Sterol-based ionizable lipids and lipid nanoparticles comprising same

Abstract

Compounds, compositions, and methods for delivering therapeutic, diagnostic, or prophylactic agents (e.g., nucleic acids) are described.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Application No. 63 / 396,125, filed on August 8, 2022, the contents of which are hereby incorporated in their entirety. Background Art

[0003] Effective targeted delivery of active agents such as diagnostic agents, small molecule drugs, proteins and nucleic acids represents an ongoing medical challenge. In particular, delivery of nucleic acids to cells is difficult due to the relative instability and low cell permeability of such species.

[0004] Lipid-containing nanoparticle compositions, liposomes, and lipid complexes have been demonstrated to be effective as delivery vehicles for active agents into cells and / or intracellular compartments. Although a variety of such lipid-containing nanoparticle compositions have been demonstrated, improvements in safety, efficacy, and specificity are still lacking.

[0005] Therefore, there is a need to develop methods and compositions that facilitate the delivery of therapeutic, diagnostic and / or prophylactic agents, such as nucleic acids, to cells.

[0006] The compounds, compositions, and methods disclosed herein address these needs and others. Summary of the invention

[0007] The present disclosure provides compounds and uses thereof. Also provided are compositions comprising the compounds of the present disclosure and medicaments. The present disclosure also provides methods of using the compositions to deliver active agents to a subject.

[0008] In one aspect, described herein are compounds of formula Y:

[0009]

[0010] or a pharmaceutically acceptable salt thereof, wherein:

[0011] R 1 is H or -CH3;

[0012] represents a carbon-carbon bond or a carbon-carbon double bond;

[0013] R 7 Is H or OR 16 ;

[0014] R 8a and R 8b Each independently is H or OR 16 , or R 8a and R 8b Together with the atoms to which they are attached, they form a cycloalkyl, aryl, heterocycloalkyl or heteroaryl group;

[0015] R 9 is H, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl;

[0016] W is CR 4a or CR 4a R 4b , where if there is a double bond between W and the adjacent carbon, then W is CR 4a ; and if there is a single bond between W and the adjacent carbon, then W is CR 4a R 4b ; R 4a and R 4b Each of is independently H, halogen or C1-C6 alkyl;

[0017] A is

[0018]

[0019] X is O or S;

[0020] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)R a C(=O)-、-NR a C(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0021] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0022] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0023] R a yes wherein n is an integer from 0 to 12;

[0024] X is 0, 1, or 2;

[0025] M is CH,

[0026] G1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0027] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0028] R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0029] R 3 yes

[0030] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0031] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl;

[0032] p and q are independently 0, 1, 2, 3 or 4;

[0033] R 35 yes

[0034] L 4 Does not exist, yes

[0035] R 10 Not present, or C1-C6 alkyl;

[0036] L 5 does not exist, or or

[0037] m is an integer 1, 2 or 3;

[0038] L 6 Does not exist, yes

[0039] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl or C2-C9 heteroaryl;

[0040] L 7 is C1-C6 alkylene;

[0041] R 13a , R 13b and R 13c Each is independently C1-C6 alkyl or C6-C 10 Aryl;

[0042] R 14 , R 16 , R 19 , R 21 , R 22 , R 24 , R 28 , R 29 and R 31 Each independently is H or C1-C6 alkyl;

[0043] R 15 Yes-OR 16 、-NR 17 R 17 or

[0044] R 17 Each independently is H, -OR 16 、C6-C 10 Aryl or C1-C6 alkyl;

[0045] R 18 Each is independently halogen or C1-C6 alkyl;

[0046] o1 is an integer from 0 to 8;

[0047] p1 and p2 are each independently an integer from 0 to 2;

[0048] Z is CH2, O, S or NR 16 ;

[0049] s and t are each independently 0 or 1;

[0050] R 23 is halo, hydroxy, C1-C6 alkyl or C1-C6 heteroalkyl;

[0051] R 20 , R 25a and R 25b , R 30a , R 30b , R 30c , R 32a , R 32b and R 34 Each is independently a C1-C6 alkyl group;

[0052] R 26a and R 26b are each independently H, C1-C6 alkyl, or R 26a and R 26b Combined with the atoms to which they are connected, Where R 26c and R 26d Each is independently H or substituted C1-C6 alkyl;

[0053] R 27a and R 26b Each is independently H, hydroxyl or C1-C6 alkyl;

[0054] u is 1, 2, or 3;

[0055] R 33a and R 33b Each is independently C1-C6 alkyl, C1-C6 heteroalkyl, halogen or hydroxy;

[0056] Q is O, S or NR 16 ;and

[0057] or a pharmaceutically acceptable salt thereof.

[0058] In some embodiments, provided herein are methods for delivering agents (e.g., polynucleotides). In some embodiments, provided herein are methods for diagnosing, treating, or preventing diseases. In some embodiments, provided herein are methods for inducing an immune response to an infectious agent. In some embodiments, provided herein are methods for treating cancer. In some embodiments, provided herein are methods for regulating the immune system to treat cancer and other immune disorders. BRIEF DESCRIPTION OF THE DRAWINGS

[0059] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate several aspects described below.

[0060] Figure 1 The synthetic route of compound target 1 is shown.

[0061] FIG. 2A to FIG. 2C Compound Target 1 1H NMR spectrum (2A), mass spectrum (2B) and HPLC spectrum (2C).

[0062] FIG. 3A to FIG. 3B Characterization of SARS-CoV-2 mRNA lipid nanoparticles (LNPs) is shown. SM-102 / DVS was formulated using commercially available SM-102 lipids, and ARV-T1 was formulated using target 1 lipids (DVS: Δ variant spike protein). Particle diameter and polydispersity index (PDI) of LNPs (3A). Surface charge (Z potential) and mRNA encapsulation efficiency of SM-102 and ARV-T1 LNPs (3B).

[0063] Figure 4 Shown is in vitro expression of the spike glycoprotein over time in 293T cells after transfection of cells with 1.0 μg DVS mRNA at 1.0 μg / mL mRNA.

[0064] FIG. 5A to FIG. 5B The in vitro transfection efficiency of 1 μg GFP mRNA (1 μg / mL) delivered into BHK cells using LNPs after 24 hours is shown. (5A) Representative fluorescence images of BHK cells after transfection. (5B) Transfection efficacy analysis of GFP expression using flow cytometry. MFI represents mean fluorescence intensity.

[0065] FIG. 6A to FIG. 6B The in vivo transfection efficiency of mRNA expressing luciferase is shown. LNPs were formulated with the indicated ionizable lipids and 1 μg of the formulated mRNA expressing luciferase was injected intramuscularly. After administration, (6A) representative imaging and (6B) quantification of luciferase expression were determined by whole body bioluminescence imaging using the IVIS spectrum in vivo imaging system at 6 hours, 24 hours, 48 ​​hours and 72 hours, respectively.

[0066] 7A to 7E Effectively elicited immunity in vivo is shown. (7A) Experimental setup. mRNA encoding the full-length spike glycoprotein of SARS-CoV-2 (Δ variant) was formulated using target 1 (ARV-T1) lipids. Commercially available lipid SM-102 was used as a comparison. 1 μg of vaccine was injected intramuscularly as planned. (7B to 7C) Spike-specific total IgG was assessed on day 14 (7B) and day 35 (7C) after the first immunization. (7D) Neutralizing antibodies in serum were assessed by pseudotyped viruses. (7E) Antigen-specific T cell responses were assessed by Elispot. Data are expressed as mean ± SD. Statistical comparisons were analyzed using one-way ANOVA and Tukey's multiple comparison test. *p<0.05, **p<0.01, ***p<0.001.

[0067] FIG. 8A to FIG. 8BSingle cell analysis of splenocytes from Ai14 / Cre tdTomato reporter mice is shown, indicating that ARV-T1-based LNPs can deliver mRNA to multiple cells in vivo. A114 mice were intravenously administered 10 μg of Cre mRNA formulated with ARV-T1 lipids. 48 hours after injection, tdTomato was confirmed by flow cytometry (8A). + CD45 + The percentage of splenocytes and (8B) CD45 + tdTomato + Distribution of splenocytes.

[0068] 9A to 9E Figure 9A shows the in vitro delivery of GFP plasmid DNA into 293T cells using ARV-T1 LNP. (9A) is a diagram showing the size and PDI characterization of GFP pDNA lipid nanoparticles (LNPs). (9B to 9E) are images of the in vivo transfection efficiency of pDNA expressing GFP (500ng DNA and 48 hours incubation). (9B) 293T only, (9C) Lipo-pDNA, (9D) LNP-pDNA, (9E) ARV-LNP-pDNA. LNP-pDNA is prepared using traditional lipid ratios, and ARV-LNP-pDNA is prepared using target 1 lipid supplements with DOTAP.

[0069] FIG. 10A to FIG. 10F It is shown that small RNA (siRNA) and large RNA (saRNA) are delivered to 293T cells in vitro using ARV-T1 LNPs, respectively. (10A) is a diagram showing the size and PDI characterization of GFP siRNA lipid nanoparticles (LNPs). T1-siRNA is prepared using the same formulation as mRNA. (10B to 10D) are images of in vivo delivery of GFP siRNA (siGFP) using ARV-T1 LNPs to inhibit GFP expression (10B) control, (10C) lipid-siGFP and (10D) ARV-T1-siGFP. (10E to 10F) are images of in vitro delivery of GFP self-amplified RNA (saRNA) using ARV-T1 LNPs, C12-200 formulations as controls (10E) C12-200-saRNA and (10F) ARV-T1-saRNA.

[0070] Fig.11 is a graph of size and PDI characterization of a small molecule drug doxorubicin (DOX) formulation using ARV-T1 LNPs.

[0071] Fig.12 The synthetic route of compound target 11 is shown.

[0072] FIG. 13A to FIG. 13B The compound target 11 is shown 1 H NMR spectrum (13A), mass spectrum (13B).

