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To investigate the efficacy of psychological interventions delivered to mothers of young children with autism spectrum disorder (ASD) in improving maternal mental health and the parent-child relationship.Electronic databases were searched from inception to December 2021. Studies were included if they were randomized controlled trials (RCTs) of psychological interventions that targeted maternal mental health (primary outcome) and/or the parent-child relationship (secondary outcome), delivered to mothers of children (mean age 60 months) with ASD. Meta-analyses of three parent mental health outcomes (stress, depressive symptomatology, and general mental health) and seven parent-child relationship outcomes (parent responsiveness, affect, directiveness, non-directiveness, child initiation, infant positive affect, and infant attentiveness) were conducted using both mean and standardized mean differences.Thirty-two RCTs met the inclusion criteria (2336 participants). Six RCTs showed improved maternal mental health (Cohen's d = 0.41-1.15), with moderate certainty of evidence for improvement in parental stress. There was low and high certainty of evidence of treatment effect on parental depressive symptoms and general mental health respectively, which did not show significant treatment effects post-intervention. Seventeen RCTs showed improved parent-child relationship (d = 0.33-2.28; low certainty of evidence).Results demonstrated that parenting interventions promoting responsiveness had the largest positive effect on the parent-child relationship and a moderate effect on overall parenting stress; however, the studies were heterogeneous, making it difficult to identify the intervention components that were responsible for the positive treatment effects. There was no significant treatment effect for studies targeting depression or general mental health, likely due to the lack of an interventional component directly targeting mental health.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is known to present with pulmonary and extra-pulmonary organ complications. In comparison with the 2009 pandemic (pH1N1), SARS-CoV-2 infection is likely to lead to more severe disease, with multi-organ effects, including cardiovascular disease. SARS-CoV-2 has been associated with acute and long-term cardiovascular disease, but the molecular changes govern this remain unknown.In this study, we investigated the landscape of cardiac tissues collected at rapid autopsy from SARS-CoV-2, pH1N1, and control patients using targeted spatial transcriptomics approaches. Although SARS-CoV-2 was not detected in cardiac tissue, host transcriptomics showed upregulation of genes associated with DNA damage and repair, heat shock, and M1-like macrophage infiltration in the cardiac tissues of COVID-19 patients. The DNA damage present in the SARS-CoV-2 patient samples, were further confirmed by γ−H2Ax immunohistochemistry. In comparison, pH1N1 showed upregulation of Interferon-stimulated genes (ISGs), in particular interferon and complement pathways, when compared with COVID-19 patients.These data demonstrate the emergence of distinct transcriptomic profiles in cardiac tissues of SARS-CoV-2 and pH1N1 influenza infection supporting the need for a greater understanding of the effects on extra-pulmonary organs, including the cardiovascular system of COVID-19 patients, to delineate the immunopathobiology of SARS-CoV-2 infection, and long term impact on health.

Multimodal analysis of circulating tumour cells (CTCs) has the potential to provide remarkable insight for cancer development and metastasis. CTCs and CTC clusters investigation using single-cell analysis, enables researchers to gain crucial information on metastatic mechanisms and the genomic alterations responsible for drug resistance, empowering treatment, and management of cancer. Despite a plethora of CTC isolation technologies, careful attention to the strengths and weaknesses of each method should be considered in order to isolate these rare cells. Here, we provide an overview of cutting-edge technologies used for single-cell isolation and analysis of CTCs. Additionally, we highlight the biological features, clinical application, and the therapeutic potential of CTCs and CTC clusters using single-cell analysis platforms for cancer management.

The discovery of immune checkpoint proteins such as PD-1/PDL-1 and CTLA-4 represents a significant breakthrough in the field of cancer immunotherapy. Therefore, humanized monoclonal antibodies, targeting these immune checkpoint proteins have been utilized successfully in patients with metastatic melanoma, renal cell carcinoma, head and neck cancers and non-small lung cancer. The US FDA has successfully approved three different categories of immune checkpoint inhibitors (ICIs) such as PD-1 inhibitors (Nivolumab, Pembrolizumab, and Cemiplimab), PDL-1 inhibitors (Atezolimumab, Durvalumab and Avelumab), and CTLA-4 inhibitor (Ipilimumab). Unfortunately, not all patients respond favourably to these drugs, highlighting the role of biomarkers such as Tumour mutation burden (TMB), PDL-1 expression, microbiome, hypoxia, interferon-γ, and ECM in predicting responses to ICIs-based immunotherapy. The current study aims to review the literature and updates on ICIs in cancer therapy.

To gain a better understanding of the complexity of gene expression in normal and diseased tissues it is important to account for the spatial context and identity of cellin situ. State-of-the-art spatial profiling technologies, such as the Nanostring GeoMx Digital Spatial Profiler (DSP), now allow quantitative spatially resolved measurement of the transcriptome in tissues. However, the bioinformatics pipelines currently used to analyse GeoMx data often fail to successfully account for the technical variability within the data and the complexity of experimental designs, thus limiting the accuracy and reliability of subsequent analysis. Carefully designed quality control workflows, that include in-depth experiment-specific investigations into technical variation and appropriate adjustment for such variation can address this issue. Here we presentstandR, a R/Bioconductor package that enables an end-to-end analysis of GeoMx DSP data. With four case studies from previously published experiments, we demonstrate how the standR workflow can enhance the statistical power of GeoMx DSP data analysis and how application of standR enables scientists to develop in-depth insights into the biology of interest.

Mucosal head and neck squamous cell carcinoma (HNSCC) are the seventh most common cancer, with approximately 50% of patients living beyond 5 years. Immune checkpoint inhibitors (ICIs) have shown promising results in patients with recurrent or metastatic (R/M) disease, however, only a subset of patients benefit from immunotherapy. Studies have implicated the tumor microenvironment (TME) of HNSCC as a major factor in therapy response, highlighting the need to better understand the TME, particularly by spatially resolved means to determine cellular and molecular components. Here, we employed targeted spatial profiling of proteins on a cohort of pre-treatment tissues from patients with R/M disease to identify novel biomarkers of response within the tumor and stromal margins. By grouping patient outcome categories into response or non-response, based on Response Evaluation Criteria in Solid Tumors (RECIST) we show that immune checkpoint molecules, including PD-L1, B7-H3, and VISTA, were differentially expressed. Patient responders possessed significantly higher tumor expression of PD-L1 and B7-H3, but lower expression of VISTA. Analysis of response subgroups indicated that tumor necrosis factor receptor (TNFR) superfamily members including OX40L, CD27, 4-1BB, CD40, and CD95/Fas, were associated with immunotherapy outcome. CD40 expression was higher in patient-responders than non responders, while CD95/Fas expression was lower in patients with partial response (PR) relative to those with stable disease (SD) and progressive disease (PD). Furthermore, we found that high 4-1BB expression in the tumor compartment, but not in the stroma, was associated with better overall survival (OS) (HR= 0.28, p-adjusted= 0.040). Moreover, high CD40 expression in tumor regions (HR= 0.27, p-adjusted= 0.035), and high CD27 expression in the stroma (HR= 0.2, p-adjusted=0.032) were associated with better survival outcomes. Taken together, this study supports the role of immune checkpoint molecules and implicates the TNFR superfamily as key players in immunotherapy response in our cohort of HNSCC. Validation of these findings in a prospective study is required to determine the robustness of these tissue signatures.

