Development of robust, indigenous ELISA for detection of IgG antibodies against CoV-2 N and S proteins: mass screening.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic has adversely affected humankind and caused millions of deaths globally since January 2020. Robust and quick serological tests such as antibody detection assays for SARS-CoV-2 provide relevant information and aid in the process of vaccine development and diagnostics, as well as in sero-epidemiological monitoring of antibody response to the virus. The receptor-binding domain (RBD) of spike and nucleocapsid protein are specific targets for detecting SARS-CoV-2 antibodies. Here, we present the development of a stable spike (S) and nucleocapsid (N) protein-based ELISA antibody detection test "CoroSuchak," with 99% sensitivity, 98% specificity, cost-effective, and detection in a minimum time for serodiagnosis and mass screening of the population for antibodies against SARS-CoV-2. Blood samples were analyzed from 374 SARS-CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) positive, 772 negative and asymptomatic, and 874 random groups of subjects. We found that the antibody titer was significantly higher (p < 0.0001) in infected and vaccinated group compared to the only vaccinated and only infected group. Using enzyme-linked immunosorbent assay (ELISA), we detected SARS-CoV-2 immunoglobulin G (IgG) antibodies in 118/123 (96%) infected individuals, 570/653 (87%) non-infected but vaccinated individuals, 231/237 (97%) individuals who were both infected and vaccinated, and 499/874 (57%) from randomly selected individuals from the first and second waves of the pandemic. Similarly in the third wave, 14/14 (100%) infected and 16/20 (80%) RT-PCR-negative but symptomatic subjects were detected. Thus, the highly sensitive and specific in-house developed ELISA antibody detection kit "CoroSuchak" is extremely useful to determine the seroprevalence of SARS-CoV-2 antibodies in the coronavirus-exposed population. KEY POINTS: •Indigenous kit using a combination of spike and nucleocapsid proteins and peptide sequences. •High sensitivity and specificity to detect variants. •Highly sensitive for mass screening.

Dual-specificity protein phosphatase DUSP4 regulates response to MEK inhibition in BRAF wild-type melanoma.

BACKGROUND: Aiming to improve treatment options for BRAF wild-type melanoma, we previously conducted the DOC-MEK study of docetaxel with MEK inhibitor (MEKi) selumetinib or placebo, revealing trends to prolongation of progression-free survival (hazard ratio 0.75, P = 0.130), and improved response rates (32% vs 14%, P = 0.059) with docetaxel plus selumetinib. NRAS status did not associate with outcome. Here, the aim was to identify novel biomarkers of response to MEKi. METHODS: A MEK 6 gene signature was quantified using NanoString and correlated with clinical outcomes. Two components of the gene signature were investigated by gene silencing in BRAF/NRAS wild-type melanoma cells. RESULTS: In melanomas of patients on the selumetinib but not the placebo arm, two gene signature components, dual-specificity protein phosphatase 4 (DUSP4) and ETS translocation variant 4 (ETV4), were expressed more highly in responders than non-responders. In vitro, ETV4 depletion inhibited cell survival but did not influence sensitivity to MEKi selumetinib or trametinib. In contrast, DUSP4-depleted cells showed enhanced cell survival and increased resistance to both selumetinib and trametinib. CONCLUSIONS: ETV4 and DUSP4 associated with clinical response to docetaxel plus selumetinib. DUSP4 depletion induced MEKi resistance, suggesting that DUSP4 is not only a biomarker but also a mediator of MEKi sensitivity. CLINICAL TRIAL REGISTRATION: DOC-MEK (EudraCT no: 2009-018153-23).

Emergency ambulatory outpatient management of immune-mediated hypophysitis.

PURPOSE: Immune-mediated hypophysitis is an important toxicity related to immune checkpoint inhibitors (ICI). Optimal management is associated with improved outcomes. It represents a wide spectrum of clinical presentations, and a proportion may be suitable for emergency ambulatory management. METHODS: Emergency ambulatory management of patients presenting with clinical features and findings consistent with ICI-induced hypophysitis was considered at a tertiary cancer/endocrinology hospital. Suitable patients were initially investigated and treated in accordance with the UK emergency management guidelines for ICI induced hypophysitis. After an initial observation period of 4 h, patients were discharged with oral hydrocortisone (20, 10, 10 mg). RESULTS: An initial cohort of 4 patients with emergency presentations of ICI-induced hypophysitis has been managed in an ambulatory fashion in the first 3 months. There were no 30-day readmissions. CONCLUSION: Carefully selected emergency presentations with immune-mediated hypophysitis may be suitable for ambulatory management.