[0073] FIG. 14A to FIG. 14E The in vitro transfection efficiency of 1 μg GFP mRNA (1 μg / mL) delivered to 293T cells using ARV-T11 LNPs and mRNA-LNPs of ARV-T11 lipids is shown. (14A) is a graph of the size and PDI characterization of GFP mRNA ARV-T11 LNPs. (14B to 14D) are representative fluorescent images of 293T cells after transfection of (14B) control, (14C) SM-102, and (14D) ARV-T11. (14E) is a graph of transfection efficacy analysis of GFP expression using flow cytometry.

[0074] Fig.15 The synthetic route of compound target 12 is shown.

[0075] FIG. 16A to FIG. 16C Compound target 12 is shown 1 H NMR spectrum (16A), mass spectrum (16B) and HPLC spectrum (16C).

[0076] FIG. 17A to FIG. 17E The in vitro transfection efficiency of 1 μg GFP mRNA (1 μg / mL) delivered to 293T cells using ARV-T12 LNPs and mRNA-LNP formulations of ARV-T12 lipids is shown. (17A) is a graph of the size and PDI characterization of GFP mRNA ARV-T12 LNPs. (17B to 17D) are representative fluorescent images of 293T cells after transfection of (17B) control, (17C) SM-102, and (17D) ARV-T12. (17E) is a graph of transfection efficacy analysis of GFP expression using flow cytometry.

[0077] Fig.18 The synthetic route of compound target 13 is shown.

[0078] FIG. 19A to FIG. 19C Compound 13 is shown 1 H NMR spectrum (19A), mass spectrum (19B) and HPLC spectrum (19C).

[0079] FIG. 20A to FIG. 20EThe in vitro transfection efficiency of 1 μg GFP mRNA (1 μg / mL) delivered to 293T cells using ARV-T13 LNPs and mRNA-LNP formulations of ARV-T13 lipids is shown. (20A) is a graph of the size and PDI characterization of GFP mRNA ARV-T13 LNPs. (20B to 20D) are representative fluorescent images of 293T cells after transfection of (20B) control, (20C) SM-102, and (20D) ARV-T12. (20E) is a graph of transfection efficacy analysis of GFP expression using flow cytometry. DETAILED DESCRIPTION

[0080] The present disclosure provides novel compounds, nanomaterials and uses thereof. Compositions comprising the compounds of the present invention and medicaments are also provided. The present disclosure also provides methods of using the compositions to deliver medicaments to subjects. These nanomaterials are used in applications such as gene therapy and drug delivery.

[0081] Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings and examples. However, the present invention may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

[0082] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The following definitions are provided for a full understanding of the terms used in this specification.

[0083] definition

[0084] General Definition

[0085] As used in this specification and the appended claims, the terms "comprising" (and forms, derivatives, or variations thereof, such as "comprising" and "comprises") and "include" (and forms, derivatives, or variations thereof, such as "including" and "includes") are inclusive (i.e., open ended) and do not exclude additional elements or steps. For example, when used in this specification, the terms "comprise and / or comprising" specify the presence of stated features, integers, steps, operations, elements, and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and / or groups thereof. Except where otherwise noted, all numbers used in the specification and claims expressing quantities of ingredients, reaction conditions, geometric shapes, dimensions, and the like should at least be understood, and are not intended to limit the application of the doctrine of equivalents to the scope of the claims, to be interpreted in light of the number of significant figures and ordinary rounding techniques.

[0086] Therefore, these terms are intended to cover not only the listed elements or steps, but also other elements or steps that are not explicitly listed. In addition, as used herein, the use of the terms "a, an" and "the" when used in conjunction with an element may mean "one", but it is also consistent with the meaning of "one or more", "at least one" and "one or more than one". Therefore, without further limitations, an element preceded by "a or an" does not exclude the presence of additional identical elements.

[0087] In this article, ranges can be expressed as from "about" a specific value and / or to "about" another specific value. "About" means within 5% of the value, such as within 4%, 3%, 2% or 1% of the value. When expressing such a range, on the other hand, it is included from a specific value and / or to another specific value. Similarly, when a value is expressed as an approximation by using the antecedent "about", it should be understood that a specific value forms another aspect. It should be further understood that the endpoints of each range are significant relative to the other endpoint, and independently of the other endpoint. It should also be understood that many values ​​are disclosed herein, and each value is also disclosed herein as "about" for the specific value in addition to the value itself. For example, if the value "10" is disclosed, "about 10" is also disclosed. Ranges can be interpreted as including the beginning and end of a range. For example, a range of 10% to 20% (i.e., a range of 10% to 20%) can include 10% and also include 20%, and include the percentage between 10% and 20%, unless otherwise explicitly stated herein.

[0088] As used herein, the terms "may," "optionally," and "may optionally" are used interchangeably and are meant to include instances where the condition occurs as well as instances where the condition does not occur. Thus, for example, the statement that a formulation "may include an excipient" is meant to include instances where the formulation includes an excipient as well as instances where the formulation does not include an excipient.

[0089] It should be understood that when combinations, subsets, groups, etc. of elements are disclosed (e.g., combinations of components in a composition or combinations of steps in a method), each element is specifically contemplated and described herein, although specific reference to each of the various individual and collective combinations and arrangements of such elements may not be explicitly disclosed.

[0090] "Administering" to a subject includes any approach of introducing or delivering a medicament to a subject. Administration can be carried out by any suitable approach, including oral, topical, percutaneous, through the dermis, intraarticular, intraarteriole, intradermal, intraventricular, intralesional, intranasal, rectal, vaginal, by suction, by implanted reservoir, parenteral (e.g., subcutaneous, intravenous, intramuscular, intraarticular, intrasynovial, intrasternal, intrathecal, intraperitoneal, intrahepatic, intralesional and intracranial injection or infusion technique) etc. As used herein, "administering simultaneously (concurrent administration)", "combination administration", "administering simultaneously (simultaneous administration)" or "administering simultaneously" means that the compound is administered at the same time point or is administered one after another substantially. In the latter case, the administration time of two compounds is close enough that the observed result is indistinguishable from the result obtained when the compound is administered at the same time point. "Systemic administration" refers to the introduction or delivery of an agent to a subject by a route that introduces or delivers the agent to a wide area of ​​the subject's body (e.g., greater than 50% of the body), such as by entering the circulatory or lymphatic system. In contrast, "local administration" refers to the introduction or delivery of an agent to a subject by a route that introduces or delivers the agent to an area or an area immediately adjacent to the point of administration and does not introduce the agent systemically in therapeutically significant amounts. For example, a locally administered agent may be easily detected in the local vicinity of the point of administration, but may not be detected or may be detected in negligible amounts in distal portions of the subject's body. Administration encompasses self-administration and administration by another person.

[0091] As used herein, the term "controlled release" or "controlled release drug delivery" or "extended release" refers to the release or administration of a drug from a given dosage form in a controlled manner to achieve a desired pharmacokinetic profile in vivo. One aspect of "controlled" drug delivery is the ability to manipulate the formulation and / or dosage form in order to establish the desired drug release kinetics.

[0092] As used herein, the terms "beneficial agent" and "active agent" are used interchangeably herein to refer to a chemical compound or composition having a beneficial biological effect. Beneficial biological effects include therapeutic effects, i.e., treating a disorder or other undesirable physiological condition, and preventive effects, i.e., preventing a disorder or other undesirable physiological condition. The term also encompasses pharmaceutically acceptable pharmacologically active derivatives of the beneficial agents specifically mentioned herein, including but not limited to salts, esters, amides, prodrugs, active metabolites, isomers, fragments, analogs, and the like. When the term "beneficial agent" or "active agent" is used, or when a specific agent is specifically identified, it should be understood that the term includes the agent itself as well as pharmaceutically acceptable pharmacologically active salts, esters, amides, prodrugs, conjugates, active metabolites, isomers, fragments, analogs, and the like.

[0093] "Therapeutic agent" refers to any composition that has a beneficial biological effect. Beneficial biological effects include therapeutic effects, such as treating a disease or other undesirable physiological conditions, and preventive effects, such as preventing a disease or other undesirable physiological conditions. The term also encompasses pharmaceutically acceptable pharmacologically active derivatives of the beneficial agents specifically mentioned herein, including but not limited to salts, esters, amides, proagents, active metabolites, isomers, fragments, analogs, and the like. When the term "therapeutic agent" is used, or when a particular agent is specifically identified, it should be understood that the term includes the agent itself as well as pharmaceutically acceptable pharmacologically active salts, esters, amides, proagents, conjugates, active metabolites, isomers, fragments, analogs, and the like.

[0094] "Reduction" can refer to any change that causes a smaller amount of symptoms, diseases, compositions, symptoms or activities. When the genetic output of the gene product utilizing material is lower than the output of the gene product not utilizing the material, the material should also be understood to reduce the genetic output of the gene. Also for example, reduction can be the change that symptoms make symptoms less than previously observed of the symptoms of the disease. Reduction can be any individual, median or average reduction of the symptoms, activities, compositions with statistically significant amounts. Therefore, the reduction can be 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 100% reduction, as long as the reduction is statistically significant.

[0095] "Inhibit," "inhibiting," and "inhibition" mean a reduction in an activity, response, condition, disease, or other biological parameter. This may include, but is not limited to, complete ablation of an activity, response, condition, or disease. This may also include, for example, a 10% reduction in an activity, response, condition, or disease compared to a native or control level. Thus, the reduction may be 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or any reduction therebetween, compared to a native or control level.

[0096] "Inactivate," "inactivating," and "inactivation" refer to the reduction or elimination of an activity, response, condition, disease, or other biological parameter as a result of chemistry (covalent bond formation) between a ligand and its biological target.

