Interventional gene targeting of cell cycle regulators identifies Cyclin E1 as a suitable target for attenuating hepatocellular carcinoma progression

Question:

Hepatocellular carcinoma (HCC) is one of the most severe tumor diseases with limited treatment options. Early and advanced stages of carcinogenesis are associated with sustained proliferation of hepatocytes and non-parenchymal cells (e.g. hepatic stellate cells, immune cells) regulated by Cyclins and Cyclin-dependent kinases (Cdks). E-Type Cyclins (E1, E2) and Cdk2 are involved in cell cycle re-entry of quiescent cells. Our previous unpublished work demonstrated that constitutive ablation of Cyclin E1 or Cdk2 in mice prevented HCC initiation. Therefore, in the present study we tested the hypothesis that Cyclin E1 or Cdk2 could be suitable targets for interventional HCC therapy.

Methods:

We used conditional Cyclin E1 (CcnE1^f/f) or Cdk2 floxed (Cdk2^f/f) mice in a C57B6/J background with inducible Cre expression controlled by the Mx-gene promoter. This approach allows efficient gene deletion in liver cells and in the hematopoietic cell compartment. For HCC induction, 14 days old male mice were intraperitoneally (i.p.) injected with a single dose of diethylnitrosamine (DEN). Cre-negative littermates were used as controls. After 22 weeks (reflecting a stage of early HCC development), interventional inactivation of Cyclin E1 or Cdk2 was performed by three i.p. injections of poly-I:poly-C. At the age of 40 weeks, mice were analyzed for number and size of HCC nodules.

Results:

Sequence analysis of 337 human HCC samples provided by the publicly available data of The Cancer Genome Atlas (TCGA) revealed that 30% of the patients had inherited genetic alterations or aberrant expression of E-type Cyclins or Cdk2, supporting the rational for this study. All DEN-treated mice developed liver tumors at the age of 40 weeks. However, interventional inactivation of Cyclin E1 resulted in a significant reduction of tumor numbers, tumor size and liver weight indices compared to DEN-treated control mice. This finding was associated with a reduced overall proliferation and down-regulation of positive cell cycle regulators. Interestingly, Cyclin E1 inactivation also changed the composition of the HCC microenvironment (e.g. T-cells) pointing to a potential new role of Cyclin E1 for immune cell regulation during liver cancer development. In sharp contrast, interventional inactivation of Cdk2 did not reveal any significant effect on tumor growth.

Conclusions:

Cdk2 is fully dispensable for advanced HCC progression. However, interventional inactivation of Cyclin E1 in an early stage of HCC development has beneficial effects and attenuates the progression of HCC. Thus, Cyclin E1 could be a promising therapeutic target for treatment of HCC patients.