A phase 1 study to assess the safety, tolerability, and pharmacokinetics of CXD101 in patients with advanced cancer.

BACKGROUND: In the current study, the authors sought to determine the maximum tolerated dose (MTD) of the novel class 1 selective histone deacetylase inhibitor CXD101 in a dose escalation study in patients with advanced solid tumors or recurrent/refractory lymphoma. METHODS: The authors escalated the dose of CXD101 from 1 mg twice daily orally for 5 days in a 21-day cycle (3+3 design). RESULTS: A total of 39 patients were enrolled, 36 of whom received CXD101. Of the 30 patients in the escalation cohort, 29 were evaluable for determination of the dose-limiting toxicity (DLT). DLTs were noted at doses of 16 mg twice daily (1 of 6 patients), 20 mg twice daily (1 of 6 patients), and 24/25 mg twice daily (2 of 5 patients, both of whom developed neutropenic fever). The MTD was 20 mg twice daily, which achieved maximal plasma concentrations (±standard deviation) of 231±76 nM to 342±126 nM, which was within the biologically active range. Six patients received 20 mg twice daily in an expansion cohort. The most frequent adverse events were fatigue, nausea, and reversible cytopenia. Key grade 3 to 4 adverse events (according to Common Terminology Criteria for Adverse Events criteria [version 4.03]) included thrombocytopenia (11%), neutropenia (17%), and neutropenic fever (2%) across the 133 CXD101 cycles given. The toxicity profile was similar to that of licensing studies with other histone deacetylase inhibitors. In 22 evaluable patients receiving a dose of ≥16 mg twice daily (17 of whom had lymphoma and 5 of whom had solid tumors), 3 partial responses (2 in patients with classic Hodgkin lymphoma after allogenic stem cell transplantation and 1 in a patient with angioimmunoblastic T-cell lymphoma) and 1 complete response (in a patient with follicular lymphoma) were noted (overall response rate of 18%) in addition to 9 patients who achieved durable stable disease. Responses were noted predominantly among patients with lymphoma (tumor reduction noted in 63% of patients on standard computed tomography). CONCLUSIONS: The MTD in the current study was found to be 20 mg twice daily. Encouraging and durable activity was observed in patients with Hodgkin lymphoma, T-cell lymphoma, and follicular lymphoma.

TORC1-signalling is down-regulated in Saccharomyces cerevisiae hsp30Δ cells by SNF1-dependent mechanisms.

Hsp30 is a plasma membrane localized heat shock protein in Saccharomyces cerevisiae whose expression is induced by numerous environmental stressors. Elucidation of its mechanism of action has remained elusive primarily because hsp30Δ cells do not show a strong phenotype. To identify cellular functions associated with Hsp30, we thus compared the transcriptome of BY4741hsp30Δ with that of its wild type counterpart. Our studies indicate down-regulation of the target of rapamycin complex 1 (TORC1)-dependent gene-expression programme in hsp30Δ cells. We further show that TORC1-signalling through its effectors (Sch9 and Tap42) was down-regulated in the deletion strain. Specifically, (a) phosphorylation levels of Sch9 were lower and nuclear exclusion of Rim15 (Sch9-downstream function) was overridden in hsp30Δ cells, (b) membrane association of Tor1 and Tap42 was lower in hsp30Δ cells, and (c) Tap42-downstream functions were abrogated in the deletion strain. Furthermore, transcription factors Rtg1, Rtg3, Gat1, and Gln3 were localized in the nucleus of the hsp30Δ as observed upon inactivation of TORC1. Studies aimed at determining how TORC1-signalling is down-regulated in hsp30Δ cells indicated that total reducing sugar levels were lower and ADP:ATP ratio was higher in hsp30Δ cells -conditions known to activate the Snf1 kinase and consequently to the inactivation of TORC1. We thus determined if TORC1-signalling could be restored in hsp30Δ cells upon the deletion of SNF1. Sch9 phosphorylation levels (TORC1-signalling) was restored to wild type levels in hsp30Δsnf1Δ cells. TORC1-signalling is thus down-regulated in hsp30Δ cells by SNF1-dependent mechanisms. A probable role for Hsp30 is discussed.

