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穿膜肽的研究及使用？
穿膜肽（Cell-Penetrating Peptides, CPPs）是短肽链，通常由5到30个氨基酸组成，具有阳离子、两亲性或疏水性特征，能够促进多种货物分子（包括蛋白质、肽、核酸和小分子）通过直接转运或内吞作用进入真核细胞。一旦进入细胞内部，这些货物可以在细胞质、细胞核或其他亚细胞区室中发挥作用。
穿膜肽的研究进展
CPPs的研究已经有三十多年的历史，其家族迅速扩大，并且对新序列的探索仍在继续。科学界已经广泛认可CPPs，并将其用于将各种分子和颗粒导入培养细胞和体内。CPPs在细胞穿透机制方面也得到了深入研究，尽管“细胞穿透”是所有CPPs的共同特征，但它们在细胞表面相互作用的伴侣、引起的细胞反应乃至摄取机制都可能存在很大差异。
穿膜肽的分类和特性
CPPs可以根据其来源、物理化学性质以及修饰和设计程度进行分类。
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阳离子和精氨酸富集CPPs：
- 许多CPPs是阳离子、两亲性或疏水性肽。
- 精氨酸富集CPPs是短阳离子肽，能够穿过真核细胞的质膜。例如，HIV TAT肽和八精氨酸（R8）都是常见的精氨酸富集CPPs。
- 精氨酸含量与CPPs产生负高斯膜曲率的能力直接相关，这种曲率是细胞穿透肽内化的必要条件。
- 有研究发现，聚精氨酸CPPs（如Arg7和Arg8）会结合并抑制SERCA2，这是一种存在于多种细胞类型中的肌浆/内质网钙ATP酶2，可能在实验和治疗环境中产生广泛的影响，因此在使用时需谨慎。相比之下，TAT肽则没有显示出这种抑制作用。
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肽- bismuth 自行车CPPs：
- 最近，研究报告了首批基于肽- bismuth 自行车的CPPs，这些CPPs可以从市售的肽前体中轻松获得，具有广泛的应用潜力。
- 这些CPPs进入人细胞的效率显著提高，比传统的CPPs（如TAT肽或八精氨酸）提高了十倍以上，尽管它们仅需三个正电荷。
- 机制研究表明，其细胞摄取是能量依赖性的内吞作用，对rottlerin抑制敏感，很可能涉及巨胞饮作用。
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折叠肽（Foldamers）CPPs：
- 折叠肽CPPs是具有细胞膜渗透性的折叠寡肽。它们作为药物递送工具，在化学生物学和药物发现领域备受关注。
穿膜肽的细胞内化机制
CPPs可以通过多种机制进入细胞，包括直接转运（direct translocation）或内吞作用（endocytosis）。
- 直接转运：某些CPPs可以直接穿过质膜。
- 内吞作用：大多数CPPs，尤其是在较高浓度下，通过内吞途径进入细胞。例如，肽- bismuth 自行车CPPs的细胞摄取机制是能量依赖性的内吞作用，最可能涉及巨胞饮作用。
穿膜肽在药物递送中的应用
CPPs因其跨越细胞膜的能力，被广泛应用于递送各种生物活性分子，从而改变药物的常规生物分布，以有利地影响毒性、患者依从性和其他治疗因素。
- 小分子药物和治疗性蛋白质：肽基药物递送系统被用作肽-药物偶联物、可注射生物降解颗粒和储库，用于递送小分子药物（<500 Da）和治疗性蛋白质。
- 核酸递送：
- CPPs可以递送核酸，例如小干扰RNA（siRNA）。siRNA虽然能够通过RNA干扰途径沉默特定蛋白质的表达，但其本身难以穿过质膜且在血液中半衰期短。
- CPPs递送siRNA的策略包括共价偶联、形成复合物和CPP修饰的纳米复合物。后两种策略因其合成简单、易用性和多功能性而成为最有前途的方法。
