以轉錄體分析lunasin胜肽與短期禁食對乳癌細胞之影響

Abstract

乳癌是全球女性最常見的惡性腫瘤之一。天然物具有多元的生理活性且低毒性副作用，已廣泛被研究於化學預防。其中lunasin是一具有生物活性的種子胜肽，其作用包括抗腫瘤；近年健康意識抬頭，改變生活型態預防疾病成為主流，其中飲食限制也備受關注，研究顯示短期禁食也可抑制癌細胞的生長。本研究欲探討lunasin與間歇性禁食 (short-term starvation, STS) 處理對於乳癌細胞生長的影響機制。實驗包含兩部分，第一部分為lunasin處理兩株乳癌細胞MCF-7與MDA-MB-231與一株正常乳腺上皮細胞MCF-10A，結果顯示lunasin可抑制MCF-7細胞之乳酸與ATP生成之趨勢，由於效應未達顯著，無法推斷透過瓦式效應。因此第二部分加入STS設計，以期提高抗癌的效果作用，分組為lunasin或STS或兩者合併處理乳癌細胞。首先建立與使用的禁食條件，在低葡萄糖培養基 (1 g/L 葡萄糖，1% FBS) 中相較於無葡萄糖培養基 (0 g/L 葡萄糖，1% FBS)，lunasin、STS與合併處理相較控制組，顯著降低兩株乳癌細胞的生長，但不影響正常細胞生長，因此選擇低糖培養基進行後續實驗。在乳癌細胞乳酸及ATP生成結果，lunasin與STS處理後沒有顯著差異，且合併組也未能有協同效應，主要變化是來自於STS的營養缺乏而非lunasin。而在轉錄體分析中，兩株乳癌細胞在STS組及合併處理組對轉錄體表現的影響皆較lunasin組顯著，顯示營養缺乏為主要影響因素。在Gene Set Enrichment Analysis (GSEA) 分析中，乳癌細胞於STS組中顯著富集細胞週期、轉錄後修飾、蛋白質合成及核輸送等路徑。根據上述，lunasin及STS處理對抑制乳癌細胞乳酸與ATP的生成效果有限，而合併處理也未展現協同效應。在轉錄體層面，STS處理是抑制乳癌細胞生長的主要因素，透過比對熱點圖篩選出具潛力的關鍵調控基因，提供未來研究相關機制的標的。

Breast cancer is the most common cancer among women worldwide. Natural compounds with bioactivity and low toxicity are widely studied for cancer prevention. Lunasin, a bioactive seed peptide, has demonstrated anti-tumor properties. In recent years, lifestyle interventions such as dietary restriction have gained attention for disease prevention. Short-term starvation (STS) has been shown to inhibit cancer cell growth. This study investigated the effects and mechanisms of lunasin and STS on breast cancer cell growth. The study included two parts, in the first part, breast cancer cells MCF-7, MDA-MB-231 and normal breast epithelial cell MCF-10A were treated with lunasin. The results showed that lunasin slightly reduced lactate and ATP production in MCF-7 cells; however, the effects can’t support inhibition of the Warburg effect. Therefore, STS was added in the second part to enhance potential anti-cancer activity. Cells were treated with lunasin, STS, or combination treatment. Low-glucose STS (1 g/L glucose with 1% FBS) was selected based on its ability to inhibit the growth to both cancer cell lines without affecting normal cells. However, lactate and ATP levels weren't significantly different, and no synergistic effect in combination treatment, suggesting that the metabolic effects were primarily driven by STS. Transcriptome analysis revealed that STS induced most of the transcriptional changes, with little added effect from lunasin. GSEA results showed significant enrichment pathways related to cell cycle, post-transcriptional modification, protein synthesis and nucleocytoplasmic transport in STS-treated groups. In conclusion, lunasin and STS treatment had limited effects on lactate and ATP production in breast cancer cells, and no synergistic in combination treatment. Additionally, no glycolysis-related pathways were enriched in the transcriptomic data. STS appeared to be the major contributor to growth inhibition in transcriptional level. Potential regulatory genes were identified from heatmap, offering targets for future mechanistic studies.