Traditional Endpoints Research Articles

Association of the AAV-PRO questionnaire with established outcome measures in AAV.

The ANCA-associated vasculitis (AAV) patient-reported outcome (AAV-PRO) questionnaire was developed to capture the impact of AAV and its treatment. We investigated the association of specific AAV-PRO domains with disease activity and extent, damage, depression, health-related quality of life, and treatment. In a prospective longitudinal study, AAV-PRO, Beck's depression inventory (BDI), Short Form 36 (SF-36), BVAS and Vasculitis Damage Index (VDI) were completed at baseline (t1) and after 3-6 months (t2). In addition, patient data (including diagnosis, therapies, relapses, and organ manifestations) were recorded. Data were analysed by t-tests and correlation-based regression analyses. A total of 156 patients with AAV participated. The mean BVAS at the time of enrolment was 1.4 ± 3.74. The median AAV-PRO domain scores were higher in patients reporting 'active disease' compared with those reporting 'in remission' (P < 0.001). In the correlation analyses, all AAV-PRO domain scores correlated strongly with the BDI (all r ≥ 0.319, all P ≤ 0.001) as well as with all eight SF-36 subdomains (all |r|≥0.267, all P ≤ 0.001). The regression analyses showed that AAV-PRO domains were strongly predicted by the BDI and SF-36 domains (|β| ≥ 0.240 for the strongest predictor of each domain). In the longitudinal comparison (t1/t2), there were no significant changes in the overall results. Our data show convergent validity for all AAV-PRO subdomains, using the established questionnaires BDI and SF-36. The AAV-PRO domains scores were not correlated with clinician-derived instruments (including the BVAS and the VDI). Thus, we regard the AAV-PRO questionnaire as a valuable measure of outcomes that might complement traditional end-points in clinical trials.

Re-Irradiation for Recurrent Head and Neck Cancer: Freedom from Cancer Recurrence Rate.

Salvage re-irradiation (rRT) for patients with locoregionally recurrent head and neck cancer (rHNC) remains challenging. A retrospective analysis was performed on 49 patients who received rRT between 2011 and 2018. The co-primary endpoint of the study was 2-year freedom from cancer recurrence rate (FCRR) and overall survival (OS), and secondary endpoints were 2-year disease-free survival (DFS), local failure (LF), regional failure (RF), distant metastases (DM), and RTOG grade 3 ≥ late toxicities. Adjuvant and definitive rRT were delivered to 22 and 27 patients, respectively. A total of 91% of patients were managed with conventional re-RT and 71% of patients received concurrent chemotherapy. The median follow-up after rRT was 30 months. The 2-year FCRR, OS, DFS, LF, RF, and DM were 64%, 51%, 28%, 32%, 9%, and 39% respectively. MVA showed that poor performance status (PS: 1-2 vs. 0) and age > 52 years were predictive of worse OS. In comparison, poor PS (1-2 vs. 0) and total dose of rRT < 60 Gy were predictive of worse DFS. Late RTOG toxicity of grade 3 ≥ was reported in nine (18.3%) patients. FCRR at 2 years after salvage rRT for rHNC was higher than other traditional endpoints and could be an important endpoint to be included in future rRT studies. rRT for rHNC at our cohort was relatively successful, with a manageable level of late severe toxicity. Replacing this approach in other developing countries is a viable option.

Patient-identified Treatment Attributes Among Older Men With Stress Urinary Incontinence: A Qualitative Look at What Matters to Patients Making Treatment Decisions

