Crystalline Endolysin Treatment for Tuberculosis in TB/HIV co-infected patients

描述（由申请人提供）：虽然结核病通常被认为是一种过去的疾病，但它仍然是全世界人民的主要杀手。截至2008年底，约有20亿人（占世界人口的三分之一）感染结核病病原结核分枝杆菌（MTb）。世界卫生组织估计，全世界每年有200万人死于结核病。结核病现在是仅次于人体免疫机能丧失病毒/后天免疫机能丧失综合症（艾滋病毒/艾滋病）的世界上第二大传染病致死原因。20世纪80年代中期，美国再次爆发结核病，重新引起人们对结核病的关注。结核病病例增加的主要原因是由于感染艾滋病毒/艾滋病的人更容易受到感染，以及结核分枝杆菌耐药菌株的发展，结核分枝杆菌导致人患结核病。世卫组织2010年4月最新报告的结果显示，在俄罗斯，每4名被诊断为耐多药结核病的结核患者中就有1人，耐多药结核病负担的约50%在中国和印度。2008年全世界有44万耐多药结核病病例，15万人死亡。根据2010年3月的报告，耐多药结核病例中约有5.4%为广泛耐药结核。如果没有新的、更有效的药物、诊断方法和疫苗，消除结核病是不切实际的。世界迫切需要更简单、更快的治疗药物；更安全有效的药物，可以治疗所有形式的结核病，特别是对艾滋病毒/结核病合并感染的免疫功能低下人群。作为抗生素治疗的替代方案，特别是对于由结核分枝杆菌引起的全身肺部感染，我们建议开发一种可以在各种配方条件和给药途径下使用的酶疗法。这种酶治疗可能和抗生素治疗一样有效，因为这种酶是高度特异性的，对目标细菌是有效的。然而，将这些酶运送到感染部位仍然是一个挑战。大多数药物蛋白在给药部位容易降解，无论是静脉、皮下、口服、局部还是通过任何其他途径给药。我们相信，一种安全有效的递送系统可能是通过使用蛋白质晶体，它可以提供高度浓缩、稳定和纯净的产品。此外，由于结核分枝杆菌存在于巨噬细胞中，因此还需要将内溶素输送到细胞内的技术。细胞穿透肽（CPP），无论是框架内工程还是非共价吸附/结合到蛋白质底物上，已被用于将各种各样的分子输送到许多细胞类型中。在本提案中，我们建议开发一种晶体配方重组细胞穿透肽（CPP）内溶素疗法，可用于各种配方条件和递送途径。CPP与内溶素结合的机会是多重的，因为它可能能够穿透巨噬细胞，从而能够杀死HIV/TB合并感染患者的结核分枝杆菌，并可能在最小的不适下延长患者的寿命。在I期研究中，我们打算研究使用内溶素（含或不含细胞穿透肽的可溶性和结晶型制剂）作为治疗结核分枝杆菌感染的体内动物模型的全身递送系统的可行性。如果成功，这种方法将导致引入新的、有效的治疗结核分枝杆菌感染的酶疗法。

DESCRIPTION (provided by applicant): Though often considered to be a disease of the past, tuberculosis (TB) is still a leading killer of people worldwide. At the end of 2008, approximately 2 billion people one-third of the world's population is infected with Mycobacterium tuberculosis (MTb), the etiologic agent of TB. The World Health Organization estimates that 2 million people worldwide die of TB each year. TB is now the second most common cause of death from infectious disease in the world after human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS). In the mid-1980s, a resurgence of outbreaks in the United States brought renewed attention to TB. Main reasons for increase in TB cases are due to higher vulnerability of people infected with HIV/AIDS and development of drug-resistant strains of MTb, which causes TB in human. In the latest 2010 April WHO report results show that one in every 4 TB patients diagnosed with MDR-TB in Russia and about 50% of MDR-TB burden is in China and India. Worldwide there are 440,000 MDR-TB cases and 150,000 deaths occurred in 2008. About 5.4% of MDR-TB cases have XDR-TB according to the report in March 2010.It is impractical to eliminate TB without novel, and more effective drugs, diagnostics and vaccines. The world immediately needs simpler and faster curative drugs; safer and effective drugs that can treat all forms of TB especially in immuno- compromised people with HIV/TB co-infections. As an alternative to antibiotic therapy, especially for systemic lung infections by M. tuberculosis, we are proposing to develop an enzyme therapy that can be used with various formulation conditions and delivery routes. The enzyme therapy may be as effective as antibiotic therapy because the enzyme is highly specific, and potent against the targeted bacteria. However, delivering these enzymes to the site of the infection remains a challenge. Most pharmaceutical proteins are susceptible to degradation at the site of administration, whether administered intravenously, subcutaneously, orally, topically or by any other route. We believe that a safe and effective delivery system for this application may be through the use of protein crystals, which provides highly concentrated, stable, and pure products. In addition, technologies are also needed to deliver endolysin to intracellular compartment since M. tuberculosis resides in macrophages. Cell penetrating peptides (CPP), either engineered in-frame or adsorbed/bound non-covalently to protein substrates, have been used to deliver a wide variety of molecules into a number of cell types. In this proposal we are proposing to develop a crystalline formulated recombinant endolysin therapy with cell-penetrating peptides (CPP) that can be used with various formulation conditions and delivery routes. The opportunities for CPP conjugated to endolysin are multiple since it may be able to penetrate the macrophages thereby able to kill M. tuberculosis in HIV/TB co-infected patients and may extend their life span with minimal discomfort. In Phase I, we intend to study the feasibility of using endolysins, both soluble and crystalline formulations with or without cell penetrating peptides, as a systemic delivery system in an in vivo animal model for M. tuberculosis infection. If successful, this approach will lead to the introduction of novel, efficient enzyme therapy for the treatment of M. tuberculosis infections. PUBLIC HEALTH RELEVANCE: Tuberculosis (TB) is one of the most devastating global health problems of our time, causing approximately 2 million deaths a year. As a preventive measure, currently BCG vaccine is used to avoid serious complications of TB and to reduce the rate of pediatric TB but It has little or no effectiveness for pulmonary TB in adults. We are proposing a novel crystalline endolysin that is pure, safe, effective even in immunocompromised patients, and unlike vaccines which needs immune system to generate the antibodies the endolysin can act directly on the Mycobacterium tuberculosis organism and thus prevent and treat tuberculosis in HIV/TB co-infected patients.