[0097] "Reduce" or other forms of the word, such as "reducing" and "reduction", refers to a decrease in an event or characteristic (e.g., tumor growth). It should be understood that this is usually related to a certain standard or expected value, in other words it is relative, but it does not always require reference to a standard or relative value. For example, "reduce tumor growth" means reducing the rate of tumor growth relative to a standard or control.

[0098] As used herein, the term "treating" or "treatment" of a subject includes administering a drug to a subject for the purpose of preventing, curing, healing, alleviating, mitigating, altering, remedying, ameliorating, improving, stabilizing, or affecting a disease or condition, or a symptom of a disease or condition. The terms "treating" and "treatment" may also refer to reducing the severity and / or frequency of symptoms, eliminating symptoms and / or underlying causes, preventing the occurrence of symptoms and / or their underlying causes, and ameliorating or remedying damage.

[0099] "Prevent" or other forms of the word (such as "preventing" or "prevention") means to prevent a specific event or characteristic, to stabilize or delay the development or progress of a specific event or characteristic, or to minimize the possibility of a specific event or characteristic occurring. Prevention does not need to be compared with a control because it is usually more absolute than, for example, reduction. As used herein, something can be reduced but not prevented, but something that is reduced can also be prevented. Similarly, something can be prevented but not reduced, but something that is prevented can also be reduced. It should be understood that, unless otherwise expressly stated, in the case of reduction or prevention used, the use of other words is also clearly disclosed. For example, the term "prevention" or "inhibition" can refer to a treatment that prevents or slows the onset of a disease or condition in advance or reduces the severity of a disease or condition. Therefore, if the treatment can treat a disease in a subject with symptoms of the disease, it can also prevent or inhibit the disease in a subject who has not yet suffered some or all of the symptoms. As used herein, the term "preventing" a condition or adverse physiological event in a subject specifically refers to preventing the occurrence of symptoms and / or their underlying causes, wherein the subject may or may not exhibit a heightened susceptibility to the condition or event.

[0100] The term "effective amount" of a therapeutic agent refers to an amount of a beneficial agent that is non-toxic but sufficient to provide the desired effect. The amount of an "effective" beneficial agent will vary from subject to subject, depending on the subject's age and general condition, the specific beneficial agent or agents, etc. Therefore, it is not always possible to specify an exact "effective amount". However, one of ordinary skill in the art can determine the appropriate "effective amount" for any subject situation using routine experimentation. Moreover, as used herein and unless otherwise expressly stated, an "effective amount" of a beneficial agent may also refer to an amount covering both a therapeutically effective amount and a prophylactically effective amount.

[0101] The "effective amount" of a drug necessary to achieve a therapeutic effect may vary according to factors such as the age, sex, and weight of the subject. The dosage regimen may be adjusted to provide the optimal therapeutic response. For example, several divided doses may be administered daily, or the dose may be proportionally reduced as indicated by the exigencies of the therapeutic situation.

[0102] As used herein, the "therapeutically effective amount" of a therapeutic agent refers to the amount that effectively achieves the desired therapeutic outcome, and the "preventive effective amount" of a therapeutic agent refers to the amount that effectively prevents adverse physiological conditions. The therapeutically effective amount and the preventive effective amount of a given therapeutic agent will generally vary according to factors such as the type and severity of the disorder or disease being treated and the age, sex, and weight of the subject. The term "therapeutically effective amount" can also refer to the amount of the therapeutic agent that effectively promotes the desired therapeutic effect or the delivery rate of the therapeutic agent (e.g., the amount that varies over time). The precise desired therapeutic effect will vary according to the condition to be treated, the tolerance of the subject, the drug to be administered and / or the drug formulation (e.g., the efficacy of the therapeutic agent (drug), the concentration of the drug in the formulation, etc.), and various other factors understood by those of ordinary skill in the art.

[0103] As used herein, the term "pharmaceutically acceptable" component may refer to a component that is not biologically or otherwise undesirable, i.e., the component may be incorporated into the pharmaceutical formulations of the present invention and administered to a subject as described herein without causing any significant undesirable biological effects or interacting in a harmful manner with any other component of the formulation containing it. When the term "pharmaceutically acceptable" is used to refer to an excipient, the term generally means that the component has met the criteria for required toxicology and manufacturing testing, or that the component is included in the Inactive Ingredient Guide formulated by the U.S. Food and Drug Administration.

[0104] "Pharmaceutically acceptable carrier" (sometimes referred to as "carrier") means a carrier or excipient that can be used to prepare a generally safe and non-toxic pharmaceutical or therapeutic composition, and includes carriers that are acceptable for veterinary and / or human pharmaceutical or therapeutic use. The term "carrier" or "pharmaceutically acceptable carrier" may include, but is not limited to, phosphate buffered saline solution, water, emulsions (such as oil / water or water / oil emulsions), and / or various types of wetting agents. As used herein, the term "carrier" encompasses, but is not limited to, any excipient, diluent, filler, salt, buffer, stabilizer, solubilizer, lipid, stabilizer, or other material known in the art for use in pharmaceutical formulations and as further described herein.

[0105] As used herein, "pharmaceutically acceptable salts" are derivatives of disclosed compounds in which the parent compound is modified by making its inorganic and organic, non-toxic acid or base addition salts. The salts of the compounds of the present invention can be synthesized from the parent compound containing an alkaline or acidic part by conventional chemical methods. Typically, such salts can be prepared by reacting the free acid forms of these compounds with a stoichiometric amount of a suitable base (such as Na, Ca, Mg or K hydroxides, carbonates, bicarbonates, etc.), or by reacting the free base forms of these compounds with a stoichiometric amount of a suitable acid. Such reactions are typically carried out in water or an organic solvent, or in a mixture of the two. In general, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are typical where feasible. The salts of the compounds of the present invention also include solvates of compounds and compound salts.

[0106] Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali metal or organic salts of acidic residues such as carboxylic acids; etc. Pharmaceutically acceptable salts include conventional non-toxic salts and quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. For example, conventional non-toxic acid salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, sulfonic acid, phosphoric acid, nitric acid, etc.; and salts prepared from organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, pamoic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, sulfanilic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isethionic acid, HOOC-(CH2)n-COOH, wherein n is 0 to 4, etc., or salts prepared using different acids that produce the same counterion. Lists of other suitable salts can be found in, for example, Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., p.1418 (1985).

[0107] In addition, as used herein, the term "pharmacologically active" (or simply "active") as in "pharmacologically active" derivative or analog may refer to a derivative or analog (e.g., salts, esters, amides, conjugates, metabolites, isomers, fragments, etc.) that has the same type of pharmacological activity as the parent compound and to approximately the same extent.

[0108] A "control" is an alternative subject or sample used for comparison purposes in an experiment. A control can be "positive" or "negative."

[0109] As used herein, "subject" refers to an individual. Therefore, "subject" can include domestic animals (e.g., cats, dogs, etc.), livestock (e.g., cattle, horses, pigs, sheep, goats, etc.), experimental animals (e.g., mice, rabbits, rats, guinea pigs, goats, sheep, pigs, dogs, cats, etc.) and birds (e.g., chickens, turkeys, songbirds, etc.). "Subject" can also include mammals, such as primates or people. Therefore, the subject can be a human or veterinary patient. The term "patient" refers to a subject under the treatment of a clinician (e.g., a physician). The administration of the therapeutic agent can be carried out at a dosage and time period effective for treating the subject. In certain embodiments, the subject is a human.

[0110] The term "nucleic acid" as used herein refers to a polymer composed of nucleotides, such as deoxyribonucleotides or ribonucleotides.

[0111] As used herein, the terms "ribonucleic acid" and "RNA" refer to a polymer composed of ribonucleotides.

[0112] As used herein, the terms "deoxyribonucleic acid" and "DNA" refer to a polymer composed of deoxyribonucleotides.

[0113] The term "oligonucleotide" refers to a single-stranded or double-stranded nucleotide polymer of about 2 to about 100 nucleotides in length. Suitable oligonucleotides can be prepared by the phosphoramidite method described by Beaucage and Carruthers, Tetrahedron Lett., 22: 1859-1862 (1981) or by the triester method according to Matteucci, et al., J. Am. Chem. Soc., 103: 3185 (1981), both of which are incorporated herein by reference, or by using a commercial automated oligonucleotide synthesizer or VLSIPS TMThe oligonucleotides are referred to as "double-stranded", and those skilled in the art will appreciate that a pair of oligonucleotides exist in a hydrogen-bonded spiral array that is typically associated with, for example, DNA. In addition to the 100% complementary form of double-stranded oligonucleotides, the term "double-stranded" as used herein also refers to forms that include such structural features such as protrusions and loops, which are more fully described in such biochemical texts such as Stryer, Biochemistry, 3rd edition, (1988), which is incorporated herein by reference for all purposes.

[0114] The term "polynucleotide" refers to a single-stranded or double-stranded polymer composed of nucleotide monomers. In some embodiments, a polynucleotide is composed of nucleotide monomers that are generally greater than 100 nucleotides in length and up to about 8,000 or more nucleotides in length.

[0115] The term "polypeptide" refers to a compound composed of a single chain of D- or L-amino acids or a mixture of D- and L-amino acids linked by peptide bonds.

[0116] The term "complementary" refers to the topological compatibility or matching together of the interaction surfaces of the probe molecule and its target. Thus, a target and its probe can be described as complementary, and further the contact surface properties complement each other.

[0117] The term "hybridization" refers to the process of establishing non-covalent, sequence-specific interactions between two or more complementary strands of nucleic acid to form a single hybrid, which in the case of two strands is called a duplex.

[0118] The term "annealing" refers to the process by which a single-stranded nucleic acid sequence pairs with a complementary sequence through hydrogen bonds to form a double-stranded nucleic acid sequence, including the reformation (renaturation) of complementary chains separated by heat (thermal denaturation).

[0119] The term "melting" refers to the denaturation of double-stranded nucleic acid sequences due to high temperatures, resulting in the separation of the double strand into two single strands by breaking the hydrogen bonds between the strands.