Clinical applicability and cost of a 46-gene panel for genomic analysis of solid tumours: Retrospective validation and prospective audit in the UK National Health Service.

BACKGROUND: Single gene tests to predict whether cancers respond to specific targeted therapies are performed increasingly often. Advances in sequencing technology, collectively referred to as next generation sequencing (NGS), mean the entire cancer genome or parts of it can now be sequenced at speed with increased depth and sensitivity. However, translation of NGS into routine cancer care has been slow. Healthcare stakeholders are unclear about the clinical utility of NGS and are concerned it could be an expensive addition to cancer diagnostics, rather than an affordable alternative to single gene testing. METHODS AND FINDINGS: We validated a 46-gene hotspot cancer panel assay allowing multiple gene testing from small diagnostic biopsies. From 1 January 2013 to 31 December 2013, solid tumour samples (including non-small-cell lung carcinoma [NSCLC], colorectal carcinoma, and melanoma) were sequenced in the context of the UK National Health Service from 351 consecutively submitted prospective cases for which treating clinicians thought the patient had potential to benefit from more extensive genetic analysis. Following histological assessment, tumour-rich regions of formalin-fixed paraffin-embedded (FFPE) sections underwent macrodissection, DNA extraction, NGS, and analysis using a pipeline centred on Torrent Suite software. With a median turnaround time of seven working days, an integrated clinical report was produced indicating the variants detected, including those with potential diagnostic, prognostic, therapeutic, or clinical trial entry implications. Accompanying phenotypic data were collected, and a detailed cost analysis of the panel compared with single gene testing was undertaken to assess affordability for routine patient care. Panel sequencing was successful for 97% (342/351) of tumour samples in the prospective cohort and showed 100% concordance with known mutations (detected using cobas assays). At least one mutation was identified in 87% (296/342) of tumours. A locally actionable mutation (i.e., available targeted treatment or clinical trial) was identified in 122/351 patients (35%). Forty patients received targeted treatment, in 22/40 (55%) cases solely due to use of the panel. Examination of published data on the potential efficacy of targeted therapies showed theoretically actionable mutations (i.e., mutations for which targeted treatment was potentially appropriate) in 66% (71/107) and 39% (41/105) of melanoma and NSCLC patients, respectively. At a cost of £339 (US$449) per patient, the panel was less expensive locally than performing more than two or three single gene tests. Study limitations include the use of FFPE samples, which do not always provide high-quality DNA, and the use of "real world" data: submission of cases for sequencing did not always follow clinical guidelines, meaning that when mutations were detected, patients were not always eligible for targeted treatments on clinical grounds. CONCLUSIONS: This study demonstrates that more extensive tumour sequencing can identify mutations that could improve clinical decision-making in routine cancer care, potentially improving patient outcomes, at an affordable level for healthcare providers.

Heat stress modulated gastrointestinal barrier dysfunction: role of tight junctions and heat shock proteins.

Increased environmental temperature exerts a visible impact on an individual's physiology. At the onset of heat stress, there is an increase in core body temperature which triggers peripheral vasodilation and sweating in an effort to dissipate the elevated body heat. The increase in peripheral circulation however reduces blood flow to the internal organs which are thus adversely affected. In particular, the gastrointestinal (GI) tract gets adversely affected during hyperthermia resulting in loosening of the tight junctions (TJs) that finally leads to higher intestinal permeability. At the cellular level, elevated levels of heat shock proteins (HSPs) induced in response to heat stress mediated cytoprotection by maintaining proper protein folding, releasing survival signals and preserving cytoskeleton integrity. Recent studies have indicated that HSPs play a crucial role in maintaining the localization of TJ proteins. Dietary supplements have also shown to have a positive effect on the maintenance of intestinal TJs. Therefore, it becomes imperative to understand the cellular, molecular and physiological alterations in response to heat stress in GI tract. In the present report, the effect of thermal stress on GI tract has been summarized. Specific role of HSPs along with mitogen activated protein (MAP) kinase signaling pathway in response to hyperthermia has also been discussed.

RADVAN: a randomised phase 2 trial of WBRT plus vandetanib for melanoma brain metastases - results and lessons learnt.