In recent years, pregnant people infected with the SARS-CoV-2 virus have shown a higher incidence of “preeclampsia-like syndrome”. Preeclampsia is a systematic syndrome that affects 5-8 % of pregnant people worldwide and is the leading cause of maternal mortality and morbidity. It is unclear what causes preeclampsia, and is characterised by placental dysfunction, leading to poor placental perfusion, maternal hypertension, proteinuria, thrombocytopenia, or neurological disturbances.In this study, we used whole-transcriptome, digital spatial profiling of placental tissues to analyse the expression of genes at the cellular level between placentae from pregnant participants who contracted SARS-CoV-2 in the third trimester of their pregnancy and those prior to the start of the pandemic. Our focused analysis of the trophoblast and villous core stromal cell populations revealed tissue-specific pathways enriched in the SARS-CoV-2 placentae that align with a pre-eclampsia signature. Most notably, we found enrichment of pathways involved in vascular tension, blood pressure, inflammation, and oxidative stress.This study illustrates how spatially resolved transcriptomic analysis of placental tissue can aid in understanding the underlying pathogenic mechanisms of SARS-CoV-2 in pregnancy that are thought to induce “preeclampsia-like syndrome”. Moreover, our study highlights the benefits of using digital spatial profiling to map the crosstalk between trophoblast and villous core stromal cells linked to pathways involved in “preeclampsia-like syndrome” presenting in pregnant people with SARS-CoV-2.

PurposeRobust biomarkers that predict disease outcomes amongst COVID-19 patients are necessary for both patient triage and resource prioritisation. Numerous candidate biomarkers have been proposed for COVID-19. However, at present, there is no consensus on the best diagnostic approach to predict outcomes in infected patients. Moreover, it is not clear whether such tools would apply to other potentially pandemic pathogens and therefore of use as stockpile for future pandemic preparedness.MethodsWe conducted a multi-cohort observational study to investigate the biology and the prognostic role of interferon alpha-inducible protein 27 (IFI27) in COVID-19 patients.ResultsWe show that IFI27 is expressed in the respiratory tract of COVID-19 patients and elevated IFI27 expression in the lower respiratory tract is associated with the presence of a high viral load. We further demonstrate that the systemic host response, as measured by blood IFI27 expression, is associated with COVID-19 infection. For clinical outcome prediction (e.g., respiratory failure), IFI27 expression displays a high sensitivity (0.95) and specificity (0.83), outperforming other known predictors of COVID-19 outcomes. Furthermore, IFI27 is upregulated in the blood of infected patients in response to other respiratory viruses. For example, in the pandemic H1N1/09 influenza virus infection, IFI27-like genes were highly upregulated in the blood samples of severely infected patients.ConclusionThese data suggest that prognostic biomarkers targeting the family of IFI27 genes could potentially supplement conventional diagnostic tools in future virus pandemics, independent of whether such pandemics are caused by a coronavirus, an influenza virus or another as yet-to-be discovered respiratory virus.

Objectives: Attaining tumour material from lung cancer patients can be challenging with limited sample availability. Therefore, non-invasive means of assessing tumour material is becoming increasingly more important. Circulating tumour DNA (ctDNA), extracted from a blood sample is appealing for the patient, and can be performed serially over the course of treatment. Materials and Methods: Here, we describe an approach for profiling the blood samples of 103 NSCLC patients for 73 variants in ctDNA across a panel of actionable lung cancer mutations using the UltraSEEK lung Panel (Agena Biosciences). Results: Our cross-sectional study showed tumour and blood concordance in the detection of KRAS mutations (G12C, G12D, G12A/V, G12R, G12RC, Q61H) in 17/27 (63%), EGFR mutations (e746_a750del, e747_A750, T790M, L861Q) in 16/20 (80%) with additional PIK3CA_p545K mutations across both cohorts. In patients without reported tumour mutations, 11/56 (19.6%) presented with plasma mutations across EGFR, KRAS and PIK3CA. Where ctDNA mutations were measured longitudinally (n = 4 patients), the individual mutations mirrored the response to therapy/progression of disease. Conclusion: Whilst preliminary, this study demonstrates the utility of detecting clinically actionable mutations in the blood samples of NSCLC patients at the time of presentation, and over the course of therapy.

The purpose of this study is to investigate the preventive strategies to reduce musculoskeletal disorders of nursing personnel. The findings of this study stated that there are three key themes of strategies to handle the research puzzle. Physical facilities such as lifts, equipment, electric beds, footwear, and other equipment provide physical infrastructure to deal with the issue. Moreover, the findings revealed that the physical facilities alone may not be effective in the long run. The second theme focuses on guidelines, procedures, or principles such as education, staffing, no lift policy, a healthy lifestyle, the culture of safety, training on manual handling, workflow management, need analysis, and stories which provide the supportive structure in addressing the issue. We argue that physical facilities, procedures, guidelines, and principles alone may not work effectively in reducing the musculoskeletal disorders of nurses and there should be a simultaneous implementation of themes one and two. We introduce the Musculoskeletal Prevention Model to deal with the research issue.

Effective and consistent engagement in personal hygiene practices is important for preventing the acquisition and transmission of communicable diseases in childhood. This study aimed to investigate trends in adherence to recommended hygiene practices and identify factors contributing to difficulties in establishing good hygiene habits with young children (0–4 years) in Australia. A self-selected community sample of parents (N = 426) completed an online survey assessing children's adherence to recommended hygiene practices and barriers and enablers of establishing good personal hygiene habits. Parents reported interest in receiving information/tips on children's personal hygiene (yes/no) and nominated topics of interest. Less than half of children in any age group consistently (always/almost always) covered coughs and sneezes, washed hands after toileting, or washed hands before meals or when dirty. Children's non-compliance (i.e. resistance, refusal) was the most commonly reported barrier to establishing good personal hygiene habits, while children's compliance (i.e. cooperative behaviour, following or complying with caregivers’ instructions) was the most commonly cited enabler. Despite low levels of adherence, less than half (41.2%) of parents wanted information/tips on children's hygiene. Results suggest a disconnect between parents’ knowledge about recommended hygiene practices and actual behaviour. Development and testing of approaches to behaviour change that incorporate evidence-based strategies to manage children's resistance and support parents to encourage the development of healthy hygiene habits is warranted.