- 非病毒基因递送系统，如Tat和HA2融合肽修饰的明胶-二氧化硅纳米颗粒（GSNPs），能够协同促进细胞内化、内涵体逃逸和细胞核靶向，从而有效地递送治疗性核酸。
- 蛋白质递送：
- CPPs能够将各种蛋白质递送至细胞内，包括酶、转录因子和肿瘤抑制蛋白等。例如，含有TAT肽转导域（PTD）的融合蛋白可以实现多种蛋白质的递送。
- 在眼科治疗中，研究人员开发了一种仿生脂蛋白纳米载体（A-PA-αCPP），利用磷脂酸（PA）和CPP功能化的apoA1模拟肽（αCPP）来非侵入性地递送阿柏西普（一种抗VEGF药物）以治疗眼底新生血管疾病，其局部给药效果与单次玻璃体内注射相当。
- 癌症治疗：
- CPPs被认为是递送治疗性生物大分子的有前景的载体，能够克服这些大分子难以有效穿透细胞质膜的障碍。
- 为了解决CPPs缺乏靶向特异性的问题，研究人员开发了将化疗药物与癌症靶向肽（CTPs）偶联的策略，利用肿瘤微环境或癌细胞的独特特征，提高癌细胞特异性。
- 多组分纳米颗粒与CTPs偶联，可以提供更高效的癌症治疗递送策略。
- 促凋亡肽（如TatBim，一种Tat CPP与Bim凋亡诱导蛋白BH3结构域的融合肽）对癌症治疗具有潜力，通过诱导癌细胞凋亡发挥作用。研究致力于最小化这些CPP-Bim肽，同时保留其有效的凋亡诱导活性。
- 免疫疗法：
- CPPs在免疫疗法中具有巨大优势，能够克服递送障碍。
- CPPs与抗原肽或蛋白质偶联，可以促进抗原呈递细胞（APCs）对MHC I类和MHC II类抗原的有效摄取和交叉呈递，从而诱导体液和细胞毒性免疫反应。
- CPP偶联的免疫调节剂可以增强抗肿瘤免疫或抗炎作用，以调节过敏和自身免疫。
- 纳米疫苗结合CPPs可以增强抗原的摄取、加工和呈递，这是启动免疫反应的基本步骤。CPPs独特的转运特性使其成为高效递送药物、基因、蛋白质和DNA/RNA疫苗的吸引力载体。
- 中枢神经系统（CNS）递送：
- CPPs已被用作跨血脑屏障（BBB）递送剂，将治疗和诊断剂输送到中枢神经系统，这对于CNS靶向放射性药物的开发是一个重大挑战。
- CPP领域的最新发展为设计下一代跨血脑屏障复合物提供了令人兴奋的机会。
- 抗细菌感染：
- CPPs可以与抗菌肽结合，形成具有细胞穿透和抗菌双重功能的自组装纳米肽，用于对抗细胞内细菌感染。例如，F3FT和N3FT纳米肽能够有效穿透细胞并消除细胞内细菌和狙击炎症，主要通过破坏细菌细胞膜和诱导活性氧物种过度积累来杀死细菌。
穿膜肽的优势与挑战
优势：
- 生物相容性好：与合成系统相比，肽基药物递送系统具有更好的生物相容性、生化和生物物理特性以及缺乏毒性。
- 分子量可控：通过固相合成和纯化，可以精确控制分子量。
- 有效递送大分子：能够高效导入多种货物，包括蛋白质、核酸等生物大分子，这些大分子通常难以穿透细胞膜。
- 降低毒性、改善药代动力学：选择性药物递送可以改善药物分布，减少给药剂量和毒性。
- 应用广泛：可用于药物递送、基因治疗、疫苗开发、诊断成像等多个领域。
挑战：
- 缺乏靶向特异性：CPPs普遍缺乏对特定细胞或组织的靶向特异性，这限制了其临床应用。为了解决这个问题，研究人员正在开发将CPPs与癌症靶向肽（CTPs）结合的策略，或利用其他靶向机制。
- 内化机制复杂性：CPPs的细胞进入机制仍然在研究中，不同的CPPs可能采用不同的途径，例如直接转运或内吞作用，这使得对其行为的预测和优化变得复杂。
- 脱靶效应：一些CPPs可能存在脱靶效应，例如聚精氨酸CPPs被发现会结合并抑制SERCA2，从而影响细胞钙稳态，这在实验和治疗环境中需要谨慎考虑。
- 体内稳定性：CPPs在体内的稳定性可能受限，容易被酶降解或快速清除，这需要通过修饰或载体系统来改善。
- 临床转化障碍：尽管CPPs在临床前研究中取得了许多 promising 成果，但将其成功转化为人类疗法仍面临挑战，包括生产成本、免疫原性以及在复杂生物环境中保持效率的问题。
