NAFLD Clinical Trials


Non-alcoholic fatty liver disease (NAFLD), a condition where fat accumulates in the liver without the excess consumption of alcohol, has become the most common cause of liver disease in the Western world and is quickly rising to become the primary cause of liver transplants. Due to the rise in obesity and diabetes, NAFLD is estimated to affect 30 % of the global population, with a high prevalence on all continents.

This increase is most likely related to the so-called Western lifestyle; fast food, lifestyle changes, and reduced physical activity. NAFLD starts out as steatosis which is an accumulation of fat in the liver and may further progress to non-alcoholic steatohepatitis (NASH), a more serious form of NAFLD where the liver has become inflamed. NASH is a potentially fatal condition that affects around 5% of the global adult population and can further develop into development of fibrotic tissue in the liver, with possible cirrhosis as a result.  


The role of keratin 18 in NASH

Keratin 18 (K18) is an intermediate filament protein forming part of the cytoskeletal structure expressed in epithelial cells. Hepatocytes, a form of epithelial cell, are the main tissue cells found in the liver.

When simple steatosis in NAFLD is accompanied by inflammation of hepatocytes, the disease is described as NASH. This prominent characteristic of the disease is mainly caused by hepatocyte cell death due to apoptosis and/or necrosis. Early on in the apoptosis of hepatocytes, caspases are activated which cleave the protein (K18), and the resulting fragments are subsequently released into the blood.

These caspase cleaved K18 (ccK18) fragments can be efficiently quantified by the unique M30 Apoptosense® ELISA. In contrast, during hepatocyte necrosis, full length K18 may also be released into the blood without being cleaved. The M65® ELISA and the M65 EpiDeath® ELISA can quantify the amount of both ccK18 fragments and full length K18, thus measuring the amount of total hepatocyte cell death.

VLVbio K18 assays in NAFLD Clinical Trials

Due to the high world wide prevalence of NAFLD and NASH, and the fact that there is no approved drug therapy for the disease, many clinical trials are underway to fulfill the need for pharmacological treatment options. The primary endpoints for NAFLD clinical trials are resolution of NASH and/or improvement in liver fibrosis by one stage. 

There are currently important challenges in evaluating novel therapies for NAFLD, which are the difficulties in cost-effective of study subjects with defined disease and the need to use surrogate biomarkers to monitor the early effects of therapy in order to predict histologic response. The early prediction of histologic response could decrease the need for biopsies which are invasive, expensive and carry the risk of subject morbidity.

The VLVbio K18 assays; M30 Apoptosense® ELISA, M65® ELISA and M65 EpiDeath® ELISA, can aid in meeting these challenges and are currently being used in multiple active NAFLD clinical trials.

VLVbio K18 assays in Pre-Screening Subjects for Inclusion in Clinical Trials

A major challenge in studying potential therapies for NAFLD is the obtaining and pre-screening of study subjects with defined disease to be included in the trials. NAFLD clinical trials generally include subjects with fibrotic NASH, defined as a NAS of ≥4 and a fibrosis stage of F≥2. However, up to 50 % of screened subjects will not meet these eligibility criteria when assessed by liver biopsy. Traditional biochemical indicators of NAFLD may be insufficient to identify disease severity leading to the preliminary inclusion of subjects lacking the required, defined characteristics of the disease. This then leads to many subjects selected for the trial being later excluded based on biopsy findings. This adds significantly to the cost and duration of clinical trials, while causing unnecessary pain and morbidity in the subjects.

Liver cell injury is a major mechanism in NAFLD progression and the VLVbio K18 assays have been shown to correlate with steatosis and inflammation, while being elevated in NAFLD patients. Thus the VLVbio K18 assays may be used as a pre-screening tool to identify subjects with progressive disease, speeding patient recruitment and reducing the number of biopsies. 

VLVbio K18 assays as an Endpoint in Clinical Trials 

Histological improvement is the gold standard in NAFLD therapeutic trials. However, it is of importance to use biomarkers of hepatic injury in order to predict histological improvement as it can take a long time to occur.

Furthermore, early phase studies are generally of limited duration (12 to 18 months), and histological endpoints may be difficult to meet in this timeframe. Sanyal et al. highlight the need for better diagnostic and therapeutic modalities for NASH, recommending a decrease in markers of hepatic injury or cell death such as the VLVbio K18 assays, as a secondary endpoint for early phase NASH trials. 

The VLVbio K18 assays may be useful as a cost-effective surrogate biomarker to demonstrate response to treatment, providing a clearer picture of the subjects’ status at the start of therapy, and to enable the results of therapy to be seen earlier and more clearly.

The VLVbio K18 assay levels have been shown to correlate with improvement in histology in many published NAFLD clinical trials.

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