Mechanism-based Changes in Anti-METH Monoclonal Antibody Therapy During Pregnancy

描述（由申请人提供）：(+)-甲基苯丙胺（冰毒）是孕妇中最容易上瘾和严重滥用的药物之一，但没有批准的药物可以保护脆弱的胎儿或母亲免受其不利的健康影响。这些研究的长期目标是开发基于抗体的药物，可以在怀孕期间给予，以保护母亲和胎儿免受甲基苯丙胺的毒性和成瘾性影响。虽然单克隆抗体（mAb）在人类妊娠期间的治疗尚未经过测试，但我们实验室在妊娠大鼠中进行的开创性研究揭示了单克隆抗体药代动力学（PKs）的新变化和治疗上的重要变化，这些变化会损害抗体的功效。具体来说，mAb的半衰期、分布量和清除率都随着妊娠期的不同而发生显著变化。类似的妊娠期特异性PK变化以前没有报道过，驱动这些变化的机制目前尚不清楚。在测试单克隆抗体功能的相关研究中，某些抗单克隆抗体药物（如mAb6H4）在给药后的头24小时内，体内甲基结合功能显著降低。这种情况发生在大约94%的单抗剂量仍然存在的情况下。重要的是，尽管所有单克隆抗体都以相同的剂量施用并且显示相似的PK特性，但对甲基安非他明的结合亲和力（KD）与体内功能无关。该建议的中心假设是，新生儿Fc受体（FcRn）挽救途径既是妊娠期间单克隆抗体清除变化的关键调节剂，也促进了单克隆抗体甲基结合功能的体内降低。FcRn是一种具有良好特征的受体，它通过ph依赖的挽救机制将IgG从分解代谢消除（清除）中拯救出来，并促进IgG从母体被动转移到胎儿。然而，妊娠期间FcRn在调节单抗PKs中的作用尚不清楚。此外，在打捞途径中发现的低pH环境表明，它可以促进对单克隆抗体的体内修饰。为了验证我们的假设，我们将执行以下目标：1 -确定大鼠妊娠期间单抗清除变化最大时FcRn组织表达水平，2 -确定FcRn挽救途径在妊娠期间调节IgG清除中的作用，以及3 -确定FcRn挽救途径在降低mAb6H4甲基化结合功能中的作用。拟议的研究将利用免疫组织化学和Western blotting技术，结合怀孕和非怀孕大鼠的体内PK实验。对这些研究有帮助的是，anti-METH mAb6H4将被重新设计为一个单一的氨基酸突变，消除FcRn结合并抑制通过救助途径。除了为医学博士候选人提供良好的培训机会外，这些创新研究将有助于单抗药物的临床前开发，以保护母亲和胎儿免受滥用药物的危险健康影响。

DESCRIPTION (provided by applicant): (+)-Methamphetamine (METH) is one of the most addictive and heavily abused drugs among pregnant women, yet there are no approved medications for protecting the vulnerable fetus or mother from its adverse health effects. The long-term objective of these studies is to develop antibody-based medications that can be given during pregnancy to protect the mother and fetus from the toxic and addictive effects of METH. While monoclonal antibody (mAb) therapy during human pregnancy is untested, seminal studies from our laboratory in pregnant rats reveal novel and therapeutically important changes to mAb pharmacokinetics (PKs) that compromise antibody efficacy. Specifically, mAb half-life, volume of distribution, and clearance all dramatically change with each gestation stage. Similar gestation-stage specific PK changes have not been reported previously, and the mechanisms that drive these changes are currently unknown. In related studies testing mAb function, certain anti-METH mAb medications (e.g., mAb6H4) show significant in vivo reductions in METH-binding function during the first 24 hrs after administration. This occurred even though >94% of the mAb doses were still present. Importantly, binding affinity for METH (KD) did not correlate with in vivo function despite all mAbs being administered at equal doses and displaying similar PK characteristics. The central hypothesis of this proposal is that the neonatal Fc receptor (FcRn) salvage pathway is both a key regulator of mAb clearance changes during pregnancy and facilitates in vivo reductions in mAb METH- binding function. FcRn is a well-characterized receptor that acts via a pH-dependent salvage mechanism to rescue IgG from catabolic elimination (clearance) and facilitate the passive transfer of IgG from mother to fetus. However, the role of FcRn in regulating mAb PKs during pregnancy is unknown. Furthermore, the low pH environment found in the salvage pathway suggests it could facilitate in vivo modifications to mAbs. To test our hypothesis, we will perform the following aims: 1 - Determine FcRn tissue expression levels during rat pregnancy at times when mAb clearance changes are maximal, 2 - Define the role of the FcRn salvage pathway in regulating IgG clearance during pregnancy, and 3 - Define the role of the FcRn salvage pathway in reducing mAb6H4 METH- binding function. The proposed studies will utilize immunohistochemistry and Western blotting techniques in conjunction with in vivo PK experiments in both pregnant and non-pregnant rats. Instrumental to these studies, anti-METH mAb6H4 will be re-engineered with a single amino acid mutation that eliminates FcRn binding and inhibits passage through the salvage pathway. In addition to providing an excellent training opportunity for a MD/PhD candidate, these innovative studies will contribute to the pre-clinical development of mAb medications for the protection of both the mother and fetus from the dangerous health effects of drugs of abuse. PUBLIC HEALTH RELEVANCE: These studies will define the role of the FcRn salvage pathway in regulating monoclonal antibody (mAb) clearance and function during pregnancy, yielding new insights into the in vivo physiologic processes that affect antibody therapeutic efficacy. This knowledge is expected to advance the preclinical development of safe and effective antibody-based medicines for use in treating pregnant women and their vulnerable fetus(es) from drugs of abuse. Furthermore, due to the ubiquitous role of FcRn in regulating both endogenous and exogenously administered antibodies of the IgG class, these studies will also contribute to our understanding of the regulation of the humoral immune system during pregnant and non-pregnant physiologic states.