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Introduction:Insulin like growth factor binding proteins (IGFBPs) are a family of growth factor modifiers, some of which are known to be independently associated with PAH survival. IGFBP7 is a unique low-affinity IGFBP that, independent of IGF, stimulates prostacyclin production.Hypothesis:Elevated IGFBP7 is associated with worse disease severity and survival in PAH.Methods:Using enzyme-linked immunosorbent assays (ELISA), we evaluated serum IGFBP7 in a multicenter PAH cohort, the NHLBI PAHBiobank (N=2583), and healthy controls (N=99). Hemodynamic and functional measures were used to assess IGFBP7’s association to PAH severity and survival. Kruskal Wallis was used for comparison and Spearman’s rank correlation for correlations. Linear and logistic regressions performed for associations. IGFBP7 was dichotomized at the median and analyzed by Kaplan-Meier survival and Cox proportional Hazard regression.Results:Serum IGFBP7 levels were significantly elevated in patients with PAH compared to controls. After adjustment, logarithmic increases in IGFBP7 were associated with a 38 meter shorter six-minute walk distance (6MWD; -37.7, 95%CI -54 to -21, p=<0.001), a higher mean right atrial pressure (2.18mmHg, 95%CI 1.51-2.89, p<0.001) and a higher REVEAL risk score (1.10, 95%CI 0.88-1.33, p <0.001). Logarithmic increases in IGFBP7 were also associated with a two-times higher likelihood of having PAH associated with Portopulmonary disease (2.39, 95%CI 1.95-2.84, p<0.001). Kaplan Meier analysis demonstrated significant survival effects of IGFBP7 above the median (see Figure 1) and an adjusted (age, sex, functional class, and 6MWD) Cox proportional hazards model (HR 3.1, 95% CI 2.2-4.4, p<0.001).Conclusions:Higher circulating IGFBP7 levels were significantly associated with worse PAH severity and decreased survival. Given its role of as a prostacyclin stimulant, IGFBP7 may also serve as a therapeutic target for disease modulation.

Introduction:Pulmonary arterial hypertension (PH) is an often fatal disease of the pre-capillary pulmonary vasculature. Recognizing high risk PH in ambulatory patients is important for optimization of therapy to improve outcomes. We previously identified insulin like growth factor binding proteins 1 and 2 (IGFBP1, IGFBP2) as prognostic markers in severe PH; here, we evaluated IGFBP1/2 as prognostic markers in PH with intermediate risk features.Methods:A multiplex ELISA assay measured serum IGFBP1/2 in the multicenter PH Biobank (N=2450), and independent validation cohorts (Johns Hopkins, N=145, Vanderbilt N= 128, 388 observations over time). IGFBP1/2 concentrations were compared to hemodynamics, six-minute walk (6MW), and WHO functional class using linear/logistic regression adjusted for age and sex. Time to death/transplant by IGFBP1/2 levels was assessed by Cox proportional hazard models. Analysis was repeated in subjects with low to intermediate risk features defined as WHO functional class 1-2, 6MW >300 m, and NT-proBNP <1500 pg/mL, based on the REVEAL and ERS risk scores.1, 2Results:In all cohorts, increased IGFBP1 and IGFBP2 were associated with lower cardiac output, higher mean pulmonary artery pressure and pulmonary vascular resistance (PVR, p<0.05 for all). In each sub-analysis, a higher IGFBP1 and IGFBP2 was associated with lower cardiac output, lower systemic blood pressure, and higher PVR (p<0.05 for all). Higher IGFBP1 and IGFBP2 were associated with shorter time to death/transplant in the overall cohort and each of the intermediate risk PH cohorts (Table 1), although underpowered in the Vanderbilt cohort.Conclusions:Identifying patients who will have adverse outcome in intermediate risk PH has proven challenging. These data suggest IGFBP1/2 are good prognostic markers in PH where functional class and exertional tolerance is preserved. IGFBP1/2 could further discriminate adverse outcomes in patients with low to intermediate risk PH.

