Development of a long term, ex vivo, patient-derived explant model of endometrial cancer.

Incidence of endometrial cancer (EC) is rising in the developed world. The current standard of care, hysterectomy, is often infeasible for younger patients and those with high body mass index. There are limited non-surgical treatment options and a lack of biologically relevant research models to investigate novel alternatives to surgery for EC. The aim of the present study was to develop a long-term, patient-derived explant (PDE) model of early-stage EC and demonstrate its use for investigating predictive biomarkers for a current non-surgical treatment option, the levonorgestrel intra-uterine system (LNG-IUS). Fresh tumour specimens were obtained from patients with early-stage endometrioid EC. Tumours were cut into explants, cultured on media-soaked gelatin sponges for up to 21 days and treated with LNG. Formalin-fixed, paraffin embedded (FFPE) blocks were generated for each explant after 21 days in culture. Tumour architecture and integrity were assessed by haematoxylin and eosin (H&E) and immunohistochemistry (IHC). IHC was additionally performed for the expression of five candidate biomarkers of LNG resistance. The developed ex vivo PDE model is capable of culturing explants from early-stage EC tumours long-term (21 Days). This model can complement existing models and may serve as a tool to validate results obtained in higher-throughput in vitro studies. Our study provides the foundation to validate the extent to which EC PDEs reflect patient response in future research.

Toku Oranga: the subjective wellbeing and psychological functioning of postgraduate and medical students in Otautahi Christchurch.

AIMS: Postgraduate and medical students are at risk of psychological distress and burnout, which can cause significant functional and occupational impairment. We aimed to report subjective wellbeing, psychological distress and burnout in postgraduate and medical students in Otautahi Christchurch, Aotearoa (New Zealand), and identify any associations between participant and course information and outcome measures including exposure to major earthquakes in 2010/2011 and the 2019 terrorist attack. METHODS: A self-report online survey was completed by 140 students between November 2019 and March 2020. Life satisfaction, psychological distress and burnout were primary outcomes. Data were analysed using univariate and multivariable analysis. RESULTS: High levels of psychological distress were present in both student groups. Burnout was reported by 78% of respondents. There were no significant associations found between exposure to the Christchurch earthquakes or terrorist attack with primary outcomes. Personality factors, resilience and perceived support and success were weakly associated with wellbeing, distress and burnout. CONCLUSIONS: Postgraduates and medical students reported high levels of psychological distress and burnout. The earthquakes and terrorist attack do not appear to be associated with negative effects in these cohorts. Personality and resilience characteristics may assist in predicting students at risk of morbidity and evaluating potentially relevant interventions.

Potential drug repurposing of ruxolitinib to inhibit the JAK/STAT pathway for the treatment of patients with epithelial ovarian cancer.

AIM: This review aimed to describe the potential for therapeutic targeting of the JAK/STAT signaling pathway by repurposing the clinically-approved JAK inhibitor ruxolitinib in the patients with epithelial ovarian cancer (OC) setting. METHODS: We reviewed publications that focus on the inhibition of the JAK/STAT pathway in hematological and solid malignancies including OC. RESULTS: Preclinical studies showed that ruxolitinib effectively reduces OC cell viability and metastasis and enhances the anti-tumor activity of chemotherapy drugs. There are a number of recent clinical trials exploring the role of JAK/STAT inhibition in solid cancers including OC. Early results have not adequately supported efficacy in solid tumors. However, there are preclinical data and clinical studies supporting the use of ruxolitinib in combination with both chemotherapy and other targeted drugs in OC setting. CONCLUSION: Inflammatory conditions and persistent activation of the JAK/STAT pathway are associated with tumourigenesis and chemoresistance, and therapeutic blockade of this pathway shows promising results. For women with OC, clinical investigation exploring the role of ruxolitinib in combination with chemotherapy agents or other targeted therapeutics is warranted.

Digital life as a cabaret, old chum: A dramaturgical analysis of older digitalised home residents and their wider caring networks.

