Nanoparticle Enabled L-asparaginase Therapy for CLL

描述（由申请人提供）：L-天冬酰胺酶已用于治疗急性淋巴细胞白血病（ALL）超过40年。然而，尽管有证据表明成人白血病也对天冬酰胺和其他氨基酸消耗敏感，但由于几个原因，天冬酰胺酶治疗尚未扩展到儿科白血病以外。最重要的是L-天冬酰胺酶的免疫原性，其可限制治疗的持续时间并导致潜在的副作用，如超敏反应和过敏反应。其他副作用，包括胰腺炎和凝血障碍，也可能限制这种形式的治疗慢性淋巴细胞白血病（CLL）患者的吸引力。该项目的长期目标是开发一种新的L-天冬酰胺酶治疗平台，克服这两个限制。该平台有两个组成部分：由我们的合作者Donald Durden博士开发的新型重组L-天冬酰胺酶，以及保护酶免受体内免疫反应和免疫中和的纳米颗粒递送载体。该酶来源于Wolinella succinogenes，缺乏导致与临床批准的酶相关的毒副作用的转氨酶活性。然而，由于免疫抑制是被我们的酶消除的那些副作用之一，如果在没有某种形式的屏蔽的情况下使用这种酶，免疫原性问题可能特别严重。我们已经开发了一种纳米颗粒递送系统，我们称之为二氧化硅空心酶载球（SHELS）。这些SHELS最初被合成为具有大孔的“纳米威夫球”，这些大孔允许它们装载未修饰的酶，随后用多孔层密封，该多孔层允许小酶底物扩散，但不渗透大分子和蛋白质，从而同时封装酶并保护其免受抗体识别。我们有初步的数据表明，原代人类CLL细胞对天冬酰胺耗竭极其敏感。我们的方法分为三个阶段，与我们的具体目标相一致。首先，我们将优化装载在SHELS中的重组Wolinella succinogenes L-天冬酰胺酶（rWSasp-SHELS）的剂量。第二，我们将评估rWSasp-SHELS在移植有单独的CLL细胞或CLL细胞与活化的T细胞共施用以促进分裂的小鼠中的功效。前者将允许在代表静息CLL群体的模型中测试颗粒，而后者将模拟颗粒对增殖区室的影响。在最终目的中，我们将比较rWSasp-SHELS与目前临床上使用的PEG-天冬酰胺酶的毒性特征。

DESCRIPTION (provided by applicant): L-asparaginase has been used to treat acute lymphoblastic leukemia (ALL) for over 40 years. However, despite evidence that adult leukemias are also sensitive to asparagine and other amino acid depletion, asparaginase therapy has not expanded beyond pediatric leukemia for several reasons. Foremost is the immunogenicity of L-asparaginase which can limit the duration of therapy and contribute to potential side effects such as hypersensitivity reactions and anaphylaxis. Other side effects, including pancreatitis and coagulopathy, may also limit the appeal of this form of therapy in Chronic Lymphocytic Leukemia (CLL) patients. The long term goal of this project is to develop a new platform for L-asparaginase therapy that overcomes both of these limitations. This platform has two components: a new recombinant L-asparaginase, developed by our collaborator Dr. Donald Durden, and a nanoparticle delivery vehicle that protects the enzyme from immune response and immune neutralization in vivo. This enzyme, derived from Wolinella succinogenes lacks glutaminase activity that contributes to the toxic side effects associated with the clinicall approved enzymes. However, since immune suppression is one of those side effects abrogated by our enzyme, the immunogenicity problem may be particularly acute if this enzyme is used without some form of shielding. We have developed a nanoparticle delivery system that we term silica hollow enzyme loaded spheres (SHELS). These SHELS are initially synthesized as "nano-wiffle balls" with large holes that allow them to be loaded with unmodified enzymes and subsequently sealed with a porous layer that allows diffusion of small enzyme substrates but is impermeable to large molecules and proteins, thus simultaneously encapsulating the enzyme and protecting it from antibody recognition. We have preliminary data that shows that primary human CLL cells are extremely sensitive to asparagine depletion. Our approach is divided into three general phases aligned with our specific aims. In the first, we will optimize the dosing of recombinant Wolinella succinogenes L-asparaginase loaded in SHELS (rWSasp-SHELS). In the second, we will evaluate the efficacy of rWSasp- SHELS in mice engrafted with either CLL cells alone or CLL cells co-administered with activated T cells to promote division. The former will allow for the testing of the particles in a model representative of the resting CLL population whereas the latter will model the impact of the particles on the proliferative compartment. In the final aim we will compare the toxicity profile of the rWSasp-SHELS with the current clinically used PEG- asparaginase.