未来展望
CPPs的研究仍在不断深入，未来的重点将放在以下几个方面：
- 提高靶向特异性：开发新型CPPs或结合靶向配体，以实现更精确的细胞或组织特异性递送，从而减少脱靶效应和提高治疗效果。
- 优化递送效率和稳定性：设计更稳定、更高效的CPPs及其偶联物，以改善体内药代动力学和生物利用度。
- 理解和调控内化机制：更深入地阐明CPPs的细胞进入机制，以便根据不同的货物和应用场景设计最佳的递送策略。
- 拓展应用领域：探索CPPs在植物研究和可持续农业等新领域的应用，例如作为递送生物活性分子的工具，以改善作物保护和生产。
- 结合纳米技术：将CPPs与纳米颗粒、脂质体等纳米载体结合，形成多功能递送系统，以克服单独使用CPPs的局限性，例如提高封装效率、稳定性和靶向性。
- 临床试验的进一步推进：鉴于CPPs在治疗急性心肌梗死等疾病的临床研究中已显示出 promising 的结果，未来将有更多CPP偶联疗法进入临床试验阶段，并有望成为新型人类疗法的重要组成部分。
总而言之，穿膜肽作为一种强大的药物递送工具，在生物医学领域展现出巨大的潜力，尤其是在克服生物膜屏障、实现细胞内药物递送方面具有独特优势。尽管仍面临一些挑战，但随着研究的深入，CPPs有望在未来的疾病治疗和生物技术应用中发挥越来越重要的作用。
References
1Cell-penetrating peptides for sustainable agriculture.PubMed
Preeti Patel, Kyle Benzle, Dehua Pei, et al.
Cell-penetrating peptides (CPPs) are short (typically 5-30 amino acids), cationic, amphipathic, or hydrophobic peptides that facilitate the cellular uptake of diverse cargo molecules by eukaryotic cells via direct translocation or endocytosis across the plasma membrane. CPPs can deliver a variety of bioactive cargos, including proteins, peptides, nucleic acids, and small molecules into the cell. Once inside, the delivered cargo may function in the cytosol, nucleus, or other subcellular compartments. Numerous CPPs have been used for studies and drug delivery in mammalian systems. Although CPPs have many potential uses in plant research and agriculture, the application of CPPs in plants remains limited. Here we review the structures and mechanisms of CPPs and highlight their potential applications for sustainable agriculture.
2Peptide-Based Drug Delivery Systems.PubMed
Dmitriy Berillo, Adilkhan Yeskendir, Zharylkasyn Zharkinbekov, et al.
