Biteghe Fa Nsole, Davids L M
Redox Laboratory, Level 6, Anatomy Building, Department of Human Biology, Faculty of Health Sciences, UCT Medical School, Observatory, 7925 Cape Town, South Africa.
Redox Laboratory, Level 6, Anatomy Building, Department of Human Biology, Faculty of Health Sciences, UCT Medical School, Observatory, 7925 Cape Town, South Africa.
J Photochem Photobiol B. 2017 Jan;166:18-27. doi: 10.1016/j.jphotobiol.2016.11.004. Epub 2016 Nov 8.
Cutaneous melanoma represents the most lethal form of skin cancer and remains refractory to current therapies. Failure of treatment has been attributed to the over-expression of ABC transporters which efflux the drugs, below their cytotoxic threshold within cells. Therefore, this study set to investigate; the efficacy of a combinatorial approach comprising chemotherapy (Dacarbazine) and photodynamic therapy (PDT) to overcome resistance in pigmented and unpigmented metastatic melanoma and potentially identify resistant mechanisms.
The cytotoxic effect of the chemotherapy, PDT and combination therapy treatment (Dacarbazine+PDT) was determined using a cell viability XTT assay. Thereafter, melanoma cells morphology, self-renewal capacity and ABCG2 protein expression, were determined using fluorescence microscopy, clonogenic assay, western blot and flow cytometry. All results were analyzed by t-test and ANOVA, followed by individual comparisons with post-tests.
This study describes possible synergism of PDT+DTIC in reducing melanoma cell viability in vitro. At 24h post-treatment, only the unpigmented melanomas were sensitive to DTIC treatment (20-25% death at 1.25mM). At 48h, a lethal dose of 50% was reached in these cells in contrast to the pigmented melanoma (20% at 48h). The same trend was observed with the combination therapy (DTIC+PDT) at both time points. Furthermore, complete morphological disruption could be observed upon PDT only and PDT+DTIC treatments. Moreover, PDT and DTIC+PDT suppressed the self-renewal capacity of both melanoma cell lines. No significant differences in ABCG2 protein expression was found at 24h post-treatment.
Overall, these results suggest that human melanomas remain heterogeneous in their phenotypes. Moreover, in our metastatic melanoma cells, ABCG2 transporters did not seem to be involved in resistance to therapies. Significantly though, a combinatorial approach of PDT and chemotherapy significantly decreases the self-renewal capacity of metastatic melanoma cells and could be a suggested adjunctive approach to post-resection treatment regimes.
皮肤黑色素瘤是皮肤癌中最致命的一种形式，目前的治疗方法对其仍然无效。治疗失败归因于ABC转运蛋白的过度表达，这些转运蛋白将药物排出细胞，使其低于细胞内的细胞毒性阈值。因此，本研究旨在调查；一种联合方法的疗效，该方法包括化疗（达卡巴嗪）和光动力疗法（PDT），以克服色素沉着和无色素转移性黑色素瘤的耐药性，并潜在地确定耐药机制。
使用细胞活力XTT测定法确定化疗、PDT和联合治疗（达卡巴嗪+PDT）的细胞毒性作用。此后，使用荧光显微镜、克隆形成测定法、蛋白质印迹法和流式细胞术确定黑色素瘤细胞的形态、自我更新能力和ABCG2蛋白表达。所有结果均通过t检验和方差分析进行分析，随后通过事后检验进行个体比较。
本研究描述了PDT+达卡巴嗪在体外降低黑色素瘤细胞活力方面可能的协同作用。治疗后24小时，只有无色素黑色素瘤对达卡巴嗪治疗敏感（在1.25mM时死亡率为20-25%）。48小时时，这些细胞达到了50%的致死剂量，而色素沉着黑色素瘤则为20%（48小时时）。在两个时间点，联合治疗（达卡巴嗪+PDT）也观察到了相同的趋势。此外，仅在PDT和PDT+达卡巴嗪治疗后可观察到完全的形态破坏。此外，PDT和达卡巴嗪+PDT抑制了两种黑色素瘤细胞系的自我更新能力。治疗后24小时，ABCG2蛋白表达未发现显著差异。
总体而言，这些结果表明人类黑色素瘤的表型仍然存在异质性。此外，在我们的转移性黑色素瘤细胞中，ABCG2转运蛋白似乎与治疗耐药性无关。不过，重要的是，PDT和化疗的联合方法显著降低了转移性黑色素瘤细胞的自我更新能力，可能是切除后治疗方案的一种建议辅助方法。
