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Technical Note
Microfluidic-Enabled Print-to-Screen (P2S) Platform for HighThroughput Screening of Combinatorial Chemotherapy Yuzhe Ding, Jiannan Li, Wenwu Xiao, Kai Xiao, Joyce S Lee, Urvashi Bhardwaj, Zijie Zhu, Philip Digiglio, Gaomai Yang, Kit S. Lam, and Tingrui Pan Anal. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.analchem.5b00826 • Publication Date (Web): 03 Sep 2015 Downloaded from http://pubs.acs.org on September 12, 2015
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Analytical Chemistry
Microfluidic-Enabled Print-to-Screen (P2S) Platform for HighThroughput Screening of Combinatorial Chemotherapy Yuzhe Ding,a,‡ Jiannan Li,a,‡ Wenwu Xiao,b Kai Xiao,b Joyce Lee,b Urvashi Bhardwaj,b Zijie Zhu,a Philip Digiglio,a Gaomai Yang,a Kit S. Lam,b and Tingrui Pan*,a a
Micro-Nano Innovations (MiNI) Laboratory, Biomedical Engineering, University of California, Davis, USA
b
Department of Biochemistry and Molecular Medicine, Division of Hematology and Oncology, UC Davis Cancer Center, University of California, Davis, USA ABSTRACT: Since the 1960s, combination chemotherapy has been widely utilized as a standard method to treat cancer. However, due to the potentially enormous number of drug candidates and combinations, conventional identification methods of the effective drug combinations are usually associated with significantly high operational costs, low throughput screening, laborious and time-consuming procedures, and ethical concerns. In this paper, we present a low-cost, high-efficiency microfluidic print-to-screen (P2S) platform, which integrates combinatorial screening with biomolecular printing for high-throughput screening of anti-cancer drug combinations. This P2S platform provides several distinct advantages and features, including automatic combinatorial printing, high-throughput parallel drug screening, modular disposable cartridge and biocompatibility, which can potentially speed up the entire discovery cycle of potent drug combinations. Microfluidic impact printing utilizing plug-and-play microfluidic cartridges is experimentally characterized with controllable droplet volume and accurate positioning. Furthermore, combinatorial print-to-screen assay is demonstrated in a proof-of-concept biological experiment which can identify the positive hits among the entire drug combination library in a parallel and rapid manner. Overall, this microfluidic print-to-screen platform offers a simple, low-cost, high-efficiency solution for high-throughput large-scale combinatorial screening, and can be applicable for various emerging applications in drug cocktail discovery.
In cancer therapy, combination of two or more chemotherapeutic drugs with different mechanisms of action is often given simultaneously or sequentially, in multiple cycles, to cancer patients.1 Although each drug is typically given at a reduced dose, the combinatorial approach can show increased efficacy with reduced toxicity compared with mono-drug therapy.2 With the advent of targeted cancer therapeutics in recent years, an increasing number of target-specific drugs, such as tyrosine kinase inhibitors, have been approved for clinical use.3 It becomes evident that these new drugs work best in combination with standard chemotherapeutic drugs.4 It has also been discovered in recent years that drugs approved for treatment of diseases other than cancer can potentiate the anticancer effects of standard chemotherapeutic agents.5 According to the National Cancer Institute, over 300 chemotherapeutic drugs and more than 50 target-specific anticancer drugs to date have received regulatory approvals for clinical cancer therapy worldwide. In addition, there are over 1000 FDA-approved non-cancer drugs, some of which might also have synergistic effect with the anticancer drugs for cancer treatment.5 Of the potentially enormous combinations of these drugs, only a very small fraction have undergone preclinical testing, which often involves manual preparation and evaluation of their anti-
cancer efficacy in cancer cell lines using conventional micro-titer plates. Once efficacious combinations are identified, they will be evaluated in syngeneic, transgenic, or xenograft tumor models in mice, which are typically associated with significantly high operation cost, low screening throughput and time-consuming procedures, and ethical concerns.6 Therefore, a low-cost highthroughput screening technique can be in high demand for rapid preparation and discovery of efficacious drug combinations prior to animal testing. Multiple recent studies have demonstrated that micropatterned drug arrays have potential to be utilized in in vitro combinatorial drug screening for high throughput, high efficiency with substantially reduced operational cost and time.7 Unfortunately, two major technical hurdles limit the potential of microarray-based drug screening: the generation, and the screening, of a large-scale combination microarrays. First, there is a lack of convenient and robust technique to generate large-scale combinatorial drug arrays from a large number of drug candidates. With the recent advances in micro- and nanofabrication, a number of biological patterning techniques have been proposed to form massive drug/cell microarrays for screening, which can generally be divided into the following three categories:
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Analytical Chemistry
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microfluidic-based patterning, contact, and non-contact printing. Specifically, microfluidic-based drug screening integrates parallel generation of drug concentrations with an on-chip cell culture, which can generate and screen different combinations of two drugs with various concentration levels in a single step.8 However, it can only deal with a limited number of combinations (e.g., 64 concentration combinations from two drugs), due to the system complexity and potential crosstalk between adjacent drug reservoirs.9 Contact printing techniques, primarily the pin-based printing utilized in microarrayers, transfer target biomolecules to the substrate through a physical contact between the “ink”-soaked pins and the substrate. Although the principle of the contact printing is straightforward, low throughput and potential crosscontamination pose serious concerns for their usage in high-throughput drug discovery.10 Non-contact printing recently becomes a preferred route to establish combinatorial microarrays. It keeps the printer head from physical contact with the substrate, eliminating the potential for cross-contamination. Various high-throughput printing mechanisms have already been established in industrial and consumer applications. In particular, inkjet printing with integrated piezoelectric-actuated cartridges can dispense more than 10,000 droplets per nozzle in a second;11 however, the cartridge design comes with extensive loading and dead volumes, and presents a significant barrier to incorporate precious chemical reagents into such a system. More recently, we have introduced a microfluidic-enabled non-contact printing technique, referred to as Microfluidic Impact Printing (MIP), adapted from both non-contact inkjet printing and dot-matrix printing. It combines many desirable features, including no cross-contamination, wide compatibility of printing solutions, and multiplexed printing with self-alignment. Importantly, the interchangeable microfluidic cartridge design permits custom loading volume and minimal dead volume (
