Identified Opportunities 确定的机会

To Be Explored. Specific opportunities for NANPs include but are not limited to the following product categories: (i) subunit and peptide vaccines; (ii) artificial antigen-presenting cells (APCs) that promote cytotoxic T lymphocyte (CTL) activation; (iii) caged and multispecific affinity platforms as CTL redirectors and modulators; (iv) monoclonal antibody (mAb) mimics; and (v) virus-like nanoparticles for targeted delivery of mRNAs, antisense oligonucleotides (ASOs), and other therapeutic nucleic acid payloads including clustered regularly interspaced short palindromic repeats (CRISPR) ribonucleoproteins.

有待探索。 NANP的特殊机会包括但不限于以下产品类别：（i）亚基和肽疫苗； （ii）促进细胞毒性T淋巴细胞（CTL）活化的人工抗原呈递细胞（APC）； （iii）笼状和多特异性亲和平台，作为CTL重定向器和调节剂； （iv）单克隆抗体（mAb）模拟物； （v）靶向mRNA，反义寡核苷酸（ASO）和其他治疗性核酸有效载荷的病毒样纳米颗粒，包括成簇的规则间隔的短回文重复序列（CRISPR）核糖核蛋白。

clinical opportunities

vaccines亚基和肽疫苗； 

mAb mimics单克隆抗体（mAb）模拟物； 

CTL activition促进细胞毒性T淋巴细胞（CTL）活化的人工抗原呈递细胞（APC）；

CTL redirectors and modulators笼状和多特异性亲和平台，作为CTL重定向器和调节剂；

In vivo dielivery of gene therpeutics

strategy to overcome transiational barriers

expand therapeutic space

demonstrate efficacy in vivo

simplify design and manufacturing

establish universal nomenclature and protocols

create working groups

unify efforts of pharma, academica and govenment

Overview of(1) clinical opportunities for nucleic acid nanoparticles (NANP) therapeutics include small interfering RNAs, antisense oligonucleotides, mRNAs, CRISPR ribonucleoproteins, and vaccine vectors; (2) translational barriers that are limiting progress from academia to the clinic to benefit patients; (3) proposed strategy to overcome these translational barriers; and (4) our recommendation of a consortium to help translate NANPs to the clinic.

（1）核酸纳米粒子（NANP）治疗剂的临床机会包括小干扰RNA，反义寡核苷酸，mRNA，CRISPR核糖核蛋白和疫苗载体; （2）转化障碍阻碍了从学术界到临床的发展，使患者受益； （3）提出克服这些转化障碍的策略； （4）我们建议的一个联盟，以帮助将NANP转换为临床。

The overall goals of prophylactic vaccines and immunotherapies are to stimulate innate and adaptive immunity and to restore immune homeostasis without affecting the patient. Although various types of immune cells are involved in mounting the response against a pathogen, APCs, and particularly dendritic cells (DCs), represent the most common targets for vaccine delivery. Substances that help APCs to recognize, to process, and to present antigenic fragments to T cells, also known as adjuvants, are commonly used to optimize immune response and to promote the generation of memory B cells. Typical cellular targets for immunotherapies are DCs, macrophages, and cytotoxic T lymphocytes. Various nanoparticles are commonly considered for vaccines and immunotherapy delivery, and the benefits of their use in this field have been extensively discussed elsewhere. Recent advances in vaccines and immunotherapies, along with beneficial properties of NANPs for these indications, have also been reviewed in detail. Herein, we briefly review several of them to highlight this opportunity.

预防性疫苗和免疫疗法的总体目标是刺激先天性和适应性免疫力，并在不影响患者的情况下恢复免疫稳态。尽管各种类型的免疫细胞参与了针对病原体的应答，但APC，尤其是树突状细胞（DC）代表了最常见的疫苗递送目标。帮助APC识别，加工和向T细胞呈递抗原性片段的物质（也称为佐剂）通常用于优化免疫反应和促进记忆B细胞的生成。免疫疗法的典型细胞靶标是DC，巨噬细胞和细胞毒性T淋巴细胞。通常考虑将各种纳米颗粒用于疫苗和免疫疗法的递送，并且在其他领域已经广泛讨论了其在该领域中使用的益处。疫苗和免疫疗法的最新进展，以及针对这些适应症的NANP的有益特性，也得到了详尽的综述。在此，我们简要回顾其中的几个以突出此机会。

