Disruption of LPA-LPAR1 pathway results in lung tumor growth inhibition by downregulating B7-H3 expression in fibroblasts.

BACKGROUND: Lysophosphatidic acids (LPAs) belong to a class of bioactive lysophospholipids with multiple functions including immunomodulatory roles in tumor microenvironment (TME). LPA exerts its biological effects via its receptors that are highly expressed in fibroblasts among other cell types. As cancer-associated fibroblasts (CAFs) are a key component of the TME, it is important to understand LPA signaling and regulation of receptors in fibroblasts or CAFs and associated regulatory roles on immunomodulation-related molecules. METHODS: Cluster analysis, immunoblotting, real-time quantitative-PCR, CRISPR-Cas9 gene editing system, immunohistochemical staining, coculture model, and in vivo xenograft model were used to investigate the effects of LPA-LPAR1 on B7-H3 in tumor promotion of CAFs. RESULTS: In this study, we found that LPAR1 and CD276 (B7-H3) were generally highly expressed in fibroblasts with good expression correlation. LPA induced B7-H3 up-expression through LPAR1, and stimulated fibroblasts proliferation that could be inhibited by silencing LPAR1 or B7-H3 as well as small molecule LPAR1 antagonist (Ki16425). Using engineered fibroblasts and non-small cell lung carcinoma (NSCLC) cell lines, subsequent investigations demonstrated that CAFs promoted the proliferation of NSCLC in vitro and in vivo, and such effect could be inhibited by knocking out LPAR1 or B7-H3. CONCLUSION: The present study provided new insights for roles of LPA in CAFs, which could lead to the development of innovative therapies targeting CAFs in the TME. It is also reasonable to postulate a combinatory approach to treat malignant fibrous tumors (such as NSCLC) with LPAR1 antagonists and B7-H3 targeting therapies.

Hormetic effects of EGC and EGCG on CES1 activity and its rescue from oxidative stress in rat liver S9.

Carboxylesterase 1 (CES1) is a hydrolytic enzyme that plays an important role in the activation or deactivation of many therapeutic agents, thus affecting their pharmacokinetic and pharmacodynamic outcomes. Using rat liver S9 as an enzyme source and enalapril as a CES1 substrate, the present study examined effects of a number of flavonoids on the formation of enalaprilat (the active form of enalapril) produced by CES1-mediated hydrolysis. While a majority of flavonoids tested showed inhibition on CES1, an unexpected hormetic effect was observed for epigallocatechin (EGC) and epigallocatechin gallate (EGCG), i.e., stimulatory effect at low concentrations and enzyme inhibition at high concentrations. Further experiments revealed that oxidative stress caused by hydrogen peroxide, arachidonic acid plus iron, and oxidized low density lipoproteins (oxLOL) reduced CES1 activity in rat liver S9 and the loss of CES1 enzyme activity could be rescued largely by EGC or EGCG. In contrast, such effects were minimal in human liver S9, probably due to the presence of a higher ratio of reduced vs oxidized forms of glutathione. The above findings suggest that the polyphenolic nature of EGC or EGCG might be responsible for rescuing CES1 activity under oxidative stress. Because of the importance of CES1 in drug activation or deactivation and rat liver S9 as a versatile in vitro system used for drug metabolism studies and drug safety assessment, caution should be exercised to avoid potential biases for data interpretation and decision making when CES1 activity in rat liver S9 is evaluated with dependency on experimental conditions.