The SLIT/ROBO Pathway in Liver Fibrosis and Cancer

Abstract

Liver fibrosis is a common outcome of most chronic liver insults/injuries that can develop into an irreversible process of cirrhosis and, eventually, liver cancer. In recent years, there has been significant progress in basic and clinical research on liver cancer, leading to the identification of various signaling pathways involved in tumorigenesis and disease progression. Slit glycoprotein (SLIT)1, SLIT2, and SLIT3 are secreted members of a protein family that accelerate positional interactions between cells and their environment during development. These proteins signal through Roundabout receptor (ROBO) receptors (ROBO1, ROBO2, ROBO3, and ROBO4) to achieve their cellular effects. The SLIT and ROBO signaling pathway acts as a neural targeting factor regulating axon guidance, neuronal migration, and axonal remnants in the nervous system. Recent findings suggest that various tumor cells differ in SLIT/ROBO signaling levels and show varying degrees of expression patterns during tumor angiogenesis, cell invasion, metastasis, and infiltration. Emerging roles of the SLIT and ROBO axon-guidance molecules have been discovered in liver fibrosis and cancer development. Herein, we examined the expression patterns of SLIT and ROBO proteins in normal adult livers and two types of liver cancers: hepatocellular carcinoma and cholangiocarcinoma. This review also summarizes the potential therapeutics of this pathway for anti-fibrosis and anti-cancer drug development.

Keywords: ROBOs; SLITs; cholangiocarcinoma; hepatocellular carcinoma; liver cancer; liver fibrosis.

Figure 1

SLIT/ROBO signaling. SLIT1-3 ligands can bind to ROBO1-3 in cell type-specific manners. Within activated HSC, SLIT2 binding to ROBO1 or ROBO2 has shown to activate downstream PI3K/Akt signaling, leading to the induction of pro-fibrotic factors, such as Ctgf and collagen. Three potential approaches that target the Slit/Robo pathway are also summarized. Approach 1 is to use Soluble RoboN Fc chimera to sequester the Slit ligand. Approach 2 is to take advantage of the RNA interference technique (microRNA or siRNA). Approach 3 is based on ROBO1 surface antigen as an immunotherapeutic target. ROBO1 may be used with immune checkpoint inhibitors, including CTLA-4, PD-L, and PD-1, for peptide-based vaccines in anti-HCC treatment. Domain structures of the vertebrates SLIT and ROBO proteins are shown. Abbreviations: SS: Signal peptide; LRR: leucine-rich repeat; EGF: epidermal growth factor; LG: laminin G; EGF: epidermal growth factor; C: cysteine knot; IgG: immunoglobulin-l; FN3: fibronectin type III domain; CC0-3: conserved cytoplasmic motif 0-3; CTLA-4: cytotoxic T-lymphocyte-associated protein 4; PD-L1: Programmed death-ligand 1; PD-1: Programmed death protein 1; miRNA: MicroRNA.

Figure 2

Expression patterns of ROBOs and SLITs transcripts in different cell types of normal human livers. (A) Graphs were generated based on maximal nTPM in Single Cell data of Human Protein Atlas (https://www.proteinatlas.org, 2 January 2023). (B) A diagram summarizes SLIT and ROBO gene product expression patterns in different liver cell types.

Figure 3

Altered mRNA levels of SLIT and ROBO in human CCA. Graph data were extracted from a GEO dataset (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE26566, accessed on 2 January 2023).