The use of smart and assistive devices for remote healthcare monitoring is becoming increasingly popular for older people in their homes. However, the lived and long-term experiences of such technology, for the older residents and their wider caring networks remains unclear. Using in-depth qualitative data collected between June 2019 and January 2020 from older people living in their own homes in rural Scotland, we highlight that although such monitoring could improve the experiences of older people and their wider caring networks, this may create additional care and surveillance. We employ the concept of dramaturgy, which understands society to be a stage on which actors perform, allowing us to explore how different residents and their networks make sense of their experiences with domestic healthcare monitoring. We found that some digitalised devices may reduce the degree to which older people and their wider caring networks can live authentic and truly independent lifestyles.

The Contribution of Environmental Science to Mental Health Research: A Scoping Review.

Mental health is influenced by multiple complex and interacting genetic, psychological, social, and environmental factors. As such, developing state-of-the-art mental health knowledge requires collaboration across academic disciplines, including environmental science. To assess the current contribution of environmental science to this field, a scoping review of the literature on environmental influences on mental health (including conditions of cognitive development and decline) was conducted. The review protocol was developed in consultation with experts working across mental health and environmental science. The scoping review included 202 English-language papers, published between 2010 and 2020 (prior to the COVID-19 pandemic), on environmental themes that had not already been the subject of recent systematic reviews; 26 reviews on climate change, flooding, air pollution, and urban green space were additionally considered. Studies largely focused on populations in the USA, China, or Europe and involved limited environmental science input. Environmental science research methods are primarily focused on quantitative approaches utilising secondary datasets or field data. Mental health measurement was dominated by the use of self-report psychometric scales. Measures of environmental states or exposures were often lacking in specificity (e.g., limited to the presence or absence of an environmental state). Based on the scoping review findings and our synthesis of the recent reviews, a research agenda for environmental science's future contribution to mental health scholarship is set out. This includes recommendations to expand the geographical scope and broaden the representation of different environmental science areas, improve measurement of environmental exposure, prioritise experimental and longitudinal research designs, and giving greater consideration to variation between and within communities and the mediating pathways by which environment influences mental health. There is also considerable opportunity to increase interdisciplinarity within the field via the integration of conceptual models, the inclusion of mixed methods and qualitative approaches, as well as further consideration of the socio-political context and the environmental states that can help support good mental health. The findings were used to propose a conceptual model to parse contributions and connections between environmental science and mental health to inform future studies.

Importance of the endometrial immune environment in endometrial cancer and associated therapies.

Endometrial cancer is rising in prevalence. The standard treatment modality of hysterectomy is becoming increasingly inadequate due primarily to the direct link between endometrial cancer and high BMI which increases surgical risks. This is an immunogenic cancer, with unique molecular subtypes associated with differential immune infiltration. Despite the immunogenicity of endometrial cancer, there is limited pre-clinical and clinical evidence of the function of immune cells in both the normal and cancerous endometrium. Immune checkpoint inhibitors for endometrial cancer are the most well studied type of immune therapy but these are not currently used as standard-of-care and importantly, they represent only one method of immune manipulation. There is limited evidence regarding the use of other immunotherapies as surgical adjuvants or alternatives. Levonorgestrel-loaded intra-uterine systems can also be effective for early-stage disease, but with varying success. There is currently no known reason as to what predisposes some patients to respond while others do not. As hormones can directly influence immune cell function, it is worth investigating the immune compartment in this context. This review assesses the immunological components of the endometrium and describes how the immune microenvironment changes with hormones, obesity, and in progression to malignancy. It also describes the importance of investigating novel pathways for immunotherapy.

Characterisation of the immune microenvironment of cutaneous squamous cell carcinoma in immunosuppression.

Cutaneous squamous cell carcinoma (cSCC) is a common cancer. Systemic immunosuppression with drugs such as Prednisone results in more aggressive disease. We hypothesise that more aggressive disease in immunosuppression is the result of immune changes in the tumor microenvironment. We characterised T cell, phagocytic and antigen presenting cell subsets in cSCC and determined if these infiltrates were altered by immunosuppressive therapy. We found a dominant "CD8 profile" in the centre of cSCC lesions, with CD8 cells correlating with Tbet, FoxP3, OX40 and "M2-like" macrophages, whereas a "Tbet and granulocyte profile" with associated inflammation predominated at the margin of the tumor. Individuals on systemic immunosuppressive therapy had lesions that were comparable in size, stage and number of vessels to immune competent individuals; however, the number of CD11c positive cells in the lesion centre was significantly reduced. We conclude that cSCC lesions are immunologically heterogeneous across the lesion and that systemically immunosuppressed individuals have reduced CD11c positive cells in the centre of the lesion. The role and detailed phenotype of CD11c positive cells in cSCC lesions warrant further investigation.

