Integrative multi-omics analysis reveals novel idiopathic pulmonary fibrosis endotypes associated with disease progression.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is characterized by the accumulation of extracellular matrix in the pulmonary interstitium and progressive functional decline. We hypothesized that integration of multi-omics data would identify clinically meaningful molecular endotypes of IPF. METHODS: The IPF-PRO Registry is a prospective registry of patients with IPF. Proteomic and transcriptomic (including total RNA [toRNA] and microRNA [miRNA]) analyses were performed using blood collected at enrollment. Molecular data were integrated using Similarity Network Fusion, followed by unsupervised spectral clustering to identify molecular subtypes. Cox proportional hazards models tested the relationship between these subtypes and progression-free and transplant-free survival. The molecular subtypes were compared to risk groups based on a previously described 52-gene (toRNA expression) signature. Biological characteristics of the molecular subtypes were evaluated via linear regression differential expression and canonical pathways (Ingenuity Pathway Analysis [IPA]) over-representation analyses. RESULTS: Among 232 subjects, two molecular subtypes were identified. Subtype 1 (n = 105, 45.3%) and Subtype 2 (n = 127, 54.7%) had similar distributions of age (70.1 +/- 8.1 vs. 69.3 +/- 7.6 years; p = 0.31) and sex (79.1% vs. 70.1% males, p = 0.16). Subtype 1 had more severe disease based on composite physiologic index (CPI) (55.8 vs. 51.2; p = 0.002). After adjusting for CPI and antifibrotic treatment at enrollment, subtype 1 experienced shorter progression-free survival (HR 1.79, 95% CI 1.28,2.56; p = 0.0008) and similar transplant-free survival (HR 1.30, 95% CI 0.87,1.96; p = 0.20) as subtype 2. There was little agreement in the distribution of subjects to the molecular subtypes and the risk groups based on 52-gene signature (kappa = 0.04, 95% CI= -0.08, 0.17), and the 52-gene signature risk groups were associated with differences in transplant-free but not progression-free survival. Based on heatmaps and differential expression analyses, proteins and miRNAs (but not toRNA) contributed to classification of subjects to the molecular subtypes. The IPA showed enrichment in pulmonary fibrosis-relevant pathways, including mTOR, VEGF, PDGF, and B-cell receptor signaling. CONCLUSIONS: Integration of transcriptomic and proteomic data from blood enabled identification of clinically meaningful molecular endotypes of IPF. If validated, these endotypes could facilitate identification of individuals likely to experience disease progression and enrichment of clinical trials. TRIAL REGISTRATION: NCT01915511.

Circulating matrix metalloproteinases and tissue metalloproteinase inhibitors in patients with idiopathic pulmonary fibrosis in the multicenter IPF-PRO Registry cohort.

BACKGROUND: Matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) play important roles in the turnover of extracellular matrix and in the pathogenesis of idiopathic pulmonary fibrosis (IPF). This study aimed to determine the utility of circulating MMPs and TIMPs in distinguishing patients with IPF from controls and to explore associations between MMPs/TIMPs and measures of disease severity in patients with IPF. METHODS: The IPF cohort (n = 300) came from the IPF-PRO Registry, an observational multicenter registry of patients with IPF that was diagnosed or confirmed at the enrolling center in the past 6 months. Controls (n = 100) without known lung disease came from a population-based registry. Generalized linear models were used to compare circulating concentrations of MMPs 1, 2, 3, 7, 8, 9, 12, and 13 and TIMPs 1, 2, and 4 between patients with IPF and controls, and to investigate associations between circulating levels of these proteins and measures of IPF severity. Multivariable models were fit to identify the MMP/TIMPs that best distinguished patients with IPF from controls. RESULTS: All the MMP/TIMPs analyzed were present at significantly higher levels in patients with IPF compared with controls except for TIMP2. Multivariable analyses selected MMP8, MMP9 and TIMP1 as top candidates for distinguishing patients with IPF from controls. Higher concentrations of MMP7, MMP12, MMP13 and TIMP4 were significantly associated with lower diffusion capacity of the lung for carbon monoxide (DLCO) % predicted and higher composite physiologic index (worse disease). MMP9 was associated with the composite physiologic index. No MMP/TIMPs were associated with forced vital capacity % predicted. CONCLUSIONS: Circulating MMPs and TIMPs were broadly elevated among patients with IPF. Select MMP/TIMPs strongly associated with measures of disease severity. Our results identify potential MMP/TIMP targets for further development as disease-related biomarkers.

