Exosomal CD44 Transmits Lymph Node Metastatic Capacity Between Gastric Cancer Cells via YAP-CPT1A-Mediated FAO Reprogramming

Mei Wang; Wanjun Yu; Xiaoli Cao; Hongbing Gu; Jiaying Huang; Chen Wu; Lin Wang; Xin Sha; Bo Shen; Ting Wang; Yongliang Yao; Wei Zhu; Feng Huang

doi:10.3389/fonc.2022.860175

Exosomal CD44 Transmits Lymph Node Metastatic Capacity Between Gastric Cancer Cells via YAP-CPT1A-Mediated FAO Reprogramming

Front Oncol. 2022 Mar 10:12:860175. doi: 10.3389/fonc.2022.860175. eCollection 2022.

Authors

Mei Wang¹, Wanjun Yu¹, Xiaoli Cao², Hongbing Gu³, Jiaying Huang¹, Chen Wu¹, Lin Wang¹, Xin Sha⁴, Bo Shen⁵, Ting Wang¹, Yongliang Yao⁶, Wei Zhu¹, Feng Huang^{1

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Affiliations

¹ Key Laboratory of Medical Science and Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, China.
² Department of Laboratory Medicine, Affiliated Tumor Hospital of Nantong University, Nantong, China.
³ Department of Laboratory Medicine, The Affiliated People's Hospital, Jiangsu University, Zhenjiang, China.
⁴ Department of Surgery, The Affiliated Hospital of Jiangsu University, Jiangsu University, Zhenjiang, China.
⁵ Department of Oncology, Jiangsu Cancer Hospital, Jiangsu Institute of Cancer Research, Nanjing Medical University Affiliated Cancer Hospital, Nanjing, China.
⁶ Department of Clinical Laboratory, Affiliated Kunshan Hospital of Jiangsu University, Suzhou, China.
⁷ Department of Clinical Laboratory, Maternal and Child Health Care Hospital of Kunshan, Suzhou, China.

Abstract

Background: Lymph node metastasis (LNM) commonly occurs in gastric cancer (GC) and is tightly associated with poor prognosis. Exosome-mediated lymphangiogenesis has been considered an important driver of LNM. Whether exosomes directly transmit the LNM phenotype between GC cells and its mechanisms remain elusive.

Methods: A highly lymphatic metastatic GC cell line (HGC-27-L) was established by serial passage of parental HGC-27 cells in BALB/c nude mice. The capacities of migration, invasion and LNM; fatty acid oxidation (FAO) levels; and the role of exosome-transferred LNM phenotype were compared among HGC-27-L, HGC-27 and primary GC cell line AGS. Exosomes derived from GC cells and sera were separately isolated using ultracentrifugation and ExoQuick exosome precipitation solution, and were characterized by transmission electron microscopy, Nanosight and western blotting. Transwell assay and LNM models were conducted to evaluate the capacities of migration, invasion and LNM of GC cells in vitro and in vivo. β-oxidation rate and CPT1 activity were measured to assess FAO. CPT1A inhibitor etomoxir was used to determine the role of FAO. Label-free LC-MS/MS proteome analysis screened the differential protein profiling between HGC-27-exosomes and AGS-exosomes. Small interference RNAs and YAP inhibitor verteporfin were used to elucidate the role and mechanism of exosomal CD44. TCGA data analysis, immunochemistry staining and ELISA were performed to analyze the expression correlation and clinical significance of CD44/YAP/CPT1A.

Results: FAO was increased in lymphatic metastatic GC cells and indispensable for sustaining LNM capacity. Lymphatic metastatic GC cell-exosomes conferred LNM capacity on primary GC cells in an FAO-dependent way. Mechanistically, CD44 was identified to be enriched in HGC-27-exosomes and was a critical cargo protein regulating exosome-mediated transmission, possibly by modulating the RhoA/YAP/Prox1/CPT1A signaling axis. Abnormal expression of CD44/YAP/CPT1A in GC tissues was correlated with each other and associated with LNM status, stages, invasion and poor survival. Serum exosomal CD44 concentration was positively correlated with tumor burden in lymph nodes.

Conclusions: We uncovered a novel mechanism: exosomal CD44 transmits LNM capacity between GC cells via YAP-CPT1A-mediated FAO reprogramming from the perspective of exosomes-transferred LNM phenotype. This provides potential therapeutic targets and a non-invasive biomarker for GC patients with LNM.

Keywords: CD44; carnitine palmitoyltransferase 1A (CPT1A); exosomes; fatty acid oxidation; gastric cancer; lymph node metastasis; yes-associated protein (YAP).