Antigen-presenting cells exposed to NANPs typically secrete type I and type III interferons (IFNs), which are known for their ability to induce DC maturation. The magnitude of IFN induction in response to NANPs depends in part on their physicochemical properties (e.g., molecular weight, CpG composition, size, and geometry); therefore, the ability of NANPs to induce an IFN response may help to personalize vaccines so that IFN levels that are optimal for a given individual may be induced. This property may be combined with subunit or peptide delivery and is particularly important because inflammation triggered by adjuvants varies between individuals.Cytokine levels in response to the same adjuvant may be insufficient for effective vaccination for one individual and too toxic for another. Customizing vaccines to the individual’s immune system is, therefore, an attractive avenue for personalized vaccination that is both efficacious and free from adverse immune-mediated effects. Both types I and III IFNs in the form of recombinant protein therapeutics are used to combat viral infections and cancer. Despite proven efficacy, systemic administration of recombinant proteins may cause side effects (e.g., fever or fever-like reactions and chills)36 and induce antidrug antibodies (ADAs). Such ADAs may affect drug efficacy and cause toxicity and, in some instances, neutralize recombinant protein therapeutics. A variety of approaches are used to minimize the immunogenicity of recombinant IFNs, but none is completely efficient at eliminating the ADA response. For example, one of the most commonly used approaches conjugation of protein to polyethylene glycol (PEG)often fails due to the immunogenicity of the PEG itself and the presence of pre-existing antiPEG antibodies in healthy donors’ blood. Due to their ability to induce IFN responses, NANPs could potentially provide a solution to these problems by directing a patient’s immune system to produce its own IFNs that are not immunogenic. However, significant research is still needed to understand the induction of ADAs by NANPs themselves, which may lead to neutralization or, in worse cases, lupus-like pathology and disease.

暴露于NANP的抗原呈递细胞通常分泌I型和III型干扰素（IFN），它们以诱导DC成熟的能力而闻名。响应于NANP的IFN诱导程度部分取决于其理化特性（例如分子量，CpG组成，大小和几何形状）;因此，NANPs诱导IFN反应的能力可能有助于个性化疫苗，从而可以诱导出对于给定个体而言最佳的IFN水平。此特性可能与亚基或肽的传递相结合，并且特别重要，因为佐剂触发的炎症在个体之间会有所不同。响应同一佐剂的细胞因子水平可能不足以对一个个体进行有效疫苗接种，而对另一个个体则毒性太大。因此，针对个人免疫系统定制疫苗是有效且无不良免疫介导作用的个性化疫苗的诱人途径。重组蛋白治疗剂形式的I型和III型IFN均用于抵抗病毒感染和癌症。尽管已证明有效，重组蛋白的全身给药可能会引起副作用（例如发烧或发烧样反应和发冷）36并诱导抗药物抗体（ADAs）。此类ADA可能会影响药物功效并引起毒性，并且在某些情况下会中和重组蛋白治疗剂。使用了多种方法来最小化重组IFN的免疫原性，但是没有一种方法能够完全有效地消除ADA反应。例如，一种最常用的方法-将蛋白质与聚乙二醇（PEG）结合-通常由于PEG本身的免疫原性和健康捐献者血液中存在的抗PEG抗体而失败。由于具有诱导IFN反应的能力，NANP可以通过指导患者的免疫系统产生自己的非免疫原性IFN，从而为这些问题提供解决方案。然而，仍需要大量研究来了解NANPs自身对ADA的诱导，这可能导致中和，或更坏的情况下，可能导致狼疮样病理和疾病。

Substances that activate Toll-like receptors (TLRs) are popular adjuvants in the vaccine and immunotherapy fields. However, some of these substances, particularly TLR7/8 agonists, are too toxic to be injected into the blood, so an alternative method of delivery must be utilized. Hong et al. demonstrated that NANPs made of RNA activate an IFN response via TLR7. When exposed to human blood cells, NANPs do not cause inflammation or an IFN response unless their uptake by the blood cells is directed by a delivery carrier.These findings emphasize an important property of RNA-based NANPs, namely, a potent TLR7 agonist ability that manifests only af ter NANPs are internalized by APCs, thus creating an opportunity for NANPs to be used as deliverycontrolled adjuvants. This agonistic activity makes NANPs an alternative to resiquimod, a well-known TLR 7/8 agonist that is only safe for use in topical applications due to the overt immunostimulation that occurs immediately when this adjuvant enters the systemic circulation.