BACKGROUND: Brain metastases occur in up to 75% of patients with advanced melanoma. Most are treated with whole-brain radiotherapy (WBRT), with limited effectiveness. Vandetanib, an inhibitor of vascular endothelial growth factor receptor, epidermal growth factor receptor and rearranged during transfection tyrosine kinases, is a potent radiosensitiser in xenograft models. We compared WBRT with WBRT plus vandetanib in the treatment of patients with melanoma brain metastases. METHODS: In this double-blind, multi-centre, phase 2 trial patients with melanoma brain metastases were randomised to receive WBRT (30 Gy in 10 fractions) plus 3 weeks of concurrent vandetanib 100 mg once daily or placebo. The primary endpoint was progression-free survival in brain (PFS brain). The main study was preceded by a safety run-in phase to confirm tolerability of the combination. A post-hoc analysis and literature review considered barriers to recruiting patients with melanoma brain metastases to clinical trials. RESULTS: Twenty-four patients were recruited, six to the safety phase and 18 to the randomised phase. The study closed early due to poor recruitment. Median PFS brain was 3.3 months (90% confidence interval (CI): 1.6-5.6) in the vandetanib group and 2.5 months (90% CI: 0.2-4.8) in the placebo group (P=0.34). Median overall survival (OS) was 4.6 months (90% CI: 1.6-6.3) and 2.5 months (90% CI: 0.2-7.2), respectively (P=0.54). The most frequent adverse events were fatigue, alopecia, confusion and nausea. The most common barrier to study recruitment was availability of alternative treatments. CONCLUSIONS: The combination of WBRT plus vandetanib was well tolerated. Compared with WBRT alone, there was no significant improvement in PFS brain or OS, although we are unable to provide a definitive result due to poor accrual. A review of barriers to trial accrual identified several factors that affect study recruitment in this difficult disease area.

Hemoglobin interaction with GP1bα induces platelet activation and apoptosis: a novel mechanism associated with intravascular hemolysis.

Intravascular hemolysis increases the risk of hypercoagulation and thrombosis in hemolytic disorders. Our study shows a novel mechanism by which extracellular hemoglobin directly affects platelet activation. The binding of Hb to glycoprotein1bα activates platelets. Lower concentrations of Hb (0.37-3 µM) significantly increase the phosphorylation of signaling adapter proteins, such as Lyn, PI3K, AKT, and ERK, and promote platelet aggregation in vitro. Higher concentrations of Hb (3-6 µM) activate the pro-apoptotic proteins Bak, Bax, cytochrome c, caspase-9 and caspase-3, and increase platelet clot formation. Increased plasma Hb activates platelets and promotes their apoptosis, and plays a crucial role in the pathogenesis of aggregation and development of the procoagulant state in hemolytic disorders. Furthermore, we show that in patients with paroxysmal nocturnal hemoglobinuria, a chronic hemolytic disease characterized by recurrent events of intravascular thrombosis and thromboembolism, it is the elevated plasma Hb or platelet surface bound Hb that positively correlates with platelet activation.

Development and usability of a virtual reality umbilical venous catheter placement simulator.

PURPOSE: Exposure to procedures varies in the neonatal intensive care unit (NICU). A method to teach procedures should be available without patient availability, expert oversight, or simulation laboratories. To fill this need, we developed a virtual reality (VR) simulation for umbilical vein catheter (UVC) placement and sought to establish its face and content validity and usability. METHODS: Engineers, software developers, graphic designers, and neonatologists developed a VR UVC placement simulator following a participatory design approach. The software was deployed on the Meta Quest 2 head-mounted display (HMD). Neonatal nurse practitioners (NNPs) from a level 4 NICU used the simulator and completed an 11-item questionnaire to establish face and content validity. Participants also completed the validated simulation task load index and system usability scale to assess the usability of the simulator. Group 1 tested the VR simulation, which was optimized based on feedback, prior to Group 2's participation. RESULTS: A total of 14 NNPs with 2-37 years of experience participated in testing. Participants scored the content and face validity of the simulator highly, with most giving scores ≥ 4/5. Usability was established with relatively high average system usability scores for both groups (Group 1: 67.14 ± 7.8, Group 2: 71 ± 14.1) and low SIM-TLX scores indicating manageable load while using the simulator. CONCLUSION: After optimization, Group 2 found the UVC simulator to be realistic and effective. Both groups felt the simulator was easy to use and did not cause physical or cognitive strain. All participants felt the UVC simulator provided a safe environment to make mistakes, and the majority would recommend this experience to trainees.

A randomised phase 2 study of intermittent versus continuous dosing of dabrafenib plus trametinib in patients with BRAFV600 mutant advanced melanoma (INTERIM).