Peptide-based drug delivery systems have many advantages when compared to synthetic systems in that they have better biocompatibility, biochemical and biophysical properties, lack of toxicity, controlled molecular weight via solid phase synthesis and purification. Lysosomes, solid lipid nanoparticles, dendrimers, polymeric micelles can be applied by intravenous administration, however they are of artificial nature and thus may induce side effects and possess lack of ability to penetrate the blood-brain barrier. An analysis of nontoxic drug delivery systems and an establishment of prospective trends in the development of drug delivery systems was needed. This review paper summarizes data, mainly from the past 5 years, devoted to the use of peptide-based carriers for delivery of various toxic drugs, mostly anticancer or drugs with limiting bioavailability. Peptide-based drug delivery platforms are utilized as peptide-drug conjugates, injectable biodegradable particles and depots for delivering small molecule pharmaceutical substances (500 Da) and therapeutic proteins. Controlled drug delivery systems that can effectively deliver anticancer and peptide-based drugs leading to accelerated recovery without significant side effects are discussed. Moreover, cell penetrating peptides and their molecular mechanisms as targeting peptides, as well as stimuli responsive (enzyme-responsive and pH-responsive) peptides and peptide-based self-assembly scaffolds are also reviewed.
3Synergistic effects of cell-penetrating peptide Tat and fusogenic peptide HA2-enhanced cellular internalization and gene transduction of organosilica nanoparticles.PubMed
She-fang Ye, Miao-miao Tian, Tian-xiao Wang, et al.
The nonviral gene delivery system is an attractive alternative to cancer therapy. A new kind of gelatin-silica nanoparticles (GSNPs) was developed through a two-step sol-gel procedure. To improve the transfection efficacy, GSNPs modified with different fusion peptides (Tat, HA2, R8, Tat/HA2, and Tat/R8) were prepared for particle size, zeta potential, cellular uptake, hemolysis activity at physiological pH (7.0) or acidic pH (5.0), and condensation of plasmid DNA. The results suggest that the sizes and zeta potentials of GS-peptide conjugates were 147 - 161 nm and 19 - 33 mV, respectively; GS-peptide conjugates exhibited low cytotoxicity; the plasmid DNA was readily entrapped at a GS-peptide/pDNA weight ratio of 50 - 200. The in vitro and in vivo studies demonstrated that the synergistic effects of cell-penetrating peptide Tat and fusogenic peptide HA2 could promote the efficient cellular internalization, endosome escape, and nucleus targeting, hence delivering the therapeutic nucleic acid efficiently.
4Unleashing cell-penetrating peptide applications for immunotherapy.PubMed
Ja-Hyun Koo, Gil-Ran Kim, Kyung-Ho Nam, et al.
With the advent of cancer immunotherapy, immunomodulation has emerged as an important strategy for the treatment of various diseases. We review recent advances in clinical trials of cell-penetrating peptide (CPP) applications for immunotherapy and also discuss their challenges and opportunities for preclinical studies in various immune diseases. CPP conjugation to antigenic peptides or proteins can enable efficient antigen uptake and cross-presentation by antigen-presenting cells (APCs), which induce both humoral and cytotoxic responses. In addition, CPP-coupled immune modulators can enhance antitumor immunity or anti-inflammatory effects to regulate allergies and autoimmunity. Given their huge advantages in overcoming delivery barriers, CPP-based strategies for immunomodulation could extend drug optimization and advance immunotherapy in various human diseases.
5Classes of Cell-Penetrating Peptides.PubMed
Margus Pooga, Ülo Langel
During the three decades of cell-penetrating peptides era the superfamily of CPPs has rapidly expanded, and the quest for new sequences continues. CPPs have been well recognized by scientific community and they have been used for transduction of a wide variety of molecules and particles into cultured cells and in vivo. In parallel with application of CPPs for delivering of active payloads, the mechanisms that such peptides take advantage of for gaining access to cells' insides have been in the focus of intense studies. Although the common denominator "cell penetration" unites all CPPs, the interaction partners on the cell surface, evoked cellular responses and even the uptake mechanisms might greatly vary between different peptide types. Here we present some possibilities for classification of CPPs based on their type of origin, physical-chemical properties, and the extent of modifications and design efforts. We also briefly analyze the internalization mechanisms with regard to their classification into groups based on physical-chemical characteristics.
6Arginine-rich cell-penetrating peptides.PubMed
Nathan Schmidt, Abhijit Mishra, Ghee Hwee Lai, et al.
