Quantitative dose–response curves from subcellular lipid multilayer microarrays

Abstract

The dose-dependent bioactivity of small molecules on cells is a crucial factor in drug discovery and personalized medicine. Although small-molecule microarrays are a promising platform for miniaturized screening, it has been a challenge to use them to obtain quantitative dose-response curves in vitro, especially for lipophilic compounds. Here we establish a small-molecule microarray assay capable of controlling the dosage of small lipophilic molecules delivered to cells by varying the sub-cellular volumes of surface supported lipid micro- and nanostructure arrays fabricated with nanointaglio. Features with sub-cellular lateral dimensions were found necessary to obtain normal cell adhesion with HeLa cells. The volumes of the lipophilic drug-containing nanostructures were determined using a fluorescence microscope calibrated by atomic-force microscopy. We used the surface supported lipid volume information to obtain EC-50 values for the response of HeLa cells to three FDA-approved lipophilic anticancer drugs, docetaxel, imiquimod and triethylenemelamine, which were found to be significantly different from neat lipid controls. No significant toxicity was observed on the control cells surrounding the drug/lipid patterns, indicating lack of interference or leakage from the arrays. Comparison of the microarray data to dose-response curves for the same drugs delivered liposomally from solution revealed quantitative differences in the efficacy values, which we explain in terms of cell-adhesion playing a more important role in the surface-based assay. The assay should be scalable to a density of at least 10,000 dose response curves on the area of a standard microtiter plate.

Enhanced cellular uptake of size-separated lipophilic silicon nanoparticles

“…This fraction-dependent effect appears to be related to the specific interaction with liposomes that can affect their encapsulation efficiency and hence the efficiency of internalization should they be used in the hydrophobic form. This work demonstrates the value of further purification and screening of synthesized nanoparticles and opens the possibility of screening lipophilic nanomaterial drugs and libraries626364.…”

Screening of Lipid Composition for Scalable Fabrication of Solvent-Free Lipid Microarrays

“…We tested 210 lipid formulations, and a 1:1 mixture of castor oil and hexanoic acid was found to be optimal in terms of droplet size, reproducibility of printed patterns, florescence intensity, and stability under immersion. Compared to phospholipid carriers (Kusi-Appiah et al, 2015), this formulation can be arrayed without the need for an additional solvent. The lipid itself can be considered the solvent for the fabrication of drug screening microarrays.…”

“…This poses a challenge for delivery to cells through aqueous solution. We use lipid multilayer (or droplet) microarrays to temporarily immobilize lipophilic compounds onto a surface, allowing cellular uptake and quantitative dose–response curves (Kusi-Appiah et al, 2012; Kusi-Appiah et al, 2015). A crucial property of lipid multilayer microarrays for drug screening applications is that the layer must be thicker than a single monolayer or bilayer in order to contain enough drug to reach biologically relevant dosages upon cellular uptake.…”

Nanobiotechnology medical applications: Overcoming challenges through innovation

“…This lipid technology was also used to develop lipid-oil droplet arrays that do not require additional solvent and can assay lipophilic drugs (87). These assays can also determine the dose-dependent bioactivity of large quantities of small molecules to determine their potential as a nanotherapeutic in nanomedicine (88,89). This lipid technology has also been used in the development of "nanointaglio, " which detects analytes in air vapors by optically sensing topographical changes to lipid grates upon the passage and binding of vapor analytes (88)(89)(90).…”

“…These assays can also determine the dose-dependent bioactivity of large quantities of small molecules to determine their potential as a nanotherapeutic in nanomedicine (88,89). This lipid technology has also been used in the development of "nanointaglio, " which detects analytes in air vapors by optically sensing topographical changes to lipid grates upon the passage and binding of vapor analytes (88)(89)(90). Overall, bioMEMS are completely revolutionizing in vitro analyte and disease diagnostics, biological sample analysis, NP characterization, and lead selection through bioactivity high throughput screenings.…”