The ability to study cancer-immune cell communication across the whole tumor section without tissue dissociation is needed, especially for cancer immunotherapy development, which requires understanding of molecular mechanisms and discovery of more druggable targets. In this work, we assembled and evaluated an integrated experimental framework and analytical process to enable genome-wide scale discovery of ligand-receptors potentially used for cellular crosstalks, followed by targeted validation. We assessed the complementarity of four different technologies: single-cell RNA sequencing and Spatial transcriptomic (measuring over >20,000 genes), RNA In Situ Hybridization (RNAscope, measuring 4-12 genes) and Opal Polaris multiplex protein staining (4-9 proteins). To utilize the multimodal data, we implemented existing methods and also developed STRISH (Spatial TRanscriptomic In Situ Hybridization), a computational method that can automatically scan across the whole tissue section for local expression of gene (e.g. RNAscope data) and/or protein markers (e.g. Polaris data) to recapitulate an interaction landscape across the whole tissue. We evaluated the approach to discover and validate cell-cell interaction in situ through in-depth analysis of two types of cancer, basal cell carcinoma and squamous cell carcinoma, which account for over 70% of cancer cases. We showed that inference of cell-cell interactions using scRNA-seq data can misdetect or detect false positive interactions. Spatial transcriptomics still suffers from misdetecting lowly expressed ligand-receptor interactions, but reduces false discovery. RNAscope and Polaris are sensitive methods for defining the location of potential ligand receptor interactions, and the STRISH program can determine the probability that local gene co-expression reflects true cell-cell interaction. We expect that the approach described here will be widely applied to discover and validate ligand receptor interaction in different types of solid cancer tumors.

Non-small cell lung cancer (NSCLC) is one of the most common types of cancer in the world and has a 5-year survival rate of ~20%. Immunotherapies have shown promising results leading to durable responses, however, they are only effective for a subset of patients. To determine the best therapeutic approach, a thorough and in-depth profiling of the tumour microenvironment (TME) is required. The TME is a complex network of cell types that form an interconnected network, promoting tumour cell initiation, growth and dissemination. The stroma, immune cells and endothelial cells that comprise the TME generate a plethora of cytotoxic or cytoprotective signalling pathways. In this review, we discuss immunotherapeutic targets in NSCLC tumours and how the TME may influence patients' response to immunotherapy.

Over the last few years, immunotherapy, in particular, immune checkpoint inhibitor therapy, has revolutionized the treatment of several types of cancer. At the same time, the uptake in clinical oncology has been slow owing to the high cost of treatment, associated toxicity profiles and variability of the response to treatment between patients. In response, personalized approaches based on predictive biomarkers have emerged as new tools for patient stratification to achieve effective immunotherapy. Recently, the enumeration and molecular analysis of circulating tumor cells (CTCs) have been highlighted as prognostic biomarkers for the management of cancer patients during chemotherapy and for targeted therapy in a personalized manner. The expression of immune checkpoints on CTCs has been reported in a number of solid tumor types and has provided new insight into cancer immunotherapy management. In this review, we discuss recent advances in the identification of immune checkpoints using CTCs and shed light on the potential applications of CTCs towards the identification of predictive biomarkers for immunotherapy.

Advances in immunotherapy have led to durable and long‐term benefits in a subset of patients across a number of solid tumor types. Understanding of the subsets of patients that respond to immune checkpoint inhibitors at the cellular level, and in the context of their tumor microenvironment (TME) is becoming increasingly important. The TME is composed of a heterogeneous milieu of tumor and immune cells. The immune landscape of the TME can inhibit or promote tumor initiation and progression; thus, a deeper understanding of tumor immunity is necessary to develop immunotherapeutic strategies. Recent developments have focused on characterizing the TME immune contexture (type, density, and function) to discover mechanisms and biomarkers that may predict treatment outcomes. This has, in part, been powered by advancements in spatial characterization technologies. In this review article, we address the role of specific immune cells within the TME at various stages of tumor progression and how the immune contexture determinants affecting tumor growth are used therapeutically.

Anthracnose caused by Colletotrichum sp. and stem-end rot (SER) caused predominantly by Lasiodiplodia sp. are common postharvest diseases affecting papaya in Sri Lanka and are currently managed by synthetic chemicals which are hazardous. Use of essential oils (EOs) is considered a non-hazardous alternative. This in vitro study investigated the antifungal potential of selected EOs against anthracnose and SER pathogens isolated from papaya var. ‘Red Lady’.A poisoned food bioassay was carried out to evaluate the antifungal effect of EOs (Cardamom, citronella, orange, mustard and lemon). Colony diameter was taken as a measure of growth. Bioassays consisted of EOs at 500, 750, 1000 μl l-1 with untreated control, three replicates and three separate trials. Cardamom (1000 μl l-1) and citronella (750 μll-1) significantly (PColletotrichum sp. and Lasiodiplodia sp. from papaya. According to GC-MS, higher percentages of antifungal geraniol, geranyl acetate are present in citronella while a-terpinyl acetate, 1, 8-cineole are present in cardamom oil.

Introduction: Metastasis results in more than 90% of cancer-related deaths globally. The process is thought to be facilitated by metastatic precursor cells, commonly termed circulating tumor cells (CTCs). CTCs can exist as single cells or cell clusters and travel through the lymphovasculature to distant organs where they can form overt metastasis. Areas covered: Studies have highlighted that CTC clusters, which may be homotypic or heterotypic in composition, have a higher metastatic potential compared to single CTCs. The characterization of CTC clusters is becoming important as heterotypic clusters can provide a mechanism for immune evasion. This review summarizes the latest advances in CTC cluster-mediated metastasis and clinical significance. Expert opinion: Comprehensive characterization of CTC clusters is needed to understand the cell types and interactions within clusters, in order to identify ways in which to reduce CTC cluster-mediated metastasis. The role of CTC clusters in prognosticating disease progression needs to be determined by documenting CTC clusters from the time of diagnosis over the course of therapy.