[0120] The term "target" refers to a molecule that has an affinity for a given probe. Targets can be naturally occurring or artificial molecules. Furthermore, they can be used in their unaltered state or as aggregates with other species.

[0121] The term "promoter" or "regulatory element" refers to a region or sequence determinant located upstream or downstream of the transcription start point and involved in the recognition and binding of RNA polymerase and other proteins to initiate transcription. The promoter need not be of bacterial origin, for example, promoters derived from viruses or other organisms can be used in the compositions, systems or methods described herein. The term "regulatory element" is intended to include promoters, enhancers, internal ribosome entry sites (IRES) and other expression control elements (e.g., transcription termination signals, such as polyadenylation signals and poly-U sequences). Such regulatory elements are described in, for example, Goeddel, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990). Regulatory elements include those sequences that direct the constitutive expression of nucleotide sequences in many types of host cells and those sequences that direct the expression of nucleotide sequences only in certain host cells (e.g., tissue-specific regulatory sequences). Tissue-specific promoters can be mainly directed to expression in desired tissues of interest, such as muscle, neurons, bones, skin, blood, specific organs (e.g., liver, pancreas) or specific cell types (e.g., lymphocytes). Regulatory elements can also direct expression in a time-dependent manner, such as in a cell cycle-dependent or developmental stage-dependent manner, which may or may not be tissue- or cell-type-specific. In some embodiments, the mediator includes one or more pol III promoters (e.g., 1, 2, 3, 4, 5 or more pol I promoters), one or more pol II promoters (e.g., 1, 2, 3, 4, 5 or more pol II promoters), one or more pol I promoters (e.g., 1, 2, 3, 4, 5 or more pol I promoters), or a combination thereof. Examples of pol III promoters include, but are not limited to, U6, H1, and T7 promoters. Examples of pol II promoters include, but are not limited to, the retroviral Rous sarcoma virus (RSV) LTR promoter (optionally with an RSV enhancer), the cytomegalovirus (CMV) promoter (optionally with a CMV enhancer) [see, e.g., Boshart et al., Cell, 41:521-530 (1985)], the SV40 promoter, the dihydrofolate reductase promoter, the β-actin promoter, the phosphoglycerol kinase (PGK) promoter, the β-globin promoter, and the EF1α promoter.The term "regulatory element" also encompasses enhancer elements such as WPRE; CMV enhancer; R-U5' fragment in the LTR of HTLV-I (Mol. Cell. Biol., Vol. 8 (1), p. 466-472, 1988); SV40 enhancer; and the intron sequence between exons 2 and 3 of rabbit β-globin (Proc. Natl. Acad. Sci. USA., Vol. 78 (3), p. 1527-31, 1981). It is understood by those skilled in the art that the design of the expression medium may depend on factors such as the choice of the host cell to be transformed, the desired expression level, etc.

[0122] The term "recombination" refers to a nucleic acid (e.g., polynucleotide) manipulated by a person or a copy or complement of a nucleic acid (e.g., polynucleotide) manipulated by a person, or if a protein (i.e., "recombinant protein") is mentioned, it refers to a protein encoded by a recombinant nucleic acid (e.g., polynucleotide). In an embodiment, a recombinant expression cassette including a promoter operably connected to a second nucleic acid (e.g., polynucleotide) may include a promoter heterologous to a second nucleic acid (e.g., polynucleotide) as a result of human manipulation (e.g., by Sambrook et al., Molecular Cloning—A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, (1989) or Current Protocols in Molecular Biology Volumes 1-3, John Wiley & Sons, Inc. (1994-1998)). In another example, a recombinant expression cassette may include nucleic acids (e.g., polynucleotides) that are combined in a manner that nucleic acids (e.g., polynucleotides) are extremely unlikely to be found in nature. For example, a restriction site or plasmid mediator sequence manipulated by a person may be located on the flank of a promoter or separate it from a second nucleic acid (e.g., polynucleotide). Those skilled in the art will recognize that nucleic acids (eg, polynucleotides) can be manipulated in a variety of ways and are not limited to the above examples.

[0123] The term "expression cassette" refers to a nucleic acid construct that, when introduced into a host cell, results in transcription and / or translation of RNA or a polypeptide, respectively. In an embodiment, an expression cassette comprising a promoter operably connected to a second nucleic acid (e.g., a polynucleotide) may include a promoter heterologous to a second nucleic acid (e.g., a polynucleotide) as a result of human manipulation (e.g., by methods described in Sambrook et al., Molecular Cloning—A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, (1989) or Current Protocols in Molecular Biology Volumes 1-3, John Wiley & Sons, Inc. (1994-1998)). In some embodiments, an expression cassette comprising a terminator (or termination sequence) operably connected to a second nucleic acid (e.g., a polynucleotide) may include a terminator heterologous to a second nucleic acid (e.g., a polynucleotide) as a result of human manipulation. In some embodiments, an expression cassette comprises a promoter operably connected to a second nucleic acid (e.g., a polynucleotide) as a structure manipulated by a person and a terminator operably connected to a second nucleic acid (e.g., a polynucleotide). In some embodiments, the expression cassette includes an endogenous promoter. In some embodiments, the expression cassette includes an endogenous terminator. In some embodiments, the expression cassette includes a synthetic (or non-natural) promoter. In some embodiments, the expression cassette includes a synthetic (or non-natural) terminator.

[0124] In the context of two or more nucleic acid or polypeptide sequences, the terms "same" or "percent identity" refer to two or more sequences or subsequences that are identical or have a specified percentage of identical amino acid residues or nucleotides when compared and aligned for maximum correspondence over a comparison window or designated region (i.e., about 60% identity over a specified region, preferably 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110%, 111%, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 15 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher identity), using BLAST or BLAST2.0 sequence comparison algorithm, with the default parameters described below or by manual alignment and visual inspection (see, e.g., NCBI web site, etc.) measurement. Such sequences are then referred to as "substantially identical". This definition also relates to or can be applied to the complement of the test sequence. The definition also includes sequences with deletions and / or additions, and sequences with substitutions. As described below, preferred algorithms can account for gaps, etc. Preferably, identity is present over a region of at least about 10 amino acids or 20 nucleotides in length, or more preferably over a region of 10-50 amino acids or 20-50 nucleotides in length. As used herein, amino acid sequence identity percentage (%) is defined as the percentage of amino acids in the candidate sequence that are identical to the amino acids in the reference sequence after aligning the sequences and introducing gaps (if necessary) to achieve maximum sequence identity percentage. Alignment for determining sequence identity percentage can be achieved in various ways well known to those skilled in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, ALIGN-2 or Megalign (DNASTAR) software. Suitable parameters for measuring alignment, including any algorithm required for achieving maximum alignment on the full-length sequences being compared, can be determined by known methods.

[0125] For sequence comparison, usually a sequence is used as a reference sequence, and the test sequence is compared with it. When using a sequence comparison algorithm, the test sequence and the reference sequence are input into a computer, subsequence coordinates are specified (if necessary), and sequence algorithm program parameters are specified. Preferably, default program parameters can be used, or replaceable parameters can be specified. Then, the sequence comparison algorithm calculates the sequence identity percentage of the test sequence relative to the reference sequence based on the program parameters.

[0126] An example of an algorithm suitable for determining percent sequence identity and percent sequence similarity is BLAST and BLAST 2.0 algorithms, which are described in Altschul et al. (1977) Nuc. Acids Res. 25: 3389-3402, and Altschul et al. (1990) J. Mol. Biol. 215: 403-410, respectively. Software for performing BLAST analysis is publicly available at the National Center for Biotechnology Information (http: / / www.ncbi.nlm.nih.gov / ). The algorithm involves first identifying high-scoring sequence pairs (HSPs) by identifying short characters of length W in the query sequence that match or satisfy a positive threshold score T when aligned with a character of the same length in the database sequence. T is referred to as a neighboring character score threshold (Altschul et al., (1990) J. Mol. Biol. 215: 403-410). These initial neighboring character hits serve as seeds for initiating searches to find longer HSPs containing them. The word hits are extended in both directions along each sequence until the cumulative alignment score can be increased. For nucleotide sequences, the cumulative score is calculated using the parameters M (reward score for a pair of matching residues; always >0) and N (penalty score for mismatched residues; always <0). For amino acid sequences, a scoring matrix is ​​used to calculate the cumulative score. Extension of the word hits in each direction will stop when the following occurs: the cumulative alignment score drops by the amount X from its maximum achieved value; the cumulative score becomes zero or below due to the accumulation of one or more negative scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses by default: a word length (W) of 11, an expectation value (E) of 10, M=5, N=-4, and a comparison of both chains. For amino acid sequences, the BLASTP program uses as defaults a wordlength of 3, an expectation value (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff and Henikoff (1989) Proc. Natl. Acad. Sci. USA 89:10915), alignments (B) of 50, expectation value (E) of 10, M=5, N=-4, and a comparison of both strands.

[0127] The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin and Altschul (1993) Proc. Natl. Acad. Sci. USA 90:5873-5787). One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability that a match between two nucleotide sequences or amino acid sequences occurs by chance. For example, if the smallest sum probability in a comparison of a test nucleic acid to a reference nucleic acid is less than about 0.2, more preferably less than about 0.01, then the nucleic acid is considered similar to the reference sequence.

[0128] The phrase "codon optimized" refers to the coding region of a gene or nucleic acid molecule used to transform a variety of hosts, and refers to changing the codons in the gene or coding region of a polynucleotide molecule to reflect the typical codon usage of the selected organism without changing the polypeptide encoded by the DNA. This optimization includes replacing at least one, or more than one, or a large number of codons with one or more codons that are more frequently used in the genes of the selected organism.