To investigate which treatment attributes matter to patients with stress urinary incontinence (SUI), why and how they matter, and the context in which patients consider treatment attributes. Nearly a quarter of older men have decisional regret following SUI treatment. Knowledge of what matters to patients when making SUI treatment decisions is necessary to improve goal-concordant care. We conducted semi-structured interviews with 36 men ≥65 years of age with SUI. Semi-structured interviews were conducted by telephone and transcribed. Four researchers (L.H., N.S., E.A., C.B.) coded the transcripts using both deductive and inductive codes to identify and describe treatment attributes. We identified 5 patient-derived treatment attributes of interest among older men with SUI who have faced treatment decisions: (1) dryness, (2) simplicity, (3) potential need for future intervention, (4) treatment regret/satisfaction, and (5) surgical avoidance. These themes reliably emerged in our patient-centered interviews from within various contexts, including prior negative healthcare experiences, the impact of incontinence on daily and quality of life, and the mental health burden of incontinence, among others. Men with SUI weigh a variety of treatment attributes in addition to dryness, a traditional clinical endpoint, and do so within the context of their individual experience. These additional attributes, such as simplicity, may run counter to the goal of dryness. This suggests that traditional clinical endpoints alone are not adequate for counseling patients. Contextualized patient-identified treatment attributes should be used to create decision-support materials to promote goal-concordant SUI treatment.

Real-World Experience of PD-1 Inhibitors for Relapsed and Refractory Hodgkin Lymphoma: A Multicenter Retrospective Analysis of Patients in China

Introduction: Despite the promising clinical trial data regarding programmed death 1 (PD-1) inhibitors in relapsed/refractory classical Hodgkin lymphoma (R/R cHL), there remains a paucity of studies describing the outcomes of patients in a real-world setting, especially for Asian cohort. Methods: We present a multicenter retrospective analysis of patients with R/R cHL who had failed ≥2 prior lines of therapies and received sintilimab or tislelizumab developed in China (sintilimab or tislelizumab monotherapy) at 3 medical centers from January 2019 to September 2021. Efficacy was evaluated with progression-free survival (PFS), overall survival, duration of response (DOR), best overall response (BOR) including objective response rate (ORR), complete response rate (CRR). Safety data were also recorded. Results: 74 patients were reviewed. The median age was 38 years (range, 14–85 years). The ORR, CRR, and disease control rate were 78.3%, 52.7%, and 91.9%, respectively. The median duration of follow-up was 22 (4–36) months. Four patients (5.4%) died of disease progression. The median PFS and DOR was 22.1 and 23.5 months. BOR as a new emergent endpoint was found to be the only independent prognostic factor for PFS in our study (HR = 6.234, p = 0.005), suggesting this endpoint carries stronger prognostic value over traditional endpoints in the immunotherapy era. 66 (89.2%) patients reported adverse event (AE) with any grade, with the majority of AEs being grade 1 or 2. Conclusion: We presented a unique real-life experience and conducted a relatively long follow-up of PD-1 antibodies developed in China for R/R HL patients which confirmed their promising effectiveness and manageable side effects given in real world in an Asian cohort. Even for those who would usually be excluded in most of clinical trials such as elderly or minor patients, anti-PD-1 monotherapy also showed a significant improvement of outcomes. Furthermore, the depth of response seemed to be a more powerful predictive tool in new era, which might serve as a basis for future immune risk-adapted strategies.

Changes in metabolic profiles of amphipods Allorchestes compressa after acute exposures to copper, pyrene, and their mixtures

Amphipods are ideal indicators for biomonitoring and ecotoxicological studies of environmental contaminants because they are extensively distributed in aquatic environments, are easy to collect and are important in nutrient cycling. Marine amphipods (Allorchestes compressa) were exposed to two concentrations of copper and pyrene, and their mixtures, for 24 and 48 h. Changes in polar metabolites were assessed using Gas Chromatography Mass Spectrometry (GC-MS)-based untargeted metabolomics. Generally, limited metabolite changes were observed for copper and pyrene single exposures (eight and two significant metabolites, respectively), while 28 metabolites had changed following exposures to mixtures. Furthermore, changes were mainly observed after 24 h but had seemingly returned to control levels after 48 h. Multiple types of metabolites were affected including amino acids, Tricarboxylic acid (TCA) cycle intermediates, sugars, fatty acids, and hormones. This study highlights the sensitivity of metabolomics in assessing the impacts of low concentrations of chemicals compared to traditional ecotoxicological endpoints.

