Objective:

2

To explore the role of the natural compound hesperidin in glycolysis

the key ratelimiting enzyme

in colorectal cancer (CRC) cell lines.

Methods:

2

In vitro

HCT116 and SW620 were treated with different doses of hesperidin (0–500 μmol/L)

cell counting kit-8 and colone formation assays were utilized to detected inhibition effect of hesperidin on CRC cell lines. Transwell and wound healing assays were performed to detect the ability of hesperidin (0

25

50 and 75 μmol/L) to migrate CRC cells. To confirm the apoptotic-inducing effect of hesperidin

apoptosis and cycle assays were employed. Western blot

glucose uptake

and lactate production determination measurements were applied to determine inhibitory effects of hesperidin (0

25 and 50 μmol/L) on glycolysis.

In vivo

according to the random number table method

nude mice with successful tumor loading were randomly divided into vehicle

low-dose hesperidin (20 mg/kg) and high-dose hesperidin (60 mg/kg) groups

with 6 mice in each group. The body weights and tumor volumes of mice were recorded during 4-week treatment. The expression of key glycolysis rate-limiting enzymes was determined using Western blot

and glucose uptake and lactate production were assessed. Finally

protein interactions were probed with DirectDIA Quantitative Proteomics

Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses.

Results:

2

Hesperidin could inhibit CRC cell line growth (

P

<

0.05 or

P

<

0.01). Moreover

hesperidin presented an inhibitory effect on the migrating abilities of CRC cells. Hesperidin also promoted apoptosis and cell cycle al

terations (

P

<

0.05). The immunoblotting results manifested that hesperidin decreased the levels of hexokinase 2

glucose transporter protein 1 (GLUT1)

GLUT3

L-lactate dehydrogenase A

6-phosphofructo-2-kinase/fructose-2

6-biphosphatase 2 (PFKFB2)

PFKFB3

and pyruvate kinase isozymes M2 (

P

<

0.01). It remarkably suppressed tumor xenograft growth in nude mice. GO and KEGG analyses showed that hesperidin treatment altered metabolic function.

Conclusion:

2

Hesperidin inhibits glycolysis and is a potential therapeutic choice for CRC treatment.