Prospective external validation of a new non-invasive test for the diagnosis of non-alcoholic steatohepatitis in patients with type 2 diabetes

Abstract

Background: One of the unmet needs in patients with type 2 diabetes mellitus (T2DM) is the prediction of non-alcoholic liver disease by non-invasive blood tests, for each of the three main histological features, fibrosis, non-alcoholic steatohepatitis (NASH) and steatosis.

Aims: To validate externally the performances of a recent panel, Nash-FibroTest, for the assessment of the severity of fibrosis stages, NASH grades and steatosis grades.

Methods: We prospectively analysed 272 patients with T2DM. Standard definitions of stages and grades were used, and analyses were centralised and blinded. The performances of the FibroTest, NashTest-2 and SteatoTest-2 were assessed using the Obuchowski measure (OM), the main outcome recommended as a summary measure of accuracy includeing all pairwise stages and grades comparisons, which is not provided par the extensively used binary area under the ROC curve.

Results: The diagnostic performance of each component of the panel was significant. OM (SE; significance) of the FibroTest, the NashTest-2 and the SteatoTest-2 was 0.862 (0.012; P < 0.001), 0.827 (0.015; P < 0.001) and 0.794 (0.020; P < 0.01), respectively. For ballooning and lobular inflammation, OM was 0.794 (0.021; P < 0.001) and 0.821 (0.017; P < 0.001), respectively. In a post hoc analysis the FibroTest outperformed VCTE by 4.1% (2.5-6.5; P < 0.001) for reliability, with a non-significant difference for OM for fibrosis staging, 0.859 (0.012) for FibroTest vs 0.870 (0.009) for VCTE.

Conclusions: From a single blood sample, the panel provides non-invasive diagnosis of the stages of fibrosis, and the grades of NASH and steatosis in patients with T2DM.

Trial registration number: NCT03634098.

© 2021 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd.

Figures

FIGURE 1

Study flow chart of core population with biopsy. Of 325 patients enrolled, 272 were eligible. Eventually among 275 patients with an interpretable biopsy, 272 had reliable FibroTest, NashTest‐2 and SteatoTest‐2. Only one patient with a non‐reliable FibroTest has been excluded

FIGURE 2

FibroTest performance in 272 type 2 diabetes patients for Fibrosis staging. (A) FibroTest was significantly different between Stage F0 (n = 54) vs F2, F3 and F4; Stage F1 (n = 65) vs F2, F3, and F4; Stage F2 (n = 50) vs F0, F1 and F4; Stage F3 (n = 74) vs F0 and F1; Stage F4 (n = 29) vs F0, F1 and F2. All 272 patients had reliable tests and centralised biopsies. The corresponding Obuchowski measure (SE; significance) was 0.862 (0.012; P < 0.001). (B) NashTest‐2 performance in 272 T2M patients for NASH grading. NashTest‐2 was significantly different between Grade A0 (n = 57) vs A2 and A3; Grade A1 (n = 51) vs A3; Grade A2 (n = 73) vs A0 and A3; Grade A3 (n = 91) vs F0, F1 and F2. The corresponding Obuchowski measure (SE; significance) was 0.827 (0.015; P < 0.001). (C) SteatoTest‐2 performance in 272 T2M patients for Steatosis grading. By definition there was no S0, and only 2 S1. SteatoTest‐2 was significantly different between grade S3 (n=207) vs S2 (n=58; P=0.03).

FIGURE 3

Spectrum of stages and grades in the original (upper row) and diabetes (lower row) subset. The spectrum of the stages of Fibrosis was not uniform in the original subset and almost uniform in diabetes. The prevalence of F3 and F4 was twice as high in diabetes as in the original subset. The difference between the mean advanced fibrosis stage and non‐advanced stage was 2.32 in diabetes and 2.11 in the original population resulting in a slight underestimation of binary AUROC for both subsets vs a perfect uniform distribution. Binary AUROCs were 0.76 and 0.84 after standardisation vs 0.72 and 0.80 before, for the diabetes and original subsets respectively

FIGURE 4

The effect of biopsy uncertainty on patient classification, due to specimen length in relation to the diagnostic performance of FibroTest (Panel A) and NashTest‐2 (Panel B). The ground truth is a large surgical liver specimen. (FP, false positive, FN, false negative). In this study with 272 patients and a median 17 mm long biopsy the expected area under the ROC curve (AUROC) of the FibroTest (or NashTest‐2) as a comparator cannot be more than 0.70 whatever its real performance due to the 30% misclassification rate of the biopsy as comparator. PPV, positive predictive value, and NPV, negative predictive value. The terms positive per cent agreement (PPA) and negative per cent agreement (NPA) are used instead of sensitivity and specificity, respectively, when the comparator is known to contain uncertainty