Arginine-rich cell-penetrating peptides are short cationic peptides capable of traversing the plasma membranes of eukaryotic cells. While successful intracellular delivery of many biologically active macromolecules has been accomplished using these peptides, their mechanisms of cell entry are still under investigation. Recent dialogue has centered on a debate over the roles that direct translocation and endocytotic pathways play in internalization of cell-penetrating peptides. In this paper, we review the evidence for the broad range of proposed mechanisms, and show that each distinct process requires negative Gaussian membrane curvature as a necessary condition. Generation of negative Gaussian curvature by cell-penetrating peptides is directly related to their arginine content. We illustrate these concepts using HIV TAT as an example.
7Cell-Penetrating Peptide-Bismuth Bicycles.PubMed
Saan Voss, Liam D Adair, Katharina Achazi, et al.
Cell-penetrating peptides (CPPs) play a significant role in the delivery of cargos into human cells. We report the first CPPs based on peptide-bismuth bicycles, which can be readily obtained from commercially available peptide precursors, making them accessible for a wide range of applications. These CPPs enter human cells as demonstrated by live-cell confocal microscopy using fluorescently labelled peptides. We report efficient sequences that demonstrate increased cellular uptake compared to conventional CPPs like the TAT peptide (derived from the transactivating transcriptional activator of human immunodeficiency virus 1) or octaarginine (R ), despite requiring only three positive charges. Bicyclization triggered by the presence of bismuth(III) increases cellular uptake by more than one order of magnitude. Through the analysis of cell lysates using inductive coupled plasma mass spectrometry (ICP-MS), we have introduced an alternative approach to examine the cellular uptake of CPPs. This has allowed us to confirm the presence of bismuth in cells after exposure to our CPPs. Mechanistic studies indicated an energy-dependent endocytic cellular uptake sensitive to inhibition by rottlerin, most likely involving macropinocytosis.
8Cell-Penetrating Peptides Delivering siRNAs: An Overview.PubMed
Luca Falato, Maxime Gestin, Ülo Langel
Cell-Penetrating Peptides (CPP) are valuable tools capable of crossing the plasma membrane to deliver therapeutic cargo inside cells. Small interfering RNAs (siRNA) are double-stranded RNA molecules capable of silencing the expression of a specific protein triggering the RNA interference (RNAi) pathway, but they are unable to cross the plasma membrane and have a short half-life in the bloodstream. In this overview, we assessed the many different approaches used and developed in the last two decades to deliver siRNA through the plasma membrane through different CPPs sorted according to three different loading strategies: covalent conjugation, complex formation, and CPP-decorated (functionalized) nanocomplexes. Each of these strategies has pros and cons, but it appears the latter two are the most commonly reported and emerging as the most promising strategies due to their simplicity of synthesis, use, and versatility. Recent progress with siRNA delivered by CPPs seems to focus on targeted delivery to reduce side effects and amount of drugs used, and it appears to be among the most promising use for CPPs in future clinical applications.
9Trojan Horse-Inspired Biomimetic Lipoprotein Nanocarrier for Noninvasive Anti-VEGF Therapy of Ocular Fundus Neovascularization.PubMed
Lin Chen, Chen Fan, Antian Wang, et al.
Pathological neovascularization is a major cause of vision loss in various ischemic and inflammatory ocular fundus diseases, with current anti-VEGF therapies requiring invasive and frequent intravitreal injections. In this study, a Trojan horse-inspired biomimetic lipoprotein nanocarrier (A-PA-αCPP) is developed for noninvasive anti-VEGF therapy via topical administration. Utilizing phosphatidic acid (PA), specifically 1,2-dioleoyl-sn-glycero-3-phosphate (DOPA), as the sole phospholipid, the nanocarrier enables electrostatic tethering of aflibercept, with a cell-penetrating peptide (CPP)-functionalized apoA1 mimetic peptide (αCPP) incorporated through self-assembly. Molecular dynamics simulations and intermolecular interaction experiments confirm effective PA-protein interactions and αCPP incorporation. The nanocarrier demonstrates high encapsulation efficiency, size uniformity, and long-term stability. It enhances transcellular and paracellular transport by promoting cellular uptake, bypassing lysosomal degradation, and modulating tight junctions. When topically administered, A-PA-αCPP efficiently delivers to the posterior segment via corneal and conjunctival-scleral-choroidal pathways. In a laser-induced choroidal neovascularization (CNV) mouse model, A-PA-αCPP eye drops show therapeutic efficacy comparable to a single intravitreal injection, while maintaining excellent ocular and systemic biocompatibility. This biomimetic platform highlights the potential of PA-mediated protein binding in nanocarrier design, offering a promising strategy for home-based anti-VEGF therapy and extending the potential to other protein-based therapeutics.