BACKGROUND: BRAF+MEK inhibitors extend life expectancy of patients with BRAFV600 mutant advanced melanoma. Acquired resistance limits duration of benefit, but preclinical and case studies suggest intermittent dosing could overcome this limitation. INTERIM was a phase 2 trial evaluating an intermittent dosing regimen. METHODS: Patients with BRAFV600 mutant advanced melanoma due to start dabrafenib+trametinib were randomised to receive either continuous (CONT), or intermittent (INT; dabrafenib d1-21, trametinib d1-14 every 28 days) dosing. A composite primary endpoint included progression-free survival (PFS) and quality of life (QoL). Secondary endpoints included response rate (ORR), overall survival (OS) and adverse events (AEs). Mutant BRAFV600E ctDNA was measured by droplet digital PCR (ddPCR), using mutant allele frequency of > 1 % as the detection threshold. RESULTS: 79 patients (39 INT, 40 CONT) were recruited; median age 67 years, 65 % AJCC (7th ed) stage IV M1c, 29 % had brain metastases. With 19 months median follow-up, INT was inferior in all efficacy measures: median PFS 8.5 vs 10.7mo (HR 1.39, 95 %CI 0.79-2.45, p = 0.255); median OS 18.1mo vs not reached (HR 1.69, 95 %CI 0.87-3.28, p = 0.121), ORR 57 % vs 77 %. INT patients experienced fewer treatment-related AEs (76 % vs 88 %), but more grade > 3 AEs (53 % vs 42 %). QoL favoured CONT. Detection of BRAFV600E ctDNA prior to treatment correlated with worse OS (HR 2.55, 95 %CI 1.25-5.21, p = 0.01) in both arms. A change to undetected during treatment did not significantly predict better OS. CONCLUSION: INTERIM findings are consistent with other recent clinical trials reporting that intermittent dosing does not improve efficacy of BRAF+MEK inhibitors.

Burden of cancer trial participation: A qualitative sub-study of the INTERIM feasibility RCT.

OBJECTIVE: A qualitative sub-study was carried out within a larger phase II feasibility trial, to identify and describe the burden experienced by advanced melanoma patients participating in a clinical trial and the factors affecting their capacity to cope with the burden. METHODS: Semi-structured interviews were conducted with fourteen patients with advanced melanoma recruited from National Health Service hospitals in the United Kingdom. Qualitative analysis was undertaken using a framework analysis approach. Normalisation process theory was applied to the concept of research participation burden in order to interpret and categorise findings. RESULTS: Burdens of participation were identified as arising from making sense of the trial and treatment; arranging transport, appointment and prescriptions; enacting management strategies and enduring side effects; reflecting on trial documents and treatment efficacy, and emotional and mental effects of randomisation and treatment side effects. Factors reported as influencing capacity include personal attributes and skills, physical and cognitive abilities and support network. DISCUSSION: This is the first study to highlight the substantial burden faced by patients with advanced melanoma in a clinical trial and factors that may lessen or worsen the burden. Consideration of identified burdens during trial design and execution will reduce the burden experienced by research participants.

Flow cytometric measurable residual disease in adult acute myeloid leukemia: a preliminary report from Eastern India.

Presence of measurable residual disease (MRD) in acute myeloid leukemia (AML) is considered to be an independent predictor of relapse and poorer survival outcomes. MRD can be measured by flow cytometric, quantitative PCR, and NGS-based assays at varying sensitivities. There is scant Indian data on different aspects of MFC-MRD in AML including analysis strategies as well as molecular spectrum, clinical correlation, etc. This retrospective observational study included all newly diagnosed patients of acute myeloid leukemia in whom complete baseline diagnostic workup was available including flow cytometry and cytogenetic and molecular studies. Among patients with cytogenetic abnormalities (n = 25), no statistically significant correlation was observed between flow cytometric MRD positivity and presence of ≥ 3 mutations as well as relapsed disease. However, in AML patients with normal karyotype (n = 32), MRD positivity correlated strongly with relapsed status (p = 0.02), although no significant correlation was found with respect to FLT3 mutation, IDH mutation, NPM1 mutation, or complex genotype. Interestingly, 90.5% of MRD-positive patients belonged to ELN (2017) intermediate to high-risk category unlike only 9.5% in the good risk category (p = 0.0002). Median relapse-free survival was 8.5 months with a follow-up range of 3-24 months. On the basis of the observations of the present study, it can be clearly inferred that MRD status affects relapse status in the normal karyotype subgroup and can delineate patients who require stem cell transplantation in addition to molecular signatures.

