Page 99 - D. Cancer biology
P. 99
Cancer Metabolism Sensitizes Metformin Treatment
by targeting the Hippo-YAP/TAZ Pathway
Jae Hyung Park and Hyun Woo Park
Department of Biochemistry, College of Life Science and Biochemistry, Yonsei University
Abstract Introduction Figure 4. Co-treatment induced YAP/TAZ cytoplasmic
The biological significance and deregulation of the Hippo sequestering, cell death and loss of cell adhesion
pathway during organ growth and tumorigenesis have A C A C Metformin
received a surge of interest in the past decade. The Hippo YAP DAPI MERGE - Glucose
pathway core kinases, MST1/2 and LATS1/2, are tumor
suppressors that inhibits the oncogenic nuclear function of YAP
YAP/TAZ and TEAD. Dysregulation of Hippo pathway leads
to tumorigenesis via hyperactivation of YAP/TAZ. Metformin
In this study, we focused on how differential metabolic
stress response (DMSR) between normal and cancer cells, Glucose Paxilliin
such as the Warburg effect, could synergize with the anti- -
cancer effect of metformin to regulate the Hippo pathway. B Paxillin DAPI MERGE
Surprisingly, metformin treatment combined with low
glucose concentration activates the Hippo pathway, which B Cell (2015) DAPI
induces YAP/TAZ phosphorylation, loss of cell adhesion, Mst1/2 -/-
and cell death in a cancer cell-specific manner. Using Metformin
CRISPR KO cells, we show that YAP/TAZ inhibition by Glucose
metformin in combination with glucose restriction is AMPK - MERGE
and p38 MAPK-independent, but Hippo-dependent. Cell (2007) PNAS (2010)
Therefore, these results demonstrate that DMSR-based A. Hippo pathway is consisted of MST1/2, LATS1/2 core-kinase and YAP/ TAZ D
glucose restriction and metformin co-treatment can be a transcriptional co-activator Metformin: 0 3 10 mM
novel therapeutic approach to treat cancer via inhibition of B. Dysregulation of Hippo pathway disrupt organ size control and increase
tumorigenesis
cancer cell-specific YAP/TAZ activity. C. Increased aerobic glycolysis, Warburg Effect, is a hallmark of cancer + Glucose
Mel624
Figure 1. Differential Metabolic Stress Response(DMSR) synergize anti-cancer effect of metformin to regulate Hippo-YAP pathway
Glucose
A B 3T3L1 MEF SVEC -
+Glu -Glu +Glu -Glu +Glu -Glu
Metformin: 0 3 10 0 3 10 mM Metformin: 0 3 10 0 3 10 mM Metformin: 0 3 10 0 3 10 mM Metformin: 0 3 10 mM
YAP YAP YAP
(Phos-tag) (Phos-tag) (Phos-tag) + Glucose
YAP YAP YAP
TAZ TAZ TAZ 3T3L1
p-AMPK p-AMPK p-AMPK
Glucose
p-ACC p-ACC p-ACC
p-p38 p-p38 p-p38
p-ERK1/2 p-ERK1/2 p-ERK1/2 -
ERK1/2 ERK1/2 ERK1/2
LATS1 LATS1 LATS1 A. Glucose restriction and metformin induce YAP cytoplasmic sequestering
p-S6K p-S6K p-S6K B. Co-treatment reduce focal adhesion
p53 p53 p53 C. Co-treatment induce YAP cytoplasmic translocation and Focal adhesion
Vinculin Vinculin Vinculin D. Co-treatment induce cell death at cancer cell(Mel624) but not normal
IMR-90 NIH3T3 C2C12 cell(3T3L1)
+Glu -Glu +Glu -Glu +Glu -Glu
Metformin: 0 3 10 0 3 10 mM Metformin: 0 3 10 0 3 10 mM Metformin: 0 3 10 0 3 10 mM
Figure 5. Glycolysis is key factor of co-treatment induced
YAP YAP YAP
(Phos-tag) (Phos-tag) (Phos-tag) YAP/TAZ inhibition
YAP YAP YAP A B
TAZ TAZ TAZ Glucose Starvation
HEK 293A HEK 293A Metformin
p-AMPK GPI KO
p-AMPK p-AMPK WT Fructose
- Glu - Glu
p-ACC
p-ACC p-ACC Metformin: 0 3 10 0 3 10 0 3 10 0 3 10 mM
p-p38 YAP
p-p38 p-p38 (Phos-tag)
p-ERK1/2 YAP
p-ERK1/2 p-ERK1/2 (Phos-tag)
ERK1/2
LATS1 LATS1 Metformin: 0 10 mM
LATS1 YAP
ITGb3 ITGb3
p-S6K TAZ
C Mel624 HaCaT Glucose
+Glu -Glu +Glu -Glu p53 p-AMPK
Metformin: 0 3 10 0 3 10 0 3 10 0 3 10 mM Vinculin -
p-ACC
YAP p-p38
(Phos-tag) Fructose: 75 100 mM
LATS1
YAP p53
TAZ (S.E) β-catenin -Glucose +metformin
TAZ (L.E) GAPDH
p-AMPK
C D
p-ACC Metformin Glucose Starvation
Metformin
p-p38 -Glucose pyruvate
Fructose
p-ERK1/2
LATS1 YAP
(Phos-tag)
ITGb3 15h
A. Glucose restriction synergize effect of metformin to induce YAP/TAZ phosphorylation in various type of cancers Glucose Starvation
B. Glucose restriction and metformin co-treatment cannot induce YAP/TAZ phosphorylation in normal cells Metformin
C. YAP/TAZ phosphorylation status is a major difference between cancer(Mel624) and normal cell(HaCaT) lines at glucose restriction and metformin co-treatment 24h Li-AcAc
condition
YAP
(Phos-tag)
Figure 2. YAP/TAZ inhibition effect of metformin is synergized with Figure 3. Effect of co-treatment on YAP/TAZ inhibition is AMPK-
glucose restriction but not glutamine or serum starvation /p38-independent but Hippo pathway dependent 36h
A B A B A. Metformin induce YAP/TAZ phosphorylation at GPI knockout cell even high
glucose condition
B. Fructose can rescue metformin and glucose restriction induced YAP
phosphorylation and cell death
C. Cancer cells are vulnerable to co-treatment but fructose can rescue its effect
D. Pyruvate and Ketone bodies cannot rescue co-treatment induced YAP
phosphorylation
Conclusion Normal Cell Cancer Cell
Glucose Glucose
Starvation Starvation
C & Metformin & Metformin
ATP decrease ATP decrease
Energy Stress Energy Stress
A-B. Co-treatment induce YAP/TAZ phosphorylation at (A)p38 α,β,γ,δ isoform AMPK activation Hippo activation
A. YAP/TAZ inhibition effect of metformin is glucose restriction dependent, but Crispr KO cell with or without JNK inhibitor and (B)AMPK α,γ subunit
glucose/serum restriction cannot show synergistic effect with metformin Crispr KO cell
B. Low glucose(0~1mM) condition combined with metformin induce YAP/TAZ C. Co-treatment cannot phosphorylates YAP at Hippo core kinase, LATS1/2 Cell Sruvival Cell Death
inhibition double KO cell