10Cancer targeting peptides.PubMed
Mona Kalmouni, Sumaya Al-Hosani, Mazin Magzoub
Despite continuing advances in the development of biomacromolecules for therapeutic purposes, successful application of these often large and hydrophilic molecules has been hindered by their inability to efficiently traverse the cellular plasma membrane. In recent years, cell-penetrating peptides (CPPs) have received considerable attention as a promising class of delivery vectors due to their ability to mediate the efficient import of a large number of cargoes in vitro and in vivo. However, the lack of target specificity of CPPs remains a major obstacle to their clinical development. To address this issue, researchers have developed strategies in which chemotherapeutic drugs are conjugated to cancer targeting peptides (CTPs) that exploit the unique characteristics of the tumor microenvironment or cancer cells, thereby improving cancer cell specificity. This review highlights several of these strategies that are currently in use, and discusses how multi-component nanoparticles conjugated to CTPs can be designed to provide a more efficient cancer therapeutic delivery strategy.
11Cell-penetrating Peptide-mediated Nanovaccine Delivery.PubMed
Jizong Jiang
Vaccination with small antigens, such as proteins, peptides, or nucleic acids, is used to activate the immune system and trigger the protective immune responses against a pathogen. Currently, nanovaccines are undergoing development instead of conventional vaccines. The size of nanovaccines is in the range of 10-500 nm, which enables them to be readily taken up by cells and exhibit improved safety profiles. However, low-level immune responses, as the removal of redundant pathogens, trigger counter-effective activation of the immune system invalidly and present a challenging obstacle to antigen recognition and its uptake via antigen-presenting cells (APCs). In addition, toxicity can be substantial. To overcome these problems, a variety of cell-penetrating peptide (CPP)-mediated vaccine delivery systems based on nanotechnology have been proposed, most of which are designed to improve the stability of antigens in vivo and their delivery into immune cells. CPPs are particularly attractive components of antigen delivery. Thus, the unique translocation property of CPPs ensures that they remain an attractive carrier with the capacity to deliver cargo in an efficient manner for the application of drugs, gene transfer, protein, and DNA/RNA vaccination delivery. CPP-mediated nanovaccines can enhance antigen uptake, processing, and presentation by APCs, which are the fundamental steps in initiating an immune response. This review describes the different types of CPP-based nanovaccines delivery strategies.
12Protein Delivery by PTDs/CPPs.PubMed
Caroline Palm-Apergi, Steven F Dowdy
The ability to deliver or transduce proteins into cells allows for the manipulation of cell biology in culture, preclinical models, and potentially human disease. Fusion proteins containing the TAT peptide transduction domain (PTD), also known as cell-penetrating peptide (CPP), allow for delivery of a wide variety of proteins, including enzymes, transcription factors, tumor suppressor proteins, and many more. TAT-fusion proteins are generated cloning in-frame into the pTAT-HA plasmid, then transformed into E. coli for expression, and purified by the 6-His affinity tag over Ni-NTA column, followed by a final IEX FPLC purification step.
