School of Pharmaceutical Sciences, University of Geneva & University of Lausanne, Geneva, Switzerland.
J Control Release. 2011 Nov 30;156(1):53-9. doi: 10.1016/j.jconrel.2011.07.024. Epub 2011 Jul 22.
The physicochemical properties and stability requirements of therapeutic proteins necessitate their parenteral administration even for local therapy; however, unnecessary systemic exposure increases the risk of avoidable side-effects. The objective of this study was to use fractional laser ablation to enable the delivery of intact, functional therapeutic antibodies into the skin in vitro and in vivo. The laser-assisted delivery of Antithymocyte globulin (ATG) and Basiliximab - FDA-approved therapeutics for the induction of immunosuppression - was investigated. In vitro delivery experiments were performed using dermatomed porcine ear and human abdominal skins; an in vitro/in vivo correlation was shown using C57 BL/10 SCSnJ mice. Antibody transport was quantified by using ELISA methods developed in-house. Results showed that increasing the pore number from 300 to 450 and 900, increased total antibody delivery (sum of amounts permeated and deposited); e.g., for ATG, from 1.18±0.10 to 3.98±0.64 and 4.97±0.83 μg/cm(2), respectively - corresponding to 19.7-, 66.3- and 82.8-fold increases over the control (untreated skin). Increasing laser fluence from 22.65 to 45.3 and 135.9J/cm(2) increased total ATG delivery from 1.70±0.65 to 4.97±0.83 and 8.70±1.55 μg/cm(2), respectively. The Basiliximab results confirmed the findings with ATG. Western blot demonstrated antibody identity and integrity post-delivery; human lymphocyte cytotoxicity assays showed that ATG retained biological activity post-delivery. Immunohistochemical staining was used to visualize ATG distribution in the epidermis. Total ATG delivery across porcine ear and human abdominal skin was statistically equivalent and an excellent in vitro/in vivo correlation was observed in the murine model. Based on published data, the ATG concentrations achieved in the laser-porated human skins were in the therapeutic range for providing local immunosuppression. These results challenge the perceived limitations of transdermal delivery with respect to biopharmaceuticals and suggest that controlled laser microporation provides a less invasive, more patient-friendly "needle-less" alternative to parenteral administration for the local delivery of therapeutic antibodies.
治疗性蛋白质的物理化学性质和稳定性要求即使是局部治疗也需要进行肠外给药；然而，不必要的全身暴露会增加避免不良反应的风险。本研究的目的是使用分束激光烧蚀使完整的、功能正常的治疗性抗体在体外和体内递送至皮肤。研究了经批准用于诱导免疫抑制的抗胸腺细胞球蛋白（ATG）和巴利昔单抗（Basiliximab）的激光辅助递送-。使用去皮的猪耳和人腹部皮肤进行了体外递送实验；使用 C57 BL/10 SCSnJ 小鼠进行了体外/体内相关性研究。使用内部开发的 ELISA 方法定量测定抗体的转运。结果表明，增加孔数从 300 增加到 450 和 900，增加了总抗体的递送量（渗透和沉积的量之和）；例如，对于 ATG，从 1.18±0.10 增加到 3.98±0.64 和 4.97±0.83 μg/cm(2)，分别对应于对照（未处理的皮肤）的 19.7、66.3 和 82.8 倍。将激光通量从 22.65 增加到 45.3 和 135.9J/cm(2)，分别将 ATG 的总递送量从 1.70±0.65 增加到 4.97±0.83 和 8.70±1.55 μg/cm(2)。巴利昔单抗的结果证实了 ATG 的发现。Western blot 显示了递送至皮肤后的抗体的身份和完整性；人淋巴细胞细胞毒性测定表明，ATG 递送至皮肤后保持了生物活性。免疫组织化学染色用于显示表皮中 ATG 的分布。猪耳和人腹部皮肤的总 ATG 递送在统计学上是等效的，并且在小鼠模型中观察到了极好的体外/体内相关性。根据已发表的数据，在激光穿孔的人皮肤上达到的 ATG 浓度处于提供局部免疫抑制的治疗范围内。这些结果挑战了关于生物制药的经皮递送的局限性，并且表明受控激光微穿孔为局部递送治疗性抗体提供了一种侵入性更小、更适合患者的“无针”替代方案。