BackgroundTumour tissue-based information is limited. Liquid biopsy can provide valuable real-time information through circulating tumour cells (CTCs). Profiling and expanding CTCs may provide avenues to study transient metastatic disease.MethodsSeventy non-small cell lung cancer (NSCLC) patients were recruited. CTCs were enriched using the spiral microfluidic chip and a RosetteSep™ using bloods from NSCLC patients. CTC cultures were carried out using the Clevers media under hypoxic conditions. CTCs were characterized using immunofluorescence and mutation-specific antibodies for samples with known mutation profiles. Exome sequencing was used to characterized CTC cultures.ResultsCTCs (>2 cells) were detected in 38/70 (54.3%) of patients ranging from 0 to 385 CTCs per 7.5 mL blood. In 4/5 patients where primary tumours harboured an EGFR exon 19 deletion, this EGFR mutation was also captured in CTCs. ALK translocation was confirmed on CTCs from a patient harbouring an ALK-rearrangement in the primary tumour. Short term CTC cultures were successfully generated in 9/70 NSCLC patients. Whole exome sequencing (WES) confirmed the presence of somatic mutations in the CTC cultures with mutational signatures consistent with NSCLC.ConclusionsWe were able to detect CTCs in >50% of NSCLC patients. NSCLC patients with >2 CTCs had a poor prognosis. The short-term CTC culture success rate was 12.9%. Further optimization of this culture methodology may provide a means by which to expand CTCs derived from NSCLC patient's bloods. CTC cultures allow for expansion of cells to a critical mass, allowing for functional characterization of CTCs with the goal of drug sensitivity testing and the creation of CTC cell lines.

Head and neck squamous cell carcinoma (HNSCC) often presents with locoregional or distant disease, despite multimodal therapeutic approaches, which include surgical resection, chemoradiotherapy, and more recently, immunotherapy for metastatic or recurrent HNSCC. Therapies often target the primary and nodal regional HNSCC sites, and their efficacy at controlling occult distant sites remains poor. While our understanding of the tumor microenvironment conducive to effective therapies is increasing, the biology underpinning locoregional sites remains unclear. Here, we applied targeted spatial proteomic approaches to primary and lymph node metastasis from an oropharyngeal SCC (OPSCC) cohort to understand the expression of proteins within tumors, and stromal compartments of the respective sites in samples of both matched and unmatched patients. In unmatched analyses of n = 43 primary and 11 nodal metastases, our data indicated that tumor cells in nodal metastases had higher levels of Ki-67, PARP, BAD, and cleaved caspase 9, suggesting a role for increased proliferation, DNA repair, and apoptosis within these metastatic cells. Conversely, in matched analyses (n = 7), pro-apoptotic markers BIM and BAD were enriched in the stroma of primary tumors. Univariate, overall survival (OS) analysis indicated CD25 in tumor regions of primary tumors to be associated with reduced survival (HR = 3.3, p = 0.003), while progesterone receptor (PR) was associated with an improved OS (HR = 0.33, p = 0.015). This study highlights the utility of spatial proteomics for delineating the tumor and stromal compartment composition, and utility toward understanding these properties in locoregional metastasis. These findings indicate unique biological properties of lymph node metastases that may elucidate further understanding of distant metastatic in OPSCC.

Non-small cell lung cancer (NSCLC) is one of the most common types of cancer in the world and has a 5-year survival rate of ~20%. Immunotherapies have shown promising results leading to durable responses, however, they are only effective for a subset of patients. To determine the best therapeutic approach, a thorough and in-depth profiling of the tumour microenvironment (TME) is required. The TME is a complex network of cell types that form an interconnected network, promoting tumour cell initiation, growth and dissemination. The stroma, immune cells and endothelial cells that comprise the TME generate a plethora of cytotoxic or cytoprotective signalling pathways. In this review, we discuss immunotherapeutic targets in NSCLC tumours and how the TME may influence patients' response to immunotherapy.

Immune checkpoint inhibitors (ICIs) Pembrolizumab and Nivolumab have shown promising results in patients with recurrent or metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). Nonetheless, only up to 20% of patients may benefit from them. Next generation predictive biomarkers of response to ICI therapies are currently needed. This retrospective study profiled pre-treatment tissue samples collected from R/M HNSCC patients (n=30) treated with Pembrolizumab or Nivolumab. Targeted spatial proteomics profiling was performed using NanoString’s GeoMx® Digital Spatial Profiler (GeoMx). To develop an informed region-of-interest (ROI) selection strategy for the GeoMx, we analyzed H&E images from serial sections and performed Oncotopix® Discovery software, which considered pathology inputs and machine learning to analyze H&E imagery and 4-channel GeoMx visualization inputs. We profiled 80 proteins simultaneously using the GeoMx across modules in cell death, immune activation, immune cell typing, immuno-oncology drug target, immune profiling, PI3K/AKT signaling, and pan-tumor markers.. Next, we performed differential expression (DE) analysis of the proteins localized in the tumor and stromal compartments against response to therapy parameters of complete, partial, stable and progressive disease (according to RECIST criteria). Four-channel GeoMx immunofluorescence images provided per-ROI cell counts (total and phenotyped based on 3 markers), and these data were used to aid in area and cell count normalization within the tumor and stromal compartments. We found that patients responsive to immunotherapy had higher expression levels of PD-L1, Bcl-2, BCLX, and BIM in the tumor, whereas VISTA, FOXP3, and CD66b were downregulated. In the stromal compartment, it was found that responders had higher expression levels of B7-H3, CD40, and SMA, but lower expression of PARP, S100B, and NY-ESO-1 in comparison to non-responders. In terms of best response analysis, patients with PR (n=5) had higher levels of PD-L1, ER-alpha, and CD68 expression, but lower levels of VISTA, CD27, and CD95/Fas in tumor regions than patients with PD (n=8). In the stroma, PD-L1, CD68, and HLA-DR were upregulated, while VISTA, BIM, and BAD were downregulated in patients with PR versus those with PD. We found that informed ROI selection strategy aided in defining key features in the tumor microenvironment for comparative analysis across samples, and for data normalization methods. Moreover, we found that immune checkpoints and proteins involved in cell death signaling play important roles in the immune responsive tumor microenvironment of HNSCC based on response to immunotherapy. Citation Format: Habib Sadeghirad, Ning Liu, James Monkman, Chin Wee Tan, Caroline Cooper, Jeni Caldera, Sarah Church, Fabian Schneider, James Robert Mansfield, Ken O'Byrne, Melissa Davis, Brett Hughes, Arutha Kulasinghe. Spatial profiling of the tumor microenvironment in head and neck cancer identifies immune checkpoints and proteins involved in cell death signaling biomarkers of immunotherapy response. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4626.

