Neuropsychological outcome after cardiac arrest: a prospective case control sub-study of the Targeted hypothermia versus targeted normothermia after out-of-hospital cardiac arrest trial (TTM2)

Background: This study is designed to provide detailed knowledge on cognitive impairment after out-of-hospital cardiac arrest (OHCA) and its relation to associated factors, and to validate the neurocognitive screening of the Targeted Hypothermia versus Targeted Normothermia after Out-of-Hospital Cardiac Arrest trial (TTM2-trial), assessing effectiveness of targeted temperature management after OHCA. Methods: This longitudinal multi-center clinical study is a sub-study of the TTM2-trial, in which a comprehensive neuropsychological examination is performed in addition to the main TTM2-trial neurocognitive screening. Approximately 7 and 24 months after OHCA, survivors at selected study sites are invited to a standardized assessment, including performance-based tests of cognition and questionnaires of emotional problems, fatigue, executive function and insomnia. At 1:1 ratio, a matched control group from a cohort of acute myocardial infarction (MI) patients is recruited to perform the same assessment. We aim to include 100 patients per group. Potential differences between the OHCA patients and the MI controls at 7 and 24 months will be analyzed with a linear regression, using composite z-scores per cognitive domain (verbal, visual/constructive, working memory, episodic memory, processing speed, executive functions) as primary outcome measures. Results from OHCA survivors on the main TTM2-trial neurocognitive screening battery will be compared with neuropsychological test results at 7 months, using sensitivity and specificity analyses. Discussion: In this study we collect detailed information on cognitive impairment after OHCA and compare this to a control group of patients with acute MI. The validation of the TTM2 neurocognitive screening battery could justify its inclusion in routine follow-up. Our results may have a potential to impact on the design of future follow-up strategies and interventions after OHCA

Effects of Hypothermia vs Normothermia on Societal Participation and Cognitive Function at 6 Months in Survivors After Out-of-Hospital Cardiac Arrest

ImportanceThe Targeted Hypothermia vs Targeted Normothermia After Out-of-Hospital Cardiac Arrest (TTM2) trial reported no difference in mortality or poor functional outcome at 6 months after out-of-hospital cardiac arrest (OHCA). This predefined exploratory analysis provides more detailed estimation of brain dysfunction for the comparison of the 2 intervention regimens.ObjectivesTo investigate the effects of targeted hypothermia vs targeted normothermia on functional outcome with focus on societal participation and cognitive function in survivors 6 months after OHCA.Design, Setting, and ParticipantsThis study is a predefined analysis of an international multicenter, randomized clinical trial that took place from November 2017 to January 2020 and included participants at 61 hospitals in 14 countries. A structured follow-up for survivors performed at 6 months was by masked outcome assessors. The last follow-up took place in October 2020. Participants included 1861 adult (older than 18 years) patients with OHCA who were comatose at hospital admission. At 6 months, 939 of 1861 were alive and invited to a follow-up, of which 103 of 939 declined or were missing.InterventionsRandomization 1:1 to temperature control with targeted hypothermia at 33 °C or targeted normothermia and early treatment of fever (37.8 °C or higher).Main outcomes and measuresFunctional outcome focusing on societal participation assessed by the Glasgow Outcome Scale Extended ([GOSE] 1 to 8) and cognitive function assessed by the Montreal Cognitive Assessment ([MoCA] 0 to 30) and the Symbol Digit Modalities Test ([SDMT] z scores). Higher scores represent better outcomes.ResultsAt 6 months, 836 of 939 survivors with a mean age of 60 (SD, 13) (range, 18 to 88) years (700 of 836 male [84%]) participated in the follow-up. There were no differences between the 2 intervention groups in functional outcome focusing on societal participation (GOSE score, odds ratio, 0.91; 95% CI, 0.71-1.17; P = .46) or in cognitive function by MoCA (mean difference, 0.36; 95% CI,−0.33 to 1.05; P = .37) and SDMT (mean difference, 0.06; 95% CI,−0.16 to 0.27; P = .62). Limitations in societal participation (GOSE score less than 7) were common regardless of intervention (hypothermia, 178 of 415 [43%]; normothermia, 168 of 419 [40%]). Cognitive impairment was identified in 353 of 599 survivors (59%).ConclusionsIn this predefined analysis of comatose patients after OHCA, hypothermia did not lead to better functional outcome assessed with a focus on societal participation and cognitive function than management with normothermia. At 6 months, many survivors had not regained their pre-arrest activities and roles, and mild cognitive dysfunction was common.Trial RegistrationClinicalTrials.gov Identifier: NCT0290830

