Search

Recently, there has been an increasing emphasis on single cell profiling for high-throughput screening workflows in drug discovery and life sciences research. However, the biology underpinning these screens is often complex and is insufficiently addressed by singleplex assay screens. Traditional single cell screening technologies have created powerful sets of 'omic data that allow users to bioinformatically infer biological function, but have as of yet not empowered direct functional analysis at the level of each individual cell. Consequently, screening campaigns often require multiple secondary screens leading to laborious, time-consuming and expensive workflows in which attrition points may not be queried until late in the process. We describe a platform that harnesses droplet microfluidics and optical electrowetting-on-dielectric (oEWOD) to perform highly-controlled sequential and multiplexed single cell assays in massively parallelised workflows to enable complex cell profiling during screening. Soluble reagents or objects, such as cells or assay beads, are encapsulated into droplets of media in fluorous oil and are actively filtered based on size and optical features ensuring only desirable droplets ( e.g. single cell droplets) are retained for analysis, thereby overcoming the Poisson probability distribution. Droplets are stored in an array on a temperature-controlled chip and the history of individual droplets is logged from the point of filter until completion of the workflow. On chip, droplets are subject to an automated and flexible suite of operations including the merging of sample droplets and the fluorescent acquisition of assay readouts to enable complex sequential assay workflows. To demonstrate the broad utility of the platform, we present examples of single-cell functional workflows for various applications such as antibody discovery, infectious disease, and cell and gene therapy.

Brain tumors are the leading cause of cancer-related death in children and are the most challenging childhood cancer in relation to diagnosis, treatment, and outcome. One potential novel strategy to improve outcomes in cancer involves the manipulation of autophagy, a fundamental process in all cells. In cancer, autophagy can be thought of as having a "Janus"-like duality. On one face, especially in the early phases of cancer formation, autophagy can act as a cellular housekeeper to eliminate damaged organelles and recycle macromolecules, thus acting as tumor suppressor. On the other face, at later stages of tumor progression, autophagy can function as a pro-survival pathway in response to metabolic stresses such as nutrient depravation, hypoxia and indeed to chemotherapy itself, and can support cell growth by supplying much needed energy. In the context of chemotherapy, autophagy may, in some cases, mediate resistance to treatment. We present an overview of the relevance of autophagy in central nervous system tumors including how its chemical modulation can serve as a useful adjunct to chemotherapy, and use this knowledge to consider how targeting of autophagy may be relevant in pediatric brain tumors., Competing Interests: CONFLICTS OF INTEREST The authors declare no conflicts of interest.

Central nervous system tumors are the most common cancer type in children and the leading cause of cancer related deaths. There is therefore a need to develop novel treatments. Large scale profiling studies have begun to identify alterations that could be targeted therapeutically, including the phosphoinositide 3-kinase (PI3K) signaling pathway, which is one of the most commonly activated pathways in cancer with many inhibitors under clinical development. PI3K signaling has been shown to be aberrantly activated in many pediatric CNS neoplasms. Pre-clinical analysis supports a role for PI3K signaling in the control of tumor growth, survival and migration as well as enhancing the cytotoxic effects of current treatments. Based on this evidence agents targeting PI3K signaling have begun to be tested in clinical trials of pediatric cancer patients. Overall, targeting the PI3K pathway presents as a promising strategy for the treatment of pediatric CNS tumors. In this review we examine the genetic alterations found in the PI3K pathway in pediatric CNS tumors and the pathological role it plays, as well as summarizing the current pre-clinical and clinical data supporting the use of PI3K pathway inhibitors for the treatment of these tumors.