Isothermal Amplification with a Target-Mimicking Internal Control and Quantitative Lateral Flow Readout for Rapid HIV Viral Load Testing in Low-Resource Settings.

Point-of-care diagnostics often use isothermal nucleic acid amplification for qualitative detection of pathogens in low-resource healthcare settings but lack sufficient precision for quantitative applications such as HIV viral load monitoring. Although viral load (VL) monitoring is an essential component of HIV treatment, commercially available tests rely on relatively high-resource chemistries like real-time polymerase chain reaction and are thus used on an infrequent basis for millions of people living with HIV in low-income countries. To address the constraints of low-resource settings on nucleic acid quantification, we describe a recombinase polymerase amplification and lateral flow detection approach that quantifies HIV-1 DNA or RNA by comparison to a competitive internal amplification control (IAC) of a known copy number, which may be set to any useful threshold (in our case, a clinically relevant threshold for HIV treatment failure). The IAC is designed to amplify alongside the HIV target with a similar efficiency, allowing for normalization of the assay to variation or inhibition and enabling an endpoint readout that is compatible with commercially available kits for nucleic acid lateral flow detection and interpretable with minimal instrumentation or by the naked eye. We find that this approach can reliably differentiate ≤600 or ≥1400 copies of HIV DNA from a 1000-copy threshold when lateral flow strips are imaged with a conventional office scanner and analyzed with free densitometry software. We further demonstrate a user-friendly adaptation of this analysis to process cell phone photographs with an automated script. Alternatively, we show via a survey that 21 minimally trained volunteers could reliably resolve ≥10-fold (log10) differences of HIV DNA or RNA by naked eye interpretation of lateral flow results. This amplification and detection workflow requires minimal instrumentation, takes just 30 min to complete, and when combined with a suitable sample preparation method, may enable HIV VL testing while the patient waits or a self-test, which has the potential to improve care. This approach may be adapted for other applications that require quantitative analysis of a nucleic acid target in low-resource settings.

Simultaneous monitoring of HIV viral load and screening of SARS-CoV-2 employing a low-cost RT-qPCR test workflow.

The COVID-19 pandemic interrupted routine care for individuals living with HIV, putting them at risk of virologic failure and HIV-associated illness. Often this population is at high risk for exposure to SARS-CoV-2 infection, and once infected, for severe disease. Therefore, close monitoring of HIV plasma viral load (VL) and screening for SARS-CoV-2 infection are needed. We developed a non-proprietary method to isolate RNA from plasma, nasal secretions (NS), or both. The extracted RNA is then submitted to RT-qPCR to estimate the VL and classify HIV/SARS-CoV-2 status (i.e., HIV virologic failure or suppressed; SARS-CoV-2 as positive, presumptive positive, negative, or indeterminate). In contrived samples, the in-house RNA extraction workflow achieved a detection limit of 200-copies per mL for HIV RNA in plasma and 100-copies per mL for SARS-CoV-2 RNA in NS. Similar detection limits were observed for HIV and SARS-CoV-2 in pooled plasma/NS contrived samples. When comparing in-house with standard extraction methods, we found high agreement (>0.91) between input and measured RNA copies for HIV LTR in contrived plasma; SARS-CoV-2 N1/N2 in contrived NS; and LTR, N1, and N2 in pooled plasma/NS samples. We further evaluated this workflow on 133 clinical specimens: 40 plasma specimens (30 HIV-positive), 67 NS specimens (31 SARS-CoV-2-positive), and 26 combined plasma/NS specimens (26 HIV-positive with 10 SARS-CoV-2-positive), and compared the results obtained using the in-house RNA extraction to those using a commercial kit (standard extraction method). The in-house extraction and standard extraction of clinical specimens were positively correlated: plasma HIV VL (R2 of 0.81) and NS SARS-CoV-2 VL (R2 of 0.95 and 0.99 for N1 and N2 genes, respectively); and pooled plasma/NS HIV VL (R2 of 0.71) and SARS-CoV-2 VL (R2 of 1 both for N1 and N2 genes). Our low-cost molecular test workflow ($1.85 per pooled sample extraction) for HIV RNA and SARS-CoV-2 RNA could serve as an alternative to current standard assays ($12 per pooled sample extraction) for laboratories in low-resource settings.

