Gene expression in circulating tumor cells reveals a dynamic role of EMT and PD-L1 during osimertinib treatment in NSCLC patients

Upon T-cell receptor stimulation, the Programmed cell Death-1 receptor (PD-1) expressed on T-cells can interact with its ligand PD-L1 expressed at the surface of cancer cells or antigen-presenting cells. Monoclonal antibodies targeting PD-1 or PD-L1 are routinely used for the treatment of cancers, but their clinical efficacy varies largely across the variety of tumor types. A part of the variability is linked to the existence of several forms of PD-L1, either expressed on the plasma membrane (mPD-L1), at the surface of secreted cellular exosomes (exoPD-L1), in cell nuclei (nPD-L1), or as a circulating, soluble protein (sPD-L1). Here, we have reviewed the different origins and roles of sPD-L1 in humans to highlight the biochemical and functional heterogeneity of the soluble protein. sPD-L1 isoforms can be generated essentially by two non-exclusive processes: (i) proteolysis of m/exoPD-L1 by metalloproteases, such as metalloproteinases (MMP) and A disintegrin and metalloproteases (ADAM), which are capable of shedding membrane PD-L1 to release an active soluble form, and (ii) the alternative splicing of PD-L1 pre-mRNA, leading in some cases to the release of sPD-L1 protein isoforms lacking the transmembrane domain. The expression and secretion of sPD-L1 have been observed in a large variety of pathologies, well beyond cancer, notably in different pulmonary diseases, chronic inflammatory and autoimmune disorders, and viral diseases. The expression and role of sPD-L1 during pregnancy are also evoked. The structural heterogeneity of sPD-L1 proteins, and associated functional/cellular plurality, should be kept in mind when considering sPD-L1 as a biomarker or as a drug target. The membrane, exosomal and soluble forms of PD-L1 are all integral parts of the highly dynamic PD-1/PD-L1 signaling pathway, essential for immune-tolerance or immune-escape.

Section: The Pd-1/pd-l1 Checkpointmentioning

confidence: 99%

Soluble Programmed Death Ligand-1 (sPD-L1): A Pool of Circulating Proteins Implicated in Health and Diseases

Bailly

Thuru

Quesnel

2021

“…After separating plasma from blood cells, an equal volume of removed plasma was replaced by adding phosphate buffered saline (PBS, pH 7.3) into the cell pellet and then samples were proceeded for CTC enrichment in the size-based microfluidic device, Par-sortix™ (ANGLE plc, Guildford, UK), using a 6.5 µm separation cassette, as previously described [36]. Captured cells were harvested in 200 µL of PBS, genomic DNA (gDNA) was extracted from the CTC fraction using the TRIZOL-LS reagent (Thermo Fisher Scientific, Inc., Waltham, MA, USA) as previously described [37] and isolated gDNA was dissolved in a final volume of 20 µL of 8 mmol/L NaOH.…”

Section: Ctc Enrichment and Genomic Dna Extractionmentioning

confidence: 99%

Detection of EGFR Mutations in Plasma cfDNA and Paired CTCs of NSCLC Patients before and after Osimertinib Therapy Using Crystal Digital PCR

Ntzifa

Κotsakis

Georgoulias

et al. 2021

Self Cite

Circulating tumor DNA (ctDNA) analysis has clinical utility in EGFR mutant NSCLC. Circulating tumor cells (CTCs) consist a unique source of information at the cellular level. Digital PCR (dPCR) is a valuable tool for accurate and valid analysis of gene mutations in liquid biopsy analysis. In the present study we detected EGFR mutations in ctDNA and paired CTCs under osimertinib therapy at two time points using crystal dPCR and the naica® system (Stilla Technologies). We quantified mutation allele frequencies (MAF) of EGFR mutations in 91 plasma cfDNA samples of 48 EGFR mutant NSCLC patients and in 64 matched CTC-derived genomic DNA samples, and the FDA-cleared cobas® EGFR mutation test in 80 identical plasma samples. Direct comparison between crystal dPCR and the cobas EGFR assay revealed a high concordance for all EGFR mutations. Our comparison of crystal dPCR results in ctDNA with the corresponding primary tissue has shown a strong correlation. EGFR mutations analysis in paired CTC-derived gDNA revealed a high heterogeneity. Crystal dPCR offers the unique advantages of high analytical sensitivity, precision, and accuracy for detecting and quantifying multiple EGFR mutations in plasma cfDNA and CTCs of NSCLC patients.

“…In a recent study by Ntzifa et al (2021), samples from 30 patients (at baseline, post first cycle with osimertinib treatment and at progression of disease, PD) were acquired and gene expression of the E markers ( CK8 , CK18 , CK19 ), M/EMT markers ( Vim , Twist1 , AXL ), and of the CSC marker ALDH-1 were analyzed. Interestingly, only gene expression of PD-L1 was significantly different between baseline and PD [ 59 ]. In addition, correlations between genes were observed at the different time points.…”

Section: Clinical Relevance Of Emt and Csc Phenotypes In Nsclc Patientsmentioning

confidence: 99%

“…CK + (CK8/CK18/CK19) CTCs were detected in 76% of the available samples. High Vim + CTC counts suggested a role for EMT during osimertinib treatment [ 59 ].…”

Section: Clinical Relevance Of Emt and Csc Phenotypes In Nsclc Patientsmentioning

confidence: 99%

Clinical Relevance of Mesenchymal- and Stem-Associated Phenotypes in Circulating Tumor Cells Isolated from Lung Cancer Patients

Pantazaka

Vardas

Roumeliotou

et al. 2021

Self Cite

Lung cancer is the leading cause of cancer-related mortality globally. Among the types of lung cancer, non-small-cell lung cancer (NSCLC) is more common, while small-cell lung cancer (SCLC) is less frequent yet more aggressive. Circulating tumor cells (CTCs), albeit rare, have been portrayed as essential players in the progression of lung cancer. CTCs are considered to adopt an epithelial-to-mesenchymal transition (EMT) phenotype and characteristics of cancer stem cells (CSCs). This EMT (or partial) phenotype affords these cells the ability to escape from the primary tumor, travel into the bloodstream, and survive extremely adverse conditions, before colonizing distant foci. Acquisition of CSC features, such as self-renewal, differentiation, and migratory potential, further reflect CTCs’ invasive potential. CSCs have been identified in lung cancer, and expression of EMT markers has previously been correlated with poor clinical outcomes. Thus far, a vast majority of studies have concentrated on CTC detection and enumeration as a prognostic tools of patients’ survival or for monitoring treatment efficacy. In this review, we highlight EMT and CSC markers in CTCs and focus on the clinical significance of these phenotypes in the progression of both non-small- and small-cell lung cancer.