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Abstract Anticancer drug development is a crucial step toward cancer treatment, that requires realistic predictions of malignant tissue development and sophisticated drug delivery. Tumors often acquire drug resistance and drug efficacy, hence cannot be accurately predicted in 2D tumor cell cultures. On the other hand, 3D cultures, including multicellular tumor spheroids (MCTSs), mimic the in vivo cellular arrangement and provide robust platforms for drug testing when grown in hydrogels with characteristics similar to the living body. Microparticles and liposomes are considered smart drug delivery vehicles, are able to target cancerous tissue, and can release entrapped drugs on demand. Microfluidics serve as a high‐throughput tool for reproducible, flexible, and automated production of droplet‐based microscale constructs, tailored to the desired final application. In this review, it is described how natural hydrogels in combination with droplet microfluidics can generate MCTSs, and the use of microfluidics to produce tumor targeting microparticles and liposomes. One of the highlights of the review documents the use of the bottom‐up construction methodologies of synthetic biology for the formation of artificial cellular assemblies, which may additionally incorporate both target cancer cells and prospective drug candidates, as an integrated “droplet incubator” drug assay platform.

Abstract High specification, polymer capsules, to produce inertial fusion energy targets, were continuously fabricated using surfactant-free, inertial centralisation, and ultrafast polymerisation, in a scalable flow reactor. Laser-driven, inertial confinement fusion depends upon the interaction of high-energy lasers and hydrogen isotopes, contained within small, spherical and concentric target shells, causing a nuclear fusion reaction at ~150 M°C. Potentially, targets will be consumed at ~1 M per day per reactor, demanding a 5000x unit cost reduction to ~$0.20, and is a critical, key challenge. Experimentally, double emulsions were used as templates for capsule-shells, and were formed at 20 Hz, on a fluidic chip. Droplets were centralised in a dynamic flow, and their shapes both evaluated, and mathematically modeled, before subsequent shell solidification. The shells were photo-cured individually, on-the-fly, with precisely-actuated, millisecond-length (70 ms), uniform-intensity UV pulses, delivered through eight, radially orchestrated light-pipes. The near 100% yield rate of uniform shells had a minimum 99.0% concentricity and sphericity, and the solidification processing period was significantly reduced, over conventional batch methods. The data suggest the new possibility of a continuous, on-the-fly, IFE target fabrication process, employing sequential processing operations within a continuous enclosed duct system, which may include cryogenic fuel-filling, and shell curing, to produce ready-to-use IFE targets.

Changes in the cellular secretome are implicated in virus infection, malignancy, and anti-tumor immunity. We analyzed the association between transcriptional signatures (TS) from 24 different immune and stromal cell types on the prognosis of HPV-infected and HPV-free head and neck squamous carcinoma (HNSCC) patients from The Cancer Genome Atlas (TCGA) cohort. We found that HPV-positive HNSCC patients have tumors with elevated immune cell TS and improved prognosis, which was specifically associated with an increased tumor abundance of memory B and activated natural killer (NK) cell TS, compared to HPV-free HNSCC patients. HPV-infected patients upregulated many transcripts encoding secreted factors, such as growth factors, hormones, chemokines and cytokines, and their cognate receptors. Analysis of secretome transcripts and cognate receptors revealed that tumor expression of IL17RB and IL17REL are associated with a higher viral load and memory B and activated NK cell TS, as well as improved prognosis in HPV-infected HNSCC patients. The transcriptional parameters that we describe may be optimized to improve prognosis and risk stratification in the clinic and provide insights into gene and cellular targets that may potentially enhance anti-tumor immunity mediated by NK cells and memory B cells in HPV-infected HNSCC patients.

Human natural killer (NK) cells can be sub-divided into two functional subsets but the clinical significance of these CD56bright and CD56dim NK cells in anti-tumour immunity remains largely unexplored. We determined the relative abundances of gene signatures for CD56bright and CD56dim NK cells along with 3 stromal and 18 other immune cell types in the patient tumour transcriptomes from the cancer genome atlas bladder cancer dataset (TCGA-BLCA). Using this computational approach, CD56bright NK cells were predicted to be the more abundant tumour-infiltrating NK subset which was also associated with improved patient prognosis. A similar favorable survival trend was projected using gene signatures for mature myeloid dendritic cells (mDC) and CD8+ effector memory T cells (TEM) and unveiled a potential CD56bright NK-mDC-CD8+T cell crosstalk in the BLCA tumour microenvironment. Expression of transcripts encoding the activating NK cell receptors, NKG2D, NKp44, CD2, and CD160, showed positive survival trends in combination with CD56bright NK cell infiltration. Transcription factors including HOBIT, IRF3, and STAT2 were also correlated with CD56bright NK cell abundance. Additionally, a HOBIT-dependent tissue-residency program correlated with the CD56bright NK and CD8+ TEM cell signatures was found to be associated with favourable BLCA patient survival. Overall, our study highlights the significance of CD56bright NK cells in BLCA patient prognosis. Our findings facilitate a better understanding of the NK cell anti-tumour responses that may ultimately lead to the development of promising NK and T cell-based therapies for BLCA.