Understanding Longitudinal Changes in Pulmonary Vascular Resistance After Left Ventricular Assist Device Implantation.

BACKGROUND: Elevated pulmonary vascular resistance (PVR) is common in patients with advanced heart failure. PVR generally improves after left ventricular assist device (LVAD) implantation, but the rate of decrease has not been quantified and the patient characteristics most strongly associated with this improvement are unknown. METHODS AND RESULTS: We analyzed 1581 patients from the Interagency Registry for Mechanically Assisted Circulatory Support registry who received a primary continuous-flow LVAD, had a baseline PVR of ≥3 Wood units (WU), and had PVR measured at least once postoperatively. Multivariable linear mixed effects modeling was used to evaluate independent associations between postoperative PVR and patient characteristics. PVR decreased by 1.53 WU (95% confidence interval [CI] 1.27-1.79 WU) per month in the first 3 months postoperatively, and by 0.066 WU (95% CI 0.060-0.070 WU) per month thereafter. Severe mitral regurgitation at any time during follow-up was associated with a 1.29 WU (95% CI 1.05-1.52 WU) higher PVR relative to absence of mitral regurgitation at that time. In a cross-sectional analysis, 15%-25% of patients had persistently elevated PVR of ≥3 WU at any given time within 36 months after LVAD implantation. CONCLUSION: The PVR tends to decrease rapidly early after implantation, and only more gradually thereafter. Residual mitral regurgitation may be an important contributor to elevated postoperative PVR. Future research is needed to understand the implications of elevated PVR after LVAD implantation and the optimal strategies for prevention and treatment.

Electrically controlled nicotine delivery through Carbon nanotube membranes via electrochemical oxidation and nanofluidically enhanced electroosmotic flow.

A promising tool for nicotine addiction treatment is a programmable nicotine delivery device coupled to smart phone-assisted behavioral therapies. Key metrics for such a device are delivery of adjustable nicotine doses tailored to individual needs, compact size and power efficiency. Reported here is a detailed optimization of carbon nanotube (CNT) membrane fabrication based on electrochemical oxidation, to improve its electrically driven performance for nicotine fluxes and switching ON (-1.5 V)-OFF (0 V) flux ratio. ON- state nicotine flux of ~ 6 µmoles/cm2/h at -1.5 V applied bias was achieved allowing ~ 6-folds decrease in the size of device (4 cm2) to attain flux equivalent to high dose nicotine gum (1.1 µmoles/cm2/h). Application of + 1.5 V bias in OFF state reduced diffusional background flux, giving an ON (-1.5 V)/OFF (+ 1.5 V) flux ratio of 68 that enabled device to deliver between the highest nicotine gum (1.1 µmoles/cm2/h) and lowest nicotine patch (0.08 µmoles/cm2/h) doses, as well as taper off nicotine doses for long term addiction treatment. The nicotine transport mechanism was studied as a function of pH and applied bias, using neutral tracer molecule, showing a mechanism of both electroosmosis and electrophoresis in the atomically smooth nanofluidic pores of CNTs. Optimal power consumption/flux efficiency of 111(µW/cm2)/µmoles/cm2/h was achieved allowing watch-battery lifetimes of 7-62 days for conventional treatment dosing regimens. Bluetooth-enabled, remotely controlled CNT membrane system has potential for treatments of nicotine, opioid and alcohol addictions that needs dose adjustment with precise temporal control.

Timing and Trends of Right Atrial Pressure and Risk of Right Heart Failure After Left Ventricular Assist Device Implantation.