Background Lung cancers are the leading cause of cancer-related deaths with a 5-year-survival of only ~20%. Whilst immunotherapies have led to durable and prolonged survival, only a subset of patients remain responsive. Additional biomarkers are thus needed to better predict if patients will respond or develop resistance against immune checkpoint inhibitor (ICI) therapies. Spatial phenotyping of the tumor microenvironment (TME) is now recognized as a proxy for ICI therapy outcomes. Our study employs an end-to-end spatial biology strategy to survey non-small-cell lung cancer (NSCLC) tissues for new biomarkers that could guide immunotherapy treatments. Methods We phenotyped pre-treatment biopsies from non-small-cell lung cancer (NSCLC) patients treated with single-agent Nivolumab. We first performed 57-plex whole-slide Single Cell Spatial Phenotyping on the PhenoCycler®-Fusion platform. Next, we developed the spatial analysis of a larger NSCLC cohort using customizable PhenoCode Signature Panels (PSP) for high-throughput immune profile (CD3/CD8/CD20/CD68/PanCK + CD4 add-in) and immuno-contexture (CD8/CD68/PD-L1/FoxP3/PanCK + PD-1 add-in) imaging. The PSP panel content combines the barcode-based antibody chemistry from the PhenoCycler platform with the signal amplification of Opal chemistry from the PhenoImager platform. PSP panels were profiled across n=27 NSCLC biopsies and provided a wider breadth of tissue profiling. Results Our whole-slide single-cell spatial phenotyping analyses revealed high phenotypic diversity in the TME of patients responsive and resistant to ICI therapy. However, higher-throughput analyses of the same tissues using targeted PSP panels revealed no significant differences in quantities of immune cell lineages, including T-cells and macrophages. On the contrary, we discovered multiple quantifiable and statistically significant spatial signatures that appear to be predictive of treatment benefit, which confirms the potential biomarker value of spatial associations in patient tissues. Conclusions This study amounts to a uniquely comprehensive Single Cell Spatial Phenotyping analysis of pre-treatment NSCLC biopsies from a single-agent Nivolumab study. Our data catalogue the diversity in the immune microenvironment of NSCLC but highlighted that immune cell quantification is insufficient to stratify patient cohorts. Single-cell spatial phenotyping, on the other hand, promises to reveal new biomarkers that may aid in better stratification of patients in pre-treatment evaluations. Citation Format: Ning Ma, Aditya Pratapa, James Monkman, Ken O’Byrne, Oliver Braubach, Arutha Kulasinghe. The potential predictive role of spatial phenotyping in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6768.

Background: Immune checkpoint inhibitors (ICI) have proven to be game-changing treatments for mucosal head and neck squamous cell cancer (HNSCC). Emerging successes with anti-PD-1/PD-L1 therapy have led to durable responses and prolonged survival in both human papillomavirus-positive (HPV+) and negative (HPV-) patients. There is now a need for predictive biomarkers to guide patient selection for highly targeted ICI therapies as currently available diagnostic biomarkers have a limited value. The tumor microenvironment (TME) composition, contexture, and cellular architecture are now recognized as key to understanding immune responsive and resistant phenotypes. Here, we are using a spatial biology approach to explore the TME in metastatic/recurrent HNSCC tumors treated with Pembrolizumab/Nivolumab. Methods: In this study, we used single cell, multiomic spatial phenotyping utilizing the PhenoCycler-Fusion spatial biology platform to characterize the TME of HNSCC tumors from a cohort of n=40 patients. The discovery cohort consisted of patients who had complete vs. partial vs. stable vs. progressive responses to ICI therapy. We first analyzed tissues using an ultrahigh-plex antibody panel of >60 antibodies that label immune cell lineages, checkpoints, activation markers as well as tissue structure and the stroma. We then conducted whole-slide, single cell resolution RNA detection with complementary markers on serial sections from the same tissue blocks; the combination of these data allowed us to obtain multiomic spatial signatures that offered uniquely comprehensive insight into the TME of our tissue cohorts. Results: Our study identified highly resolved tissue immune contexture analysis and metabolic tissue signatures associated with resistance and sensitivity to immunotherapy. Most notably, multiomic profiling of HNSCC tumours provided deeper insights into ICI therapy resistance than the single omics based approaches alone. Conclusions: Our study demonstrates the power of unbiased, multiomic spatial phenotyping with whole-slide imaging to identify biomarkers associated with response to ICI therapy in HNSCC. Citation Format: HaYeun Ji, Niyati Jhaveri, Ning Ma, Bassem B. Cheikh, Aditya Pratapa, James Monkman, Ken O’Byrne, Brett Hughes, Arutha Kulasinghe, Oliver Braubach. Single cell, multiomic spatial phenotyping of immunotherapy responses in head and neck cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5647.

Background: Cutaneous squamous cell carcinoma (cSCC) is the second most common skin cancer and represents a major global health burden. Approximately 50% of cSCC patients develop primary resistance and 20% will develop secondary resistance to immune checkpoint inhibitors (ICI). There are limited biomarkers that reflect the tumor dynamics predictive of response to ICI therapy, and therefore we need to explore new biomarkers that can better understand the cSCC tumor microenvironment (TME). Methods: Our retrospective study profiled pre-treatment cutaneous skin cancer tissues from patients with immunotherapy sensitive and resistant diseases. In this exploratory study, we designed a high-dimensional > 50-plex antibody panel identifying cell lineages, activation states, and immune checkpoints. In addition, we deployed a combination of antibodies for a novel multiplexed metabolic readout, to enhance our in situ profiling beyond well-known TME targets. Whole-slide spatial phenotyping was conducted on the PhenoCycler-Fusion spatial biology platform. Highplex image analysis was performed using Phenoplex (Visiopharm), including tissue segmentation and cellular phenotyping. Here, we used a deep learning approach to discriminate tumor regions and margins based on their morphology. Spatial analyses including phenotypic composition, cell-to-cell localization, and cellular neighborhoods were performed and compared to clinicopathological findings and responses to ICI therapy. Results: We have profiled the TME of cSCC via ultrahigh-plex, single-cell spatial analyses of whole tissues. Our data have revealed unique cell phenotype compositions and metabolic states within the TME of different patient cohorts. Areas of metabolic activity were visualized and compartmentalized to determine spatial distributions of inherent sensitivity and resistance to ICI therapy. Additionally, a comparison of the phenotypic expression patterns between immunotherapy sensitive and resistant cohorts in the tumor, stromal and margin regions showed relevant differences in the immune content. Taken together, this high-dimensional imaging approach of the cSCC TME has revealed new tissue insights. Conclusion: There is a need to understand the contexture of the cSCC immune microenvironment. Here we identify distinct cellular phenotypic compositions and metabolic states in tissues from immune-competent and immunocompromised cSCC. We thereby confirm biological differences in tissue composition and cellular interactions between these tissues and provide putative new biomarkers for cSCC patient stratification. Citation Format: Niyati Jhaveri, Dmytro Klymyshyn, Bassem B. Cheikh, James Monkman, Dan Winkowski, James Mansfield, Subham Basu, Michael Prater, Nadine Nelson, Gabrielle Belz, Oliver Braubach, Arutha Kulasinghe. The development of a spatial metabolic map of immunotherapy sensitive and resistant cutaneous skin carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6774.