Electrochemical detection of the p53 gene using exponential amplification reaction (EXPAR) and CRISPR/Cas12a reactions.

Animproved electrochemical sensor has been developed for sensitive detection of the p53 genebased on exponential amplification reaction (EXPAR) and CRISPR/Cas12a. Restriction endonuclease BstNI is introduced to specifically identify and cleave the p53 gene, generating primers to trigger the EXPAR cascade amplification. A large number of amplified products are then obtained to enable the lateral cleavage activity of CRISPR/Cas12a. For electrochemical detection, the amplified product activates Cas12a to digest the designed block probe, which allows the signal probe to be captured by the reduced graphene oxide-modified electrode (GCE/RGO), resulting in an enhanced electrochemical signal. Notably, the signal probe is labeled with large amounts of methylene blue (MB). Compared with traditional endpoint decoration, the special signal probe effectively amplifies the electrochemical signalsby a factor of about 15.Experimental results show that the electrochemical sensor exhibits wide ranges from 500 aM to 10pM and 10pM to 1nM, as well as a relatively low limit detection of 0.39 fM, which is aboutan order of magnitude lowerthan that of fluorescence detection. Moreover, the proposed sensor shows reliable application capability in real human serum, indicating that this work has great prospects for the construction of a CRISPR-based ultra-sensitive detection platform.

Daphnia magna sub-lethal exposure to phthalate pollutants elicits disruptions in amino acid and energy metabolism

Phthalic acid esters (PAEs) are a class of chemicals that are usually incorporated as additives in the manufacturing of plastics. PAEs are not covalently bound to the material matrix and can, consequently, be leached into the environment. PAEs have been reported to act as endocrine disruptors, neurotoxins, metabolic stressors, and immunotoxins to aquatic organisms but there is a lack of information regarding the impact of sub-lethal concentrations to target organisms. The freshwater crustacean Daphnia magna, a commonly used model organism in aquatic toxicity, was exposed to four phthalate pollutants: dimethyl phthalate (DMP), diethyl phthalate (DEP), monomethyl phthalate (MMP), and monoethyl phthalate (MEP). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed in a targeted metabolomic approach to quantify polar metabolites extracted from a single Daphnia body. Individual metabolite percent changes and hierarchical clustering heatmap analysis showed unique metabolic profiles for each phthalate pollutant. Metabolite percent changes were mostly downregulated or presented opposing responses for the low and high concentrations tested. Meanwhile, pathway analyses suggest the disruption of related and unique pathways, mostly connected with amino acid and energy metabolism. The pathways aminoacyl-tRNA biosynthesis, arginine biosynthesis, and glutathione metabolism were disrupted by most selected PAEs. Overall, this study indicates that although phthalate pollutants can elicit distinct metabolic perturbations to each PAE, they still impacted related biochemical pathways. These chemical-class based responses could be associated with a common toxic mechanism of action. The reported findings show how targeted metabolomic approaches can lead to a better understanding of sub-lethal exposure to pollutants, revealing metabolomic endpoints do not hold a close relationship with traditional acute toxicity endpoints.

Derivation of metabolic point of departure using high-throughput in vitro metabolomics: investigating the importance of sampling time points on benchmark concentration values in the HepaRG cell line

