2021

Liquid Biopsy for Active Surveillance of Fibrolamellar Hepatocellular Carcinoma

Goal: Develop a blood-based screening method to detect relapse in FLC patients

Principal Investigator: Gary S. Stein, PhD

Grant length: One year

Study overview: This study is focused on developing a blood-based screening method to detect relapse in FLC patients that have been diagnosed and treated by surgery. This screening protocol is intended to be non-invasive, to overcome limitations of current imaging strategies, and to be accessible and affordable for patients and their families. Currently, treatment of patients with FLC is limited by the understanding of how this cancer responds to therapies. Cancer physicians currently have limited ability to test for, and monitor patients’ response to therapy, and to detect disease progression. It is therefore important to have effective monitoring strategies for physicians to determine if patients are successfully remaining in remission, or if they are progressing towards the earliest stages of relapse. Catching relapsed tumors early, prior to spreads throughout the body, is critically important to increase FLC patient survival.

The study is evaluating the use of DnaJ-PKAc transcripts in circulating nucleated cells as a proxy for detecting the presence of circulating tumor cells (CTCs). A major challenge to detecting and quantitating CTCs is their extremely low abundance – roughly 1 to >100 per 7.5 mL of whole blood, compared to the billions of other cells typically found in blood. The objective of this initiative is to test whether the DnaJ-PKAc transcript can be detected in blood samples and to establish the threshold of transcript detection. We will utilize an engineered model of FLC that possesses the DnaJ-PKAc fusion gene as a substitute for actual CTCs. These pseudo-CTCs will be spiked into blood samples from normal donors to simulate whole blood from a metastatic or progressive disease scenario.

2016

Characterizing enzyme inhibition of the DnaJPKAc chimeric protein derived from fibrolamellar hepatocellular carcinomas

Goal: Characterize inhibition of the DnaJ-PKAc chimeric protein in fibrolamellar carcinoma

Principal Investigator: Hibba tul Rehman, MD

Grant length: One year

Study overview: The DNAJB1-PRKACA fusion gene produces the DNAJ-PKAc fusion kinase protein, which is present in nearly all FLC tumors and promotes liver tumor formation in mice. Kinases, including fusion kinases, have been successful drug targets in numerous cancer types. Inhibition of DnaJ-PKAc may provide a targeted therapy for FLC. This study proposed a two pronged approach towards identifying therapeutic inhibitors of the fusion:

  1. Screening a previously developed peptide library to identify peptides that preferentially bind chimeric DnaJ-PKAc over normal, wide-type protein in vitro.
  2. Developing a library of inhibitory peptides that would preferentially inhibit DnaJ-PKAc.

The aim of these efforts was to develop an understanding that will support the development of inhibitors that regulate the function of the chimeric kinase without affecting the wild-type kinase, thus selectively targeting cancer cells without affecting healthy tissue.

Key Findings: The study analyzed a library of small inhibitor peptides derived from endogenous PKI (which inhibit wild-type PKA in the body), which showed no preferential competitive inhibition of wild-type PKA versus DnaJ-PKAc.  This result was due to the overwhelming structural similarities observed at the active sites of wtPKA and DnaJ-PKAc.  A survey of publicly-available data sets to determine the predominant isoform(s) of PKI expressed in normal liver and FLC tumors showed that PKIβ was likely to be the predominant isoform expressed in human liver.  Upon testing of the inhibitory properties of full-length PKIβ, identical inhibition of both wtPKAc and DnaJ-PKAc was observed.

Details of the analysis was published in April 2019 in The Journal of Cellular Biochemistry. The full article can be read or downloaded here.

Implications: This study showed the importance of adopting a targeted approach to inhibiting DNAJ-PKAc in FLC.