Comparison of gene expression and flow cytometry for immune profiling in chronic lymphocytic leukaemia

Sharpe, Chia; Davis, Joanne E.; Mason, Kylie D.; Tam, Constantine S.; Ritchie, David; Koldej, Rachel

doi:10.1016/j.jim.2018.09.013

Cited by 5 publications

(3 citation statements)

References 22 publications

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“…To determine if the elevated chemokine profile was associated with increased immune cell infiltration in the kidney tissue, we used immune cell gene signatures to compare cell scores among AIN, ATN and HTN groups. These scores are indicative of the relative abundance of cells in the tissue (17,18).…”

Section: Identification Of Differentially Expressed Genes and Infiltr...mentioning

confidence: 99%

Tertiary lymphoid structure signatures are associated with immune checkpoint inhibitor related acute interstitial nephritis

Singh¹,

Long²,

Tchakarov³

et al. 2022

JCI Insight

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Tertiary lymphoid structures (TLSs) are associated with anti-tumor response following immune checkpoint inhibitor (ICI) therapy, but a commensurate observation of TLS is absent for immune related adverse events (irAEs) i.e. acute interstitial nephritis (AIN). We hypothesized that TLS-associated inflammatory gene signatures are present in AIN and performed NanoString-based gene expression and multiplex 12-chemokine profiling on paired kidney tissue, urine and plasma specimens of 36 participants who developed acute kidney injury (AKI) on ICI therapy: AIN (18), acute tubular necrosis (9), or HTN nephrosclerosis (9). Increased T and B cell scores, a Th1-CD8 + T cell axis accompanied by interferon-g and TNF superfamily signatures were detected in the ICI-AIN group. TLS signatures were significantly increased in AIN cases and supported by histopathological identification. Furthermore, urinary TLS signature scores correlated with ICI-AIN diagnosis but not paired plasma. Urinary CXCL9 correlated best to tissue CXCL9 expression (rho 0.75, p < 0.001) and the ability to discriminate AIN vs. non-AIN (AUC 0.781, p-value 0.003). For the first time, we report the presence of TLS signatures in irAEs, define distinctive immune signatures, identify chemokine markers distinguishing ICI-AIN from common AKI etiologies and demonstrate that urine chemokine markers may be used as a surrogate for ICI-AIN diagnoses.

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Section: Identification Of Differentially Expressed Genes and Infiltr...mentioning

confidence: 99%

Tertiary lymphoid structure signatures are associated with immune checkpoint inhibitor related acute interstitial nephritis

Singh¹,

Long²,

Tchakarov³

et al. 2022

JCI Insight

View full text Add to dashboard Cite

show abstract

“…Pathways and cell types are predefined using specific gene sets, as described previously [13]. The definition of cell types using these gene sets were found to be strongly correlated to cell types measured by flow cytometry in different tissue types, including cancers [14][15][16]. Differences in pathway scores and cell type scores were tested with Mann-Whitney U tests and Benjamin-Hochberg procedures were used to correct for multiple testing.…”

Section: Discussionmentioning

confidence: 99%

RNA from stabilized whole blood enables more comprehensive immune gene expression profiling compared to RNA from peripheral blood mononuclear cells

Sijde

Schraauwen

et al. 2020

PLoS ONE

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Monitoring changes in the immune profile in blood samples can help identifying changes in tumor biology and therapy responsiveness over time. Immune-related gene expression profiles offer a highly reproducible method to monitor changes of the immune system. However, measuring gene expression profiles in whole blood samples can be complicated because of the high protein and enzyme abundancy that affect the stability and quality of the RNA. Peripheral blood mononuclear cells (PBMCs) are one the most commonly used source for immune cell RNA extraction, though, this method does not reflect all components of the peripheral blood. The aim of this study was to determine the differences in immune-related gene expression between RNA isolated from stabilized whole blood and RNA isolated from PBMCs. Whole blood samples from 12 pancreatic cancer patients were collected before and after chemotherapy (n = 24). Blood samples were collected in both EDTA tubes, and Tempus tubes containing an RNA stabilizer (total n = 48). PBMCs were isolated from EDTA samples using Ficoll and were snap frozen. Subsequently, immune-related gene expression was profiled using the PanCancer Immune Profiling Panel of NanoString technology. Gene expression profiles of PBMCs were compared to that of Tempus tubes using the Advanced Analysis module of nSolver software. Both types of samples provided good quality RNA and gene expression measurements. However, RNA isolated from Tempus tubes resulted in significantly higher gene counts than PBMCs; 107/730 genes were exclusively detected in Tempus samples, while under the detection limit in PBMCs. In addition, 192/730 genes showed significantly higher gene counts in Tempus samples, 157/730 genes showed higher gene counts in PBMCs. Thus, RNA isolated from whole blood stabilizing blood tubes, such as Tempus tubes, enable higher gene counts and more comprehensive measurements of gene expression profiles compared to RNA isolated from PBMCs.

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“…Currently, flow cytometry is the gold standard of PBMC counting due to its proven sensitivity and reliability; however, it relies on highly specialized and bulky equipment in lab facilities and thus is logistically challenging to be used for frequently monitoring the trajectory of the immune functions under an evolving disease status. In addition, it often requires complex sample preparation procedures, including fluorescent labeling (which mostly relies on a panel of eight fluorescent markers and is often challenged by spectral overlap [6,7]) and technical expertise, which adds extra difficulties and costs for implementing them as point-of-care (POC) tests that allow PBMC monitoring on a regular basis to better triage patients for the most suitable care options and allocate medical resources in an effective manner [8]. Therefore, there is an unmet need for developing an easy-to-operate platform that can quantitatively detect PBMCs in a rapid and sensitive manner.…”

Section: Introductionmentioning

confidence: 99%

A Digital Microfluidic Device Integrated with Electrochemical Impedance Spectroscopy for Cell-Based Immunoassay

Zhang

Liu

2022

Biosensors

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The dynamic immune response to various diseases and therapies has been considered a promising indicator of disease status and therapeutic effectiveness. For instance, the human peripheral blood mononuclear cell (PBMC), as a major player in the immune system, is an important index to indicate a patient’s immune function. Therefore, establishing a simple yet sensitive tool that can frequently assess the immune system during the course of disease and treatment is of great importance. This study introduced an integrated system that includes an electrochemical impedance spectroscope (EIS)-based biosensor in a digital microfluidic (DMF) device, to quantify the PBMC abundance with minimally trained hands. Moreover, we exploited the unique droplet manipulation feature of the DMF platform and conducted a dynamic cell capture assay, which enhanced the detection signal by 2.4-fold. This integrated system was able to detect as few as 104 PBMCs per mL, presenting suitable sensitivity to quantify PBMCs. This integrated system is easy-to-operate and sensitive, and therefore holds great potential as a powerful tool to profile immune-mediated therapeutic responses in a timely manner, which can be further evolved as a point-of-care diagnostic device to conduct near-patient tests from blood samples.

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Comparison of gene expression and flow cytometry for immune profiling in chronic lymphocytic leukaemia

Cited by 5 publications

References 22 publications

Tertiary lymphoid structure signatures are associated with immune checkpoint inhibitor related acute interstitial nephritis

Tertiary lymphoid structure signatures are associated with immune checkpoint inhibitor related acute interstitial nephritis

RNA from stabilized whole blood enables more comprehensive immune gene expression profiling compared to RNA from peripheral blood mononuclear cells

A Digital Microfluidic Device Integrated with Electrochemical Impedance Spectroscopy for Cell-Based Immunoassay

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