Amongst omics technologies, metabolomics should have particular value in regulatory toxicology as the measurement of the molecular phenotype is the closest to traditional apical endpoints, whilst offering mechanistic insights into the biological perturbations. Despite this, the application of untargeted metabolomics for point-of-departure (POD) derivation via benchmark concentration (BMC) modelling is still a relatively unexplored area. In this study, a high-throughput workflow was applied to derive PODs associated with a chemical exposure by measuring the intracellular metabolome of the HepaRG cell line following treatment with one of four chemicals (aflatoxin B1, benzo[a]pyrene, cyclosporin A, or rotenone), each at seven concentrations (aflatoxin B1, benzo[a]pyrene, cyclosporin A: from 0.2048 μM to 50 μM; rotenone: from 0.04096 to 10 μM) and five sampling time points (2, 6, 12, 24 and 48 h). The study explored three approaches to derive PODs using benchmark concentration modelling applied to single features in the metabolomics datasets or annotated metabolites or lipids: (1) the 1st rank-ordered unannotated feature, (2) the 1st rank-ordered putatively annotated feature (using a recently developed HepaRG-specific library of polar metabolites and lipids), and (3) 25th rank-ordered feature, demonstrating that for three out of four chemical datasets all of these approaches led to relatively consistent BMC values, varying less than tenfold across the methods. In addition, using the 1st rank-ordered unannotated feature it was possible to investigate temporal trends in the datasets, which were shown to be chemical specific. Furthermore, a possible integration of metabolomics-driven POD derivation with the liver steatosis adverse outcome pathway (AOP) was demonstrated. The study highlights that advances in technologies enable application of in vitro metabolomics at scale; however, greater confidence in metabolite identification is required to ensure PODs are mechanistically anchored.

Surface Defects Regulate the in Vivo Bioenergetic Response of Earthworm Eisenia fetida Coelomocytes to Molybdenum Disulfide Nanosheets.

Two-dimensional molybdenum disulfide (2D MoS2) nanomaterials are seeing increased use in several areas, and this will lead to their inevitable release into soils. Surface defects can occur on MoS2 nanosheets during synthesis or during environmental aging processes. The mechanisms of MoS2 nanosheet toxicity to soil invertebrates and the role of surface defects in that toxicity have not been fully elucidated. We integrated traditional toxicity end points, targeted energy metabolomics, and transcriptomics to compare the mechanistic differences in the toxicity of defect-free and defect-rich MoS2 nanosheets (DF-MoS2 and DR-MoS2) to Eisenia fetida using a coelomocyte-based in vivo assessment model. After organism-level exposure to DF-MoS2 for 96 h at 10 and 100 mg Mo/L, cellular reactive oxygen species (ROS) levels were elevated by 25.6-96.6% and the activity of mitochondrial respiratory electron transport chain (Mito-RETC) complex III was inhibited by 9.7-19.4%. The tricarboxylic acid cycling and glycolysis were also disrupted. DF-MoS2 preferentially up-regulated subcellular component motility processes related to microtubules and caused mitochondrial fission. Unlike DF-MoS2, DR-MoS2 triggered an increased degree of mitochondrial fusion, as well as more severe oxidative stress. The activities of Mito-RETC complexes (I, III, IV, V) associated with oxidative phosphorylation were significantly inhibited by 22.8-68.6%. Meanwhile, apoptotic pathways were activated upon DR-MoS2 exposure, which together with the depolarization of mitochondrial membrane potential, mediated significant apoptosis. In turn, genes related to cellular homeostasis and energy release were up-regulated to compensate for DR-MoS2-induced energy deprivation. Our study indicates that MoS2 nanosheets have nanospecific effects on E. fetida and also that the role of surface defects from synthesis or that accumulate from environmental impacts needs to be fully considered when evaluating the toxicity of these 2D materials.

Tough times never last too long: the future of psychopharmacology.

Tough times never last too long: the future of psychopharmacology.

Modeling spontaneous opioid withdrawal in male and female outbred mice using traditional endpoints and hyperalgesia.

