Nanoparticle Enabled L-asparaginase Therapy for CLL

描述（由申请人提供）：l -天冬酰胺酶用于治疗急性淋巴细胞白血病（ALL）已有40多年的历史。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缺乏敏感，但由于几个原因，天冬酰胺酶治疗尚未扩展到儿童白血病。首先是l -天冬酰胺酶的免疫原性，它可以限制治疗的持续时间，并有助于潜在的副作用，如过敏反应和过敏反应。其他副作用，包括胰腺炎和凝血功能障碍，也可能限制这种治疗形式对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l -天冬酰胺酶治疗的新平台，克服这两个限制。这个平台有两个组成部分：一种新的重组l -天冬酰胺酶，由我们的合作者Donald Durden博士开发，以及一种纳米颗粒递送载体，可以保护酶免受体内免疫反应和免疫中和。这种酶来源于琥珀酸Wolinella succinogenes，缺乏谷氨酰胺酶活性，导致与临床批准的酶相关的毒副作用。然而，由于免疫抑制是我们的酶消除的副作用之一，如果这种酶在没有某种形式的屏蔽的情况下使用，免疫原性问题可能会特别严重。我们已经开发了一种纳米颗粒输送系统，我们称之为二氧化硅空心酶负载球（SHELS）。这些外壳最初被合成为带有大孔的“纳米威夫球”，允许它们装载未修饰的酶，随后用多孔层密封，允许小酶底物扩散，但大分子和蛋白质无法渗透，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，人类原发性CLL细胞对天冬酰胺耗竭极为敏感。我们的方法根据我们的具体目标分为三个一般阶段。首先，我们将优化重组琥珀酸Wolinella succinogenes l -天冬酰胺酶（rWSasp-SHELS）负载SHELS的剂量。在第二项研究中，我们将评估rWSasp- SHELS在单独移植CLL细胞或CLL细胞与活化T细胞共同移植以促进分裂的小鼠中的功效。前者将允许在代表静止CLL群体的模型中测试颗粒，而后者将模拟颗粒对增殖室的影响。在最后的目的中，我们将比较rwasp - shels与目前临床使用的PEG-天冬酰胺酶的毒性。

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.