Dr Rath Research Institute, Santa Clara, CA 95050, USA.
Int J Oncol. 2012 Dec;41(6):1996-2004. doi: 10.3892/ijo.2012.1639. Epub 2012 Sep 24.
Head and neck squamous cell carcinoma (HNSCC) and acute myeloid leukemia are the major causes of mortality and morbidity in Fanconi anemia (FA) patients. The objective of this study was to investigate the antineoplastic activity of a novel antineoplastic nutrient mixture (NM) (containing lysine, proline, ascorbic acid and green tea extract) in human FA-associated HNSCC (FA HNSCC) in vitro and in vivo. The human FA HNSCC cell line, OHSU-974 (Fanconi Anemia Research Fund), was cultured in RPMI medium supplemented with 20% FBS and antibiotics. At near confluence, cells were treated in triplicate with various concentrations of NM: 0, 50, 100, 250, 500 and 1,000 µg/ml. Cells were also treated with phorbol 12-myristate 13-acetate (PMA) to induce matrix metalloproteinase (MMP)-9 activity. Cell proliferation was detected by MTT assay, the secretion of MMPs by gelatinase zymo-graphy, cell invasion through Matrigel, cell migration by a scratch test and morphology by hematoxylin and eosin (H&E) staining. In vivo, athymic male nude mice (n=12) were inoculated with 3x106 OHSU‑974 cells subcutaneously and randomly divided into 2 groups: group A was fed a regular diet and group B a regular diet supplemented with 1% NM. Four weeks later, the mice were sacrificed and their tumors were excised, weighed and processed for histological analysis. NM inhibited the growth of OHSU-974 tumors by 47% and tumor burden by 50%. At lower concentrations, NM demonstrated no effect on proliferation, but at 1,000 µg/ml a 40% toxicity was observed. Zymography revealed the MMP-2 and PMA-induced MMP-9 secretion. NM suppressed the secretion of both MMPs in a dose-dependent manner, with a virtual inhibition at 500 µg/ml. NM inhibited OHSU-974 cell invasion through Matrigel in a dose-dependent manner with a complete block at 1,000 µg/ml. H&E staining showed no morphological changes below 500 µg/ml. These results suggest that NM has potential therapeutic use in the treatment of human FA HNSCC.
头颈部鳞状细胞癌（HNSCC）和急性髓性白血病是范可尼贫血（FA）患者死亡和发病的主要原因。本研究的目的是研究一种新型抗肿瘤营养混合物（NM）（含有赖氨酸、脯氨酸、抗坏血酸和绿茶提取物）在体外和体内对人 FA 相关 HNSCC（FA HNSCC）的抗肿瘤活性。人 FA HNSCC 细胞系 OHSU-974（范可尼贫血研究基金）在补充有 20% FBS 和抗生素的 RPMI 培养基中培养。当接近汇合时，将细胞用不同浓度的 NM（0、50、100、250、500 和 1000μg/ml）处理 3 次。还使用佛波醇 12-肉豆蔻酸 13-乙酸酯（PMA）处理细胞以诱导基质金属蛋白酶（MMP）-9 活性。通过 MTT 测定法检测细胞增殖，通过明胶酶谱法检测 MMPs 的分泌，通过 Matrigel 检测细胞侵袭，通过划痕试验检测细胞迁移，通过苏木精和伊红（H&E）染色检测细胞形态。在体内，将 3x106 OHSU-974 细胞皮下接种到 12 只无胸腺雄性裸鼠中，并将它们随机分为 2 组：A 组喂食常规饮食，B 组喂食常规饮食+1%NM。4 周后，处死小鼠并取出肿瘤，称重并进行组织学分析。NM 抑制 OHSU-974 肿瘤生长 47%，肿瘤负担减轻 50%。在较低浓度下，NM 对增殖没有影响，但在 1000μg/ml 时观察到 40%的毒性。明胶酶谱显示 MMP-2 和 PMA 诱导的 MMP-9 分泌。NM 以剂量依赖性方式抑制两种 MMP 的分泌，在 500μg/ml 时几乎完全抑制。NM 以剂量依赖性方式抑制 OHSU-974 细胞穿过 Matrigel 的侵袭，在 1000μg/ml 时完全阻断。H&E 染色显示在 500μg/ml 以下没有形态变化。这些结果表明，NM 在治疗人类 FA HNSCC 方面具有潜在的治疗用途。