Opioid withdrawal significantly impacts drug dependence cycles as hyperalgesia associated with withdrawal is often a reason for continued drug use. Animal models of addiction are important tools for studying how drug dependence and withdrawal impact not only normal neurocircuitry but also the effectiveness of potential treatments for dependence and withdrawal. We conducted a study of the time course of spontaneous morphine withdrawal in outbred male and female mice that can be used to examine sex differences in male and female mice using both traditional somatic endpoints and mechanical hyperalgesia as an endpoint of withdrawal. Male and female national institute of health (NIH) Swiss mice were made dependent upon morphine using an escalating dosing schedule. Injections were stopped after 5 days. Withdrawal behavior was assessed at time intervals up to 106 h after the final injection. Numbers of forepaw tremors, wet-dog shakes, jumps and other behaviors were scored to create a global score. Paw pressure readings were then also taken to track changes in sensitivity to a painful stimulus over time. Male and female mice had approximately similar withdrawal severity peaking at 24 h after the final injection as measured by composite global scores. Females did exhibit an earlier and greater frequency of tremors than males. Although males and females showed similar hyperalgesia during withdrawal, females recovered faster. Spontaneous opioid withdrawal peaking at 24 h was demonstrated in male and female NIH Swiss mice. We also successfully demonstrated that hyperalgesia is an endpoint that varies over the course of withdrawal.

Understanding hazardous concentrations of microplastics in fresh water using non-traditional toxicity data

Microplastic pollution has become a major environmental problem, indicating the need to implement quantitative governance standards in combination with reducing or banning single-use plastic. Previous studies have predicted no-effect concentrations for limited microplastic-based toxicity data but have not considered environmentally relevant sizes, shapes, or polymers. To provide high quantity and quality data for microplastics of different sizes, shapes, or polymer compositions, non-traditional and traditional toxicity data may need to be considered in combination. In this study, we reviewed toxicity data for microplastics in freshwaters from 2018 to 2022 and analyzed the toxicity data using traditional and non-traditional methods. Based on 166 chronic traditional toxicity data points, the hazard concentration (HC) values calculated from non-traditional toxicity endpoints or all toxicity endpoints were lower than those calculated from traditional toxicity endpoints. Based on 398 chronic traditional plus non-traditional toxicity data points, the HC values calculated from traditional plus non-traditional values were higher than those calculated from traditional toxicity values. With these results, we developed a new framework for deriving microplastic-specific hazardous concentrations, one that especially considers non-traditional toxicity endpoints and values for microplastics. Overall, this study offers a basis for future management strategies and associated frameworks for mitigating microplastic toxicity.

HPR34 Emerging Role of Patient Reported Outcomes in Drug Approval in Europe

Rapid development in digital technology has offered vast metrics to strengthen patient-reported outcome (PRO) databases. Globally, PRO endpoints have gained momentum in addition to traditional clinical trial endpoints, thereby integrating patients’ perspective into drug approvals. The aim of this research is to evaluate the role of PRO endpoints in decision making process of European Medicines Agency (EMA) for drug approval.

Instrumental Activities of Daily Living Tools in Very-Low Vision: Ready for Use in Trials?

Traditional endpoints assessing visual function are limited by their responsiveness to interventions restoring or maintaining vision. An alternative concept is assessing instrumental activities of daily living (IADL). Herein, we review all available vision-specific IADL instruments relevant for vision restoration trials and report data for the most promising instrument. Six relevant instruments exist: The Low Vision Functional Status Evaluation (LVFSE), Timed IADL (TIADL), Melbourne Low-Vision Activities of Daily Living Index (MLVAI), Assessment of Disability Related to Vision (ADREV), Functional Low-Vision Observer Rated Assessment (FLORA), and Very Low Vision IADL (IADL-VLV). Both internal consistency and test-retest data were available for the LVFSE, MLVAI, and IADL-VLV. In a sample from a low-vision clinic (n = 51; age 57 ± 16 years), we report additional validation data on the IVI-VLV including test-retest reliability (intraclass correlation coefficient 0.981 [0.961; 0.991]). The LVSFE was noticeably less reliable than the MLVAI and the IADL-VLV. Content and construct validity data were available for the LVFSE, TIADL, MLVAI, ADREV, and IADL-VLV, but only the MLVAI and IADL-VLV were developed for an ultra-low vision context. Ceiling effects were present across instruments. Thus, of all appropriate IADL instruments related to vision, the IADL-VLV and MLVAI best meet existing requirements for use in vision restoration trials, e.g., in gene therapies or visual prostheses in inherited retinal diseases, but require further validation.