BACKGROUND: Elevated right atrial pressure (RAP) is associated with poor outcomes after left ventricular assist device (LVAD) implantation. However, the optimal time for RAP measurement and the importance of resolution of right heart congestion prior to LVAD implantation remain unclear. METHODS AND RESULTS: We performed a retrospective cohort study of 134 consecutive LVAD recipients from our institution. Congestion was defined as RAP ≥ 14 mmHg and was assessed at hospital admission and implant. The primary outcome was death or right ventricular assist device (RVAD) implantation. When stratified by congestion status at admission, congested and non-congested patients had similar event-free survival rates (hazard ratio [HR]: 1.2, 95% confidence interval [CI]: 0.6-2.6). However, when stratified at implant, congested patients had a higher rate death or RVAD implantation (HR: 2.5, 95% CI: 1.1-5.6). Patients were then divided into 4 groups based on their trajectory of congestion status: no congestion, resolved congestion, new congestion, or persistent congestion. Patients with no congestion and resolved congestion had similar outcomes, whereas patients with persistent congestion had a markedly increased rate of death or RVAD implantation (HR: 3.1, 95% CI: 1.3-7.6). CONCLUSION: RAP at LVAD implantation is more strongly associated with postoperative outcomes than admission RAP. Patients not responsive to decongestive therapies, with persistently elevated RAP, represent a high-risk cohort for adverse outcomes following LVAD implantation.

Heart failure management in dialysis patients: Many treatment options with no clear evidence.

Heart failure with reduced ejection fraction (HFrEF) impacts approximately 20% of dialysis patients and is associated with high mortality rates. Key issues discussed in this review of HFrEF management in dialysis include dialysis modality choice, vascular access, dialysate composition, pharmacological therapies, and strategies to reduce sudden cardiac death, including the use of cardiac devices. Peritoneal dialysis and more frequent or longer duration of hemodialysis may be better tolerated due to slower ultrafiltration rates, leading to less intradialytic hypotension and better volume control; dialysate cooling and higher dialysate calcium may also have benefits. While high-quality evidence exists for many drug classes in the non-dialysis population, dialysis patients were excluded from major trials, and only limited data exist for many medications in kidney failure patients. Despite limited evidence, beta blocker and angiotensin-converting enzyme inhibitor or angiotensin receptor blocker use is common in dialysis. Similarly, devices such as implantable cardiac defibrillators (ICDs) and cardiac resynchronization therapy that have proven benefits in non-dialysis HFrEF patients have not consistently been beneficial in the limited dialysis studies. The use of leadless pacemakers and subcutaneous ICDs can mitigate future hemodialysis access limitations. Additional research is critical to address knowledge gaps in treating maintenance dialysis patients with HFrEF.

Discovery of SERPINA3 as a candidate urinary biomarker of lupus nephritis activity.

Objectives: We used an unbiased proteomics approach to identify candidate urine biomarkers (CUBMs) predictive of LN chronicity and pursued their validation in a larger cohort. Methods: In this cross-sectional pilot study, we selected urine collected at kidney biopsy from 20 children with varying levels of LN damage (discovery cohort) and performed proteomic analysis using isobaric tags for relative and absolute quantification (iTRAQ). We identified differentially excreted proteins based on degree of LN chronicity and sought to distinguish markers exhibiting different relative expression patterns using hierarchically clustered log10-normalized relative abundance data with linked and distinct functions by biological network analyses. For each CUBM, we performed specific ELISAs on urine from a validation cohort (n = 41) and analysis of variance to detect differences between LN chronicity, with LN activity adjustment. We evaluated for CUBM expression in LN biopsies with immunohistochemistry. Results: iTRAQ detected 112 proteins in urine from the discovery cohort, 51 quantifiable in all replicates. Simple analysis of variance revealed four differentially expressed, chronicity-correlated proteins (P-values < 0.05). Further correlation and network analyses led to selection of seven CUBMs for LN chronicity. In the validation cohort, none of the CUBMs distinguished LN chronicity degree; however, urine SERPINA3 demonstrated a moderate positive correlation with LN histological activity. Immunohistochemistry further demonstrated SERPINA3 staining in proximal tubular epithelial and endothelial cells. Conclusion: We identified SERPINA3, a known inhibitor of neutrophil cathepsin G and angiotensin II production, as a potential urine biomarker to help quantify LN activity.