[0129] When a nucleic acid is in a functional relationship with another nucleic acid sequence, it is "operably linked". For example, if the DNA of a presequence or secretory leader sequence is expressed as a preprotein that participates in the secretion of a polypeptide, the DNA is operably linked to the DNA of the polypeptide; if a promoter or enhancer affects the transcription of a sequence, the promoter or enhancer is operably linked to a coding sequence; or, if a ribosome binding site is positioned to promote translation, the ribosome binding site is operably linked to a coding sequence. Generally, "operably linked" means that the DNA sequences being linked are close to each other, and in the case of a secretory leader sequence, are continuous and in the reading phase. However, operably linked nucleic acids (such as enhancers and coding sequences) do not have to be continuous. Connection is accomplished by connecting at convenient restriction sites. If there is no such site, a synthetic oligonucleotide adapter or joint is used according to conventional practice. In an embodiment, when a promoter is able to affect (e.g., regulate relative to the absence of a promoter) the expression of a protein from a coding sequence (i.e., the coding sequence is under the transcriptional control of the promoter), the promoter is operably linked to the coding sequence.

[0130] The term "nucleobase" refers to the portion of a nucleotide that carries out Watson / Crick base pairing. The most common natural nucleobases, adenine (A), guanine (G), uracil (U), cytosine (C), and thymine (T) carry out hydrogen bonding, binding one nucleic acid strand to another in a sequence-specific manner.

[0131] A nucleic acid sequence is "heterologous" to a second nucleic acid sequence when it is derived from a foreign species, or, when from the same species, has been modified from its original form by human activity. For example, a heterologous promoter (or heterologous 5' untranslated region (5'UTR)) operably linked to a coding sequence refers to a coding sequence from a species different from that from which the promoter is derived, or, if from the same species, to a coding sequence that is different from a naturally occurring allelic variant (e.g., a 5'UTR or 3'UTR from a different gene is operably linked to a nucleic acid encoding a costimulatory molecule).

[0132] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous population of antibodies, i.e., the individual antibodies within the population are identical except for possible naturally occurring mutations that may be present in a smaller subset of the antibody molecules. The monoclonal antibodies herein particularly include "chimeric" antibodies, in which a portion of the heavy and / or light chain is identical or homologous to corresponding sequences in antibodies derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain is identical or homologous to corresponding sequences in antibodies derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, as long as they exhibit the desired antagonistic activity.

[0133] The disclosed monoclonal antibodies can be prepared using any procedure for producing monoclonal antibodies. For example, the disclosed monoclonal antibodies can be prepared using a hybridoma method such as the method described in Kohler and Milstein, Nature, 256:495 (1975). In the hybridoma method, mice or other appropriate host animals are typically immunized with an immunizing agent to stimulate lymphocytes that produce or are capable of producing antibodies that will specifically bind to the immunizing agent. Alternatively, lymphocytes can be immunized in vitro.

[0134] Monoclonal antibodies can also be prepared by recombinant DNA methods. Conventional methods (e.g., by using oligonucleotide probes that can bind specifically to the heavy and light chains of encoding mouse antibodies) can be easily separated and sequenced to encode the disclosed monoclonal antibody DNA. Phage display technology (e.g., as described in U.S. Pat. No. 5,804,440 to Burton et al. and U.S. Pat. No. 6,096,441 to Barbas et al.) can also be used to produce and screen libraries of antibodies or active antibody fragments.

[0135] In vitro methods are also applicable to the preparation of monovalent antibodies. Digestion of antibodies to produce fragments thereof, particularly Fab fragments, can be accomplished using conventional techniques known in the art. For example, papain can be used for digestion. Examples of papain digestion are described in WO 94 / 29348 and U.S. Patent No. 4,342,566, issued on December 22, 1994. Papain digestion of antibodies typically produces two identical antigen-binding fragments (called Fab fragments, each of which has an antigen-binding site) and a residual Fc fragment. Pepsin treatment obtains a fragment having two antigen-binding sites and still capable of cross-linking antigens.

[0136] As used herein, the term "antibody or antigen-binding fragment thereof" or "antibody or fragment thereof" encompasses chimeric antibodies and hybrid antibodies with dual or multiple antigen or epitope specificity, as well as fragments such as F(ab')2, Fab', Fab, Fv, sFv, scFv, etc., including hybrid fragments. Thus, antibody fragments that retain the ability to bind to the specific antigen of the antibody are provided. For example, antibody fragments that retain binding activity are included within the meaning of the term "antibody or antigen-binding fragment thereof". Such antibodies and fragments can be prepared by techniques known in the art, and can be screened for specificity and activity according to the examples and methods set forth in the general methods for producing antibodies and screening antibodies for specificity and activity (see Harlow and Lane. Antibodies, A Laboratory Manual. Cold Spring Harbor Publications, New York, (1988)).

[0137] The meaning of "antibody or antigen-binding fragment thereof" also includes conjugates of antibody fragments and antigen-binding proteins (single-chain antibodies). The meaning of "antibody or antigen-binding fragment thereof" also includes immunoglobulin single variable domains, such as, for example, nanobodies.

[0138] Fragments, whether attached to other sequences or not, may also include insertion, deletion, substitution or other selected modifications of specific regions or specific amino acid residues, provided that the activity of the antibody or antibody fragment is not significantly changed or impaired compared to unmodified antibodies or antibody fragments. These modifications may provide some additional characteristics, such as removal / addition of amino acids capable of forming disulfide bonds, increase of their biological lifespan, changes in their secretion characteristics, etc. In any case, antibodies or antibody fragments must have biologically active properties, such as specific binding to their cognate antigens. The function or active region of an antibody or antibody fragment may be identified by mutagenesis of a specific region of a protein, subsequent expression and testing of the expressed polypeptide. Such methods are apparent to those skilled in the art, and may include site-specific mutagenesis of nucleic acids encoding antibodies or antibody fragments. (Zoller, MJ Curr. Opin. Biotechnol. 3: 348-354, 1992).

[0139] As used herein, the term "antibody" or "antibodies" may also refer to human antibodies and / or humanized antibodies. Many non-human antibodies (e.g., antibodies derived from mice, rats, or rabbits) have natural antigenicity in humans and therefore may cause an undesirable immune response when administered to humans. Therefore, human or humanized antibodies are used in the method to reduce the chance that antibodies administered to humans cause an undesirable immune response.

[0140] Chemical Definition

[0141] As used herein, the term "substituted" is intended to include all permissible substituents of organic compounds. In a broad sense, permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, and aromatic and non-aromatic substituents of organic compounds. Exemplary substituents include, for example, those described below. For suitable organic compounds, permissible substituents may be one or more and may be the same or different. For the purposes of this disclosure, heteroatoms such as nitrogen may have hydrogen substituents and / or any permissible substituents of organic compounds described herein that satisfy the valence of heteroatoms. This disclosure is not intended to be limited in any way to permissible substituents of organic compounds. In addition, the term "substituted" or "substituted by ... " includes implicit conditions, i.e., such substitution is consistent with the permissible valence of the substituted atom and the substituent, and the substitution produces a stable compound, such as a compound that does not spontaneously undergo transformations such as by rearrangement, cyclization, elimination, etc.

[0142] "Z 1 ”, “Z 2 ”, “Z 3 ” and “Z 4" are used herein as general symbols to represent various specific substituents. These symbols can be any substituent, not limited to those disclosed herein, and when they are defined as certain substituents in one instance, they can be defined as some other substituents in another instance.

[0143] The term "aliphatic" as used herein refers to non-aromatic hydrocarbon groups and includes branched and unbranched alkyl, alkenyl or alkynyl groups.

[0144] As used herein, term "alkyl" is a branched or unbranched saturated hydrocarbon radical of 1 to 24 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, amyl, hexyl, heptyl, octyl, nonyl, decyl, dodecyl, tetradecyl, hexadecyl, eicosyl, tetracosyl etc. Alkyl can also be substituted or unsubstituted. Alkyl groups can be substituted by one or more groups, including but not limited to alkyl, halogenated alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxyl, ketone, nitro, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide or mercaptan, as described below.

[0145] Throughout the specification, "alkyl" is generally used to refer to unsubstituted alkyl groups and substituted alkyl groups; however, substituted alkyl groups are also specifically referred to herein by identifying specific substituents on the alkyl group. For example, the term "halogenated alkyl" specifically refers to an alkyl group substituted with one or more halides (e.g., fluorine, chlorine, bromine or iodine). The term "alkoxyalkyl" specifically refers to an alkyl substituted with one or more alkoxy groups, as described below. The term "alkylamino" specifically refers to an alkyl group substituted with one or more amino groups, as described below, etc. When "alkyl" is used in one case and a specific term such as "alkyl alcohol" is used in another case, it is not meant to imply that the term "alkyl" also does not refer to a specific term such as "alkyl alcohol".

[0146] This practice also applies to the other groups described herein. That is, although terms such as "cycloalkyl" refer to unsubstituted and substituted cycloalkyl moieties, the substituted moieties may be otherwise specifically indicated herein; for example, a specific substituted cycloalkyl may be referred to as, for example, "alkylcycloalkyl". Similarly, a substituted alkoxy may be specifically referred to as, for example, "haloalkoxy", a specific substituted alkenyl may be, for example, "alkenyl alcohol", etc. In addition, the practice of using a general term such as "cycloalkyl" and a specific term such as "alkylcycloalkyl" is not meant to imply that the general term also does not include the specific term.

[0147] As used herein, the term "alkoxy" is an alkyl group bonded through a single terminal ether bond; that is, an "alkoxy" group may be defined as -OZ 1 , where Z 1is alkyl as defined above.

[0148] The term "alkenyl" as used herein is a hydrocarbon group of 2 to 24 carbon atoms, the structural formula of which contains at least one carbon-carbon double bond. Asymmetric structures, such as (Z 1 Z 2 )C=C(Z 3 Z 4 ), both E and Z isomers are intended to be included. This may be assumed in the structural formulae herein where an asymmetric olefin is present, or it may be explicitly indicated by the bond symbol C=C. The alkenyl group may be substituted with one or more groups including, but not limited to, alkyl, halogenated alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxy, ketone, nitro, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide, or thiol as described herein, as described below.