A holistic NMR framework to understand environmental impact: Examining the impacts of superparamagnetic iron oxide nanoparticles (SPIONs) in Daphnia magna via imaging, spectroscopy, and metabolomics.

Superparamagnetic iron oxide nanoparticles (SPIONs) are a contaminant of emerging interest, often used in the medical field as an imaging contrast agent, with additional uses in wastewater treatment and as food additives. Although the use of SPIONs is increasing, little research has been conducted on the toxic impacts to living organisms beyond traditional lethal concentration endpoints. Daphnia magna are model organisms for aquatic toxicity testing with a well understood metabolome and high sensitivity to SPIONs. Thus, as environmental concentrations continue to increase, it is becoming critical to understand their sub-lethal toxicity. Due to the paramagnetic nature of SPIONs, a range of potential nuclear magnetic resonance spectroscopy (NMR) experiments are possible, offering the potential to probe the physical location (via imaging), binding (via relaxation weighted spectroscopy), and the biochemical pathways impacted (via in vivo metabolomics). Results indicate binding to carbohydrates, likely chitin in the exoskeleton, along with a decrease in energy metabolites and specific biomarkers of oxidative stress. The holistic NMR framework used here helps provide a more comprehensive understanding of SPIONs impacts on D. magna and showcases NMR's versatility in providing physical, chemical, and biochemical insights.

Machine-learning-based video analysis of grasping behavior during recovery from cervical spinal cord injury

Comprehensive characterizations of hand grasping behaviors after cervical spinal cord injuries are fundamental for developing rehabilitation strategies to promote recovery in spinal-cord-injured primates. We used the machine-learning-based video analysis software, DeepLabCut, to sensitively quantify kinematic aspects of grasping behavioral deficits in squirrel monkeys with C5-level spinal cord injuries. Three squirrel monkeys were trained to grasp sugar pellets from wells of varying depths before and after a left unilateral lesion of the cervical dorsal column. Using DeepLabCut, we identified post-lesion deficits in kinematic grasping behavior that included changes in digit orientation, increased variance in vertical and horizontal digit movement, and longer time to complete the task. While video-based analyses of grasping behavior demonstrated deficits in fine-scale digit function that persisted through at least 14 weeks post-injury, traditional end-point behavioral analyses showed a recovery of global hand function as evidenced by recovery of the proportion of successful retrievals by approximately 14 weeks post-injury. The combination of traditional end-point and video-based kinematic analyses provides a more comprehensive characterization of grasping behavior and highlights that global grasping performance may recover despite persistent fine-scale kinematic deficits in digit function. Machine-learning-based video analysis of kinematic digit function, in conjunction with traditional end-point behavioral analyses of grasping behavior, provide sensitive and specific indices for monitoring recovery of fine-grained hand sensorimotor behavior after spinal cord injury that can aid future studies that seek to develop targeted therapeutic interventions for improving behavioral outcomes.

Incidence of heart failure and renal endpoints in heart failure patients with and without severe chronic kidney disease: a nationwide real-life study of 21,959 patients

