Peripheral cytokine levels as a prognostic indicator in gastric cancer : a review of existing literature

Although strong connections exist between the carcinogenesis of gastric cancer and chronic inflammation, gastric cancer is unique in that the chronic gastritis which frequently precedes carcinogenesis is strongly associated with H. pylori infection. The interplay between H. pylori virulence factors and host immune cells is complex but culminates in the activation of inflammatory pathways and transcription factors such as NF-κB, STAT3, and AP-1, all of which upregulate cytokine production. Due to the key role of cytokines in modulating the immune response against tumour cells as well as possibly stimulating tumour growth and proliferation, different patterns of cytokine secretion may be associated with varying patient outcomes. In relation to gastric cancer, interleukin-6, 8, 10, 17A, TNF, and IFN-γ may have pro-tumour properties, although interleukin-10, TNF, and IFN-γ may have anti-tumour effects. However, due to the lack of studies investigating patient outcomes, only a link between higher interleukin-6 levels and poorer prognosis has been demonstrated. Further investigations which link peripheral cytokine levels to patient prognosis may elucidate important pathological mechanisms in gastric cancer which adversely impact patient survival and allow treatments targeting these processes to be developed

The multiple potential biomarkers for predicting immunotherapy response : finding the needle in the haystack

Immune checkpoint inhibitors (ICIs) are being increasingly utilised in a variety of advanced malignancies. Despite promising outcomes in certain patients, the majority will not derive benefit and are at risk of potentially serious immune-related adverse events (irAEs). The development of predictive biomarkers is therefore critical to personalise treatments and improve outcomes. A number of biomarkers have shown promising results, including from tumour (programmed cell death ligand 1 (PD-L1), tumour mutational burden (TMB), stimulator of interferon genes (STING) and apoptosis-associated speck-like protein containing a CARD (ASC)), from blood (peripheral blood mononuclear cells (PBMCs), circulating tumour DNA (ctDNA), exosomes, cytokines and metal chelators) and finally the microbiome

A robust approach to differentiate human monocyte-derived microglia from peripheral blood mononuclear cells

Microglia are implicated in most neurodegenerative diseases. Here, we present a robust and efficient protocol to differentiate monocyte-derived microglia-like cells (MDMi) from whole blood. The protocol consists of three parts. The first part will describe two methods for PBMC isolation. This will be followed by MDMi differentiation, and lastly, the characterization of MDMi by immunocytochemistry. MDMi can be used to investigate microglial-related responses in various age-related neurodegenerative diseases and can be applied to drug testing on a personalized basis. For complete details on the use and execution of this protocol, please refer to Quek et al

Molecular biomarkers in glioblastoma : a systematic review and meta-analysis

BACKGROUND: Glioblastoma (GBM) is a highly aggressive cancer with poor prognosis that needs better treatment modalities. Moreover, there is a lack of reliable biomarkers to predict the response and outcome of current or newly designed therapies. While several molecular markers have been proposed as potential biomarkers for GBM, their uptake into clinical settings is slow and impeded by marker heterogeneity. Detailed assessment of prognostic and predictive value for biomarkers in well-defined clinical trial settings, if available, is scattered throughout the literature. Here we conducted a systematic review and meta-analysis to evaluate the prognostic and predictive significance of clinically relevant molecular biomarkers in GBM patients. MATERIAL AND METHODS: A comprehensive literature search was conducted to retrieve publications from 3 databases (Pubmed, Cochrane and Embase) from January 2010 to December 2021, using specific terms. The combined hazard ratios (HR) and confidence intervals (95% CI) were used to evaluate the association of biomarkers with overall survival (OS) in GBM patients. RESULTS: Twenty-six out of 1831 screened articles were included in this review. Nineteen articles were included in the meta-analyses, and 7 articles were quantitatively summarised. Fourteen studies with 1231 GBM patients showed a significant association of MGMT methylation with better OS with the pooled HR of 1.66 (95% CI 1.32-2.09, p < 0.0001, random effect). Five studies including 541 GBM patients analysed for the prognostic significance of IDH1 mutation showed significantly better OS in patients with IDH1 mutation with a pooled HR of 2.37 (95% CI 1.81-3.12; p < 0.00001]. Meta-analysis performed on 5 studies including 575 GBM patients presenting with either amplification or high expression of EGFR gene did not reveal any prognostic significance with a pooled HR of 1.31 (95% CI 0.96-1.79; p = 0.08). CONCLUSIONS: MGMT promoter methylation and IDH1 mutation are significantly associated with better OS in GBM patients. No significant associations were found between EGFR amplification or overexpression with OS