Introduction:Right ventricular (RV) diastolic dysfunction in pulmonary arterial hypertension (PAH) remains poorly understood. In this study, we used RV pressure-volume (PV) loops to assess RV diastology as a function of afterload in PAH.Methods:RV PV loops were prospectively measured in control (n=9) and PAH (n=39) patients at rest and during supine bike exercise. PAH patients were stratified by effective arterial elastance (Ea), a lumped measure of total afterload, into two groups: high afterload (HA, n=20) and severe afterload (SA, n=19).Results:PAH groups were matched with respect to age, sex, and PAH medication usage. Ea at rest and exercise was higher in SA PAH (1.8 vs. 0.6 mmHg/ml, P <0.0001 at rest; 2.3 vs 0.8 mmHg/ml, P < 0.0001 with exercise). In the SA group, RV end-diastolic pressure (RVEDP) was higher at rest (13.3 vs 7.6 mmHg, P = 0.0017) and rose markedly with exertion (23.4 vs 13.1 mmHg, P=0.0001). Because RV volumes changed similarly in both groups, SA patients exhibited stiffer RVs (dP/dV 0.30 vs 0.11 mmHg/mL, P=.024). To understand pericardial-interventricular interactions, we calculated the RVEDP-to-pulmonary capillary wedge pressure (PCWP) ratio and the left ventricular (LV) transmural filling pressure (LVTMFP). RVEDP/PCWP was substantially higher in SA (2.1 vs 1.0, P=.005). Therefore, even though PCWP rose considerably in SA (mean 16 mmHg), exercise LVTMFP was significantly reduced in SA vs HA PAH (-7.5 vs 2.55 mmHg, P=0.002). Along with reduced LV size seen by concomitant echo, we thus revealed blunted LV filling specifically in SA PAH. Accordingly, exercise cardiac index, stroke volume index, and LV stroke work index were all markedly reduced in SA PAH.Conclusions:Severe afterload in PAH results in high RV diastolic pressures and RV stiffening, which in turn blunt LV filling and LV output with exertion. Due to ventricular interactions, exercise PCWP may rise considerably in PAH, thus reducing its ability to differentiate from left heart disease.

Introduction:Pulmonary arterial hypertension (PAH) results in progressive elevation of pulmonary vascular resistance and right heart failure. Insulin like growth factor binding proteins (IGFBPs) are a family of growth factor modifiers. As increased IGFBP4 expression is associated with vascular proliferation in lung cancer, it may also be associated with PAH severity.Hypothesis:Elevated IGFBP4 is associated with worse disease severity and survival in PAH.Methods:Using enzyme-linked immunosorbent assays (ELISA), we evaluated serum IGFBP4 in a multicenter PAH cohort, the NHLBI PAHBiobank (N=2571), and healthy controls (N=125). Hemodynamic and functional measures were used to assess IGFBP4’s association to PAH severity and survival. Spearman’s rank correlation test and Kruskal Wallis test were used for correlations. IGFBP4 was dichotomized at the median and analyzed by Kaplan-Meier survival and Cox proportional Hazard regression.Results:Serum IGFBP4 levels were significantly elevated (p<0.0001) compared to controls. After adjustment for age and sex, IGFBP4 above mean was associated with a 40 meter shorter six minute walk distance (95%CI, -55 to -25, p=<0.001). In the IPAH subtype only, higher IGFBP4 was associated with higher mean right atrial pressures (2 mmHg; 95%CI, 1.2 to 2.9, p=<0.001), mean pulmonary arterial pressures (2.0 mmHg; 95%CI, 0.09 to 4.0, p=0.041) and pulmonary capillary wedge pressures (0.75mmHg; 95%CI, 0.18 to 1.3, p=0.010). Cox multivariable analysis demonstrated higher serum IGFBP4 as an independent predictor of survival in the overall PAHB cohort (See Figure 1; HR, 3.6; 95% CI, 2.6-5.0), with worse outcomes for CHD-APAH (HR, 9.9; CI, 2.2-45) and IPAH (HR, 5.3; 95% CI, 2.9-9.7).Conclusions:Higher circulating IGFBP4 levels are significantly associated with worse PAH severity and shorter survival. Dysregulation of IGF metabolism/growth axis could play a significant role in PAH cardio-pulmonary pathobiology.

Background:Pulmonary arterial hypertension (PH) is a progressive disease of the pulmonary vasculature. Risk assessment to guide treatment is fundamental to improved outcomes. Current risk models rely on invasive (cardiac cath) and subjective features, prompting need for a non-invasive risk assessment tool in PH.Hypothesis:A multi-marker protein model will accurately predict PH survivalMethods:Using a custom multiplex ELISA (based on prior models), we measured proteins targeting pathobiologic pathways in PAH; NTproBNP (cardiac function), ST2 (cardiac fibrosis), IL-6 (inflammation), endostatin (angiogenesis), and HDGF (vascular growth). Subjects were enrolled from the PH Biobank (PAHB, N=2335, age>15) and a validation cohort from Johns Hopkins and Vanderbilt Universities (N=250, age>18). Unsupervised k-means clustering classified subjects by biomarkers. Transplant free survival by cluster were examined using Kaplan-Meier and Cox proportional hazard modeling. Risk by cluster was compared to current clinical models (REVEAL and ESC/ERS Risk) using Harrell’s C Index.Results:The algorithm generated 4 clusters with excellent risk discrimination in both cohorts (Figure 1). The HR for the severe group (vs lowest risk) in the PAHB was 10.1, (C Index 0.72), while in the validation the HR was 8.6 (C Index 0.74). Addition of genetic mutations, age, sex, and comorbidities did not improve model fit. When compared to the REVEAL and ERS/ESC scores (C Index 0.69 and 0.59 respectively), the proteomic model had better discrimination.Conclusions:This multimarker proteomic model non-invasively assessed PH severity and prognosis with improved accuracy compared to current clinical models. A simple blood test may enable frequent assessment of PH patients in a routine clinic setting to support therapeutic decision-making. In addition, this non-invasive model may provide a surrogate endpoint for future clinical trial designs (after prospective validation).