Immune checkpoint inhibitor-mediated hypophysitis: no place like home.

Ambulatory emergency care forms a fundamental part of the strategy of trying to ensure safe and sustainable acute care services. Immune checkpoint inhibitor(ICI)-mediated hypophysitis is an important life-threatening complication of therapy. Patients presenting with clinical features and findings consistent with ICI-mediated hypophysitis were considered in the current study. In the absence of severe features (sodium <125 mmol/L, hypotension, reduced consciousness, hypoglycaemia and/or visual field defect), patients were administered a single intravenous dose of hydrocortisone (100 mg), observed for at least 4 h and then discharged on oral hydrocortisone (20 mg, 10 mg and 10 mg). Patients were then seen urgently in the endocrinology outpatient setting for further management. Fourteen patients (median age 64, 10 male) were managed using the pathway. All patients had biochemically confirmed adrenocorticotropic hormone (ACTH) deficiency. Seven of the 14 were treated with combination ICI therapy, with four having pan-anterior hypopituitarism. There were no 30-day readmissions or any associated hypophysitis-related mortality. All patients continued ICI therapy without interruption.

Dabrafenib plus trametinib versus anti-PD-1 monotherapy as adjuvant therapy in BRAF V600-mutant stage III melanoma after definitive surgery: a multicenter, retrospective cohort study.

Background: Both dabrafenib/trametinib (D/T) and anti-PD-1 monotherapy (PD-1) are approved adjuvant therapies for patients with stage III BRAF V600-mutant melanoma. However, there is still a lack of head-to-head comparative data. We aimed to describe efficacy and toxicity outcomes for these two standard therapies across melanoma centers. Methods: This multicenter, retrospective cohort study was conducted in 15 melanoma centers in Australia, China, Germany, Italy, Japan, UK, and US. We included adult patients with resected stage III BRAF V600-mutant melanoma who received either adjuvant D/T or PD-1 between Jul 2015 and Oct 2022. The primary endpoint was relapse-free survival (RFS). Secondary endpoints included overall survival (OS), recurrence pattern and toxicity. Findings: We included 598 patients with stage III BRAF V600-mutant melanoma who received either adjuvant D/T (n = 393 [66%]) or PD-1 (n = 205 [34%]) post definitive surgery between Jul 2015 and Oct 2022. At a median follow-up of 33 months (IQR 21-43), the median RFS was 51.0 months (95% CI 41.0-not reached [NR]) in the D/T group, significantly longer than PD-1 (44.8 months [95% CI 28.5-NR]) (univariate: HR 0.66, 95% CI 0.50-0.87, P = 0.003; multivariate: HR 0.58, 95% CI 0.39-0.86, P = 0.007), with comparable OS with PD-1 (multivariate, HR 0.90, 95% CI 0.48-1.70, P = 0.75). Similar findings were observed using a restricted-mean-survival-time model. Among those who experienced recurrence, the proportion of distant metastases was higher in the D/T cohort. D/T had a higher incidence of treatment modification due to adverse events (AEs) than PD-1, but fewer persistent AEs. Interpretation: In patients with stage III BRAF V600-mutant melanoma post definitive surgery, D/T yielded better RFS than PD-1, with higher transient but lower persistent toxicity, and comparable OS. D/T seems to provide a better outcome compared with PD-1, but a longer follow-up and ideally a large prospective trial are needed. Funding: Dr. Xue Bai was supported by the Beijing Hospitals Authority Youth Programme (QMS20211101) for her efforts devoted to this study. Dr. Keith T. Flaherty was funded by Adelson Medical Research Foundation for the efforts devoted to this study.

Severe heat stress modulated nuclear factor erythroid 2-related factor 2 and macrophage migration inhibitory factor pathway in rat liver.