Urine biomarkers of chronic kidney damage and renal functional decline in childhood-onset systemic lupus erythematosus.

OBJECTIVES: To delineate urine biomarkers that reflect kidney structural damage and predict renal functional decline in pediatric lupus nephritis (LN). METHODS: In this prospective study, we evaluated kidney biopsies and urine samples of 89 patients with pediatric LN. Urinary levels of 10 biomarkers [adiponectin, ceruloplasmin, kidney injury molecule-1, monocyte chemotactic protein-1, neutrophil gelatinase-associated lipocalin, osteopontin, transforming growth factor-ß (TGFß), vitamin-D binding protein, liver fatty acid binding protein (LFABP), and transferrin] were measured. Regression analysis was used to identify individual and combinations of biomarkers that determine LN damage status [NIH-chronicity index (NIH-CI) score ≤ 1 vs. ≥ 2] both individually and in combination, and biomarker levels were compared for patients with vs. without renal functional decline, i.e., a 20% reduction of the glomerular filtration rate (GFR) within 12 months of a kidney biopsy. RESULTS: Adiponectin, LFABP, and osteopontin levels differed significantly with select histological damage features considered in the NIH-CI. The GFR was associated with NIH-CI scores [Pearson correlation coefficient (r) = - 0.49; p < 0.0001] but not proteinuria (r = 0.20; p > 0.05). Similar to the GFR [area under the ROC curve (AUC) = 0.72; p < 0.01], combinations of osteopontin and adiponectin levels showed moderate accuracy [AUC = 0.75; p = 0.003] in discriminating patients by LN damage status. Renal functional decline occurred more commonly with continuously higher levels of the biomarkers, especially of TGFß, transferrin, and LFABP. CONCLUSION: In combination, urinary levels of adiponectin and osteopontin predict chronic LN damage with similar accuracy as the GFR. Ongoing LN activity as reflected by high levels of LN activity biomarkers heralds renal functional decline. KEY MESSAGES: • Levels of osteopontin and adiponectin measured at the time of kidney biopsy are good predictors of histological damage with lupus nephritis. • Only about 20% of children with substantial kidney damage from lupus nephritis will have an abnormally low urine creatinine clearance. • Continuously high levels of biomarkers reflecting lupus nephritis activity are risk factors of declining renal function.

Programmable carbon nanotube membrane-based transdermal nicotine delivery with microdialysis validation assay.

To evaluate the performance of switchable carbon nanotubes (CNT) membrane devices for transdermal nicotine delivery, we have developed an in-vitro microdialysis method that allow us to detect variable transdermal fluxes of nicotine through CNT devices and can be applied directly to in-vivo studies. Microdialysis membranes were placed beneath the porcine skin and its nicotine levels increased 6-8 times when the CNT membrane on skin was turned from OFF to ON state by application of bias. Fluxes in the ON state were approximately 3 times that of commercial nicotine patches and switching times were less than two hours, thus suggesting the improved therapeutic potential of our device. Blue tooth enabled CNT devices that can be programmed by smartphone and coupled with remote counseling application for enhanced smoking cessation treatments.

Detection of Myocardial Dysfunction in Septic Shock: A Speckle-Tracking Echocardiography Study.