Despite efforts to improve earlier diagnosis of non-small cell lung cancer (NSCLC), most patients present with advanced stage disease, which is often associated with poor survival outcomes with only 15% surviving for 5 years from their diagnosis. Tumour tissue biopsy is the current mainstream for cancer diagnosis and prognosis in many parts of the world. However, due to tumour heterogeneity and accessibility issues, liquid biopsy is emerging as a game changer for both cancer diagnosis and prognosis. Liquid biopsy is the analysis of tumour-derived biomarkers in body fluids, which has remarkable advantages over the use of traditional tumour biopsy. Circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA) are two main derivatives of liquid biopsy. CTC enumeration and molecular analysis enable monitoring of cancer progression, recurrence, and treatment response earlier than traditional biopsy through a minimally invasive liquid biopsy approach. CTC-derived ex-vivo cultures are essential to understanding CTC biology and their role in metastasis, provide a means for personalized drug testing, and guide treatment selection. Just like CTCs, ctDNA provides opportunity for screening, monitoring, treatment evaluation, and disease surveillance. We present an updated review highlighting the prognostic and therapeutic significance of CTCs and ctDNA in NSCLC.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that has few effective treatment options due to its lack of targetable hormone receptors. Whilst the degree of tumour infiltrating lymphocytes (TILs) has been shown to associate with therapy response and prognosis, deeper characterization of the molecular diversity that may mediate chemotherapeutic response is lacking. Here we applied targeted proteomic analysis of both chemotherapy sensitive and resistant TNBC tissue samples by the Nanostring GeoMx Digital Spatial Platform (DSP). By quantifying 68 targets in the tumour and tumour microenvironment (TME) compartments and performing differential expression analysis between responsive and non-responsive tumours, we show that increased ER-alpha expression and decreased 4-1BB and MART1 within the stromal compartments is associated with adjuvant chemotherapy response. Similarly, higher expression of GZMA, STING and fibronectin and lower levels of CD80 were associated with response within tumour compartments. Univariate overall-survival (OS) analysis of stromal proteins supported these findings, with ER-alpha expression (HR=0.19, p=0.0012) associated with better OS while MART1 expression (HR=2.3, p=0.035) was indicative of poorer OS. Proteins within tumour compartments consistent with longer OS included PD-L1 (HR=0.53, p=0.023), FOXP3 (HR=0.5, p=0.026), GITR (HR=0.51, p=0.036), SMA (HR=0.59, p=0.043), while EPCAM (HR=1.7, p=0.045), and CD95 (HR=4.9, p=0.046) expression were associated with shorter OS. Our data provides early insights into the levels of these markers in the TNBC tumour microenvironment, and their association with chemotherapeutic response and patient survival.

The complexity of the cellular and acellular players within the tumor microenvironment (TME) allows for significant variation in TME constitution and role in anticancer treatment response. Spatial alterations in populations of tumor cells and adjacent non-malignant cells, including endothelial cells, fibroblasts and tissue-infiltrating immune cells, often have a major role in determining disease progression and treatment response in cancer. Many current standard systemic antineoplastic treatments target the cancer cells and could be further refined to directly target commonly dysregulated cell populations of the TME. Recent developments in immuno-oncology and bioengineering have created an attractive potential to model these complexities at the level of the individual patient. These developments, along with the increasing momentum in precision medicine research and application, have catalysed exciting new discoveries in understanding drug-TME interactions, target identification, and improved efficacy of therapies. While rapid progress has been made, there are still many challenges to overcome in the development of accurate

Head and neck squamous cell carcinoma (HNSCC) represents a heterogeneous group of tumors. While significant progress has been made using multimodal treatment, the 5-year survival remains at 50%. Developing effective therapies, such as immunotherapy, will likely lead to better treatment of primary and metastatic disease. However, not all HNSCC tumors respond to immune checkpoint blockade therapy. Understanding the complex cellular composition and interactions of the tumor microenvironment is likely to lead to new knowledge for effective therapies and treatment resistance. In this review, we discuss HNSCC characteristics, predictive biomarkers, factors influencing immunotherapy response, with a focus on the tumor microenvironment.

Despite efforts to improve earlier diagnosis of non-small cell lung cancer (NSCLC), most patients present with advanced stage disease, which is often associated with poor survival outcomes with only 15% surviving for 5 years from their diagnosis. Tumour tissue biopsy is the current mainstream for cancer diagnosis and prognosis in many parts of the world. However, due to tumour heterogeneity and accessibility issues, liquid biopsy is emerging as a game changer for both cancer diagnosis and prognosis. Liquid biopsy is the analysis of tumour-derived biomarkers in body fluids, which has remarkable advantages over the use of traditional tumour biopsy. Circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA) are two main derivatives of liquid biopsy. CTC enumeration and molecular analysis enable monitoring of cancer progression, recurrence, and treatment response earlier than traditional biopsy through a minimally invasive liquid biopsy approach. CTC-derived ex-vivo cultures are essential to understanding CTC biology and their role in metastasis, provide a means for personalized drug testing, and guide treatment selection. Just like CTCs, ctDNA provides opportunity for screening, monitoring, treatment evaluation, and disease surveillance. We present an updated review highlighting the prognostic and therapeutic significance of CTCs and ctDNA in NSCLC.

