酮體生成酵素HMGCS2對於癌症進程之效應

Abstract

中文摘要 實驗目的：目前已知3-Hydroxy-3-methylglutaryl-CoA Synthase 2 (HMGCS2)基因表現量與癌症病患之癌症惡性度成正相關，但目前機制並不明確。本實驗為探討酮體生成酵素HMGCS2對於大腸癌與口腔癌進成之效應，及釐清HMGCS2透過何種訊息傳遞影響大腸癌與口腔癌之腫瘤進程。 實驗設計：利用即時定量聚合酶連鎖反應(Q-PCR)檢測癌症病人檢體中HMGCS2基因之表現與臨床病徵之關聯性。藉由西方墨點法(Western bloot)探討HMGCS2內生性蛋白質之表現與癌細胞惡性度之相關性。使用細胞移行與浸襲實驗(Migration and Invasion Assay)探討HMGCS2對於癌細胞移行與浸襲之影響。建立肝轉移小鼠動物模式探討HMGCS2對於癌細胞轉移之效應。 實驗結果：臨床檢體分析結果發現，HMGCS2基因表現量與大腸癌與口腔癌病患之癌症惡性度、癌症轉移能力與癌症復發成正相關。細胞實驗中發現，HMGCS2內生性蛋白質表現量與大腸癌與口腔癌細胞株之惡性程度成正相關。高度表現內生性HMGCS2蛋白質之細胞株中，將其剔除會抑制癌細胞轉移能力與浸襲能力；低表現內生性HMGCS2蛋白質之細胞株中，將其過度表現則會促進癌細胞轉移能力與浸襲能力；但HMGCS2蛋白質表現量不會影響癌細胞之細胞增生能力。動物實驗中，將DLD1細胞株之HMGCS2剔除會降低其在肝轉移小鼠動物模式中之轉移能力。利用微小核醣核酸陣列與生物資訊學認為Src可能為HMGCS2之下游主要調控因子。將Src在穩定表現HMGCS2之癌細胞中剔除，會抑制HMGCS2促進之細胞移行和浸襲能力，但不會影響癌細胞之細胞增生能力。 結論：HMGCS2可以透過Src訊息傳遞促進大腸癌與口腔癌之移行能力與浸襲能力。HMGCS2可以當作大腸癌與口腔癌之臨床預後指標，為大腸癌與口腔癌之治療方針。 關鍵字:大腸直腸癌、口腔鱗狀上皮細胞癌、移行、浸襲、酮體生成酵素HMGCS2

Abstract Purpose： Mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase (HMGCS2) is the rate-limiting enzyme of ketogenesis. This metabolic reaction is highly related to advanced TNM stage and early recurrence in colorectal cancer (CRC) and oral squamous cell carcinoma (OSCC). However, the effects of HMGCS2 in CRC and OSCC progression are largely unknown. We hypotheses that HMGCS2 could interfere cancer metabolism and regulate tumor progression. Material and Method： Colon cancer and oral cancer cell lines were used as in vitro models to investigate cell phenotypes, including migration, invasion, and proliferation by HMGCS2 shRNA knockdown. Boyden chamber assay were performed to check migration and invasion abilities. Proliferation ability was identified by MTT assay. 3-Hydroxybutyrate was measured by ketone body assay. Mice were splenic injected with shHMGCS2-silencing or pLKO control DLD1 cells to metastasis ability. Results： Here, we showed that HMGCS2 enhanced cancer cell migration and invasion abilities via ketogenesis-independent manner in vitro. Knocked-down HMGCS2 significantly decreased CRC metastasis ability in hepatic animal model. Using high through-put microarray assay and bioinformatic analysis, we identified Src as a crucial downstream effecter of HMGCS2-promoted CRC and OSCC progression. Src mRNA and protein expression levels were significantly diminished in silenced-HMGCS2 clones, and knocked-down Src could re-inhibited migration and invasion abilities in HMGCS2 stable transfectants. Conclusion： Taken together, we discovered a novel role of HMGCS2 which could promote CRC and OSCC cell migration and cell invasion abilities through Src signaling pathway. This target oncogene, HMGCS2, may play as a prognosis marker in CRC and OSCC, and will be beneficial for developing therapeutic strategy against advanced CRC and OSCC in the future. Key words：Oral cancer, Colorectal cancer, Migration, Invasion, HMGCS2, Src