Urban-Rural Differences in Cardiac Arrest Outcomes: A Retrospective Population-Based Cohort Study.

Background: Approximately 10% of people who suffer an out-of-hospital cardiac arrest (OHCA) treated by paramedics survive to hospital discharge. Survival differs by up to 19.2% between urban centres and rural areas. Our goal was to investigate the differences in OHCA survival between urban centres and rural areas. Methods: This was a retrospective cohort study of OHCA patients treated by Nova Scotia Emergency Medical Services (EMS) in 2017. Cases of traumatic, expected, and noncardiac OHCA were excluded. Data were collected from the Emergency Health Service electronic patient care record system and the discharge abstract database. Geographic information system analysis classified cases as being in urban centres (population > 1000 people) or rural areas, using 2016 Canadian Census boundaries. The primary outcome was survival to hospital discharge. Multivariable logistic regression covariates were age, sex, bystander resuscitation, whether the arrest was witnessed, public location, and preceding symptoms. Results: A total of 510 OHCAs treated by Nova Scotia Emergency Medical Services were included for analysis. A total of 12% (n = 62) survived to discharge. Patients with OHCAs in urban centres were 107% more likely to survive than those with OHCAs in rural areas (adjusted odds ratio = 2.1; 95% confidence interval = 1.1 to 3.8; P = 0.028). OHCAs in urban centres had a significantly shorter mean time to defibrillation of shockable rhythm (11.2 minutes ± 6.2) vs those in rural areas (17.5 minutes ± 17.3). Conclusions: Nova Scotia has an urban vs rural disparity in OHCA care that is also seen in densely populated OHCA centres. Survival is improved in urban centres. Further improvements in overall survival, especially in rural areas, may arise from community engagement in OHCA recognition and optimized healthcare delivery.

Contexte: Environ 10 % des personnes qui subissent un arrêt cardiaque en milieu extrahospitalier (ACEH), traité par des intervenants paramédicaux, survivent jusqu'à leur congé de l'hôpital. Le taux de survie peut différer de 19,2 % entre les centres urbains et les régions rurales. Notre étude visait à étudier les différences en matière de survie après un ACEH entre les centres urbains et les régions rurales. Méthodologie: Il s'agissait d'une étude de cohorte rétrospective portant sur des patients ayant subi un ACEH traité par les services médicaux d'urgence de la Nouvelle-Écosse en 2017. Les cas d'ACEH traumatique, prévu et non cardiaque ont été exclus. Les données ont été recueillies à partir du système de dossiers électroniques de soins aux patients des services médicaux d'urgence et de la Base de données sur les congés des patients. L'analyse du système d'information géographique a classé les cas selon qu'ils sont survenus dans un centre urbain (population de plus de 1 000 personnes) ou dans une région rurale, en utilisant les limites du recensement canadien de 2016. Le principal paramètre d'évaluation était la survie à la sortie de l'hôpital. Les covariables utilisées dans la régression logistique multivariée étaient l'âge, le sexe, la réanimation effectuée par des témoins si présents lors de l'arrêt cardiaque, l'emplacement public et les symptômes précédents. Résultats: Au total, 510 ACEH traités par les services médicaux d'urgence de la Nouvelle-Écosse ont été inclus aux fins de l'analyse. En tout, 12 % (n = 62) des sujets ont survécu jusqu'à leur congé hospitalier. Les patients ayant subi un ACEH dans un centre urbain étaient 107 % plus susceptibles de survivre que ceux ayant subi un ACEH dans une région rurale (rapport de cotes ajusté : 2,1; intervalle de confiance à 95 % : 1,1 ­ 3,8; p = 0,028). Le temps moyen de délivrance d'un choc lors d'un ACEH avec rythme défibrillable est significativement plus court (11,2 ± 6,2 minutes) dans un centre urbain que dans une région rurale (17,5 ± 17,3 minutes). Conclusions: La Nouvelle-Écosse fait état d'une disparité dans les soins de l'ACEH entre les régions urbaines et les régions rurales, que l'on observe également dans les villes densément peuplées. La survie est plus longue dans les centres urbains. Il est possible de prolonger davantage la survie globale, en particulier dans les régions rurales, en sensibilisant la communauté à l'ACEH et en optimisant la prestation des soins de santé.