BackgroundRobust biomarkers that predict disease outcomes amongst COVID-19 patients are necessary for both patient triage and resource prioritisation. Numerous candidate biomarkers have been proposed for COVID-19. However, at present, there is no consensus on the best diagnostic approach to predict outcomes in infected patients. Moreover, it is not clear whether such tools would apply to other potentially pandemic pathogens and therefore of use as stockpile for future pandemic preparedness.MethodsWe conducted a multi-cohort observational study to investigate the biology and the prognostic role of interferon alpha-inducible protein 27 (IFI27) in COVID-19 patients.FindingsWe show that IFI27 is expressed in the respiratory tract of COVID-19 patients and elevated IFI27 expression is associated with the presence of a high viral load. We further demonstrate that systemic host response, as measured by blood IFI27 expression, is associated with COVID-19 severity. For clinical outcome prediction (e.g. respiratory failure), IFI27 expression displays a high positive (0.83) and negative (0.95) predictive value, outperforming all other known predictors of COVID-19 severity. Furthermore, IFI27 is upregulated in the blood of infected patients in response to other respiratory viruses. For example, in the pandemic H1N1/09 swine influenza virus infection, IFI27-like genes were highly upregulated in the blood samples of severely infected patients.InterpretationThese data suggest that prognostic biomarkers targeting the family of IFI27 genes could potentially supplement conventional diagnostic tools in future virus pandemics, independent of whether such pandemics are caused by a coronavirus, an influenza virus or another as yet-to-be discovered respiratory virus.Research in contextEvidence before this studyWe searched the scientific literature using PubMed to identify studies that used the IFI27 biomarker to predict outcomes in COVID-19 patients. We used the search terms “IFI27”, “COVID-19, “gene expression” and “outcome prediction”. We did not identify any study that investigated the role of IFI27 biomarker in outcome prediction. Although ten studies were identified using the general terms of “gene expression” and “COVID-19”, IFI27 was only mentioned in passing as one of the identified genes. All these studies addressed the broader question of the host response to COVID-19; none focused solely on using IFI27 to improve the risk stratification of infected patients in a pandemic.Added value of this studyHere, we present the findings of a multi-cohort study of the IFI27 biomarker in COVID-19 patients. Our findings show that the host response, as reflected by blood IFI27 gene expression, accurately predicts COVID-19 disease progression (positive and negative predictive values; 0.83 and 0.95, respectively), outperforming age, comorbidity, C-reactive protein and all other known risk factors. The strong association of IFI27 with disease severity occurs not only in SARS-CoV-2 infection, but also in other respiratory viruses with pandemic potential, such as the influenza virus. These findings suggest that host response biomarkers, such as IFI27, could help identify high-risk COVID-19 patients - those who are more likely to develop infection complications - and therefore may help improve patient triage in a pandemic.Implications of all the available evidenceThis is the first systemic study of the clinical role of IFI27 in the current COVID-19 pandemic and its possible future application in other respiratory virus pandemics. The findings not only could help improve the current management of COVID-19 patients but may also improve future pandemic preparedness.

Lung cancer is the leading cause of cancer morbidity and mortality worldwide and early diagnosis is crucial for the management and treatment of this disease. Non-invasive means of determining tumour information is an appealing diagnostic approach for lung cancers as often accessing and removing tumour tissue can be a limiting factor. In recent years, liquid biopsies have been developed to explore potential circulating tumour biomarkers which are considered reliable surrogates for understanding tumour biology in a non-invasive manner. Most common components assessed in liquid biopsy include circulating tumour cells (CTCs), cell-free DNA (cfDNA), circulating tumour DNA (ctDNA), microRNA and exosomes. This review explores the clinical use of circulating tumour biomarkers found in liquid biopsy for screening, early diagnosis and prognostication of lung cancer patients.

Risk stratification of COVID‐19 patients is essential for pandemic management. Changes in the cell fitness marker, hFwe‐Lose, can precede the host immune response to infection, potentially making such a biomarker an earlier triage tool. Here, we evaluate whether hFwe‐Lose gene expression can outperform conventional methods in predicting outcomes (e.g., death and hospitalization) in COVID‐19 patients. We performed a post‐mortem examination of infected lung tissue in deceased COVID‐19 patients to determine hFwe‐Lose’s biological role in acute lung injury. We then performed an observational study (n = 283) to evaluate whether hFwe‐Lose expression (in nasopharyngeal samples) could accurately predict hospitalization or death in COVID‐19 patients. In COVID‐19 patients with acute lung injury, hFwe‐Lose is highly expressed in the lower respiratory tract and is co‐localized to areas of cell death. In patients presenting in the early phase of COVID‐19 illness, hFwe‐Lose expression accurately predicts subsequent hospitalization or death with positive predictive values of 87.8–100% and a negative predictive value of 64.1–93.2%. hFwe‐Lose outperforms conventional inflammatory biomarkers and patient age and comorbidities, with an area under the receiver operating characteristic curve (AUROC) 0.93–0.97 in predicting hospitalization/death. Specifically, this is significantly higher than the prognostic value of combining biomarkers (serum ferritin, D‐dimer, C‐reactive protein, and neutrophil–lymphocyte ratio), patient age and comorbidities (AUROC of 0.67–0.92). The cell fitness marker, hFwe‐Lose, accurately predicts outcomes in COVID‐19 patients. This finding demonstrates how tissue fitness pathways dictate the response to infection and disease and their utility in managing the current COVID‐19 pandemic., A post‐mortem examination of COVID‐19 infected lung tissues and an observational study were performed in order to evaluate whether expression of cell fitness marker hFwe‐Lose in patient's nasopharyngeal swabs could predict hospitalization or death from COVID‐19.

Coronavirus disease 2019 (COVID-19) has resulted in a global pandemic, challenging both the medical and scientific community for the development of novel vaccines and a greater understanding of the effects of the SARS-CoV-2 virus. COVID-19 has been associated with a pronounced and out-of-control inflammatory response. Studies have sought to understand the effects of inflammatory response markers to prognosticate the disease. Herein, we aimed to review the evidence of 11 groups of systemic inflammatory markers for risk-stratifying patients and prognosticating outcomes related to COVID-19. Numerous studies have demonstrated the effectiveness of neutrophil to lymphocyte ratio (NLR) in prognosticating patient outcomes, including but not limited to severe disease, hospitalization, intensive care unit (ICU) admission, intubation, and death. A few markers outperformed NLR in predicting outcomes, including 1) systemic immune-inflammation index (SII), 2) prognostic nutritional index (PNI), 3) C-reactive protein (CRP) to albumin ratio (CAR) and high-sensitivity CAR (hsCAR), and 4) CRP to prealbumin ratio (CPAR) and high-sensitivity CPAR (hsCPAR). However, there are a limited number of studies comparing NLR with these markers, and such conclusions require larger validation studies. Overall, the evidence suggests that most of the studied markers are able to predict COVID-19 prognosis, however NLR seems to be the most robust marker.

In families of children with autism spectrum disorder (ASD) the mother-child relationship can be affected by many factors; however, the impacts of maternal broad autism phenotype (BAP) and attachment styles are unknown. This study investigated predictors of emotional availability in the mother-child relationship in families of children with ASD.Mothers (N = 231) of children with ASD living in Australia completed questionnaires assessing emotional availability in the parent-child relationship (mutual attunement, child involvement, affect quality) and mothers' BAP, mental health, adult attachment style and parenting experiences. Hierarchical multiple regressions and a mediation analysis were conducted.Parent-reported positive parenting experiences was the greatest predictor of mutual attunement, child involvement and affect quality in the parent-child relationship. Maternal stress and anxiety predicted poorer mutual attunement, stress and depressive symptoms predicted less child involvement and anxiety predicted worse affect quality. Negative parenting experiences, maternal anxiety and stress mediated the relationship between BAP and mutual attunement.Interventions combining mental health and parenting support for mothers of children with ASD to bolster emotional availability within the mother-child relationship may be warranted.