Peripheral blood proteomic profiling of idiopathic pulmonary fibrosis biomarkers in the multicentre IPF-PRO Registry.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease for which diagnosis and management remain challenging. Defining the circulating proteome in IPF may identify targets for biomarker development. We sought to quantify the circulating proteome in IPF, determine differential protein expression between subjects with IPF and controls, and examine relationships between protein expression and markers of disease severity. METHODS: This study involved 300 patients with IPF from the IPF-PRO Registry and 100 participants without known lung disease. Plasma collected at enrolment was analysed using aptamer-based proteomics (1305 proteins). Linear regression was used to determine differential protein expression between participants with IPF and controls and associations between protein expression and disease severity measures (percent predicted values for forced vital capacity [FVC] and diffusion capacity of the lung for carbon monoxide [DLco]; composite physiologic index [CPI]). Multivariable models were fit to select proteins that best distinguished IPF from controls. RESULTS: Five hundred fifty one proteins had significantly different levels between IPF and controls, of which 47 showed a |log2(fold-change)| > 0.585 (i.e. > 1.5-fold difference). Among the proteins with the greatest difference in levels in patients with IPF versus controls were the glycoproteins thrombospondin 1 and von Willebrand factor and immune-related proteins C-C motif chemokine ligand 17 and bactericidal permeability-increasing protein. Multivariable classification modelling identified nine proteins that, when considered together, distinguished IPF versus control status with high accuracy (area under receiver operating curve = 0.99). Among participants with IPF, 14 proteins were significantly associated with FVC % predicted, 23 with DLco % predicted, 14 with CPI. Four proteins (roundabout homolog-2, spondin-1, polymeric immunoglobulin receptor, intercellular adhesion molecule 5) demonstrated the expected relationship across all three disease severity measures. When considered in pathways analyses, proteins associated with the presence or severity of IPF were enriched in pathways involved in platelet and haemostatic responses, vascular or platelet derived growth factor signalling, immune activation, and extracellular matrix organisation. CONCLUSIONS: Patients with IPF have a distinct circulating proteome and can be distinguished using a nine-protein profile. Several proteins strongly associate with disease severity. The proteins identified may represent biomarker candidates and implicate pathways for further investigation. TRIAL REGISTRATION: ClinicalTrials.gov (NCT01915511).

Effects of BI 655064, an antagonistic anti-CD40 antibody, on clinical and biomarker variables in patients with active rheumatoid arthritis: a randomised, double-blind, placebo-controlled, phase IIa study.

OBJECTIVE: To evaluate the safety, efficacy and therapeutic mechanism of BI 655064, an antagonistic anti-CD40 monoclonal antibody, in patients with rheumatoid arthritis (RA) and an inadequate response to methotrexate (MTX-IR). METHODS: In total, 67 patients were randomised to receive weekly subcutaneous doses of 120 mg BI 655064 (n=44) or placebo (n=23) for 12 weeks. The primary endpoint was the proportion of patients who achieved 20% improvement in American College of Rheumatology criteria (ACR20) at week 12. Safety was assessed in patients who received at least one dose of study drug. RESULTS: At week 12, the primary endpoint was not met, with 68.2% of patients treated with BI 655064 achieving an ACR20 vs 45.5% with placebo (p=0.064); using Bayesian analysis, the posterior probability of seeing a difference greater than 35% was 42.9%. BI 655064 was associated with greater changes in CD40-CD40L pathway-related markers, including reductions in inflammatory and bone resorption markers (interleukin-6, matrix metalloproteinase-3, receptor activator of nuclear factor-κB ligand), concentration of autoantibodies (immunoglobulin [Ig]G rheumatoid factor [RF], IgM RF, IgA RF) and CD95+ activated B-cell subsets. No serious adverse events (AEs) related to BI 655064 treatment or thromboembolic events occurred; reported AEs were mainly of mild intensity. CONCLUSION: Although blockade of the CD40-CD40L pathway with BI 655064 in MTX-IR patients with RA resulted in marked changes in clinical and biological parameters, including reductions in activated B-cells, autoantibody production and inflammatory and bone resorption markers, with a favourable safety profile, clinical efficacy was not demonstrated in this small phase IIa study. TRIAL REGISTRATION NUMBER: NCT01751776.

Psoriatic skin molecular and histopathologic profiles after treatment with risankizumab versus ustekinumab.

