Purpose: Pancreatic ductal adenocarcinoma (PDAC) is the third-highest cause of cancer-related death in the United States. PDAC represents one of the most challenging cancer due to its pathological characteristics, such as dense desmoplastic (DS) tissue with >90 % tumor stroma. Among all epithelial tumors, PDAC has high abundance DS that contributes to chemotherapy resistance and retards drug delivery efficacy in the core of the tumor. The Overexpression of multiple tumorigenic biomarkers such as CD24, CD44, ESA, and c-Met contribute to forming PDAC stroma, resulting in the development of drug-resistant features. Thus, dual targeting CD44 and c-Met biomarker is a smart approach for enhancing drug delivery efficacy in PDAC. We developed a combination drug delivery system to improve DS penetration, thus achieving profound therapeutic benefits in PDAC. We successfully synthesize a Gemcitabine-pro-drug nanoparticle that can selectively target CD44 and c-Met biomarker and synergistically kills PDAC in combination with Everolimus.
Methods: The synthesis of dual-targeted non-lipid-based nanoparticles (DTNP) is performed by using EDC/NHS coupling and click chemistry. The final product was evaluated using NMR and MS spectrometry. DTNP has been utilized as a universal construct for conjugating an imaging agent and chemotherapeutic drugs, Gemcitabine. To investigate the efficacy of the dual-targeted imaging agent, the AsPc-1 cells were orthotopically injected in NOD/SCID mice. The tumor-bearing mice were intravenously treated with DTNP conjugated-near Infrared (NIR), namely DTNP-NIR dye for 24 and 48h. The bio-distribution of DTNP-NIR agent was evaluated in tumor and vital organs. The immunohistology of DTNP-NIR treated tumor section was performed to evaluate the CD44+ and c-Met targetability of the nanoparticle. The combination of Gemcitabine with Everolimus was determined using MTT assay and combination index value was evaluated using CompuSyn software to demonstrate the synergistic anti-PDAC effect.
Results: In vivo study of near Infrared (NIR) dye conjugated DTNP showed high PDAC tumor accumulation compared to healthy organs. Interestingly, the NIR-dye accumulation in the tumor site of DTNP increased with time of treatment, indicating the retention of DTNP-NIR in the tumor milieu. The 7-fold higher tumor/liver uptake ratio of DTNP-NIR compared to non-targeted non-lipid nanoparticles (NTNP) supports the safety of nanoparticle. In IHC study DTNP treated tumor section proved the higher tumor core penetration of nanoparticle compared to control, suggesting the efficacy DTNP for PDAC selective delivery of Gemcitabine. The preliminary results of in vitro combination index of Gemcitabine in combination with Everolimus revealed synergistic AsPC-1 cell killing with combination index (CI) value < 1. This data builds a strong rationale of developing Gemcitabine-DNPT for therapeutic benefit in PDAC. The cell-killing effect of Gemcitabine-DTNP is retained as compared with commercial gemcitabine with an IC50 of Gemcitabine is12 μM in AsPC-1. Currently, tumor efficacy studies are being carried out to investigate the activity of Gemcitabine-DTNP.
Conclusion: The obtained results indicate that the DTNP opens a new avenue for imaging-guided diagnosis and therapy in PDAC tumor. Moreover, the DTNP can be utilized as a universal payload delivery vehicle for chemotherapeutic drugs that can potentially overcome the current challenges of PDAC therapy.