Biomarkers of brain injury after cardiac arrest; a statistical analysis plan from the TTM2 trial biobank investigators

Background: Several biochemical markers in blood correlate with the magnitude of brain injury and may be used to predict neurological outcome after cardiac arrest. We present a protocol for the evaluation of prognostic accuracy of brain injury markers after cardiac arrest. The aim is to define the best predictive marker and to establish clinically useful cut-off levels for routine implementation. Methods: Prospective international multicenter trial within the Targeted Hypothermia versus Targeted Normothermia after Out-of-Hospital Cardiac Arrest (TTM2) trial in collaboration with Roche Diagnostics International AG. Samples were collected 0, 24, 48, and 72 hours after randomisation (serum) and 0 and 48 hours after randomisation (plasma), and pre-analytically processed at each site before storage in a central biobank. Routine markers neuron-specific enolase (NSE) and S100B, and neurofilament light, total-tau and glial fibrillary acidic protein will be batch analysed using novel Elecsys® electrochemiluminescence immunoassays on a Cobas e601 instrument. Results: Statistical analysis will be reported according to the Standards for Reporting Diagnostic accuracy studies (STARD) and will include comparisons for prediction of good versus poor functional outcome at six months post-arrest, by modified Rankin Scale (0–3 vs. 4–6), using logistic regression models and receiver operating characteristics curves, evaluation of mortality at six months according to biomarker levels and establishment of cut-off values for prediction of poor neurological outcome at 95–100% specificities. Conclusions: This prospective trial may establish a standard methodology and clinically appropriate cut-off levels for the optimal biomarker of brain injury which predicts poor neurological outcome after cardiac arrest

Origins and genetic legacy of prehistoric dogs

Dogs were the first domestic animal, but little is known about their population history and to what extent it was linked to humans. We sequenced 27 ancient dog genomes and found that all dogs share a common ancestry distinct from present-day wolves, with limited gene flow from wolves since domestication but substantial dog-to-wolf gene flow. By 11,000 years ago, at least five major ancestry lineages had diversified, demonstrating a deep genetic history of dogs during the Paleolithic. Coanalysis with human genomes reveals aspects of dog population history that mirror humans, including Levant-related ancestry in Africa and early agricultural Europe. Other aspects differ, including the impacts of steppe pastoralist expansions in West and East Eurasia and a near-complete turnover of Neolithic European dog ancestry

Cell-free DNA profiling of metastatic prostate cancer reveals microsatellite instability, structural rearrangements and clonal hematopoiesis.

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.BACKGROUND: There are multiple existing and emerging therapeutic avenues for metastatic prostate cancer, with a common denominator, which is the need for predictive biomarkers. Circulating tumor DNA (ctDNA) has the potential to cost-efficiently accelerate precision medicine trials to improve clinical efficacy and diminish costs and toxicity. However, comprehensive ctDNA profiling in metastatic prostate cancer to date has been limited. METHODS: A combination of targeted and low-pass whole genome sequencing was performed on plasma cell-free DNA and matched white blood cell germline DNA in 364 blood samples from 217 metastatic prostate cancer patients. RESULTS: ctDNA was detected in 85.9% of baseline samples, correlated to line of therapy and was mirrored by circulating tumor cell enumeration of synchronous blood samples. Comprehensive profiling of the androgen receptor (AR) revealed a continuous increase in the fraction of patients with intra-AR structural variation, from 15.4% during first-line metastatic castration-resistant prostate cancer therapy to 45.2% in fourth line, indicating a continuous evolution of AR during the course of the disease. Patients displayed frequent alterations in DNA repair deficiency genes (18.0%). Additionally, the microsatellite instability phenotype was identified in 3.81% of eligible samples (≥ 0.1 ctDNA fraction). Sequencing of non-repetitive intronic and exonic regions of PTEN, RB1, and TP53 detected biallelic inactivation in 47.5%, 20.3%, and 44.1% of samples with ≥ 0.2 ctDNA fraction, respectively. Only one patient carried a clonal high-impact variant without a detectable second hit. Intronic high-impact structural variation was twice as common as exonic mutations in PTEN and RB1. Finally, 14.6% of patients presented false positive variants due to clonal hematopoiesis, commonly ignored in commercially available assays. CONCLUSIONS: ctDNA profiles appear to mirror the genomic landscape of metastatic prostate cancer tissue and may cost-efficiently provide somatic information in clinical trials designed to identify predictive biomarkers. However, intronic sequencing of the interrogated tumor suppressors challenges the ubiquitous focus on coding regions and is vital, together with profiling of synchronous white blood cells, to minimize erroneous assignments which in turn may confound results and impede true associations in clinical trials.The Belgian Foundation Against Cancer (grant number C/2014/227); Kom op tegen Kanker (Stand up to Cancer), the Flemish Cancer Society (grant number 00000000116000000206); Royal College of Surgeons/Cancer Research UK (C19198/A1533); The Cancer Research Funds of Radiumhemmet, through the PCM program at KI (grant number 163012); The Erling-Persson family foundation (grant number 4-2689-2016); the Swedish Research Council (grant number K2010-70X-20430-04-3), and the Swedish Cancer Foundation (grant number 09-0677)