The genomic landscape of thyroid cancer tumourigenesis and implications for immunotherapy

Thyroid cancer is the most prevalent endocrine malignancy that comprises mostly indolent differentiated cancers (DTCs) and less frequently aggressive poorly differentiated (PDTC) or anaplastic cancers (ATCs) with high mortality. Utilisation of next-generation sequencing (NGS) and advanced sequencing data analysis can aid in understanding the multi-step progression model in the development of thyroid cancers and their metastatic potential at a molecular level, promoting a targeted approach to further research and development of targeted treatment options including immunotherapy, especially for the aggressive variants. Tumour initiation and progression in thyroid cancer occurs through constitutional activation of the mitogen-activated protein kinase (MAPK) pathway through mutations in BRAF, RAS, mutations in the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) pathway and/or receptor tyrosine kinase fusions/translocations, and other genetic aberrations acquired in a stepwise manner. This review provides a summary of the recent genetic aberrations implicated in the development and progression of thyroid cancer and implications for immunotherapy

Tumour immune microenvironment biomarkers predicting cytotoxic chemotherapy efficacy in colorectal cancer

The role of the local tumour and stromal immune landscape is increasingly recognised to be important in cancer development, progression and response to therapy. The composition, function, spatial orientation and gene expression profile of the infiltrate of the innate and adaptive immune system at the tumour and surrounding tissue has an established prognostic role in colorectal cancer (CRC). Multiple studies have confirmed that a tumour immune microenvironment (TIME) reflective of a type 1 adaptive immune response is associated with improved prognosis. There have been significant efforts to evolve these observations into validated, histopathology-based prognostic biomarkers, such as the Immunoscore. However, the clinical need lies much more in the development of predictive, not prognostic, biomarkers which have the potential to improve patient outcomes. This is particularly pertinent to help guide cytotoxic chemotherapy use in CRC, which remains the standard of care. Cytotoxic chemotherapy has recognised immunomodulatory activity distinct from its antimitotic effects, including mechanisms such as immunogenic cell death (ICD) and induction/inhibition of key immune players. Response to chemotherapy may differ with regard to molecular subtype of CRC, which are strongly associated with immune phenotypes. Thus, immune markers are potentially useful, though under-reported, predictive biomarkers. In this review, we discuss the impact of the TIME on response to cytotoxic chemotherapy in CRC, with a focus on baseline immune markers, and associated genomic and transcriptomic signatures

A metatranscriptomic approach to explore longitudinal tissue specimens from non-healing diabetes related foot ulcers

Cellular mechanisms and/or microbiological interactions which contribute to chronic diabetes related foot ulcers (DRFUs) were explored using serially collected tissue specimens from chronic DRFUs and control healthy foot skin. Total RNA was isolated for next-generation sequencing. We found differentially expressed genes (DEGs) and enriched hallmark gene ontology biological processes upregulated in chronic DRFUs which primarily functioned in the host immune response including: (i) Inflammatory response; (ii) TNF signalling via NFKB; (iii) IL6 JAK-STAT3 signalling; (iv) IL2 STAT5 signalling and (v) Reactive oxygen species. A temporal analysis identified RN7SL1 signal recognition protein and IGHG4 immunoglobulin protein coding genes as being the most upregulated genes after the onset of treatment. Testing relative temporal changes between healing and non-healing DRFUs identified progressive upregulation in healed wounds of CXCR5 and MS4A1 (CD20), both canonical markers of lymphocytes (follicular B cells/follicular T helper cells and B cells, respectively). Collectively, our RNA-seq data provides insights into chronic DRFU pathogenesis

Droplet digital PCR based detection of EGFR mutations in advanced lung cancer patient liquid biopsies : a comparison of circulating tumour DNA extraction kits