Abstract Background Renal endpoints like end stage renal disease (ESRD) and a 50% decline in estimated glomerular filtration rate (eGFR) have been used in recent randomized clinical trials in heart failure (HF). However, the distribution of these events in larger real-life HF cohorts is unknown Purpose We examined the risk of traditional HF endpoints like death and worsening HF and the risk of these new renal endpoints in HF with and without severe chronic kidney disease (+/−eGFR 30 ml/min/1.73 m2) at the time of diagnosis of HF. Methods From nationwide registries, we identified patients receiving a diagnosis of HF from 2014–2018. Patients were included if they had creatinine available in proximity of the diagnosis (90 days before inclusion). Outcomes comprised of: (i) all-cause mortality, (ii) HF worsening, (iii) end stage renal disease and (iv) sustained 50% eGFR decline. 4-year rates of the primary outcomes were estimated using cumulative incidence function adjusted for competing risk of death, and multivariable Cox proportional hazards models were used to examine the association of covariates with a combined endpoint of the primary outcomes. Results We included 21,959 patients with HF and known CKD status. Median age was 73.9 years, and 7% had an eGFR &amp;lt;30 mL/min/1.73 m2. There were few differences in baseline characteristics for patients with and without eGFR below 30. Patients with eGFR below 30 were elder and had more comorbidities. The mortality rate was 33.9% for patients with eGFR≥30 and 70.4% for patients with eGFR &amp;lt;30. For patients with eGFR &amp;lt;30 at the time of HF diagnosis renal outcomes were more pronounced as the first event as compared to patients with eGFR ≥30. The risk of all-cause mortality and HF hospitalization as first event was comparable between eGFR ≥30 and eGFR&amp;lt;30 (22.7% vs 22.5% and 25.8% vs 22.5%, respectively). The risk of end-stage renal disease as first event was significantly higher for patients with eGFR &amp;lt;30 (0.03% vs. 33.8%). The risk of a sustained 50% decline in eGFR was 4.6% for patients with eGFR ≥30 and 1.0% for patients with eGFR&amp;lt;30 (figure 1). Across all important subgroups an eGFR &amp;lt;30 ml/min/1.73 m2 was associated with an increased rate of the composite endpoint of death, worsening HF or a renal endpoint, and no interactions were observed (P&amp;gt;0.01 for all) (figure 2). Conclusion The risks of newly introduced renal trial-endpoints in a large real-life cohort of HF patients was low compared to the risks of death or worsening HF if eGFR ≥30 ml/min/1.73 m2 at the time of diagnosis of HF. However, this pattern changed in HF patients with a eGFR &amp;lt;30 ml/min/1.73 m2 at the time of diagnosis time of HF. The present study provides important insights into cardiorenal epidemiology in HF and have implications for planning of future trials. Funding Acknowledgement Type of funding sources: Public hospital(s). Main funding source(s): Herlev/Gentofte Hospital

Evaluating the time toxicity of cancer treatment in the CCTG CO.17 randomized clinical trial (RCT).

248 Background: Most individual treatments for advanced cancer are associated with modest survival benefits. The time spent in pursuing these treatments can be substantial. We have previously developed a pragmatic and patient-centered metric of these time costs, which we term ‘’time toxicity’’, as any day with physical healthcare system contact. This includes outpatient visits (e.g., for bloodwork, scans, infusions, urgent care), emergency room visits, and overnight stays in a healthcare facility. Herein we sought to assess time toxicity in a completed RCT. Methods: We conducted a secondary analysis of the Canadian Cancer Trials Group CO.17 RCT that evaluated weekly cetuximab infusions vs supportive care alone in 572 patients with advanced colorectal cancer. CO.17 recruited participants in Canada, Australia and New Zealand. Initial results reported a 6-week improvement in median overall survival (OS) with cetuximab (6.1 vs 4.6 months, p = 0.005). Subsequent analyses reported that OS benefit was restricted to and significantly more in patients with K-ras wild-type tumors. We calculated patient-level time toxicity by analyzing treatment, follow-up, and resource utilization forms. We considered a day without physical healthcare contact as a ‘’home day’’. Thus, for a patient, OS was time toxic days + home days. We compared medians of time measures across arms, and stratified results by K-ras status. Results: In the overall population, median time toxic days were higher in the cetuximab arm (28, vs 10, p &lt; 0.001), although median home days were not statistically different (140, vs 121, p = 0.09). The proportion of time toxic days (time toxic days/OS) were significantly more with cetuximab (18%, vs 6%, p &lt; 0.001). Of the 28 time toxic days in the overall cetuximab arm, 14 (50%) were protocol-related (e.g., scheduled infusions etc). Stratified results are in the table. Conclusions: Time toxicity can be extracted through secondary analyses of RCTs. In CO.17, despite an overall OS-benefit with cetuximab, home days were statistically similar across arms. In the K-ras-mutated group, cetuximab was associated with numerically similar OS but more time toxic days. In the K-ras-wild type group, cetuximab was associated with higher home days. Thus, time toxicity data need not always be sobering. Time toxicity measures can supplement traditional survival endpoints in RCTs to guide patient-oncologist decision-making. Cooperative group RCTs are well positioned for such analyses. Clinical trial information: NCT00079066. [Table: see text]