Strategies to improve cancer radioimmunotargeting

Radioimmunotherapy (RIT) and radioimmunolocalisation (RIL) are developing and promising technologies to diagnose and treat tumours by use of radiolabelled antibodies targeting tumour specific antigens. The major reason why RIL and RIT not are efficient enough, is the comparatively low accumulation of radiolabelled antibodies in the tumours. Irrespective of the antigen - antibody system used, the maximal tumour uptake in humans is often limited to below 0.1 % of the total injected dose, with significant radionuclide remaining in the blood pool and extravascular fluid. In the present thesis, the following putative improvement techniques for radioimmunotargeting have been evaluated in an experimental model using HeLa cell-xenografted nude mice: 1) Repetitive, simultaneous targeting of different antigens, 2) Removal of non-targeting antibodies using secondary antiidiotypic antibodies, 3) Preinjection of unlabelled antibody to remove shedded antigen and 4) Use of fractionated antibody administration. By use of multiple injections of mixtures of two different 131I-labelled monoclonal antibodies targeting placental alkaline phosphatase (H7) and cytokeratin 8 (TS1), respectively, a significant tumour growth inhibition compared to controls, was obtained. In the treated group, a negligible increase in tumour volume was seen compared to the control group, in which a 20-fold increase was observed. Quantitative determinations of volume densities of viable tumour cells, necrotic cells and connective tissue demonstrated no significant differences in the relative proportions between the groups, indicating that the irradiation caused decelerated growth. Using hybridoma technology, monoclonal antiidiotypic antibodies were generated against both TS1 and H7. The in vitro and in vivo effects of these antibodies, aH7 and aTSl, were investigated. Both these antiidiotypes were found to generate stable complexes with the radiolabelled idiotypic antibody, as revealed by gel-electrophoresis and autoradiography. Using biosensor technology (BIAcore, Pharmacia) the interactions were followed in real time and the association rate-, dissociation rate-, and affinity constants between the reactants were determined. In vivo, the antiidiotypes promoted a rapid dose dependent clearance of the 125I-labelled idiotypes with a decrease in total body radioactivity and concomitant dramatic increase in non-protein bound 125I excreted in the urine. The syngeneic monoclonal antiidiotypic antibody αTSl, was furthermore evaluated as a secondary clearing antibody at radioimmunolocalisation. Injection of αTSl in a molar ratio of 0.5-0.75:1 to TS1, 24 hours after the 125I-labelled TS1 improved the tumour to normal tissue ratio 2-3 fold. This was due to a decreased level of total body radioactivity as well as a slight decrease in tumour-radioactivity. A model describing the kinetics of the involved components, i.e. the antigen, the idiotype and the antiidiotype was presented. It is concluded that high affinity monoclonal antiidiotypes can be used as tools to regulate the levels of idiotypic antibodies in vivo. This strategy, combined with preinjection of non­labelled idiotypic antibodies, caused accumulated doses of 3 Gy to the tumour and 0.9 Gy to non tumour tissues as calculated for 125I-labelled antibodies (80 MBq/mg) by MIRD formalism based on repetitive quantitative radioimmunoscintigraphies. By approaching the maximal tolerated whole body radiation dose for mice (i.e. 6 Gy), it can be estimated that doses up to 20 Gy are possible to obtain following one single injection of labelled antibody. It was furthermore demonstrated that a single bolus injection of antibody is to be preferred, compared to exactly the same dose divided into three or ten fractions. Thus, not only the dose of radioactivity, but also the amount of antibody should be considered for fractionated RIT. In summary, the thesis demonstrates that several techniques can be used to improve radioimmunolocalisation and to approach the proposed 70 Gy required to sterilise tumours at [email protected]

