A targeted near-infrared nanoprobe for deep-tissue penetration and imaging of prostate cancer

Fluorescent guided surgery (FGS) has been highlighted as a potential to increase precision of diseased tissue whilst minimising the removal of surrounding healthy tissue, particularly relevant for increasingly prevalent prostate cancer (PCa). There remain challenges in the design of imaging probes presenting high selectivity for tumour tissue, clear visualization and minimal toxicity. Here we report the design and development of a novel NIR-nanoprobe and evaluate its potential in penetration inside PCa tumour tissues. Prostate-specific membrane antigen (PSMA) receptor-targeted near infrared emitting quantum dots (PSMA-NIR-QDot) are demonstrated as deep tissue imaging agents for intraoperative navigation during surgery and improving detection specificity for PCa. Probes were designed and synthesized by conjugating functionalized amino-PEG QDots through a heterobifunctional linker to a DUPA targeted polypeptide construct. Nanoprobes were evaluated in vitro in PSMA+ PCa cell lines for specificity and affinity determined by flow cytometric analysis. The penetration efficacy was tested further on large PCa 3D tumour spheroids (dia ~ 1.2 mm, width ~ 250 µm) by deep tissue multiphoton imaging. The PSMA-NIR-Qdot was found to be an efficient deep tissue penetrating intra-operative guided surgical tool with high affinity (KD = 15 nM) and penetrative capacity. The results have been demonstrated in vitro in 2D and 3D tissue models, mimicking cancer lesions in vivo. The presented deep-tissue imaging NIR nanoprobes target prostatic lesions that (i) bind to PSMA+ tumour with sub-nanomolar affinity and high specificity, (ii) show excellent safety profile in primary cell lines in vitro, and (iii) show high penetrative capacity in 3D prostate tumour model (~ 250 µm tissue depth). These probes may potentially offer vastly improved surgical accuracy for diseased tissue removal.</p

Use of Folate-Conjugated Imaging Agents To Target Alternatively Activated Macrophages in a Murine Model of Asthma

Pro-inflammatory macrophages play a prominent role in such autoimmune diseases as rheumatoid arthritis, Crohn’s disease, psoriasis, sarcoidosis, and atherosclerosis. Because pro-inflammatory macrophages have also been shown to overexpress a receptor for the vitamin folic acid (i.e., folate receptor beta; FR-β), folate-linked drugs have been explored for use in imaging and treatment of these same diseases. To determine whether allergic inflammatory disorders might be similarly targeted with folate-linked drugs, we have examined the characteristics of macrophages that are prominent in the pathogenesis of asthma. We report here that macrophages from the lungs of mice with experimental allergic asthma express FR-β. We further document that these FR-β⁺ macrophages coexpress markers of alternatively activated (M2-type) macrophages, including the mannose receptor and arginase-1. Finally, we demonstrate that folate-conjugated fluorescent dyes and radioimaging agents can be specifically targeted to these asthmatic lung macrophages, with little uptake by macrophages present in healthy lung tissue. These data suggest strategies for the development of novel diagnostic agents for the imaging of asthma and other diseases involving alternatively activated macrophages

Development of Tumor-Targeted Near Infrared Probes for Fluorescence Guided Surgery

Complete surgical resection of malignant disease is the only reliable method to cure cancer. Unfortunately, quantitative tumor resection is often limited by a surgeon’s ability to locate all malignant disease and distinguish it from healthy tissue. Fluorescence-guided surgery has emerged as a tool to aid surgeons in the identification and removal of malignant lesions. While nontargeted fluorescent dyes have been shown to passively accumulate in some tumors, the resulting tumor-to-background ratios are often poor, and the boundaries between malignant and healthy tissues can be difficult to define. To circumvent these problems, our laboratory has developed high affinity tumor targeting ligands that bind to receptors that are overexpressed on cancer cells and deliver attached molecules selectively into these cells. In this study, we explore the use of two tumor-specific targeting ligands (i.e., folic acid that targets the folate receptor (FR) and DUPA that targets prostate specific membrane antigen (PSMA)) to deliver near-infrared (NIR) fluorescent dyes specifically to FR and PSMA expressing cancers, thereby rendering only the malignant cells highly fluorescent. We report here that all FR- and PSMA-targeted NIR probes examined bind cultured cancer cells in the low nanomolar range. Moreover, upon intravenous injection into tumor-bearing mice with metastatic disease, these same ligand–NIR dye conjugates render receptor-expressing tumor tissues fluorescent, enabling their facile resection with minimal contamination from healthy tissues