Impact of pre-analytic procurement and processing variables on the detection of HCC DNA in urine Hepatocellular carcinoma (HCC) is the world’s 2nd leading cause of cancer-related death and one of the fastest-growing cancers in the US; 85% of patients die within 5 years, mainly due to late detection, limited treatment options, and high recurrence. Currently, HCC recurrence and treatment responses are monitored by serum AFP and serial imaging. Unfortunately, serum AFP can only be used for ~50% of HCC cases due to low sensitivity. While MRI/CT imaging is the gold standard for diagnosis, it is expensive and less accessible. It also has limited utility in the detection of small tumors (< 2 cm) and is challenging in the presence of dysplastic nodules. Early detection of recurrent HCC is ineffective with the current available methods. Genetic liquid biopsies have been routinely used in clinics for almost all cancer types as part of the National Comprehensive Cancer Network (NCCN) guideline. However, its usage in HCC is limited even though its promises have been well documented in the literature, mostly for blood and some for urine. Our hypothesis is that by establishing an evidence-based biospecimen practice for urine collection, storage, and transrenal DNA (trDNA) isolation, urine HCC DNA analysis with tailored sensitive assays can provide HCC genetics with more frequent tumor surveillance and treatment response assessment compared to current standard of care for prognosis. As a pioneer in developing urine cancer liquid biopsies, we have established the standard operating procedure (SOP) for urine collection for the Early Detection Research Network colon cancer validation study (protocol ID: 320) and other non-urinary tract cancers. We have identified pre-analytic variables that influence urine HCC DNA assay performance in the process of developing an HCC urine DNA test for HCC liquid biopsies. In this application, we propose to use a liver-specific trDNA marker, hepatitis B virus (HBV) DNA, which is found in urine, from patients with chronic HBV infection to validate and mitigate these pre-analytic variables. Aims 1 and 2, will identify, validate, and mitigate pre-analytic factors, associated with urine collection, storage and processing for trDNA isolation, and characterize of the size of trDNA that influences clinical assay results and reproducibility. As a result, an evidence-based urine biospecimen practice can be established to help support urine HCC DNA assay development, validation for HCC genetic liquid biopsy and assessment of treatment response. In Aim 3, we will then demonstrate the reproducibility and clinical validation of the utility of urine HCC DNA in assessing responses to HCC treatment. Developing a robust evidence-based urine biospecimen practice for sample collection, processing, and long-term storage will lead to the validation of urine tests for assessing HCC treatment in clinical trials.

Hepatocellular carcinoma (HCC) is the world’s 2nd leading cause of cancer-related death and one of the fastest-growing cancers in the US; 85% of patients die within 5 years, mainly due to late detection, limited treatment options, and high recurrence. This project is to develop a robust evidence-based urine biospecimen practice for sample collection, processing, and long-term storage will lead to the validation of urine tests for assessing HCC treatment in clinical trials.