BACKGROUND: Patients with septic shock are at increased risk of myocardial dysfunction. However, the left ventricular ejection fraction (EF) typically remains preserved in septic shock. Strain measurement using speckle-tracking echocardiography may quantify abnormalities in myocardial function not detected by conventional echocardiography. To investigate whether septic shock results in greater strain changes than sepsis alone, we evaluated strain in patients with sepsis and septic shock. METHODS: We prospectively identified 35 patients with septic shock and 15 with sepsis. These patients underwent serial transthoracic echocardiograms at enrollment and 24 hours later. Measurements included longitudinal, radial, and circumferential strain in addition to standard echocardiographic assessments of left ventricular function. RESULTS: Longitudinal strain worsened significantly over 24 hours in patients with septic shock (P < 0.0001) but did not change in patients with sepsis alone (P = 0.43). No significant changes in radial or circumferential strain or EF were observed in either group over the 24-hour measurement period. In patients with septic shock, the significant worsening in longitudinal strain persisted after adjustment for left ventricular end-diastolic volume and vasopressor use (P < 0.0001). In patients with sepsis, adjustment for left ventricular end-diastolic volume and vasopressor use did not alter the finding of no significant differences in longitudinal strain (P = 0.48) or EF (P = 0.96). CONCLUSIONS: In patients with septic shock, but not sepsis, myocardial strain imaging using speckle-tracking echocardiography identified myocardial dysfunction in the absence of changes in EF. These data suggest that strain imaging may play a role in cardiovascular assessment during septic shock.

Cancer and cardiovascular disease: the use of novel echocardiography measures to predict subsequent cardiotoxicity in breast cancer treated with anthracyclines and trastuzumab.

As the population of breast cancer survivors grows, it has become evident that chemotherapy has significant cardiotoxic side effects. Echocardiography is a noninvasive, cost-effective, and widely available imaging tool that is well positioned to serve as a primary modality for monitoring chemotherapy-induced cardiotoxicity. Although left ventricular ejection fraction is a standard measurement by which to monitor chemotherapy-induced cardiotoxicity, its predictive value in identifying subsequent cardiotoxicity is limited. More sophisticated echocardiography modalities may offer improved sensitivity and specificity for detecting chemotherapy-induced cardiotoxicity. These include tissue Doppler imaging measures, newer techniques based upon two- and three-dimensional strain and torsion analysis, and three-dimensional measures of cardiac size. While these modalities are not all currently part of clinical practice, a body of data supporting their use is steadily building. More research remains to be performed, and noninvasively detecting cancer therapy-induced cardiac dysfunction at the earliest stages is of increasing interest.

Pleural pressure and optimal positive end-expiratory pressure based on esophageal pressure versus chest wall elastance: incompatible results*.

OBJECTIVES: 1) To compare two published methods for estimating pleural pressure, one based on directly measured esophageal pressure and the other based on chest wall elastance. 2) To evaluate the agreement between two published positive end-expiratory pressure optimization strategies based on these methods, one targeting an end-expiratory esophageal pressure-based transpulmonary pressure of 0 cm H2O and the other targeting an end-inspiratory elastance-based transpulmonary pressure of 26 cm H2O. DESIGN: Retrospective study using clinical data. SETTING: Medical and surgical ICUs. PATIENTS: Sixty-four patients mechanically ventilated for acute respiratory failure with esophageal balloons placed for clinical management. METHODS: Esophageal pressure and chest wall elastance-based methods for estimating pleural pressure and setting positive end-expiratory pressure were retrospectively applied to each of the 64 patients. In patients who were ventilated at two positive end-expiratory pressure levels, chest wall and respiratory system elastances were calculated at each positive end-expiratory pressure level. MEASUREMENTS AND MAIN RESULTS: The pleural pressure estimates using both methods were discordant and differed by as much as 10 cm H2O for a given patient. The two positive end-expiratory pressure optimization strategies recommended positive end-expiratory pressure changes in opposite directions in 33% of patients. The ideal positive end-expiratory pressure levels recommended by the two methods for each patient were discordant and uncorrelated (R = 0.05). Chest wall and respiratory system elastances grew with increases in positive end-expiratory pressure in patients with positive end-expiratory esophageal pressure-based transpulmonary pressures (p < 0.05). CONCLUSIONS: Esophageal pressure and chest wall elastance-based methods for estimating pleural pressure do not yield similar results. The strategies of targeting an end-expiratory esophageal pressure-based transpulmonary pressure of 0 cm H2O and targeting an end-inspiratory elastance-based transpulmonary pressure of 26 cm H2O cannot be considered interchangeable. Finally, chest wall and respiratory system elastances may vary unpredictably with changes in positive end-expiratory pressure.