Introduction:Pulmonary arterial hypertension (PAH) is a progressive disease involving inflammation, endothelial dysfunction, and vascular remodeling. Inflammation is increasingly recognized as part of the pathobiology of PAH, so noninvasive serum inflammatory biomarkers may offer insight into disease management and survival.Hypothesis:We hypothesized that circulating levels of interleukin 6 (IL-6), a pro-inflammatory cytokine, are associated with PAH severity and survival.Methods:Serum IL-6 levels were measured by ELISA using samples and clinical data from enrollees (N=2017) in the NHLBI PAH Biobank. Relationships between IL-6 levels, clinical variables, and mortality were analyzed with regression and Cox proportional hazard models adjusted for demographic and hemodynamic variables.Results:The cohort was 80% female and had a mean age of 55 and median 6-minute walk distance (6MWD) of 348 meters. The majority had idiopathic (43%) or collagen vascular disease-associated (31%) PAH. Each log-unit higher IL-6 was associated with a 16.0 meter shorter 6MWD (95% CI 10.2-21.7, p<0.001), 0.50 mmHg higher right atrial pressure (95% CI 0.33-0.67, p<0.001), 17% greater odds of having dyspnea at rest (OR 1.17, 95% CI 1.07-1.28, p=0.001), 8% greater odds of requiring treatment with prostacyclin analogs (OR 1.08, 95% CI 1.01-1.15, p=0.019), and greater odds of having PAH associated with collagen vascular disease (OR 1.22, 95% CI 1.13-1.31, p<0.001) or portal hypertension (OR 1.37, 95% CI 1.18-1.59, p<0.001). IL-6 was also associated with a higher REVEAL score (p<0.001), a predictive algorithm for 1-year PAH survival, and a 21% greater risk of death for each log-unit higher IL-6 (HR 1.21, 95% CI 1.06-1.37, p=0.003).Conclusions:Serum IL-6 levels are significantly associated with PAH severity and survival. As a result, IL-6 may offer a tool to guide disease management, especially for patients with collagen vascular disease and portal hypertension-associated PAH.

Introduction:Pediatric pulmonary hypertension (PH) is a severe disease defined by sustained elevation of pulmonary vascular resistance (PVRi). Noninvasive diagnostic markers, as well as predictors of clinical changes over time, have been elusive.Hypothesis:We hypothesize that ST2 is associated with hemodynamic and functional changes in pediatric pulmonary hypertension.Methods:Two cohorts of patients were studied; a cross-sectional cohort from the NHLBI National Biological Sample and Data Repository for PAH (PAH Biobank, N=175), and a second longitudinal cohort from Children?s Hospital of Colorado (CHC, N=61). Serum samples were tested using a custom robotically spotted ST2 ELISA. Association of ST2 with clinical variables was evaluated using adjusted linear regression and Kaplan Meier analysis.Results:Adjusted Linear regression (age, race, gender) was employed to evaluate ST2 concentration as a predictor of pulmonary vascular resistance (PVRi), cardiac output (CO), and six-minute walk distance (6MWD) in each cohort at enrollment. ST2 was associated with shorter 6MWD, and increased PVRi in both cohorts (Biobank: 6MWD p=0.03, PVRi p=0.019; CHC: 6MWD p=0.012, PVRi p=0.001). An adjusted longitudinal regression analysis was used on the CHC cohort to evaluate the performance of ST2 over time. ST2 was significantly associated with higher PVRi (p<0.001), shorter 6MWD (p=0.023), and higher mean pulmonary artery pressure (p<0.001) over time. Kaplan Meier analysis demonstrated that ST2 was predictive of death, transplant, or palliative surgery (p=0.036).Conclusions:Two successive pediatric PAH cohorts showed ST2 elevation in association with unfavorable hemodynamic and functional measures. ST2 concentration further shows significant association with hemodynamic and functional changes over time. This suggests that ST2 serum elevation reflects PH severity, can be used diagnostically, and as a longitudinal marker of clinical worsening.