激活Toll样受体（TLR）的物质是疫苗和免疫治疗领域的常用佐剂。但是，这些物质中的某些物质，特别是TLR7 / 8激动剂，毒性太大，无法注入血液，因此必须采用另一种递送方法。 Hong等。证明由RNA制成的NANP通过TLR7激活IFN应答。当暴露于人类血细胞中时，NANPs不会引起炎症或IFN反应，除非它们被血细胞摄取是由递送载体指导的。这些发现强调了基于RNA的NANPs的重要特性，即强大的TLR7激动剂能力。这表明只有在ANP使NANP内化之后，NANP才可以用作递送控制的佐剂。这种激动作用使NANPs替代了雷西莫德（一种著名的TLR 7/8激动剂），因为该佐剂进入全身循环后会立即发生明显的免疫刺激，因此只能安全地用于局部应用。

The ability of NANPs to act as scaffolds for in vivo delivery of TNAs can be leveraged to alter the expression of genes that are responsible for altered immune responses against tumors, the so-called cancer immunity cycle. Moreover, NANPs may be designed to possess split functionalities whereby they carry no function when used independently but acquire functionality only after co-delivery into the same cell. This modality enables researchers to use NANPs to turn on and off biological responses when such controls are needed. The advantage of such dual-functionality NANPs is the elimination of off-target toxicity when an individual NANP is delivered into the off-target cell. One design of the dualfunctional NANPs includes so-called split functionality in which a first NANP aimed at performing one function (e.g., activation of the immune response) is administered as a single treatment and then a second NANP with alternative functionality (e.g., inhibition of the immune response) is administered to neutralize the effect of the first NANP. This modality enables additional control over the immune cells activated by immunotherapy (e.g., CTLs) and, therefore, is instrumental in immunotherapy to avoid adverse effects arising due to activation of the immune system, which is often overstimulated and may lead to autoimmunity. Another example of controlled immune activation is via NANPs releasing NF-kB decoy oligonucleotides. Because altered NF-kB function is often observed in various types of cancer,the inhibition of NF-kB by decoy-releasing NANPs has the potential to create a new therapeutic modality for cancer therapy. The same property would also benefit vaccines that contain TLR agonists inducing a robust tumor necrosis factor (TNF) response. Although TNF is needed to activate APCs, high levels of this cytokine may lead to tissue necrosis at the site of vaccine injection. Having NANPs release NF-kB decoy oligonucleotide and activating type I IFN in such vaccines would help to control undesirable cytokine levels and provide optimal levels of desirable cytokines. Proof of efficacy of NF-kB-controlling NANPs has been recently reported and warrants further research in this area.

可以利用NANP作为体内递送TNA的支架的能力来改变负责改变针对肿瘤的免疫应答的基因的表达，即所谓的癌症免疫周期。此外，可以将NANP设计为具有分开的功能，由此当独立使用它们时，它们不具有任何功能，而仅在共同递送至同一细胞后才具有功能。这种方式使研究人员可以在需要此类控制时使用NANP来开启和关闭生物反应。这种双功能NANP的优点是当将单个NANP递送到脱靶细胞中时消除脱靶毒性。双功能NANP的一种设计包括所谓的拆分功能，其中将旨在执行一种功能（例如，免疫应答的激活）的第一NANP作为单一治疗施用，然后将具有替代功能的第二NANP（例如，抑制NPN）给药。免疫应答）以中和第一个NANP的作用。该方式使得能够对由免疫疗法（例如，CTL）激活的免疫细胞进行额外的控制，因此在免疫疗法中起着重要作用，避免了由于免疫系统激活而引起的不良反应，这种不良反应常常被过度刺激并可能导致自身免疫。受控免疫激活的另一个例子是通过NANPs释放NF-kB诱饵寡核苷酸。由于经常在各种类型的癌症中观察到NF-kB功能的改变，因此通过诱饵释放的NANP对NF-kB的抑制作用有可能为癌症治疗创造新的治疗方式。相同的特性也将有益于包含TLR激动剂的疫苗，后者可诱导强大的肿瘤坏死因子（TNF）反应。尽管需要TNF来激活APC，但是高水平的这种细胞因子可能会在疫苗注射部位导致组织坏死。使NANP释放此类疫苗中的NF-kB诱饵寡核苷酸并激活I型IFN将有助于控制不良细胞因子水平并提供理想水平的理想细胞因子。最近已经报道了控制NF-kB的NANP的功效证明，值得在这一领域进行进一步的研究。