Activation of natural killer (NK) cell function is regulated by cytokines, such as IL-2, and secreted factors upregulated in the tumor microenvironment, such as platelet-derived growth factor D (PDGF-DD). In order to elucidate a clinical role for these important regulators of NK cell function in antitumor immunity, we generated transcriptional signatures representing resting, IL-2-expanded, and PDGF-DD-activated, NK cell phenotypes and established their abundance in The Cancer Genome Atlas bladder cancer (BLCA) dataset using CIBERSORT. The IL-2-expanded NK cell phenotype was the most abundant in low and high grades of BLCA tumors and was associated with improved prognosis. In contrast, PDGFD expression was associated with numerous cancer hallmark pathways in BLCA tumors compared with normal bladder tissue, and a high tumor abundance of PDGFD transcripts and the PDGF-DD-activated NK cell phenotype were associated with a poor BLCA prognosis. Finally, high tumor expression of transcripts encoding the activating NK cell receptors, KLRK1 and the CD160–TNFRSF14 receptor–ligand pair, was strongly correlated with the IL-2-expanded NK cell phenotype and improved BLCA prognosis. The transcriptional parameters we describe may be optimized to improve BLCA patient prognosis and risk stratification in the clinic and potentially provide gene targets of therapeutic significance for enhancing NK cell antitumor immunity in BLCA.

The binding of platelet-derived growth factor D (PDGF-DD) to the NKp44 receptor activates a distinct transcriptional program in primary IL-2 expanded human natural killer (NK) cells. We were interested in knowing if the PDGF-DD-NKp44 pathway of NK cell activation might play a clinically relevant role in anti-tumor immunity. In order to address this question, we determined transcriptional signatures unique to resting, IL-2 expanded, and PDGF-DD activated, NK cells, in addition to different T cell subsets, and established the abundance of these immune cell phenotypes in The Cancer Genome Atlas (TCGA) low-grade glioma (LGG) dataset using CIBERSORT. Our results show that LGG patient tumors enriched for either the PDGF-DD activated NK cell or memory CD8+ T cell phenotypes are associated with a more favorable prognosis. Combined cell phenotype analyses revealed that patients with LGG tumors enriched for the PDGF-DD activated NK cell phenotype and the CD4+ T helper cell phenotype had a more favorable prognosis. High expression of transcripts encoding members of the killer cell lectin-like receptor (KLR) family, such as KLRK1 and KLRC2, KLRC3 and KLRC4 in LGG tumors were associated with more favorable prognosis, suggesting that these NK cell family receptors may play a prominent role in LGG anti-tumor immunity. Finally, many of the TCGA findings were reciprocated in LGG patients from the Chinese Glioma Genome Atlas (CGGA) dataset. Our results provide transcriptomic evidence that PDGF-DD activated NK cells and KLR family receptors may play an important clinical role in immune surveillance of LGG.

The Natural Cytotoxicity Receptors (NCRs), NKp46, NKp44, and NKp30, were some of the first human activating Natural Killer (NK) cell receptors involved in the non-MHC-restricted recognition of tumor cells to be cloned over 20 years ago. Since this time many host- and pathogen-encoded ligands have been proposed to bind the NCRs and regulate the cytotoxic and cytokine-secreting functions of tissue NK cells. This diverse set of NCR ligands can manifest on the surface of tumor or virus-infected cells or can be secreted extracellularly, suggesting a remarkable NCR polyfunctionality that regulates the activity of NK cells in different tissue compartments during steady state or inflammation. Moreover, the NCRs can also be expressed by other innate and adaptive immune cell subsets under certain tissue conditions potentially conferring NK recognition programs to these cells. Here we review NCR biology in health and disease with particular reference to how this important class of receptors regulates the functions of tissue NK cells as well as confer NK cell recognition patterns to other innate and adaptive lymphocyte subsets. Finally, we highlight how NCR biology is being harnessed for novel therapeutic interventions particularly for enhanced tumor surveillance.