BACKGROUND: IL-23 contributes to the activation, maintenance, and proliferation of TH17 cells and plays a major role in psoriasis pathophysiology. IL-23p19 inhibition with risankizumab resulted in superior clinical responses in patients with psoriasis compared with ustekinumab (dual IL-12/IL-23 inhibitor), but comparative molecular effects have not been established. OBJECTIVE: We investigated the similarities and differences in molecular and histopathologic profiles in skin lesions from patients with psoriasis receiving risankizumab versus ustekinumab at an early time point. METHODS: Lesional skin biopsy samples from 81 patients with moderate-to-severe plaque psoriasis participating in 2 different studies (a phase I risankizumab study and a phase II study of risankizumab vs ustekinumab) were analyzed by using histopathology, immunohistochemistry, and RNA sequencing. RESULTS: Risankizumab induced a rapid decrease in levels of proteins and transcriptomic biomarkers associated with the IL-23 pathway, which were maintained through 8 weeks. At week 4, risankizumab decreased histopathologic expression of biomarkers, including K16, Ki67, CD3, lipocalin-2, CD11c, dendritic cell lysosome-associated membrane glycoprotein, ß-defensin 2, and S100A7; global histopathologic scoring revealed that 54% and 69% of patients treated with 90 or 180 mg of risankizumab, respectively, were graded as experiencing "excellent improvement" versus 29% of patients treated with ustekinumab. At week 4, there was a common decrease in expression of 2645 genes expressed in lesional skin between patients receiving risankizumab and ustekinumab and a significant decrease in 2682 genes unique to risankizumab treatment. Risankizumab more strongly downregulated expression of genes associated with keratinocytes, epidermal cells, and monocytes, versus ustekinumab. CONCLUSION: Risankizumab demonstrated more pronounced changes in the molecular and histopathologic profile of psoriatic skin lesions compared with ustekinumab at week 4.

Selective IL-23 Inhibition by Risankizumab Modulates the Molecular Profile in the Colon and Ileum of Patients With Active Crohn's Disease: Results From a Randomised Phase II Biopsy Sub-study.

BACKGROUND AND AIMS: We aimed to investigate the underlying mechanism of action of risankizumab, a monoclonal antibody targeting the IL-23 p19 subunit, previously reported to induce clinical and endoscopic remission in a randomised phase II study in patients with active Crohn's disease. METHODS: Ileum and colon biopsies obtained at screening and Week 12 from a subgroup of patients [n = 106] in the risankizumab phase II study were analysed by transcriptome-wide RNA-Seq profiling. Univariate associations were assessed using linear modelling. RESULTS: By Week 12, risankizumab significantly decreased [p < 0.005] the expression of 1880 and 765 genes in the colon [false-discovery rate = 0.02] and ileum [false-discovery rate = 0.05], respectively. These genes were associated with the IL-23/IL-17 axis, Th1 pathway, innate immunity, and tissue turnover. Colonic transcriptomic profiles following risankizumab treatment reflected the transcriptomic changes observed in patients achieving endoscopic response and remission at Week 12 and were significantly different from placebo [p < 0.005]. The colonic transcriptomic profile, significantly modulated by risankizumab at Week 12, was indicative of suppression of pathways associated with epithelial biology. Furthermore, pathways associated with Crohn's disease modulated by risankizumab treatment included second messenger-mediated signalling, immune response, lymphocyte and leucocyte activation, lymphocyte differentiation and cell-cell adhesion. CONCLUSIONS: Endoscopic remission and response observed with risankizumab in patients with active Crohn's disease was associated with significant transcriptomic changes in the colon, compared with placebo. Differentiated expression of genes associated with the IL-23/IL-17 axis was observed in the colon and ileum 12 weeks after risankizumab treatment.

An open-label, single-arm, phase 2 trial of the Polo-like kinase inhibitor volasertib (BI 6727) in patients with locally advanced or metastatic urothelial cancer.