Long-Term Changes in Estimated Glomerular Filtration Rate in Left Ventricular Assist Device Recipients: A Longitudinal Joint Model Analysis.

Background Advanced kidney disease is often a relative contraindication to left ventricular assist device (LVAD) implantation because of concerns for poor outcomes including worsening kidney disease. Data are lacking on long-term changes and sex-based differences in estimated glomerular filtration rate (eGFR), with published data limited by potential bias introduced by the competing risks of death and heart transplantation. Methods and Results We conducted a longitudinal analysis of 288 adults receiving durable continuous-flow LVADs from January 2010 to December 2017 at a single center. A joint model was constructed to evaluate change in eGFR over 2 years, the prespecified primary outcome, adjusted for the competing risks of death and heart transplantation. Median baseline eGFR was 60 mL/min per 1.73 m2 (interquartile range 42-78). At 2 years, 74 patients died and 104 received a heart transplant. In unadjusted analysis, LVAD recipients had a modest initial increase in eGFR of ≈2 mL/min per 1.73 m2 within the first 6 months after implantation, followed by a decrease in eGFR below baseline values at 1 and 2 years. Men experienced an eGFR decline of 5 to 10 mL/min per 1.73 m2 over the first year which then stabilized, while women had an ≈5 mL/min per 1.73 m2 increase in eGFR within the first 6 months followed by decline towards baseline eGFR levels (interaction P=0.005). Conclusions Estimated GFR remains relatively stable in most patients following LVAD implantation. Larger studies are needed to investigate sex-based differences in eGFR and to evaluate eGFR trajectory and mortality in LVAD recipients with lower eGFR.

Cardiac Cachexia in Left Ventricular Assist Device Recipients and the Implications of Weight Gain Early After Implantation.

Background Severe cardiac cachexia or malnutrition are commonly considered relative contraindications to left ventricular assist device (LVAD) implantation, but post-LVAD prognosis for patients with cachexia is uncertain. Methods and Results Intermacs (Interagency Registry for Mechanically Assisted Circulatory Support) 2006 to 2017 was queried for the preimplantation variable cachexia/malnutrition. Cox proportional hazards modeling examined the relationship between cachexia and LVAD outcomes. Of 20 332 primary LVAD recipients with available data, 516 (2.54%) were reported to have baseline cachexia and had higher risk baseline characteristics. Cachexia was associated with higher mortality during LVAD support (unadjusted hazard ratio [HR], 1.36 [95% CI, 1.18-1.56]; P<0.0001), persisting after adjustment for baseline characteristics (adjusted HR, 1.23 [95% CI, 1.0-1.42]; P=0.005). Mean weight change at 12 months was +3.9±9.4 kg. Across the cohort, weight gain ≥5% during the first 3 months of LVAD support was associated with lower mortality (unadjusted HR, 0.90 [95% CI, 0.84-0.98]; P=0.012; adjusted HR, 0.89 [95% CI, 0.82-0.97]; P=0.006). Conclusions The proportion of LVAD recipients recognized to have cachexia preimplantation was low at 2.5%. Recognized cachexia was independently associated with higher mortality during LVAD support. Early weight gain ≥5% was independently associated with lower mortality during subsequent LVAD support.

Epidemiology and management of right ventricular-predominant heart failure and shock in the cardiac intensive care unit.

Cardiogenic shock from left ventricular failure is a common presentation in the intensive care unit. In contrast, right ventricular (RV)-predominant heart failure (HF) causing shock is less well recognized. We review the epidemiology and mechanisms of RV-predominant HF and discuss pharmacologic and device-based approaches for the management of this challenging clinical problem.

