Department of Paediatrics & Adolescent Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
Exp Hematol Oncol. 2013 Apr 10;2(1):11. doi: 10.1186/2162-3619-2-11.
Relapsed T-lineage acute lymphoblastic leukemia (T-ALL) has been an incurable disease. Recent reports showed that an L-arginine depleting enzyme, pegylated arginase (BCT-100) may be effective against T-ALL cells. On the other hand, studies including ours had shown the symbiosis of ALL blasts and human mesenchymal stromal cells (hMSCs) in bone marrow microenvironment during L-asparaginase treatment. As L-asparaginase and BCT-100 both act by depleting lymphoid cells of specific amino acid, we hypothesized that hMSCs may also protect T-ALL blasts from BCT-100 treatment in co-culture and such protection may be abrogated by pre-treating hMSCs with vincristine (VCR).
XTT assay was used to test sensitivities of T-ALL cell lines and hMSCs to BCT-100. Apoptosis of T-ALL cell lines with or without BCT-100 treatment were tested by annexin V / propidium iodide (AV/PI) assay using flow cytometer. Western blotting was performed to analyze the expression of ornithine transcarbamylase (OTC), an enzyme involved in L-arginine metabolism which may account for BCT-100 resistance.
hMSCs were resistant to BCT-100 while CCRF-CEM, Jurkat and MOLT-4 were very sensitive to it. hMSCs could protect all the three cell lines from BCT-100 treatment in transwell co-culture. All the 3 T-ALL cell lines were also found to be rescued by an L-arginine precursor citrulline, while the breakdown product of BCT-100, ornithine only had limited salvaging effect on CCRF-CEM but not Jurkat and MOLT-4. Both hMSCs and 3 T-ALL cell lines express citrulline synthesis enzyme, ornithine transcarbamylase (OTC) at basal level while only hMSCs could express OTC at relatively higher level under BCT-100 treatment. Treating hMSCs with vincristine before co-culturing with T-ALL could resume the cytotoxicity of BCT-100 to CCRF-CEM and MOLT-4 cells.
Our results suggest a possible strategy to overcome resistance to BCT-100 from cancer microenvironments by suppressing hMSCs either in marrow or in the perivascular niche using vincristine.
复发 T 系急性淋巴细胞白血病（T-ALL）一直是一种无法治愈的疾病。最近的报告表明，一种精氨酸耗竭酶，聚乙二醇化精氨酸酶（BCT-100）可能对 T-ALL 细胞有效。另一方面，包括我们在内的研究表明，在 L-天冬酰胺酶治疗期间，ALL blasts 和人间质基质细胞（hMSCs）在骨髓微环境中存在共生关系。由于 L-天冬酰胺酶和 BCT-100 都通过耗尽淋巴细胞的特定氨基酸而起作用，我们假设 hMSCs 也可能在共培养中保护 T-ALLblasts 免受 BCT-100 治疗，并且这种保护可以通过用长春新碱（VCR）预处理 hMSCs 来消除。
使用 XTT 测定法测试 T-ALL 细胞系和 hMSCs 对 BCT-100 的敏感性。通过流式细胞术用膜联蛋白 V/碘化丙啶（AV/PI）测定法检测 T-ALL 细胞系在有或没有 BCT-100 治疗时的凋亡情况。通过 Western 印迹分析参与 L-精氨酸代谢的酶，鸟氨酸氨甲酰基转移酶（OTC）的表达，这可能是 BCT-100 耐药的原因。
hMSCs 对 BCT-100 具有抗性，而 CCRF-CEM、Jurkat 和 MOLT-4 则对其非常敏感。hMSCs 可以在 Transwell 共培养中保护所有三种细胞系免受 BCT-100 治疗。还发现所有三种 T-ALL 细胞系均被 L-精氨酸前体瓜氨酸拯救，而 BCT-100 的分解产物鸟氨酸仅对 CCRF-CEM 具有有限的挽救作用，而对 Jurkat 和 MOLT-4 则没有。hMSCs 和三种 T-ALL 细胞系在基础水平上均表达瓜氨酸合成酶，鸟氨酸氨甲酰基转移酶（OTC），而只有 hMSCs 在 BCT-100 治疗下才能相对较高水平表达 OTC。在用长春新碱预处理 hMSCs 后再与 T-ALL 共培养，可以恢复 BCT-100 对 CCRF-CEM 和 MOLT-4 细胞的细胞毒性。
我们的结果表明，通过使用长春新碱抑制骨髓或血管周围龛中的 hMSCs，可能是克服癌症微环境中对 BCT-100 耐药的一种策略。
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