13Therapeutic applications of cell-penetrating peptides.PubMed
Randolph M Johnson, Stephen D Harrison, Derek Maclean
Since the discovery over 15 years ago of a protein transcription factor that possessed the ability to cross the plasma membrane, cell-penetrating peptides (CPPs) have been evaluated for the ability to transport diverse cargoes into cells, tissues, and organs. Certain CPPs have been used for the intracellular delivery of information-rich molecules to modulate protein-protein interactions and thereby inhibit key cellular mechanisms of disease. The ability to introduce drugs into cells allows the conventional biodistribution of drugs to be altered in order to favorably impact toxicity, patient compliance, and other treatment factors. In this monograph, we present the current status and future prospects for the application of CPPs to the development of human therapeutics. We discuss some of the advantages and disadvantages of using CPPs in the in vivo setting, and review the current status of a number of preclinical and human clinical studies of CPP-mediated delivery of therapeutics. These include CPP-conjugated moieties directed against a growing variety of targets and disease areas, including cancer, cardiology, pain, and stroke. Our discussion focuses on those therapeutics that have been tested in humans, including a CPP conjugate for the treatment of acute myocardial infarction. The promising results obtained in a number of these studies indicate that CPPs may have an important role in the development of novel therapeutics.
14Peptides for trans-blood-brain barrier delivery.PubMed
Reuben Blades, Lars M Ittner, Ole Tietz
Trans-blood-brain barrier (BBB) delivery of therapeutic and diagnostic agents is a major challenge in the development of central nervous system (CNS) targeted radiopharmaceuticals. This review is an introduction to the use of peptides as delivery agents to transport cargos into the CNS. The most widely used BBB-penetrating peptides are reviewed here, with a particular emphasis on the broad range of cargos delivered into the CNS using these. Cell-penetrating peptides (CPPs) have been deployed as trans-BBB delivery agents for some time; new developments in the CPP field offer exciting opportunities for the design of next generation trans-BBB complexes. Many of the peptides highlighted here are ready to be combined with diagnostic and therapeutic radiopharmaceuticals to develop highly effective CNS-targeted agents.
15Polyarginine Cell-Penetrating Peptides Bind and Inhibit SERCA2.PubMed
Per Kristian Lunde, Ornella Manfra, Thea Parsberg Støle, et al.
Cell-penetrating peptides (CPPs) are short peptide sequences that have the ability to cross the cell membrane and deliver cargo. Although it is critical that CPPs accomplish this task with minimal off-target effects, such actions have in many cases not been robustly screened. We presently investigated whether the commonly used CPPs TAT and the polyarginines Arg and Arg exert off-target effects on cellular Ca homeostasis. In experiments employing myocytes and homogenates from the cardiac left ventricle or soleus muscle, we observed marked inhibition of Ca recycling into the sarcoplasmic reticulum (SR) following incubation with polyarginine CPPs. In both tissues, the rate of SR Ca leak remained unchanged, indicating that protracted Ca removal from the cytosol stemmed from inhibition of the SR Ca ATPase 2 (SERCA2). No such inhibition occurred following treatment with TAT, or in preparations from the SERCA1-expressing extensor digitorum longus muscle. Experiments in HEK cells overexpressing individual SERCA isoforms confirmed that polyarginine incubation specifically inhibited the activity of SERCA2a and 2b, but not SERCA1 or 3. The attenuation of SERCA2 activity was not dependent on the presence of phospholamban, and ELISA-based analyses rather revealed direct interaction between the polyarginines and the actuator domain of the protein. Surface plasmon resonance experiments confirmed strong binding within this region of SERCA2, and slow dissociation between the two species. Based on these observations, we urge caution when employing polyarginine CPPs. Indeed, as SERCA2 is expressed in diverse cell types, the wide-ranging consequences of SERCA2 binding and inhibition should be anticipated in both experimental and therapeutic settings.