[0149] As used herein, the term "alkynyl" is a hydrocarbon radical of 2 to 24 carbon atoms, the structural formula of which contains at least one carbon-carbon triple bond. The alkynyl group may be substituted with one or more groups, including but not limited to alkyl, alkyl halide, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxyl, ketone, nitro, silyl, sulfo-oxy, sulfonyl, sulfone, sulfoxide or thiol, as described below.

[0150] The term "aryl" as used herein is a group containing any carbon-based aromatic group, including but not limited to benzene, naphthalene, phenyl, biphenyl, phenoxybenzene, etc. The term "heteroaryl" is defined as a group containing an aromatic group having at least one heteroatom incorporated into the ring of an aromatic group. Examples of heteroatoms include but are not limited to nitrogen, oxygen, sulfur and phosphorus. The term "non-heteroaryl" included in the term "aryl" defines a group containing an aromatic group that does not contain heteroatoms. An aryl or heteroaryl group can be substituted or unsubstituted. An aryl or heteroaryl group can be substituted by one or more groups, including but not limited to alkyl, halogenated alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxyl, ketone, nitro, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide or thiol as described herein. The term "biaryl" is a specific type of aryl group and is included in the definition of aryl. Biaryl refers to two aryl groups held together by a fused ring structure, as in naphthalene, or two aryl groups linked by one or more carbon-carbon bonds, as in biphenyl.

[0151] The term "cycloalkyl" as used herein is a non-aromatic carbonyl ring consisting of at least three carbon atoms. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc. The term "heterocycloalkyl" is a cycloalkyl group as defined above, wherein at least one carbon atom in the carbon atoms of the ring is replaced by a heteroatom, such as but not limited to nitrogen, oxygen, sulfur or phosphorus. Cycloalkyl and heterocycloalkyl can be substituted or unsubstituted. Cycloalkyl groups and heterocycloalkyl groups can be substituted by one or more groups, including but not limited to alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxyl, ketone, nitro, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide or thiol as described herein.

[0152] The term "cycloalkenyl" as used herein is a non-aromatic carbonyl ring consisting of at least three carbon atoms and containing at least one double bond (i.e., C=C). Examples of cycloalkenyl groups include, but are not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl, etc. The term "heterocycloalkenyl" is a cycloalkenyl group as defined above, and is included in the meaning of the term "cycloalkenyl", wherein at least one of the carbon atoms of the ring is replaced by a heteroatom, such as, but not limited to, nitrogen, oxygen, sulfur or phosphorus. Cycloalkenyl and heterocycloalkenyl groups can be substituted or unsubstituted. Cycloalkenyl groups and heterocycloalkenyl groups can be substituted by one or more groups, including, but not limited to, alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, aldehyde, amino, carboxylic acid, ester, ether, halide, hydroxyl, ketone, nitro, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide or thiol as described herein.

[0153] The term "cyclic group" is used herein to refer to an aryl group, a non-aryl group (i.e., a cycloalkyl, heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl group), or both. The cyclic group has one or more ring systems that may be substituted or unsubstituted. The cyclic group may contain one or more aryl groups, one or more non-aryl groups, or one or more aryl groups and one or more non-aryl groups.

[0154] The term "aldehyde" as used herein is represented by the formula— C(O)H. Throughout the specification, "C(O)" or "CO" is a shorthand notation for C=O.

[0155] As used herein, the term "amine" or "amino group" is represented by the formula: NZ 1 Z 2 Indicates that Z 1 and Z 2 Each may be a substituent group as described herein, such as a hydrogen, alkyl, halogenated alkyl, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl group.

[0156] The term "carboxylic acid" as used herein is represented by the formula -C(O)OH. A "carboxylate" or "carboxyl" group as used herein is represented by the formula -C(O)OH. - express.

[0157] As used herein, the term "ester" is a compound of the formula -OC(O)Z 1 or—C(O)OZ 1 Indicates that Z 1 The alkyl group, halogenated alkyl group, alkenyl group, alkynyl group, aryl group, heteroaryl group, cycloalkyl group, cycloalkenyl group, heterocycloalkyl group or heterocycloalkenyl group described above may be mentioned.

[0158] As used herein, the term "ether" is a 1 OZ 2 Indicates that Z 1 and Z 2 and alkyl, alkyl, alkyl, alkyl, alkylene, alkylene, alkylene, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl groups as described above.

[0159] As used herein, the term "ketone" is a 1 C(O)Z 2 Indicates that Z 1 and Z 2 The groups may independently be the alkyl, halogenated alkyl, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl groups described above.

[0160]

[0046] The term "halide" or "halogen" as used herein refers to fluorine, chlorine, bromine and iodine.

[0161] The term "hydroxyl" as used herein is represented by the formula— OH.

[0162] As used herein, the term "nitro" is represented by the formula— NO2.

[0163] As used herein, the term "silyl group" is represented by the formula: SiZ 1 Z 2 Z 3 Indicates that Z 1 , Z 2 and Z 3 The groups may independently be the alkyl, halogenated alkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl groups described above.

[0164] The term "sulfonyl" as used herein refers to a sulfonyl group of the formula -S(O)2Z 1 A sulfo-oxo group represented by 1It may be hydrogen, alkyl, halogenated alkyl, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or the above-mentioned heterocycloalkenyl group as described above.

[0165] The term "sulfonylamino" or "sulfonamide" as used herein is represented by the formula -S(O)2NH-.

[0166] The term "phosphono" is used herein to refer to a group of the formula -P(O)(OZ 1 )2 represents a phosphorus-oxo group, wherein Z 1 It may be a hydrogen, alkyl, halogenated alkyl, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl group as described above.

[0167] As used herein, the term "thiol" is represented by the formula -SH.

[0168] The term "thio" as used herein is represented by the formula —S—.

[0169] As used herein, “R 1 ”, “R 2 ”, “R 3 ”, “R n " etc. (where n is an integer) may independently have one or more of the groups listed above. For example, if R 1 For a straight chain alkyl group, one of the hydrogen atoms of the alkyl group can be optionally substituted by a hydroxyl group, an alkoxy group, an amine group, an alkyl group, a halide, etc. Depending on the selected group, the first group can be incorporated into the second group, or alternatively, the first group can be suspended (i.e., connected) to the second group. For example, for the phrase "an alkyl group containing an amino group", the amino group can be incorporated into the main chain of the alkyl group. Alternatively, the amino group can be attached to the main chain of the alkyl group. The nature of the selected group will determine whether the first group is embedded or attached to the second group.

[0170] As used herein, "ionizable lipids" refer to lipids with groups or atoms that can be charged (ionized) depending on the pH of the environment. Typically, the charge of primary, secondary, and tertiary amines depends on the pH of the system. At low pH levels, these amines tend to be strongly cationic. At high pH levels, these amines will not ionize.

[0171] Unless stated to the contrary, formulas with chemical bonds shown only as solid lines, rather than wedges or dashes, contemplate every possible isomer, e.g., each enantiomer, diastereomer, and meso compound, as well as mixtures of isomers, such as racemic or scalemic mixtures.

[0172] Reference will now be made in detail to specific aspects of the disclosed materials, compounds, compositions, articles, and methods, examples of which are illustrated in the accompanying Examples and Figures.

[0173] Compound

[0174] Described herein are compounds of formula Y:

[0175]

[0176] or a pharmaceutically acceptable salt thereof, wherein:

[0177] R 1 is H or -CH3;

[0178] represents a carbon-carbon bond or a carbon-carbon double bond;

[0179] R 7 Is H or OR 16 ;

[0180] R 8a and R 8b Each independently is H or OR 16 , or R 8a and R 8b Together with the atoms to which they are attached, they form a cycloalkyl, aryl, heterocycloalkyl or heteroaryl group;

[0181] R 9 is H, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl;

[0182] W is CR 4a or CR 4a R 4b , where if there is a double bond between W and the adjacent carbon, then W is CR 4a ; and if there is a single bond between W and the adjacent carbon, then W is CR 4a R 4b ; R 4a and R 4b Each of is independently H, halogen or C1-C6 alkyl;

[0183] A is

[0184]

[0185] X is O or S;

[0186] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)R a C(=O)-、-NR aC(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0187] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0188] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0189] R a yes wherein n is an integer from 0 to 12;

[0190] x is 0, 1, or 2;

[0191] M is CH,

[0192] G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0193] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0194] R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0195] R 3 yes

[0196] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0197] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl;

[0198] p and q are independently 0, 1, 2, 3 or 4;

[0199] R 35 yes

[0200] L 4 Does not exist, yes

[0201] R 10 Not present, or C1-C6 alkyl;

[0202] L 5 does not exist, or or

[0203] m is an integer 1, 2 or 3;

[0204] L 6 Does not exist, yes

[0205] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl or C2-C9 heteroaryl;

[0206] L 7 is C1-C6 alkylene;

[0207] R 13a , R 13b and R 13c Each is independently C1-C6 alkyl or C6-C 10 Aryl;

[0208] R 14 , R 16 , R 19 , R 21 , R 22 , R 24 , R 28 , R 29 and R 31Each independently is H or C1-C6 alkyl;

[0209] R 15 Yes-OR 16 、-NR 17 R 17 or

[0210] R 17 Each independently is H, -OR 16 、C6-C 10 Aryl or C1-C6 alkyl;

[0211] R 18 Each is independently halogen or C1-C6 alkyl;

[0212] o1 is an integer from 0 to 8;

[0213] p1 and p2 are each independently an integer from 0 to 2;

[0214] Z is CH2, O, S or NR 16 ;

[0215] s and t are each independently 0 or 1;

[0216] R 23 is halo, hydroxy, C1-C6 alkyl or C1-C6 heteroalkyl;

[0217] R 20 , R 25a and R 25b , R 30a , R 30b , R 30c , R 32a , R 32b and R 34 Each is independently a C1-C6 alkyl group;

[0218] R 26a and R 26b are each independently H, C1-C6 alkyl, or R 26a and R 26b Combined with the atoms to which they are connected, Where R 26c and R 26d Each is independently H or substituted C1-C6 alkyl;

[0219] R 27a and R 26b Each is independently H, hydroxyl or C1-C6 alkyl;

[0220] u is 1, 2, or 3;

[0221] R 33aand R 33b Each is independently C1-C6 alkyl, C1-C6 heteroalkyl, halogen or hydroxy;

[0222] Q is O, S or NR 16 ;and

[0223] or a pharmaceutically acceptable salt thereof.