Social Isolation in Older Adults: A Qualitative Study on the Social Dimensions of Group Outdoor Health Walks.

Physical distancing practices during the COVID-19 global pandemic contributed to a high degree of social isolation among older adults. To reduce loneliness and other ill effects of social isolation, public health experts recommended outdoor social gathering, with physical distancing. Adopting a case study approach, we explored how social aspects of group outdoor health walks (GOHWs) mitigate social isolation for older adults and improve individual social wellbeing. We used semi-structured interviews to understand the experiences of social isolation and social relationships in nine older (50-80 s) adults participating in a GOHW in Scotland, United Kingdom (UK). Verbatim transcripts were analysed through an iterative process of thematic analysis carried out by an interdisciplinary team of qualitative researchers from environmental psychology, medicine, and geography. Themes provide insight into the social dimensions of GOHWs, the mediating effects of social experiences, and the contribution these make to individual social wellbeing. GOHWs provide opportunities to be part of a group and attend to the needs of inexperienced or physically challenged individuals. Being part of the group walk fosters casual interpersonal interactions through spontaneous mixing during and after the walk. This programmatic structure counters loneliness, engenders pleasurable anticipation of regular contact with others, supports physical activity, and fosters group cohesion. These in turn contribute to individual social wellbeing, including expanding social networks, meaningful relationships, a sense of belonging, and acting on empathy for others. GOWHs may be beneficial for mitigation of social isolation as we emerge from the COVID-19 pandemic. Findings were used to propose a conceptual model to parse social constructs and inform selection or development of quantitative social measures for future studies of nature-based interventions such as GOHWs.

Exercise in People With Cancer: A Spotlight on Energy Regulation and Cachexia.

Exercise is increasingly becoming a standard of cancer care, with well-documented benefits for patients including improved mental wellbeing and reduced treatment-related side effects. However, important gaps in knowledge remain about how to optimise exercise prescription for people with cancer. Importantly, it remains unclear how exercise affects the progression of cancer cachexia (a wasting disease stemming from energy imbalance, and a common manifestation of advanced malignant disease), particularly once the condition has already developed. It was recently suggested that the anti-tumour effect of exercise might come from improved energetic capacity. Here, we highlight the possible effect of exercise on energetic capacity and energy regulation in the context of cancer, and how this might affect the progression of cancer cachexia. We suggest that due to the additional energy demand caused by the tumour and associated systemic inflammation, overreaching may occur more easily in people with cancer. Importantly, this could result in impaired anti-tumour immunity and/or the exacerbation of cancer cachexia. This highlights the importance of individualised exercise programs for people with cancer, with special consideration for the regulation of energy balance, ongoing monitoring and possible nutritional supplementation to support the increased energy demand caused by exercise.

Responding to Cardiac Arrest in the Community in the Digital Age.

Sudden cardiac arrest (SCA) is a common event, affecting almost 400,000 individuals annually in North America. Initiation of cardiopulmonary resuscitation (CPR) and early defibrillation using an automated external defibrillator (AED) are critical for survival, yet many bystanders are reluctant to intervene. Digital technologies, including mobile devices, social media, and crowdsourcing might help play a role to improve survival from SCA. In this article we review the current digital tools and strategies available to increase rates of bystander recognition of SCA, prompt immediate activation of emergency medical services (EMS), initiate high-quality CPR, and to locate, retrieve, and operate AEDs. Smartphones can help to educate and connect bystanders with EMS dispatchers, through text messaging or video calling, to encourage the initiation of CPR and retrieval of the closest AED. Wearable devices and household smart speakers could play a future role in continuous vital signs monitoring in individuals at risk of lethal arrhythmias and send an alert to either chosen contacts or EMS. Machine learning algorithms and mathematical modelling might aid EMS dispatchers with better recognition of SCA as well as policymakers with where to best place AEDs for optimal accessibility. There are challenges with the use of digital tech, including the need for government regulation and issues with data ownership, accessibility, and interoperability. Future research will include smart cities, e-linkages, new technologies, and using social media for mass education. Together or in combination, these emerging digital technologies might represent the next leap forward in SCA survival.

