Introduction:
Hallmarks of cancer development, progression and emergence of therapy resistance are
associated with embryonic and/or stem cell characteristics. Lin 28 homolog A (LIN28A)
is a proto-oncogene and RNA binding protein, which represses the let-7 family of miRNAs
in embryonic stem cells. The microRNA (miR) let-7 family comprises the most prominent
tumorsuppressive microRNAs. However, the exact role of the embryonic stem cell marker
LIN28A is unclear in hepatocellular carcinoma (HCC) progression and acquired sorafenib-resistance.
Moreover, it is unknown whether LIN28A is only an upstream-regulator of miRs, or if
LIN28A itself could also be regulated by miRs.
Methods:
Primary human hepatocytes and several human HCC cell lines (PLC, Hep3B, HepG2 and
Huh 7) as well as sorafenib-resistant HCC cells were used for expression and functional
analysis. In vivo expression analysis was performed using paired tumor and corresponding non-tumor
liver tissues from HCC patients. Gene expression and cellular localization was analyzed
qRT-PCR, Western blot, and Fluorescence microscopy. A microRNA mimic/inhibitor for
miR-622 was used for functional analysis. For exploration of stem cell properties,
clonogenicity assays were performed.
Results:
LIN28A expression was increased in HCC cell lines as compared to primary human hepatocytes.
Tissue micro array analysis revealed enhanced LIN28A protein expression in HCC in vivo and correlated with advanced tumor stages. Fluorescence-based microscopic analysis
showed that LIN28A can both localize to the cytoplasm and the nucleus in HCC cells
and tissues. Furthermore, LIN28A levels were higher in sorafenib-resistant HCC cell
lines as compared to non-resistant cells. Clonogenicity assays revealed that LIN28A
induces stem cell properties in HCC cells. Multi-step based in silico and experimental analysis revealed a novel feedback regulation of LIN28A expression
in HCC mediated by the tumorsuppressive microRNA-622.
Conclusions:
We found that the embryonic stem cell marker LIN28A is overexpressed in HCC in vitro and in vivo and correlated with tumor stages. LIN28A levels were also enhanced in sorafenib-resistance
and LIN28A was sufficient to induce stem cell properties. Further, we detected a novel
feedback inhibition of LIN28A expression which was mediated by the tumorsuppressive
microRNA-622 in hepatocellular carcinoma.