CD38: an ecto-enzyme with functional diversity in T cells.

CD38, a nicotinamide adenine dinucleotide (NAD)+ glycohydrolase, is considered an activation marker of T lymphocytes in humans that is highly expressed during certain chronic viral infections. T cells constitute a heterogeneous population; however, the expression and function of CD38 has been poorly defined in distinct T cell compartments. We investigated the expression and function of CD38 in naïve and effector T cell subsets in the peripheral blood mononuclear cells (PBMCs) from healthy donors and people with HIV (PWH) using flow cytometry. Further, we examined the impact of CD38 expression on intracellular NAD+ levels, mitochondrial function, and intracellular cytokine production in response to virus-specific peptide stimulation (HIV Group specific antigen; Gag). Naïve T cells from healthy donors showed remarkably higher levels of CD38 expression than those of effector cells with concomitant reduced intracellular NAD+ levels, decreased mitochondrial membrane potential and lower metabolic activity. Blockade of CD38 by a small molecule inhibitor, 78c, increased metabolic function, mitochondrial mass and mitochondrial membrane potential in the naïve T lymphocytes. PWH exhibited similar frequencies of CD38+ cells in the T cell subsets. However, CD38 expression increased on Gag-specific IFN-γ and TNF-α producing cell compartments among effector T cells. 78c treatment resulted in reduced cytokine production, indicating its distinct expression and functional profile in different T cell subsets. In summary, in naïve cells high CD38 expression reflects lower metabolic activity, while in effector cells it preferentially contributes to immunopathogenesis by increasing inflammatory cytokine production. Thus, CD38 may be considered as a therapeutic target in chronic viral infections to reduce ongoing immune activation.

Antigen-agnostic microfluidics-based circulating tumor cell enrichment and downstream molecular characterization.

Circulating tumor cells (CTC) isolated from the peripheral blood of cancer patients by a minimally invasive procedure provide surrogate markers of the tumor that can be repeatedly sampled. However, the selection and enumeration of CTCs by traditional methods based on surface proteins like EPCAM may not detect CTCs with a mesenchymal phenotype. Here, we employed an antibody-agnostic platform, the Parsortix® PR1 system, which enriches CTCs based on cell size and membrane deformability. We evaluated the linearity, sensitivity, and specificity of the Parsortix PR1 system in tandem with 3 downstream molecular characterization techniques using healthy donor blood spiked with cultured cell lines. Signal amplification of mRNA using a QuantiGene 25-gene assay was able to quantitate multiple epithelial genes, including CDH1, EGFR, ERBB2, KRT18, and MUC1, from high numbers of spiked cells and was able to detect KRT18 when only 50 MCF-7 or SUM190 cells were spiked into healthy donor blood. However, target amplification of mRNA by quantitative polymerase chain reaction (qPCR) showed better sensitivity; qPCR without pre-amplification was able to detect CTC-related genes in Parsortix PR1-enriched cells when as few as 5 SKBR3 cells were spiked into blood. Finally, the HTG EdgeSeq nuclease protection assay was able to profile mRNA expression of over 2,560 cancer-related genes from Parsortix PR1 enriched cells, showing enrichment in cancer signaling pathways and ERBB2, KRT19, and KRT7. Overall, the Parsortix PR1 platform may be amenable to transition into routine clinical workflows.