Effect of immune modulation on the skeletal muscle mitochondrial exercise response: An exploratory study in mice with cancer.

Cancer causes mitochondrial alterations in skeletal muscle, which may progress to muscle wasting and, ultimately, to cancer cachexia. Understanding how exercise adaptations are altered by cancer and cancer treatment is important for the effective design of exercise interventions aimed at improving cancer outcomes. We conducted an exploratory study to investigate how tumor burden and cancer immunotherapy treatment (anti-PD-1) modify the skeletal muscle mitochondrial response to exercise training in mice with transplantable tumors (B16-F10 melanoma and EO771 breast cancer). Mice remained sedentary or were provided with running wheels for ~19 days immediately following tumor implant while receiving no treatment (Untreated), isotype control antibody (IgG2a) or anti-PD-1. Exercise and anti-PD-1 did not alter the growth rate of either tumor type, either alone or in combination therapy. Untreated mice with B16-F10 tumors showed increases in most measured markers of skeletal muscle mitochondrial content following exercise training, as did anti-PD-1-treated mice, suggesting increased mitochondrial content following exercise training in these groups. However, mice with B16-F10 tumors receiving the isotype control antibody did not exhibit increased skeletal muscle mitochondrial content following exercise. In untreated mice with EO771 tumors, only citrate synthase activity and complex IV activity were increased following exercise. In contrast, IgG2a and anti-PD-1-treated groups both showed robust increases in most measured markers following exercise. These results indicate that in mice with B16-F10 tumors, IgG2a administration prevents exercise adaptation of skeletal muscle mitochondria, but adaptation remains intact in mice receiving anti-PD-1. In mice with EO771 tumors, both IgG2a and anti-PD-1-treated mice show robust skeletal muscle mitochondrial exercise responses, while untreated mice do not. Taken together, we postulate that immune modulation may enhance skeletal muscle mitochondrial response to exercise in tumor-bearing mice, and suggest this as an exciting new avenue for future research in exercise oncology.

Effects of exercise and anti-PD-1 on the tumour microenvironment.

Immune checkpoint inhibition is highly effective in treating a subset of patients with certain cancers, such as malignant melanoma. However, a large proportion of patients will experience treatment resistance, and other tumour types, such as breast cancer, have thus far proven largely refractory to immune checkpoint inhibitors as single agents. Exercise has been associated with improved cancer patient survival, has known immune-modulatory effects, may improve anti-tumour immunity and may normalise tumour blood vessels. Therefore, we hypothesised that post-implant exercise would boost the effect of concurrent immunotherapy by enhancing anti-tumour immune responses and improving tumour blood flow. To investigate this, mice with EO771 breast tumours or B16-F10 melanomas received anti-PD-1, an isotype control antibody or no treatment. Mice were randomised to exercise (voluntary wheel running) or no exercise at tumour implant. Exercise reduced the number of CD8+T cells in EO771 (p = 0.0011) but not B16-F10 tumours (p = 0.312), and reduced the percentage of CD8+T cells within the total T cell population in both tumour types (B16-F10: p = 0.0389; EO771: p = 0.0015). In contrast, the combination of exercise and anti-PD-1 increased the percentage of CD8+T cells in EO771 (p = 0.0339) but not B16-F10 tumours. Taken together, our results show that exercise and anti-PD-1 induce changes in the tumour immune microenvironment which are dependant on tumour type.

RNAscope compatibility with image analysis platforms for the quantification of tissue-based colorectal cancer biomarkers in archival formalin-fixed paraffin-embedded tissue.

