Shape effects of filaments versus spherical particles in flow and drug delivery

Abstract

Interaction of spherical particles with cells and within animals has been studied extensively, but the effects of shape have received little attention. Here we use highly stable, polymer micelle assemblies known as filomicelles to compare the transport and trafficking of flexible filaments with spheres of similar chemistry. In rodents, filomicelles persisted in the circulation up to one week after intravenous injection. This is about ten times longer than their spherical counterparts and is more persistent than any known synthetic nanoparticle. Under fluid flow conditions, spheres and short filomicelles are taken up by cells more readily than longer filaments because the latter are extended by the flow. Preliminary results further demonstrate that filomicelles can effectively deliver the anticancer drug paclitaxel and shrink human-derived tumours in mice. Although these findings show that long-circulating vehicles need not be nanospheres, they also lend insight into possible shape effects of natural filamentous viruses.

Figure 1. Filomicelles and their persistent circulation

a, Filomicelles are self-assembled from di-block copolymers: yellow/green in cross section indicates hydrophobic polymer, orange/blue is hydrophilic, and aqua is water. Electron microscopy demonstrates the nanometre-scale diameter of the filomicelles, and fluorescence microscopy shows a single filomicelle. Distributions of filomicelle length are shown for two samples. b, Injection of fluorescent filomicelles into rodents, followed by fluorescent imaging of blood samples showed that filomicelles circulated in vivo for up to one week. c, Relative numbers of filomicelles in the circulation show that inert filomicelles (of OE7′) persist when compared with stealth polymersomes and λ-phage. d, Degradable filomicelles (of OCL3) also persist, and filomicelles with longer initial lengths (L_o) circulate longer up to a limiting length. The error bars in c, d show the standard deviation for four or more animals.

Figure 2. Kinetics of filomicelle length reduction in vivo

a, Inert filomicelles shorten, with the rate of shortening decreasing as they shorten. The grey region represents the optical limit of L measurements. b, Degradable filomicelles (OCL3) shorten at a rate that depends on initial length. The inset plots the length-dependent shrinkage rate. c, Filomicelles show a saturable increase in half-life of circulating mass, fitting a cooperative clearance model with τ_max = 5.2 days, m = 2.1 and L_α = 2.5 μm. d, Distribution of inert and degradable filomicelles in clearance organs for L_o = 4 or 8 μm after four days in the circulation of rats. All error bars show the standard deviation for three or more animals.

Figure 3. In vitro interactions between filomicelles and phagocytes (P)

a, Fluorescent filomicelles of varying contour length were incubated with activated macrophages for 24 h in static culture. The fluorescence intensity of cells is proportional to the phagocytosis of filomicelles and proves to be a strong function of L_o. The Hill exponent, n = 6, suggests strongly cooperative binding along the length of the cylinder to the cell surface. L_β = 1.9 μm. The error bars in the right-hand plot show the standard deviation. b, In a flow chamber with immobilized phagocytes, long filomicelles (right) flow past the cells, and occasionally leave a fragment, but smaller micelles and vesicles are captured (left, arrows point to small micelles and vesicles). Flow velocity is ∼25 μm s⁻¹, which is similar to that in the spleen. The scale bars represent 5 μm.

Figure 4. Internalization and fragmentation of filomicelles in vitro by human lung-derived epithelial cells

a, Lung cells, prelabelled with a green fluorescent phospholipid probe, were incubated in static cultures with red fluorescent filomicelles. Cell nuclei were later labelled with blue Hoechst dye prior to imaging. b, Cellular uptake of filomicelles increases with incubation time, reaching saturation within 1 h. c, Remaining filomicelles in the culture supernatant shrink in length on a similar timescale.

Figure 5. Filomicelles mediate paclitaxel (TAX) delivery to rapidly growing tumour xenografts on nude mice

Tumour-bearing mice were injected with either saline or OCL3 filomicelles as controls, TAX as free drug in ethanol (1 mg kg⁻¹ is its maximum tolerated dose), or TAX loaded at two doses into the hydrophobic cores of filomicelles of two lengths. a, Apoptosis was measured one week later by quantitative imaging of TUNEL-stained tumour sections and shows little effect of free drug but increasing cell death with increasing L_o and increasing paclitaxel dose. b, Tumour size decreases with increasing apoptosis, with tumour shrinkage clear for the longest filomicelles at the highest TAX dose. All data shows the average from four mice. The error bars show the standard deviation.