRIPTION (provided by applicant): We seek to use a novel cell-internalization SELEX (Systemic Evolution of Ligands by Exponential enrichment) to isolate thioaptamer-conjugated liposomal nanoparticles (TA-NP) that internalize and deliver its siRNA into the cytosol of tumor initiating cells (TIC) of oral squamous cell carcinoma, minimizing off-target delivery. In vivo delivery and release of siRNA from the endosomes to cytosol remain the two biggest obstacles for translating anticancer siRNA drugs to the clinic. Oral squamous cell carcinoma commonly known as oral cancer (OC) is the most common malignancy of head and neck with high global public health impact. Fanconi anemia (FA) is a hereditary cancer syndrome that predisposes one to OC. Evidence points to TIC as the key driver of field cancerization, resistance to therapy and disease relapse. A therapeutic vehicle that selectively targets and delivers anticancer drugs to TIC offers great promise for OC treatment. Our data and published studies show: (1).TIC is more enriched in FA-OC than sporadic OC; (2) Chk1 and CD147 promote TIC survival and chemoresistance and its overexpression contribute to poor prognosis in OC. The main goal of this application is to use FA- OC-TIC as target cells to develop TA- NP for the cytosolic delivery of siRNA. We hypothesize that isolated TA-NP will facilitate targeted delivery of siRNA to OC-TIC cytosol, and silence CD-147/Chk1 in OC-TIC's, disrupting their niche and rendering them chemosensitive and susceptible to elimination by chemotherapy with gemcitabine. Aims:(1): Elucidate the effect of RNAi mediated silencing of CD147/Chk1 in FA-OC-TIC; (2A): Select a list of TA-Liposomal NP (TA-NP) specific for TIC fraction of FA-OC cells, while avoiding hepatocyte uptake, using a positive and negative cell-uptake based SELEX; (2B): Determine the selected TA-NP's targeting specificity and therapeutic efficacy in in vitro and in murine OC xenografts. We will isolate ALDH+ TIC in FA-OC cell lines and examine the effects of siRNA-mediated silencing of CD147 and Chk1. (2) We will use a modified conjugate SELEX with positive and negative selections to identify OC-TIC-specific internalizing TA-NP. (3) We will validate TIC-targeting specificity and silencing efficacy of isolated TA-NP-sRNA in vitro and in murine orthotopic FA-OC xenografts. RNAi mediated inhibition of CD147 and Chk1 in OC-TIC will yield valuable insight into their potential role in the tumor propagating and chemoresistant properties of OC-TIC. The proposed TA-NP will have important clinical potentials for in vivo delivery of therapeutic and imaging agents targeting the OC-TIC.