RNAscope®, has emerged as an important in-situ hybridisation method to validate mRNA expression within single cells whilst preserving tissue morphology in histological samples. The aim of this research was to compare the utility of various open-source and commercial image analysis methods, to quantify mRNA transcripts identified by RNAscope within formalin fixed paraffin embedded (FFPE) histological samples and cell monolayer preparations. Examination of MLH1 expression from 10 histological FFPE colorectal cancer specimens using four image analysis tools (Colour Deconvolution, SpotStudio, WEKA and the LEICA RNA-ISH algorithm) showed the WEKA tool as having the greatest level of agreement with manual quantification. Comparing image analysis methods to qRT-PCR for quantifying MLH1, GFI1 and TNFRSF11A expression within two colorectal cell lines results suggest that these image analysis methods perform at a similar level to qRT-PCR. Furthermore, we describe the strengths and limitations for each image analysis method when used in combination with RNAscope assays. Our study concludes that there are several freely available and commercial image analysis tools that enable reliable RNA in situ expression analysis, however operators need to consider factors, such as expected expression levels of target genes, software usability and functionality.

Outdoor Recreation for Older Adults in Scotland: Qualitatively Exploring the Multiplicity of Constraints to Participation.

Active ageing can lead to better health outcomes in older people. Examining constraints to outdoor recreation for older people, including outdoor physical activity, may therefore assist with developing strategies for active ageing. Findings are presented from a study seeking to understand the constraints to older peoples' access to outdoor recreation in Scotland, and this paper aimed to examine the multitude of constraints that discourage or prevent older people from accessing the outdoors and the ways in which these constraints are hierarchical (or not). This paper adopted a qualitative methodology using the hierarchical leisure constraints model (HLCM) as a lens to analyse the data, presenting the data in three vignettes. The paper identified multiple co-occurring constraints and considered these in relation to expectations based on the HLCM. Recognising that constraints to outdoor recreation for older people are multiple, co-occurring and mutually reinforcing may enable more effective solutions to be developed to overcome them.

Gene and Protein Expression Is Altered by Ascorbate Availability in Murine Macrophages Cultured under Tumour-Like Conditions.

Tumour-associated macrophages (TAMs) are ubiquitously present in tumours and commonly associated with poor prognosis. In immune cells, ascorbate affects epigenetic regulation, differentiation and phenotype via its co-factor activity for the 2-oxoglutarate dependent dioxygenase enzymes. Here, we determined the effect of ascorbate on TAM development in response to tumour microenvironmental cues. Naïve murine bone marrow monocytes were cultured with Lewis Lung Carcinoma conditioned media (LLCM) or macrophage colony-stimulating factor (MCSF) to encourage the development into tumour-associated macrophages. Cells were stimulated with hypoxia (1% O2), with or without ascorbate (500 µM) supplementation. Cells and media were harvested for gene, cell surface marker and protein analyses. LLCM supported bone marrow monocyte growth with >90% of cells staining CD11b+F4/80+, indicative of monocytes/macrophages. LLCM-grown cells showed increased expression of M2-like and TAM genes compared to MCSF-grown cells, which further increased with hypoxia. In LLCM-grown cells, ascorbate supplementation was associated with increased F4/80 cell surface expression, and altered gene expression and protein secretion. Our study shows that ascorbate modifies monocyte phenotype when grown under tumour microenvironmental conditions, but this was not clearly associated with either a pro- or anti-tumour phenotype, and reflects a complex and nuanced response of macrophages to ascorbate. Overall, ascorbate supplementation clearly has molecular consequences for TAMs, but functional and clinical consequences remain unknown.

Influence of serum inflammatory cytokines on cytochrome P450 drug metabolising activity during breast cancer chemotherapy: a patient feasibility study.

Individual response to chemotherapy in patients with breast cancer is variable. Obesity and exercise are associated with better and worse outcomes, respectively, and it is known that both impact the systemic cytokine milieu. Cytochrome P450 (CYP) enzymes are responsible for the metabolism of many chemotherapy agents, and CYP enzyme activity has been shown to be modified by inflammatory cytokines in vitro and in vivo. Cytokine-associated changes in CYP metabolism may alter chemotherapy exposure, potentially affecting treatment response and patient survival. Therefore, better understanding of these biological relationships is required. This exploratory single arm open label trial investigated changes in in vivo CYP activity in twelve women treated for stage II or III breast cancer, and demonstrated for the first time the feasibility and safety of utilising the Inje phenotyping cocktail to measure CYP activity in cancer patients receiving chemotherapy. Relative CYP activity varied between participants, particularly for CYP2C9 and CYP2D6, and changes in serum concentrations of the inflammatory cytokine monocyte chemoattractant protein 1 inversely correlated to CYP3A4 activity during chemotherapy. Future use of phenotyping cocktails in a clinical oncology setting may help guide drug dosing and improve chemotherapy outcomes.Clinical Trial Registration: Trial was retrospectively registered to the Australia New Zealand Clinical Trial Registry (ANZCTR). ACTRN12620000832976, 21 Aug 2020, https://www.anzctr.org.au/ACTRN12620000832976.aspx .