[0224] In some embodiments, R 35 yes in

[0225] L 4 Does not exist, yes

[0226] R 10 Not present, or C1-C6 alkyl;

[0227] L 5 does not exist, or or

[0228] m is an integer 1, 2 or 3;

[0229] L 6 Does not exist, yes and

[0230] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl or C2-C9 heteroaryl;

[0231] or a pharmaceutically acceptable salt thereof.

[0232] In one aspect, there is provided herein a compound of formula I:

[0233]

[0234] or a pharmaceutically acceptable salt thereof, wherein:

[0235] R 1 is H or -CH3;

[0236] represents a carbon-carbon bond or a carbon-carbon double bond;

[0237] L 4 Does not exist, yes

[0238] R 10 Not present, or C1-C6 alkyl;

[0239] L 5 does not exist, or or

[0240] m is an integer 1, 2 or 3;

[0241] L 6 Does not exist, yes

[0242] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl, C2-C9 heteroaryl;

[0243] R 7 Is H or OR 16 ;

[0244] R 8a and R 8b Each independently is H or OR 16 , or R 8a and R 8b Together with the atoms to which they are attached, they form a cycloalkyl, aryl, heterocycloalkyl or heteroaryl group;

[0245] R 16 is H or C1-C6 alkyl;

[0246] R 9 is H, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl;

[0247] W is CR 4a or CR 4a R 4b , where if there is a double bond between W and the adjacent carbon, then W is CR 4a ; and if there is a single bond between W and the adjacent carbon, then W is CR 4a R 4b ; R 4a and R 4b Each of is independently H, halogen or C1-C6 alkyl;

[0248] A is

[0249]

[0250] X is O or S;

[0251] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)R a C(=O)-、-NR a C(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0252] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0253] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0254] R a yes wherein n is an integer from 0 to 12;

[0255] x is 0, 1, or 2;

[0256] M is CH,

[0257] G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0258] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0259] R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0260] R 3 yes

[0261] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0262] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and

[0263] p and q are independently 0, 1, 2, 3 or 4;

[0264] or a pharmaceutically acceptable salt thereof.

[0265] In some embodiments, described herein are compounds of Formula Ia:

[0266]

[0267] or a pharmaceutically acceptable salt thereof, wherein:

[0268] R 1 is H or -CH3;

[0269] represents a carbon-carbon bond or a carbon-carbon double bond;

[0270] L 4 Does not exist, yes

[0271] L 5 does not exist, or or

[0272] Where m is an integer 1, 2 or 3;

[0273] L 6 Does not exist, yes

[0274] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl, C2-C9 heteroaryl;

[0275] R 10Not present, or C1-C6 alkyl;

[0276] A is

[0277]

[0278] X is O or S;

[0279] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)R a C(=O)-、-NR a C(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0280] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0281] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0282] R a yes wherein n is an integer from 0 to 12;

[0283] x is 0, 1, or 2;

[0284] M is CH,

[0285] G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0286] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0287] R 2 Yes-CR b R c、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0288] R 3 yes

[0289] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0290] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and

[0291] p and q are independently 0, 1, 2, 3 or 4;

[0292] or a pharmaceutically acceptable salt thereof.

[0293] In some embodiments, the compound of formula Y can be:

[0294]

[0295]

[0296] or a pharmaceutically acceptable salt thereof; wherein A, R 1 , R 7 , R 8a , R 8b , R 9 , W is the same as formula Y.

[0297] In some embodiments, the compound of formula Y can be:

[0298]

[0299]

[0300]

[0301]

[0302]

[0303]

[0304]

[0305]

[0306]

[0307]

[0308]

[0309] or a pharmaceutically acceptable salt thereof; wherein A is the same as Formula Y.

[0310] In some embodiments, described herein are compounds of Formula II or Formula III:

[0311]

[0312] or a pharmaceutically acceptable salt thereof, wherein:

[0313] R 1 is H or -CH3;

[0314] X is O or S;

[0315] represents a carbon-carbon bond or a carbon-carbon double bond;

[0316] L 4 Does not exist, yes

[0317] R 10 Not present, or C1-C6 alkyl;

[0318] L 5 does not exist, or or

[0319] m is an integer 1, 2 or 3;

[0320] L 6 Does not exist, yes

[0321] R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl or its derivatives;

[0322] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)Ra C(=O)-、-NR a C(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0323] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0324] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0325] R a yes wherein n is an integer from 0 to 12;

[0326] x is 0, 1, or 2;

[0327] M is CH,

[0328] G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0329] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0330] R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0331] R 3 yes

[0332] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b、-C(=O)NR b ,

[0333] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and

[0334] p and q are independently 0, 1, 2, 3 or 4;

[0335] or a pharmaceutically acceptable salt thereof.

[0336] In some embodiments, R 1 is -CH3. In some embodiments, X is O. In some embodiments, L 4 yes In some embodiments, L 5 yes In some embodiments, L 5 yes and m is 3. In some embodiments, L 6 In some embodiments, R 4 is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, hexyl, heptyl, octyl, nonyl or decyl. In some embodiments, R 4 In some embodiments, M is CH.

[0337] In some embodiments, R 3 yes

[0338] In some embodiments, R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0339] In some embodiments, described herein are compounds of Formula IV or Formula V:

[0340]

[0341] or a pharmaceutically acceptable salt thereof, wherein:

[0342] L 1 is -O(C=O)-, -C(=O)-, -S(O) x -、-C(=O)R aC(=O)-、-NR a C(=O)-、-NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-;

[0343] L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -、-SS-、-NR a C(=O)- or -C(=O)R a N-;

[0344] L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-;

[0345] R a yes wherein n is an integer from 0 to 12;

[0346] x is 0, 1, or 2;

[0347] M is CH,

[0348] G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0349] G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl;

[0350] R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl;

[0351] R 3 yes

[0352] R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NRb ,

[0353] R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and

[0354] p and q are independently 0, 1, 2, 3 or 4.

[0355] In some embodiments, L 1 is -C(=O)-. In some embodiments, L 2 is -(C=O)O-. In some embodiments, L 3 In some embodiments, G 1 is a C5 alkyl group. In some embodiments, G 2 is a C6 alkyl group. In some embodiments, G 3 is a C2 alkyl group. In some embodiments, G 4 Does not exist.

[0356] In some embodiments, R 2 Yes-CR b R c In some embodiments, R b and R c is a C2 alkyl group. In some embodiments, R b and R c is a C4 alkyl group. In some embodiments, R b and R c is a C6 alkyl group. In some embodiments, R b and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b and R c It is C 12 In some embodiments, R 2 Yes-CR b R c , and R b It is C 12 Alkyl and R c It is C 10 In some embodiments, R2 Yes-CR b R c , and R b It is C 10 Alkyl and R c It is C 12 In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkyl and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkyl and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkyl and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CRb R c , and R b It is C 10 Alkyl and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkyl and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b Rc , and R b is a C8 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and Rc is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2Yes-CR b R c , and R b is a C6 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C4 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C2 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C8 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and Rb is a C4 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C1 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C8 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C6 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c is a C4 alkyl group. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkyl group and R c It is a C2 alkyl group.

[0357] In some embodiments, R 2 Yes-CR b R c In some embodiments, R b and R c is C2 alkenyl. In some embodiments, R b and R c is C4 alkenyl. In some embodiments, R b and R c is C6 alkenyl. In some embodiments, R b and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b and R c It is C 10 In some embodiments, R 2 Yes-CRb R c , and R b and R c It is C 12 In some embodiments, R 2 Yes-CR b R c , and R b It is C 12 Alkenyl and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkenyl and R c It is C 12 In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c It is C 10 In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 And R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b Rc , and R b It is C 10 Alkenyl and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkenyl and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkenyl and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b It is C 10 Alkenyl and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C6 alkenyl. In some embodiments, R 2 Yes-CRb R c , and R b is C8 alkenyl and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C6 and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R bis a C6 alkenyl group and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C2 alkenyl. In some embodiments, R2 Yes-CR b R c , and R b is C4 alkenyl and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C8 alkenyl and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C2 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c, and R b is C8 alkenyl and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C6 alkenyl group and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is C4 alkenyl and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C2 alkenyl group and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C1 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C8 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C6 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c is C4 alkenyl. In some embodiments, R 2 Yes-CR b R c , and R b is a C1 alkenyl group and R c It is a C2 alkenyl group.

[0358] In some embodiments, R 2 Yes - C6H 13 、-C7H 15 、-C8H 17 , -C 10 H 21 , -C 11 H 23 , -C 12 H 25, -C 13 H 27 , -C 14 H 29 , -C 15 H 31 , -C 16 H 33 , -C 17 H 35 , -C 18 H 36 , -C 19 H 39 , -C 20 H 41 , -C 21 H 43 , -C 22 H 45 , -C 23 H 47 , -C 24 H 49 、-C6H 12 、-C7H 14 、-C8H 16 , -C 10 H 20 , -C 11 H 21 , -C 12 H 24 , -C 13 H 26 , -C 14 H 28 , -C 15 H 30 , -C 16 H 32 , -C 17 H 34 , -C 18 H 35 , -C 19 H 38 , -C 20 H 40 , -C 21 H 42 , -C 22 H 44 , -C 23 H 46 or -C 24 H 48 .