Heat stress impairs physiology and overall functionality of the body at tissue and organ level in animals. Liver being a vital organ performs more than hundreds regulatory functions of the body. Present study investigates the modulation of molecular pathways that are responsible for liver damage triggered by heat stress. Male Sprague dawley rats were exposed to heat stress (45 °C) in heat simulation chamber till core temperature reaches 40 °C and 42 °C in 25 and 42 min respectively. For in-depth evaluation of liver functions during severe heat stress, hepatic transcriptome and proteome were analysed by microarray and two dimensional gel electrophoresis respectively. Results revealed major alterations in redox status, inflammation, mitochondrial dysfunction and proteostasis related pathways. Data of molecular pathway analysis demonstrate that nuclear factor erythroid 2-related factor 2 (NRF-2) mediated oxidative stress response and macrophage migration inhibitory factor (MIF) regulated inflammatory pathways were upregulated in severe heat stressed liver. Expression levels of downstream molecules of above pathways such as heat shock protein 90AB 1, peroxiredoxin 5, Jun N-terminal kinases 1/2, heme-oxygenase 1, apolipoprotein 1 and interleukin 10 were examined and result suggested the upregulation of these genes modulates the NRF-2 and MIF regulated pathways in heat stressed liver. Irregularity in molecular signalling networks lead to mitochondrial dysfunction indicated by upregulation of ATP synthase ß and peroxiredoxin 1 along with decreased levels of glucose-6-phosphate dehydrogenase and enhanced activity of cytochrome c in liver mitochondria. Thus, current study demonstrated heat induced alterations in key liver functions were regulated by NRF-2 and MIF pathways.

Delphi panel for consensus on the optimal management of dabrafenib plus trametinib-related pyrexia in patients with melanoma.

Purpose: Dabrafenib and trametinib combination therapy (dab + tram) is indicated to treat BRAF V600 mutation-positive unresectable/metastatic melanoma and as adjuvant treatment for resected stage III disease. Dab + tram-related pyrexia may require early therapy discontinuation. A modified Delphi panel was conducted to develop consensus on the optimal management of dab + tram-related pyrexia in patients with melanoma. Methods: In all, 10 UK oncologists experienced in melanoma management participated in a three-round modified Delphi study (Round 1: one-to-one interview; Rounds 2 and 3: email survey). In each round, participants rated the extent of their agreement with statements about defining and managing dab + tram-related pyrexia. Consensus was defined as >80% agreement for critical management (CM) and >60% for non-critical management (NCM) statements. Results: All 10 participants completed Round 1; 9 completed Rounds 2 and 3. Consensus was reached on 42/66 statements (20 CM and 22 NCM). Drug-related pyrexia was agreed as being strictly an elevation of body temperature, although other symptoms may be present (89% agreement). Panelists agreed on the need for simple and generic guidance on dab + tram-related pyrexia management that does not differentiate between patient groups (100%), and that management of first and second dab + tram-related pyrexia episodes should be the same regardless of treatment intent (100%). Regarding CM, participants agreed that both dab and tram should be interrupted for pyrexia (100%) without considering the use of steroids (89%); patients on dab + tram presenting to non-oncology services with pyrexia should be directed to an oncology-specific service as soon as possible and assessed for infection (100%). NCM statements on steroid use following dab + tram interruption and when to restart dab + tram did not reach consensus. Conclusions: These consensus statements provide a framework on optimal management of dab + tram-related pyrexia in patients with melanoma which should inform future guidelines.

Measurement of diffusion lung capacity (DLCO) in silicosis patients: Correlation with radiographic abnormalities on high-resolution CT scan chest.

Objective: This study was conducted to evaluate diffusion capacity of lung for carbon monoxide (DLCO) in patients with simple and complicated silicosis and to correlate abnormal findings detected, if any, with the computed tomography abnormalities in these patients. Methods: This study included 56 patients with simple and complicated silicosis and without tuberculosis, in whom we performed DLCO as per standard technique. Various computed tomography findings, that is, presence, size and distribution of nodules associated with relative parenchymal and vascular markings, were recorded in the study subjects and classified into standard grading to be finally compared with DLCO. Visual grading score system was used to describe the extent of emphysematous changes based on the area of abnormally low attenuation, vascular disruption, bullae and so on and data were recorded. Results: Results showed that 85.7% patients had small opacities of varying grades and 28.5% showed large opacities, with 16% of them having type 'C' large opacities. The mean DLCO (% predicted) of patients with category '0' high-resolution computed tomography (HRCT) abnormality was 92.3 ± 6.8 (within normal limits), and this gradually decreased with increasing HRCT category to 44.2 ± 11.2 in grade '4' of progressive massive fibrosis (PMF) patients in this study (P < 0.01). This reflects a significant inverse correlation between visual HRCT category and the DLCO % predicted (r > -0.89, P < 0.001). The mean DLCO (% predicted) was 51 ± 12.6 in patients with grade '1' emphysema in HRCT, 53 ± 13.5 in grade '2', 43 ± 6.4 in grade '3' and 37.7 ± 6.3 in grade '4'; however, there was no correlation between emphysema grading and pulmonary functional index (r = -0.33, P = 0.15). Conclusion: This study observed significant abnormality in DLCO among silicosis patients and its strong correlation with the extent of radiological abnormality. HRCT finding of large opacities could be an important indicator of the severity of silicosis, as reflected by significantly reduced DLCO in such patients.