Pre‐existing cardiovascular disease (CVD) increases the morbidity and mortality of COVID‐19 and is strongly associated with poor disease outcomes. However, SARS‐CoV‐2 infection can also trigger de novo acute and chronic cardiovascular disease. Acute cardiac complications include arrhythmia, myocarditis and heart failure, which are significantly associated with higher in‐hospital mortality. The possible mechanisms by which SARS‐CoV‐2 causes this acute cardiac disease include direct damage caused by viral invasion of cardiomyocytes as well as indirect damage through systemic inflammation. The long‐term cardiac complications associated with COVID‐19 are incompletely characterised and thought to include hypertension, arrhythmia, coronary atherosclerosis and heart failure. Although some cardiac‐related symptoms can last over 6 months, the effect of these complications on long‐term patient health remains unclear. The risk factors associated with long‐term cardiovascular disease remain poorly defined. Determining which patients are most at‐risk of long‐term cardiovascular disease is vital so that targeted follow‐up and patient care can be provided. The aim of this review was to summarise the current evidence of the acute and long‐term cardiovascular consequences of SARS‐CoV‐2 infection and the mechanisms by which SARS‐CoV‐2 may cause cardiovascular disease., This review summarises the current evidence of the acute and long‐term cardiovascular consequences of SARS‐CoV‐2 infection and the mechanisms by which SARS‐CoV‐2 may cause cardiovascular disease.

IntroductionImmunotherapies, such as immune checkpoint inhibitors (ICI) have shown durable benefit in a subset of non-small cell lung cancer (NSCLC) patients. The mechanisms for this are not fully understood, however the composition and activation status of the cellular milieu contained within the tumour microenvironment (TME) is becomingly increasingly recognised as a driving factor in treatment-refractory disease.MethodsHere, we employed multiplex IHC (mIHC), and digital spatial profiling (DSP) to capture the targeted immune proteome and transcriptome of tumour and TME compartments of pre-treatment samples from a 2nd line NSCLC ICI-treated cohort (n=41 patients; n=25 responders, n=16 non-responders).ResultsWe demonstrate by mIHC that the interaction of CD68+ macrophages with PD1+, FoxP3+ cells is significantly enriched in ICI refractory tumours (p=0.012). Our study revealed that patients sensitive to ICI therapy expressed higher levels of IL2 receptor alpha (CD25, p=0.028) within the tumour compartments, which corresponded with the increased expression of IL2 mRNA (p=0.001) within their stroma, indicative of key conditions for ICI efficacy prior to treatment. IL2 mRNA levels within the stroma positively correlated with the expression of pro-apoptotic markers cleaved caspase 9 (p=2e-5) and BAD (p=5.5e-4) and negatively correlated with levels of memory T cells (CD45RO) (p=7e-4). Immuno-inhibitory markers CTLA-4 (p=0.021) and IDO-1 (p=0.023) were also supressed in ICI-responsive patients. Of note, tumour CD44 (p=0.02) was depleted in the response group and corresponded inversely with significantly higher stromal expression of its ligand SPP1 (osteopontin, p=0.008). Analysis of differentially expressed transcripts indicated the potential inhibition of stromal interferon-gamma (IFNγ) activity, as well as estrogen-receptor and Wnt-1 signalling activity within the tumour cells of ICI responsive patients. Cox survival analysis indicated tumour CD44 expression was associated with poorer prognosis (HR=1.61, p=0.01), consistent with its depletion in ICI sensitive patients. Similarly, stromal CTLA-4 (HR=1.78, p=0.003) and MDSC/M2 macrophage marker ARG1 (HR=2.37, p=0.01) were associated with poorer outcome while levels of apoptotic marker BAD (HR=0.5, p=0.01) appeared protective. Interestingly, stromal mRNA for E-selectin (HR=652, p=0.001), CCL17 (HR=70, p=0.006) and MTOR (HR=1065, p=0.008) were highly associated with poorer outcome, indicating pro-tumourigenic features in the tumour microenvironment that may facilitate ICI resistance.ConclusionsThrough multi-modal approaches, we have dissected the characteristics of NSCLC and provide evidence for the role of IL2 and stromal activation by osteopontin in the efficacy of current generations of ICI therapy. The enrichment of SPP1 in the stroma of ICI sensitive patients in our data is a novel finding, indicative of stromal activation that may aid immune cell survival and activity despite no clear association with increased levels of immune infiltrate.

Scientific collaboration has been a critical aspect of the development of all fields of science, particularly clinical medicine. It is well understood that myriads of benefits can be yielded by interdisciplinary and international collaboration. For instance, our rapidly growing knowledge on COVID-19 and vaccine development could not be attained without expanded collaborative activities. However, achieving fruitful results requires mastering specific tactics in collaborative efforts. These activities can enhance our knowledge, which ultimately benefits society. In addition to tackling the issue of the invisible border between different countries, institutes, and disciplines, the border between the scientific community and society needs to be addressed as well. International and transdisciplinary approaches can potentially be the best solution for bridging science and society. The Universal Scientific Education and Research Network (USERN) is a non-governmental, non-profit organization and network to promote professional, scientific research and education worldwide. The fifth annual congress of USERN was held in Tehran, Iran, in a hybrid manner on November 7-10, 2020, with key aims of bridging science to society and facilitating borderless science. Among speakers of the congress, a group of top scientists unanimously agreed on The USERN 2020 consensus, which is drafted with the goal of connecting society with scientific scholars and facilitating international and interdisciplinary scientific activities in all fields, including clinical medicine.

Non-small cell lung cancer (NSCLC) is one of the most common cancers globally and has a 5-year survival rate ~20%. Immunotherapies have demonstrated long-term and durable responses in NSCLC patients, although they appear to be effective in only a subset of patients. A more comprehensive understanding of the underlying tumour biology may contribute to identifying those patients likely to achieve optimal outcomes. Profiling the tumour microenvironment (TME) has shown to be beneficial in addressing fundamental tumour-immune cell interactions. Advances in multiplexing immunohistochemistry and molecular barcoding has led to recent advances in profiling genes and proteins in NSCLC. Here, we review the recent advancements in spatial profiling technologies for the analysis of NSCLC tissue samples to gain new insights and therapeutic options for NSCLC. The combination of spatial transcriptomics combined with advanced imaging is likely to lead to deep insights into NSCLC tissue biology, which can be a powerful tool to predict likelihood of response to therapy.