Folic Acid-Conjugated Cellulose Nanocrystals Show High Folate-Receptor Binding Affinity and Uptake by KB and Breast Cancer Cells

Abstract

The study evaluates cellulose nanocrystals (CNCs) as nanocarriers for targeted, intracellular delivery of molecular agents. CNCs were labeled with fluorescein-5′-isothiocyanate as an imaging agent and conjugated to folic acid (FA) as a targeting ligand. The CNC conjugates were characterized by UV–vis spectroscopy, ζ-potential analysis, dynamic light scattering, and atomic force microscopy. Cellular binding/uptake of the FA-conjugated CNCs by KB and MDA-MB-468 cells was quantified with cellular uptake assays. Internalization of the particles was confirmed by confocal microscopy. Uptake mechanisms were determined by inhibition studies with chlorpromazine and genistein. Binding affinity was qualitatively assessed with a free folate inhibition assay. Both KB and MDA-MB-468 cells exhibited significant and folate-receptor specific binding/uptake of FA-conjugated CNCs. Clathrin-mediated endocytosis was a significant uptake mechanism in both cell types, whereas caveolae-mediated endocytosis only played a significant role in MDA-MB-468 cells. Uptake inhibition of FA-conjugated CNCs by KB cells required high concentrations (>1 mM) of free FA. The observed FR-specific internalization of FA-conjugated CNCs by FR-positive cancer cells and tumors and their remarkable high affinity for the FR demonstrate the great potential of CNCs as novel nanocarriers for imaging agents and chemotherapeutics in the early detection and treatment of cancer.

Cellulose Nanocrystals/Chitosan-Based Nanosystems: Synthesis, Characterization, and Cellular Uptake on Breast Cancer Cells

“…As an example, Roman and co-workers have looked extensively into the use of CNCs as nanocarriers by the functionalization of these elongated nanostructures solely with fluorescein isothiocyanate (FITC) as an imaging agent [20][21][22] or with both FITC and folic acid (FA) as imaging and targeting agents [23][24][25], respectively. These authors showed the potential of these nanosystems for the active targeted delivery of chemotherapeutic agents to folate receptor (FR)-positive cancer cells, such as human (DBTRG-05MG, H4) and rat (C6) brain tumor cells [23], KB and human breast cancer cells (MDA-MB-468) [24,25].…”

Folic acid conjugated polymeric drug delivery vehicle for targeted cancer detection in hepatocellular carcinoma

“…The synthesis of a polymeric nanosystem offers several challenges, including the inefficiency associated with chemical conjugation of the targeting folate ligand with the polymer core, as well as the potential cytotoxicity of the vehicle itself. Folate‐targeted intracellular delivery of therapeutic load to pathogenic tissues has been the subject of multiple studies (Bittleman, Dong, Roman, & Lee, ; Saroj & Rajput, ; Stella et al, ; Xia et al, ; Zhang et al, ). All of these studies show conjugation of folic acid into a PEG polymer (Stella et al, ; Zhang et al, ), Silica/Selenium nanoparticle (Saroj & Rajput, ; Xia et al, ), or cellulose nanocrystals (Bittleman et al, ) for selective delivery.…”

“…However, chemical characterization of folic acid in the nano molecule, and/or biological evaluation of the synthesized nanocarriers in non‐folate expressing (negative) cell types are often limited. While these carriers show great promise in cell culture‐based assays, their efficacy and targetability need to be evaluated in an in vivo HCC model (Bittleman et al, ; Saroj & Rajput, ; Stella et al, ; Zhang et al, ). A folate targeting selenium nanoparticle delivering therapeutic siRNA has shown promise in restricting tumors in a xenograft mouse model (Xia et al, ).…”

Nano-Crystallization: An Efficient Technology for Targeting Chemotherapeutic Drugs

“…It showed selective targeting and higher internalization in the cell lines. This proves them as potential diagnostic imaging agents for tumor detection at the early stages [94]. Elongated folic acid modified cellulose nanocrystals and unmodified cellulose crystals were evaluated for their cellular uptake to human brain tumor cells DBTRG-05MG, H4 and C6 rat brain tumor cells.…”