Patient and treatment characteristics of emergency presentations due to immune-mediated toxicities.

BACKGROUND: The prevalence of immune-mediated toxicities from immune checkpoint inhibitors (ICIs) is well described. However, the characteristics and treatment patterns for patients with emergency presentations owing to immune-mediated toxicity are less well known. MATERIALS AND METHODS: This study reviews all emergency presentations in patients treated with ICI at a single centre between May 2018 and March 2020. The aims were to describe and quantify patient and treatment characteristics, toxicity type and outcomes. RESULTS: 1165 patients were treated with ICI, and there were 597 emergency presentations in 370 patients. Of these, 191/597 (32%) were owing to an immune-mediated toxicity, median age was 64 years, and 127/191 (67%) were men. The most common tumour types were melanoma (53%) and lung cancer (22%), and the most common ICI received was ipilimumab + nivolumab combination immunotherapy (42%), followed by pembrolizumab monotherapy (21%) and nivolumab monotherapy (20%). The median number of cycles received was three (range 1-54), and 75/191 (39%) had previous ≥ grade 2 immune-mediated toxicity. 29% patients required second-line immunosuppression. The median time in the hospital was four days. There was a rising number of emergency presentations reflecting overall increasing use of ICI. CONCLUSIONS: Over a quarter of patients treated with ICI had an emergency presentation, and immune-mediated toxicity accounted for 32% of these. A diagnosis of melanoma, treatment with combination immunotherapy and previous ≥ grade 2 immune-mediated toxicity were common in patients with immune-mediated toxicity. These data allow better identification of patients likely to require admission and forward planning for acute oncology services.

Improved pyrexia-related outcomes associated with an adapted pyrexia adverse event management algorithm in patients treated with adjuvant dabrafenib plus trametinib: Primary results of COMBI-APlus.

BACKGROUND: COMBI-AD demonstrated long-term benefit of adjuvant dabrafenib plus trametinib in patients with resected stage III BRAF V600E/K-mutant melanoma; however, 9% of patients permanently discontinued therapy due to pyrexia. COMBI-APlus evaluated whether an adapted pyrexia management algorithm reduces high-grade pyrexia and pyrexia-related adverse outcomes. METHODS: COMBI-APlus is an open-label, phase IIIb trial evaluating an adapted pyrexia management algorithm in patients with high-risk resected stage III BRAF V600E/K-mutant melanoma treated with up to 12 months of adjuvant dabrafenib plus trametinib. Both drugs were interrupted for pyrexia (temperature ≥38°C) or the occurrence of pyrexia syndrome for suspected recurrent pyrexia. Treatment was restarted at the same dose once patients were symptom free for ≥24 h. The primary endpoint was the composite rate of grade 3/4 pyrexia, hospitalisation due to pyrexia, or permanent discontinuation due to pyrexia versus historical COMBI-AD control (20.0%; 95% confidence interval [CI], 16.3%-24.1%). RESULTS: At data cutoff (5 October 2020), COMBI-APlus met its primary endpoint of significant improvement in the composite rate of pyrexia (8.0% [95% CI, 5.9%-10.6%]), with rates of 3.8% for grade 3/4 pyrexia, 4.3% for hospitalisation due to pyrexia, and 2.4% for discontinuation due to pyrexia. Estimated 12-month relapse-free survival was 91.8% (95% CI, 89.0%-93.9%). The most common adverse events were consistent with those in COMBI-AD, and 14.7% of patients permanently discontinued treatment due to adverse events. CONCLUSIONS: The adapted pyrexia management algorithm appears to reduce the incidence of severe pyrexia outcomes, enables patients to manage pyrexia at home, and helps patients remain on treatment. CLINICAL TRIAL REGISTRATION: NCT03551626.