Augmented antitumor activity of 5‐fluorouracil by double knockdown of MDM4 and MDM2 in colon and gastric cancer cells

Abstract

Inactivation of the TP53 tumor suppressor gene is essential during cancer development and progression. Mutations of TP53 are often missense and occur in various human cancers. In some fraction of wild‐type (wt) TP53 tumors, p53 is inactivated by upregulated murine double minute homolog 2 (MDM2) and MDM4. We previously reported that simultaneous knockdown of MDM4 and MDM2 using synthetic DNA‐modified siRNAs revived p53 activity and synergistically inhibited in vitro cell growth in cancer cells with wt TP53 and high MDM4 expression (wtTP53/highMDM4). In the present study, MDM4/MDM2 double knockdown with the siRNAs enhanced 5‐fluorouracil (5‐FU)‐induced p53 activation, arrested the cell cycle at G1 phase, and potentiated the antitumor effect of 5‐FU in wtTP53/highMDM4 human colon (HCT116 and LoVo) and gastric (SNU‐1 and NUGC‐4) cancer cells. Exposure to 5‐FU alone induced MDM2 as well as p21 and PUMA by p53 activation. As p53‐MDM2 forms a negative feedback loop, enhancement of the antitumor effect of 5‐FU by MDM4/MDM2 double knockdown could be attributed to blocking of the feedback mechanism in addition to direct suppression of these p53 antagonists. Intratumor injection of the MDM4/MDM2 siRNAs suppressed in vivo tumor growth and boosted the antitumor effect of 5‐FU in an athymic mouse xenograft model using HCT116 cells. These results suggest that a combination of MDM4/MDM2 knockdown and conventional cytotoxic drugs could be a promising treatment strategy for wtTP53/highMDM4 gastrointestinal cancers.

Apolipoprotein M promotes growth and inhibits apoptosis of colorectal cancer cells through upregulation of ribosomal protein S27a

“…Among these genes, MDM4 has been reported to play an important role in negative regulation of the tumor suppressor p53 through its interaction with MDM2 (Gansmo et al, 2015[ 7 ]). MDM4/ MDM2 double knockdown with siRNAs enhanced 5-fluorouracil (5-FU)-induced p53 activation, arrested cell cycle at G1 phase, and potentiated the antitumor effect of 5-FU in wtTP53/highMDM4 human colon cancer cells (Imanishi et al, 2019[ 10 ]). Suda et al showed that MDM4 was abnormally expressed in CRC tumor tissues compared with levels in matched normal mucosa and speculated that MDM4 might play a role in colorectal carcinogenesis (Suda et al, 2011[ 32 ]).…”

Isoforms of the p53 Family and Gastric Cancer: A Ménage à Trois for an Unfinished Affair

“…For instance, MDM4 has an elevated expression that correlates with lymph node metastasis [ 164 ]. Inversely, the downregulation of MDM4 increases slightly the activity of cytotoxic drugs (5-FU) in GC cells [ 168 ]. Another example is the LIM domain protein ENIGMA , whose overexpression is caused by SRF [ 165 ].…”

MDM4 as a Prognostic Factor for Patients With Gastric Cancer With Low Expression of p53

“…Immunoblot analysis: Immunoblot analysis was performed as described previously (16). To circumvent the low linear dynamic range of immunoblot analysis, serially diluted total protein extracts from MDM4-NUGC4 cells and undiluted extracts from EGFP-NUGC4 cells were electrophoresed and probed for MDM4 levels.…”