Introducing oxygen transfer rate measurements as a novel method for time-resolved cytotoxicity assessment in shake flasks

BackgroundDetermining the cytotoxicity of test substances is essential for the safety assessment of chemicals. To quantify the cytotoxicity, dose–response curves are determined and the half-maximum inhibitory concentration (IC50) is subsequently calculated. Results often rely on a single endpoint evaluation which typically requires manual sampling and subsequent sample analysis to determine the IC50. Hence, no information on culture behavior are available during treatment. Here, measurement of the oxygen transfer rate (OTR) was applied as a method for time-resolved assessment of the cytotoxicity to (a) provide information on culture behavior during treatment and (b) quantitatively assess the cytotoxic effect of a test substance.ResultsTo investigate the applicability of the method, different types and concentrations of test substances were added and the OTR was measured for two different CHO suspension cell lines in shake flasks: (a) For CHO cell line one, culture behavior was not affected when 0.1 mM ZnCl2 was added. However, for both cell lines, adding up to 0.2 mM CoCl2 led to a decrease in the OTR that was concentration dependent. For CHO cell line two, adding up to 5% DMSO led to a concentration-dependent decrease in the OTR and in the slope of the OTR; (b) From the measured OTR, dose–response curves were established and used to calculate the IC50. For cell line one, the IC50 for CoCl2 was 0.03 mM while the IC50 from a comparative measurement based on trypan blue exclusion was 0.06 mM. Cell line two was less susceptible to CoCl2 as the IC50 was not reached at the concentrations tested. For DMSO, single OTR values and determined slopes were used for evaluation. In both cases, the IC50 was calculated to 2.3%.ConclusionsOur approach provides a method for analyzing the cytotoxicity of a test substance based on OTR measurements. This method provides (i) insights on culture behavior, (ii) information on the progression of cytotoxicity, (iii) dose–response curves, and (iv) a first indication of IC50 values. Quantitative assessment of cytotoxicity is possible non-invasively and in real-time during treatment. Compared to traditional endpoint evaluation, our method simplifies experimentations and enhances the comparison of assay results.

Attomolar sensitivity microRNA detection using real-time digital microarrays

MicroRNAs (miRNAs) are a family of noncoding, functional RNAs. With recent developments in molecular biology, miRNA detection has attracted significant interest, as hundreds of miRNAs and their expression levels have shown to be linked to various diseases such as infections, cardiovascular disorders and cancers. A powerful and high throughput tool for nucleic acid detection is the DNA microarray technology. However, conventional methods do not meet the demands in sensitivity and specificity, presenting significant challenges for the adaptation of miRNA detection for diagnostic applications. In this study, we developed a highly sensitive and multiplexed digital microarray using plasmonic gold nanorods as labels. For proof of concept studies, we conducted experiments with two miRNAs, miRNA-451a (miR-451) and miRNA-223-3p (miR-223). We demonstrated improvements in sensitivity in comparison to traditional end-point assays that employ capture on solid phase support, by implementing real-time tracking of the target molecules on the sensor surface. Particle tracking overcomes the sensitivity limitations for detection of low-abundance biomarkers in the presence of low-affinity but high-abundance background molecules, where endpoint assays fall short. The absolute lowest measured concentration was 100 aM. The measured detection limit being well above the blank samples, we performed theoretical calculations for an extrapolated limit of detection (LOD). The dynamic tracking improved the extrapolated LODs from femtomolar range to sim 10 attomolar (less than 1300 copies in 0.2 ml of sample) for both miRNAs and the total incubation time was decreased from 5 h to 35 min.