Low molecular-weight chitosan as a pH-sensitive stealth coating for tumor-specific drug delivery

Abstract

When a nanoparticle is developed for systemic application, its surface is typically protected by poly(ethylene glycol) (PEG) to help prolonged circulation and evasion of immune clearance. On the other hand, PEG can interfere with interactions between nanocarriers and target cells and negatively influence the therapeutic outcomes. To overcome this challenge, we propose low molecular-weight chitosan (LMWC) as an alternative surface coating, which can protect the nanomedicine in neutral pH but allow cellular interactions in the weakly acidic pH of tumors. LMWCs with a molecular weight of 2-4 kDa, 4-6.5 kDa, and 11-22 kDa were produced by hydrogen peroxide digestion and covalently conjugated with poly(lactic-co-glycolic acid) (PLGA). Nanoparticles created with PLGA-LMWC conjugates showed pH-sensitive cell interactions, which enabled specific drug delivery to cells in a weakly acidic environment. The hydrophilic LMWC layer reduced opsonization and phagocytic uptake. These properties qualify LMWCs as a promising biomaterial for pH-sensitive stealth coating.

Fig. 1

pH-dependent transmittance change of chitosans

Fig. 2

FTIR spectroscopy of PLGA-LMWCs, PLGA and LMWC_4–6.5k. The presence of amide and amine bands (arrow, 1500–1690 cm⁻¹) and a broad N-H signal (brace, 3000–3700 cm⁻¹) confirms the conjugation of LMWC to PLGA.

Fig. 3

Scanning electron microscope images of PLGA NPs and PLGA-LMWC_2–4k NPs.

Fig. 4

pH dependence of cellular association of PLGA* NPs and PLGA*-LMWC NPs. Confocal images of (a) SKOV-3 ovarian cancer cells or (b) NCI/ADR-RES multidrug resistant ovarian cancer cells were taken after 3-hour incubation with the NPs (overlaid images of NP (green), nuclei (blue), and transmission images).

Fig. 4

pH dependence of cellular association of PLGA* NPs and PLGA*-LMWC NPs. Confocal images of (a) SKOV-3 ovarian cancer cells or (b) NCI/ADR-RES multidrug resistant ovarian cancer cells were taken after 3-hour incubation with the NPs (overlaid images of NP (green), nuclei (blue), and transmission images).

Fig. 5

J774A.1 macrophages incubated with PLGA* NPs or PLGA*-LMWC_2–4k NPs for 3 hours at pH 7.4. Overlaid images of NP (green), nuclei (blue), and transmission images.

Fig. 6

Viability of SKOV-3 cells exposed to PTX or PTX/NPs at different pHs for (a) 72 hours or (b) 3 hours. Data are expressed as averages with standard deviations of 4 identically and independently prepared samples.

Fig. 7

In vitro release of PTX from NPs. There was no difference across the samples at each time point (p>0.05 with ANOVA). Data are expressed as averages with standard deviations of 3 identically and independently prepared samples. For the clarity of presentation, error bars are shown in one direction.

Fig. 8

Protein adsorbed to 1 mg PLGA mPs or PLGA-LMWC mPs. All samples are significantly different from each other with p values less than 0.001, except for PLGA-LMWC_2–4k mPs vs. PLGA-LMWC_11–22k mPs (p<0.05). Data are expressed as averages with standard deviations of 6 identically and independently prepared samples.

Scheme 1

Synthesis of PLGA-LMWC and schematic diagram of PLGA-LMWC NP