Association between postoperative hemodynamic metrics of pulmonary hypertension and right ventricular dysfunction and clinical outcomes after left ventricular assist device implantation.

BACKGROUND: While preoperative hemodynamic risk factors associated with early right heart failure (RHF) following left ventricular assist device (LVAD) surgery are well-established, the relationship between postoperative hemodynamic status and subsequent outcomes remains poorly defined. METHODS: We analyzed adult CF-LVAD patients from the STS-INTERMACS registry surviving at least 3 months without evidence of early RHF and with hemodynamic data available at 3 months after LVAD implant. The association between metrics of RV afterload and function and the subsequent risk of death, right heart failure (RHF), gastrointestinal bleeding (GIB), or stroke were assessed using multivariable Cox proportional hazards modeling. RESULTS: Among 1,050 patients with available 3-month hemodynamics, pulmonary hypertension was common, with 585 (55.7%) having mPAP ≥ 20 mm Hg and 164 (15.6%) having PVR ≥ 3 WU. Pulmonary artery pulsatility index (PAPi, HR 0.62 per log-increase for values < 3, 95% CI 0.43-0.89) and PVR (HR 1.19 per 1 WU-increase for values > 1.5 WU, 95% CI 1.03-1.38) were independently associated with the composite of death or RHF. Postoperative RAP (HR 1.18 per 5 mm Hg increase, 95% CI 1.04-1.33), RAP:PCWP (HR 1.46 per log-increase, 95% CI 1.12-1.91), and PAPi (HR 0.76 per log-increase, 95% CI 0.61-0.95) were each associated with GIB risk. Postoperative hemodynamics was not associated with stroke risk. CONCLUSIONS: Hemodynamic metrics of postoperative RV dysfunction and elevated RV afterload are independently associated with RHF, mortality and GIB. Whether strategies targeting postoperative optimization of RV function and afterload can reduce the burden of these adverse events requires prospective study.

Generalizability of Cardiovascular Disease Clinical Prediction Models: 158 Independent External Validations of 104 Unique Models.

BACKGROUND: While clinical prediction models (CPMs) are used increasingly commonly to guide patient care, the performance and clinical utility of these CPMs in new patient cohorts is poorly understood. METHODS: We performed 158 external validations of 104 unique CPMs across 3 domains of cardiovascular disease (primary prevention, acute coronary syndrome, and heart failure). Validations were performed in publicly available clinical trial cohorts and model performance was assessed using measures of discrimination, calibration, and net benefit. To explore potential reasons for poor model performance, CPM-clinical trial cohort pairs were stratified based on relatedness, a domain-specific set of characteristics to qualitatively grade the similarity of derivation and validation patient populations. We also examined the model-based C-statistic to assess whether changes in discrimination were because of differences in case-mix between the derivation and validation samples. The impact of model updating on model performance was also assessed. RESULTS: Discrimination decreased significantly between model derivation (0.76 [interquartile range 0.73-0.78]) and validation (0.64 [interquartile range 0.60-0.67], P<0.001), but approximately half of this decrease was because of narrower case-mix in the validation samples. CPMs had better discrimination when tested in related compared with distantly related trial cohorts. Calibration slope was also significantly higher in related trial cohorts (0.77 [interquartile range, 0.59-0.90]) than distantly related cohorts (0.59 [interquartile range 0.43-0.73], P=0.001). When considering the full range of possible decision thresholds between half and twice the outcome incidence, 91% of models had a risk of harm (net benefit below default strategy) at some threshold; this risk could be reduced substantially via updating model intercept, calibration slope, or complete re-estimation. CONCLUSIONS: There are significant decreases in model performance when applying cardiovascular disease CPMs to new patient populations, resulting in substantial risk of harm. Model updating can mitigate these risks. Care should be taken when using CPMs to guide clinical decision-making.