Background: Mutations in the epidermal growth factor receptor gene, EGFR, predict response or resistance to first generation tyrosine kinase inhibitors in non-small cell lung cancer. These biomarkers can now be conveniently detected from liquid biopsies, however technical details of these assays are still being refined. Objective: To compare detection of four different non-small cell lung cancer (NSCLC) associated EGFR mutations from patient ctDNA isolated with five different ctDNA isolation kit. Methods: Droplet digital PCR (ddPCR) assays detecting four EGFR mutations were developed. ctDNA was isolated with five kits from plasma samples, one pleural and one ascites fluid from nine NSCLC patients with known EGFR mutations. ctDNA fragment sizes and concentrations were also assessed. Results: Each kit isolated DNA from all samples which contained an expected dominant DNA fragment of ~ 170 base pairs. Normalised for plasma input, one kit produced ctDNA extracts which consistently enabled the highest cop n umber detection for all EGFR variants, and importantly was able to validate mutations in all patient samples. Other kits stood out in regards to cost economy as well as ease and speed of processing but were less efficient and one kit was found to be incompatible with ddPCR. Conclusion: This study demonstrated successful ctDNA isolation from plasma, pleural fluid and ascites by four of five ctDNA isolation kits. The QIAmp circulating nucleic acid kit produced consistently the most sensitive detection of EGFR variants. While other kits allow for lower volume plasma input down to 0.1 ml, are faster, more economical and simpler to use, they are challenged by very low ctDNA concentrations in plasma

Plasma next generation sequencing and droplet digital PCR-based detection of epidermal growth factor receptor (EGFR) mutations in patients with advanced lung cancer treated with subsequent-line osimertinib

Background: Gene mutation analysis from plasma circulating tumor DNA (ctDNA) can provide timely information regarding the mechanism of resistance that could translate to personalised treatment. We compared concordance rate of next generation sequencing (NGS) and droplet digital polymerase chain reaction (ddPCR) in the detection of the EGFR activating and T790M mutation from plasma ctDNA with diagnostic tissue biopsy‐based assays. The second objective was to test whether putative osimertinib resistance associated mutations were detectable from plasma using NGS. Methods: From January 2016 to December 2017, we prospectively collected plasma samples from patients prior to commencement of second‐ or third‐line osimertinib therapy and upon disease progression, in a single tertiary hospital in South Western Sydney, Australia. Amplicon‐based NGS and ddPCR assays were used to detect activating epidermal growth factor receptor (EGFR) and T790M mutations in 18 plasma samples from nine patients; all patients were required to have tissue biopsies with known EGFR status. Results: High concordance of allelic fractions were seen in matched plasma NGS and ddPCR for activating EGFR mutations and T790M mutations (R2 = 0.92, P < 0.0001). Using tissue biopsies as reference standard, sensitivity was 100% for NGS and 94% for ddPCR. Several possible osimertinib resistance associated mutations, including PIK3CA, BRAF and TP53 mutations, were detected by NGS in samples upon progression on osimertinib therapy. Conclusion: ddPCR assays for EGFR mutations appear to be as sensitive and highly concordant as amplicon‐based NGS. NGS has the ability to detect novel resistance mutations

Harnessing liquid biopsies to guide immune checkpoint inhibitor therapy

Immunotherapy (IO), involving the use of immune checkpoint inhibition, achieves improved response-rates and significant disease-free survival for some cancer patients. Despite these beneficial effects, there is poor predictability of response and substantial rates of innate or acquired resistance, resulting in heterogeneous responses among patients. In addition, patients can develop life-threatening adverse events, and while these generally occur in patients that also show a tumor response, these outcomes are not always congruent. Therefore, predicting a response to IO is of paramount importance. Traditionally, tumor tissue analysis has been used for this purpose. However, minimally invasive liquid biopsies that monitor changes in blood or other bodily fluid markers are emerging as a promising cost-effective alternative. Traditional biomarkers have limitations mainly due to difficulty in repeatedly obtaining tumor tissue confounded also by the spatial and temporal heterogeneity of tumours. Liquid biopsy has the potential to circumvent tumor heterogeneity and to help identifying patients who may respond to IO, to monitor the treatment dynamically, as well as to unravel the mechanisms of relapse. We present here a review of the current status of molecular markers for the prediction and monitoring of IO response, focusing on the detection of these markers in liquid biopsies. With the emerging improvements in the field of liquid biopsy, this approach has the capacity to identify IO-eligible patients and provide clinically relevant information to assist with their ongoing disease management