16Recent CPP-based applications in medicine.PubMed
Kaido Kurrikoff, Ülo Langel
: Cell-penetrating peptides (CPP) offer versatile tools for the field of drug delivery and development of macromolecular therapeutics. These tools have matured into applications that allow the use of proteins, nucleic acid, peptides, imaging agents, and low molecular weight drugs as therapeutic entities. : The progress of the field is discussed in the current review, with the examples from a recent couple of years, in the utilization of CPPs in medicine. Specific focus is on the research articles that include applications that have direct translational value. Progress with protein mimicry and its recent leap in medicine is discussed with special attention, but also achievements with nanoformulation and drug targeting is presented. : The opinion section discusses some of the limitations and shortcomings, proposing the areas where more effort should be invested in order to increase the translational value of the current preclinical research.
17Self-assembling peptide with dual function of cell penetration and antibacterial as a nano weapon to combat intracellular bacteria.PubMed
Yongjie Zhu, Wanying Xu, Wenwen Chen, et al.
Intracellular bacterial infections and antimicrobial resistance are threatening global public health systems. Antimicrobial peptides are a potential solution to combat bacterial resistance, but the design of self-assembled nanopeptides with dual functions of cell penetration and antibacterial properties to combat intracellular bacteria remains a challenge. Here, we propose a strategy to develop self-assembled nanopeptides with dual functions through the chimerization of self-assembled core, hydrophobic motif, and cell-permeable unit. The optimal nanopeptides, F3FT and N3FT, exhibited potent antibacterial activity and excellent biocompatibility. Crucially, F3FT and N3FT are able to efficiently penetrate cells and eliminate intracellular bacteria and sniping inflammation. Moreover, F3FT and N3FT kill bacteria mainly by disrupting bacterial cell membranes and inducing excessive accumulation of reactive oxygen species. F3FT and N3FT have exhibited good safety and potent therapeutic potential in vivo. This scheme of constructing nanopeptides through multifunctional domains design provides a paradigm for dealing with escalating of intracellular bacteria and antimicrobial resistance.
18Cell-Penetrating Peptide Foldamers: Drug-Delivery Tools.PubMed
Makoto Oba
Highly efficient drug-delivery tools for membrane-impermeable compounds, proteins, and nucleic acids in living cells are useful in the fields of chemical biology and drug discovery, and such tools have been widely studied. One strategy in the development of novel drug-delivery tools is to utilize cell-penetrating peptide (CPP) foldamers. CPP foldamers are folded oligopeptides that possess cell membrane permeability. In recent decades, a wide variety of CPP foldamers have been reported by many groups. Herein, CPP foldamers are introduced and discussed from the viewpoints of component monomers (amino acids) and their application as drug-delivery tools.
19Minimization of apoptosis-inducing CPP-Bim peptide.PubMed
Shengli Zhou, Kazunori Watanabe, Seiichiro Koide, et al.
Pro-apoptotic peptides may be promising agents for cancer therapy owing to their ability to induce apoptosis in cancer cells. TatBim, a fusion peptide of Tat cell-penetrating peptide (CPP) and the BH3 domain derived from Bim apoptosis-inducing protein, is a pro-apoptotic peptide. In this study, based on the TatBim sequence, we attempted to minimize the CPP-Bim peptide while retaining apoptosis-inducing activity. The CPP and Bim parts were systematically shortened, and the pro-apoptotic activities of the shortened peptides were examined. We obtained TatBim-N1C2 and R8Bim-N1C2 as minimized peptides with efficient apoptotic activity. These peptides may have potential applications in future biomedical studies, such as cancer therapeutics.
20Therapeutic potential of cell-penetrating peptides.PubMed
Luís Vasconcelos, Kalle Pärn, Ulo Langel
The ability of cell-penetrating peptides to cross plasma membranes has been used for various applications, including the delivery of bioactive molecules to inhibit disease-producing cellular mechanisms. Selective drug delivery into target cells improves drug distribution and decreases dosing and toxicity. In this review, the authors outline the main challenges in the field, namely clarification of mechanisms of entry into cells, as well as current and future perspectives regarding cell-penetrating peptides application for human therapeutics. Here, the authors discuss some of the factors that influence efficacy of delivery and review the current status of preclinical studies and clinical trials involving the use of cell-penetrating peptide-mediated delivery of therapeutics.