Strategies to improve cancer radioimmunotargeting

Radioimmunotherapy (RIT) and radioimmunolocalisation (RIL) are developing and promising technologies to diagnose and treat tumours by use of radiolabelled antibodies targeting tumour specific antigens. The major reason why RIL and RIT not are efficient enough, is the comparatively low accumulation of radiolabelled antibodies in the tumours. Irrespective of the antigen - antibody system used, the maximal tumour uptake in humans is often limited to below 0.1 % of the total injected dose, with significant radionuclide remaining in the blood pool and extravascular fluid. In the present thesis, the following putative improvement techniques for radioimmunotargeting have been evaluated in an experimental model using HeLa cell-xenografted nude mice: 1) Repetitive, simultaneous targeting of different antigens, 2) Removal of non-targeting antibodies using secondary antiidiotypic antibodies, 3) Preinjection of unlabelled antibody to remove shedded antigen and 4) Use of fractionated antibody administration. By use of multiple injections of mixtures of two different 131I-labelled monoclonal antibodies targeting placental alkaline phosphatase (H7) and cytokeratin 8 (TS1), respectively, a significant tumour growth inhibition compared to controls, was obtained. In the treated group, a negligible increase in tumour volume was seen compared to the control group, in which a 20-fold increase was observed. Quantitative determinations of volume densities of viable tumour cells, necrotic cells and connective tissue demonstrated no significant differences in the relative proportions between the groups, indicating that the irradiation caused decelerated growth. Using hybridoma technology, monoclonal antiidiotypic antibodies were generated against both TS1 and H7. The in vitro and in vivo effects of these antibodies, aH7 and aTSl, were investigated. Both these antiidiotypes were found to generate stable complexes with the radiolabelled idiotypic antibody, as revealed by gel-electrophoresis and autoradiography. Using biosensor technology (BIAcore, Pharmacia) the interactions were followed in real time and the association rate-, dissociation rate-, and affinity constants between the reactants were determined. In vivo, the antiidiotypes promoted a rapid dose dependent clearance of the 125I-labelled idiotypes with a decrease in total body radioactivity and concomitant dramatic increase in non-protein bound 125I excreted in the urine. The syngeneic monoclonal antiidiotypic antibody αTSl, was furthermore evaluated as a secondary clearing antibody at radioimmunolocalisation. Injection of αTSl in a molar ratio of 0.5-0.75:1 to TS1, 24 hours after the 125I-labelled TS1 improved the tumour to normal tissue ratio 2-3 fold. This was due to a decreased level of total body radioactivity as well as a slight decrease in tumour-radioactivity. A model describing the kinetics of the involved components, i.e. the antigen, the idiotype and the antiidiotype was presented. It is concluded that high affinity monoclonal antiidiotypes can be used as tools to regulate the levels of idiotypic antibodies in vivo. This strategy, combined with preinjection of non­labelled idiotypic antibodies, caused accumulated doses of 3 Gy to the tumour and 0.9 Gy to non tumour tissues as calculated for 125I-labelled antibodies (80 MBq/mg) by MIRD formalism based on repetitive quantitative radioimmunoscintigraphies. By approaching the maximal tolerated whole body radiation dose for mice (i.e. 6 Gy), it can be estimated that doses up to 20 Gy are possible to obtain following one single injection of labelled antibody. It was furthermore demonstrated that a single bolus injection of antibody is to be preferred, compared to exactly the same dose divided into three or ten fractions. Thus, not only the dose of radioactivity, but also the amount of antibody should be considered for fractionated RIT. In summary, the thesis demonstrates that several techniques can be used to improve radioimmunolocalisation and to approach the proposed 70 Gy required to sterilise tumours at [email protected]