BACKGROUND: Polo-like kinases (Plks) control multiple steps during the cell cycle, and Plk1 is overexpressed in urothelial cancer (UC). Volasertib (BI 6727), a Plk inhibitor, has demonstrated antitumor activity in several malignancies, including UC. In this phase 2 trial, the authors investigated volasertib as a second-line treatment in advanced/metastatic UC. METHODS: Patients who progressed within 2 years of 1 prior chemotherapy regimen received 300 mg volasertib on day 1 every 3 weeks. The dose was escalated to 350 mg in cycle 2 if volasertib was tolerated in cycle 1. The primary endpoint was tumor response, which was assessed every 6 weeks; secondary endpoints were progression-free survival, overall survival, duration of response, safety, and pharmacokinetics. RESULTS: Fifty patients were enrolled, and the median patient age was 68.5 years (range, 52-83 years). All patients had received prior platinum, 94% of patients had relapsed ≤2 years after prior therapy, 36% had liver metastases, and 54% had lung metastases. The median number of treatment cycles was 2 (range, 1-27 treatment cycles), and 23 patients were dose escalated at cycle 2. Seven patients (14%) had a partial response, 13 (26%) had stable disease, and 30 (60%) progressed within 6 weeks. The median response duration was 41 weeks (range, 29.1-77.3 weeks). The median progression-free survival was 1.4 months, and the median overall survival was 8.5 months. The most frequent grade 3 and 4 adverse events were neutropenia (28%), thrombocytopenia (20%), and anemia (16%). No cumulative toxicity was observed. CONCLUSIONS: Volasertib as second-line treatment for advanced/metastatic UC had an acceptable safety profile but demonstrated insufficient antitumor activity for further evaluation as a monotherapy.

Types of stroke recurrence in patients with ischemic stroke: a substudy from the PRoFESS trial.

BACKGROUND: Risk profiles for stroke recurrence are poorly characterized. AIMS: We determined the variation in the risk and type of recurrent stroke among index ischemic stroke subtypes, and whether index stroke subtype and conventional stroke risk factors were predictors of stroke recurrence. METHODS: Patients enrolled in the Prevention Regimen for Effectively Avoiding Second Strokes trial were included in this study. RESULTS: In 1794 patients' recurrent stroke subtypes were the same as the index stroke in: 48·3% of patients with large artery atherothrombosis stroke; 50% of patients with cardioembolic stroke; 48·7% of patients with small artery occlusion stroke; 8·1% of patients with stroke of other etiology, and 45·3% of patients with undetermined etiology stroke. Patients with cardioembolic stroke, who were unwilling or unable to take oral anticoagulants, had the greatest risk of stroke recurrence. Predictors of stroke recurrence in multivariable analysis were: older age and previous stroke among large artery atherothrombosis strokes; older age, male sex, previous stroke, previous transient ischemic attack, hypertension, diabetes, and tobacco use among small artery occlusion strokes; older age among cardioembolic strokes; atrial fibrillation and anti-diabetic medications among other etiology strokes; older age, previous stroke and atrial fibrillation among undetermined etiology strokes. Predictors of brain hemorrhage as recurrent stroke were index small artery occlusion stroke, older age, previous stroke, and antiplatelet treatment with aspirin plus extended-release dipyridamole. CONCLUSIONS: Risk predictors for stroke recurrence and for brain hemorrhage differ by index ischemic stroke subtype, information that is important when initiating secondary prevention therapy.

A phase II study of afatinib (BIBW 2992), an irreversible ErbB family blocker, in patients with HER2-positive metastatic breast cancer progressing after trastuzumab.

Afatinib is an oral, ErbB family blocker, which covalently binds and irreversibly blocks all kinase-competent ErbB family members. This phase II, open-label, single-arm study explored afatinib activity in human epidermal growth factor receptor 2 (HER2)-positive breast cancer patients progressing after trastuzumab treatment. Patients had stage IIIB/IV HER2-positive metastatic breast cancer, with progression following trastuzumab or trastuzumab intolerance and an Eastern Cooperative Oncology Group (ECOG) performance status of 0-2. Patients received 50 mg afatinib once-daily until disease progression. Primary endpoint was objective response rate (Response Evaluation Criteria in Solid Tumors 1.0), with tumor assessments every 8 weeks. Forty-one patients were treated. Patients had received a median of three prior chemotherapy lines (range, 0-15) and 68.3% had received trastuzumab for >1 year. Four patients (10% of 41 treated; 11% of evaluable patients) had partial response. Fifteen patients (37% of 41) had stable disease as best response and 19 (46% of 41) achieved clinical benefit. Median progression-free survival was 15.1 weeks (95% confidence interval [CI]: 8.1-16.7); median overall survival was 61.0 weeks (95% CI: 56.7-not evaluable). Most frequent common terminology criteria for adverse events grade 3 treatment-related adverse events were diarrhea (24.4%) and rash (9.8%). Afatinib monotherapy was associated with promising clinical activity in extensively pretreated HER2-positive breast cancer patients who had progressed following trastuzumab treatment.