Functional effects of immune complexes formed between pembrolizumab and patient-generated anti-drug antibodies.

The PD-1-targeting IgG4 antibody pembrolizumab has significant anti-tumor activity in a proportion of stage IV melanoma patients. A subset of patients develop anti-drug antibodies (ADA) which can form immune complexes (IC) with pembrolizumab. Although IC can induce powerful, Fc-mediated, immune-regulatory effects, their functional impact during pembrolizumab treatment is unclear. The functional effects of IC generated in vitro using pembrolizumab and patient-derived ADA was, therefore, investigated. Screening identified a patient whose trough serum samples from three treatment cycles contained both ADA with neutralizing activity and low levels of pembrolizumab. This patient responded well to therapy over 2 years and had ongoing, infusion-related, hypersensitivity reactions despite the later absence of detectable ADA. The components of IC were mimicked by forming a complex of pembrolizumab by absorption onto a solid phase with or without subsequent exposure to the ADA+ patient sera. Complexes comprised of pembrolizumab alone significantly inhibited TLR4 (LPS)-driven IL-10 production by PBMC and stimulated the generation of reactive oxygen species by granulocytes. In contrast, soluble and solid-phase F(ab´)2 fragments of pembrolizumab had no effect demonstrating the requirement for cross-linked Fc regions. IC containing pembrolizumab and ADA could additionally induce complement and NK activation. The results of this study demonstrate that, when oligomerized, the Fc region of pembrolizumab alone can provide immuno-regulatory signals. Furthermore, IC containing both pembrolizumab and patient-generated ADA can induce additional signals. These Fc-mediated signals may modulate both hypersensitivity reactions and anti-tumor responses associated with pembrolizumab therapy.

Effect of post-implant exercise on tumour growth rate, perfusion and hypoxia in mice.

Preclinical studies have shown a larger inhibition of tumour growth when exercise begins prior to tumour implant (preventative setting) than when training begins after tumour implant (therapeutic setting). However, post-implantation exercise may alter the tumour microenvironment to make it more vulnerable to treatment by increasing tumour perfusion while reducing hypoxia. This has been shown most convincingly in breast and prostate cancer models to date and it is unclear whether other tumour types respond in a similar way. We aimed to determine whether tumour perfusion and hypoxia are altered with exercise in a melanoma model, and compared this with a breast cancer model. We hypothesised that post-implantation exercise would reduce tumour hypoxia and increase perfusion in these two models. Female, 6-10 week old C57BL/6 mice were inoculated with EO771 breast or B16-F10 melanoma tumour cells before randomisation to either exercise or non-exercising control. Exercising mice received a running wheel with a revolution counter. Mice were euthanised when tumours reached maximum ethical size and the tumours assessed for perfusion, hypoxia, blood vessel density and proliferation. We saw an increase in heart to body weight ratio in exercising compared with non-exercising mice (p = 0.0008), indicating that physiological changes occurred with this form of physical activity. However, exercise did not affect vascularity, perfusion, hypoxia or tumour growth rate in either tumour type. In addition, EO771 tumours had a more aggressive phenotype than B16-F10 tumours, as inferred from a higher rate of proliferation (p<0.0001), a higher level of tumour hypoxia (p = 0.0063) and a higher number of CD31+ vessels (p = 0.0005). Our results show that although a physiological training effect was seen with exercise, it did not affect tumour hypoxia, perfusion or growth rate. We suggest that exercise monotherapy is minimally effective and that future preclinical work should focus on the combination of exercise with standard cancer therapies.