[0359] In some embodiments, R 3 for In some embodiments, p and q are 0. In some embodiments, p and q are 1. In some embodiments, p and q are 2. In some embodiments, p and q are 3. In some embodiments, p and q are 4. In some embodiments, p is 0 and q is 1. In some embodiments, p is 0 and q is 2. In some embodiments, p is 0 and q is 3. In some embodiments, p is 0 and q is 4. In some embodiments, p is 1 and q is 2. In some embodiments, p is 1 and q is 3. In some embodiments, p is 1 and q is 4. In some embodiments, p is 2 and q is 1. In some embodiments, p is 2 and q is 3. In some embodiments, p is 2 and q is 4. In some embodiments, p is 3 and q is 1. In some embodiments, p is 3 and q is 2. In some embodiments, p is 3 and q is 4. In some embodiments, p is 4 and q is 1. In some embodiments, p is 4 and q is 2. In some embodiments, p is 4 and q is 3.

[0360] In some embodiments, M is CH. In some embodiments, M is In some embodiments, M is In some embodiments, M is In some embodiments, M is In some embodiments, M is In some embodiments, M is In some embodiments, M is

[0361] In some embodiments, R 3 yes

[0362] In some embodiments, R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b ,

[0363] In some embodiments, the compound of Formula Y, I, Ia or II-V can be selected from:

[0364]

[0365]

[0366]

[0367]

[0368]

[0369]

[0370]

[0371] or a pharmaceutically acceptable salt thereof.

[0372] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0373]

[0374]

[0375] or a pharmaceutically acceptable salt thereof.

[0376] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0377]

[0378] or a pharmaceutically acceptable salt thereof, or any combination thereof.

[0379] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0380] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0381]

[0382] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0383]

[0384] or a pharmaceutically acceptable salt thereof.

[0385] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0386]

[0387] In some embodiments, the compound of Formula Y, I, Ia or II-V can be:

[0388]

[0389] or a pharmaceutically acceptable salt thereof.

[0390] Described herein are compositions comprising a compound of Formula Y, I, Ia, II, III, IV, aV, or any combination thereof; and an active agent.

[0391] Composition

[0392] The compositions described herein include compounds of formula Y, I, Ia, II, III, IV, V, or any combination thereof, and an active agent. The composition may also include an excipient that can be used for a variety of medical and non-medical applications. For example, the pharmaceutical compositions described herein can be used to deliver an effective amount of an agent to a subject in need thereof. The compositions described herein can be used for non-medical applications, such as emulsions or emulsifiers, and can be used, for example, as food ingredients, for fire extinguishing, for disinfecting surfaces, for oil removal, etc.

[0393] In some embodiments, the composition can be a lipid nanoparticle dispersion, a liposomal formulation, a lipid emulsion, or any combination thereof, comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof.

[0394] In one aspect, the present invention provides a composition comprising: a lipid nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0395] In certain embodiments, the composition may include an agent as described herein. For example, in certain embodiments, the agent is any chemical compound to be administered to a subject that can be delivered using particles or nanoparticles as described herein. The agent can be an organic molecule (e.g., therapeutic agent, drug), an inorganic molecule, a small molecule, an organometallic compound, a metal, a nucleic acid, a protein, an amino acid, a peptide, a polypeptide, a polynucleotide, a targeting agent, an isotope-labeled organic or inorganic molecule, a vaccine, an immune agent, or an agent for bioprocessing. In certain embodiments, the agent is a polynucleotide. In certain embodiments, the agent includes an mRNA encoding at least one antigenic polypeptide or an immunogenic fragment thereof that can induce an immune response to the antigenic polypeptide.

[0396] In some embodiments, mRNA encoding at least one antigenic polypeptide or an immunogenic fragment thereof capable of inducing an immune response against the antigenic polypeptide is encapsulated by the nanoparticle.

[0397] In some aspects, disclosed herein is a pharmaceutical composition comprising a pharmaceutically acceptable carrier and nanoparticles, wherein the nanoparticles comprise a compound of Formula I, Ia, II, III, IV, V, or any combination thereof; and mRNA encoding at least one antigenic polypeptide or an immunogenic fragment thereof that is capable of inducing an immune response against the antigenic polypeptide.

[0398] Nanoparticles

[0399] Described herein are lipid nanoparticles comprising 20% ​​to 80% of a compound described herein (e.g., a compound of Formula I, Ia, II, III, IV, V, or any combination thereof); greater than 0% to 5% of a polyethylene glycol-lipid; greater than 0% to 40% of a helper lipid; 0% to 80% of a sterol; and

[0400] An active agent is encapsulated within the nanoparticles.

[0401] Also described herein are compositions comprising an effective amount of the lipid nanoparticles described herein and a pharmaceutically acceptable carrier.

[0402] In one aspect, the present disclosure provides a nanoparticle comprising: A, I, Ia II, III, IV, V compound, or any combination thereof; and optionally a helper lipid; a polyethylene glycol-lipid; a sterol or any combination thereof; and

[0403] An active agent is encapsulated within the nanoparticles.

[0404] In one aspect, the present disclosure provides a nanoparticle comprising: A, I, Ia II, III, IV, V compound, or any combination thereof; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0405] An active agent is encapsulated within the nanoparticles.

[0406] In one aspect, the present disclosure provides a nanoparticle comprising: A, I, Ia II, III, IV, V compound, or any combination thereof; a helper lipid; a polyethylene glycol-lipid; and a sterol; and

[0407] An active agent is encapsulated within the nanoparticles.

[0408] In one aspect, the present disclosure provides a nanoparticle comprising: a compound of formula Y; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0409] An active agent is encapsulated within the nanoparticles.

[0410] In one aspect, the present invention provides a nanoparticle comprising: a compound of formula I; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0411] An active agent is encapsulated within the nanoparticles.

[0412] In one aspect, the present disclosure provides a nanoparticle comprising: a compound of formula Ia; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0413] An active agent is encapsulated within the nanoparticles.

[0414] In one aspect, the present disclosure provides a nanoparticle comprising: a compound of formula II; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0415] An active agent is encapsulated within the nanoparticles.

[0416] In one aspect, the present disclosure provides a nanoparticle comprising: a compound of formula III; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0417] An active agent is encapsulated within the nanoparticles.

[0418] In one aspect, the present disclosure provides a nanoparticle comprising: a compound of formula IV; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0419] An active agent is encapsulated within the nanoparticles.

[0420] In one aspect, the present invention provides a nanoparticle comprising: a compound of formula V; and a helper lipid; a polyethylene glycol-lipid; and / or a sterol; and

[0421] An active agent is encapsulated within the nanoparticles.

[0422] Various compounds of formula Y, I, Ia, II, III, IV, or V are described in the Compounds section above. In some embodiments, the nanoparticles comprise 20% to 80% by molar ratio of a compound of formula Y, I, Ia II, III, IV, V, or any combination thereof; and

[0423] An active agent is encapsulated within the nanoparticles.

[0424] In some embodiments, the nanoparticles include helper lipids. In some embodiments, the helper lipids may be non-cationic lipids. In some embodiments, the non-cationic lipids may include, but are not limited to, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1-stearoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (SOPE), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dioleyl-sn-glycero-3-phosphatidylcholine (DOPE). In one embodiment, the non-cationic lipid is 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DPPE), 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1,2-dioleoyl-5 / 7-glycero-3-phospho-(l'-rac-glycerol) (DOPG), 1-palmitoyl-2-linoleoyl-sn-glycero-3-phosphocholine (PLPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). 1-stearoyl-2-myristoyl-sn-glycero-3-phosphocholine (SMPC) or a combination thereof. In one embodiment, the non-cationic lipid is 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). In one embodiment, the non-cationic lipid is 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE). In one embodiment, the non-cationic lipid is 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC). In one embodiment, the non-cationic lipid is 1-stearoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (SOPE). Although several non-cationic lipids are described herein, additional non-cationic lipids can be used in combination with the compounds disclosed herein.

[0425] In some embodiments, the helper lipid can be present in a molar ratio of at least 0% (e.g., at least 5%, at least 10%, at least 20%, at least 30%, or at least 40%). In some embodiments, the helper lipid can be present in a molar ratio of 40% or less (e.g., 30% or less, 20% or less, 10% or less, 5% or less, 1% or less, or 0.5% or less).

[0426] Helper lipids can exist in a molar ratio within the range of any of the above-mentioned minimum values ​​to any of the above-mentioned maximum values. For example, in some embodiments, helper lipids can exist in a molar ratio of 0% to 40% (e.g., greater than 0% to 30%, greater than 0% to 20%, greater than 0% to 10%, greater than 0% to 5%, greater than 0% to 1%, greater than 0% to 0.5%, 1% to 30%, 1% to 20%, 1% to 10%, 1% to 5%, 5% to 30%, 5% to 20%, 5% to 10%, 10% to 30%, 10%% to 20%, 20% to 30%, 20% to 40%, or 30% to 40%).

[0427] In some embodiments, the nanoparticles include polyethylene glycol-lipids (PEG-lipids). PEG-lipids are incorporated to form a hydrophilic outer layer and stabilize the particles. Non-limiting examples of polyethylene glycol-lipids include PEG-modified lipids, such as PEG-modified phosphatidylethanolamine, PEG-modified phosphatidic acid, PEG-modified ceramides, PEG-modified dialkylamines, PEG-modified diacylglycerols and PEG-modified dialkylglycerols. Representative polyethylene glycol-lipids include DMG-PEG, DLPE-PEG, DMPE-PEG, DPPC-PEG and DSPE-PEG. In one embodiment, polyethylene glycol-lipids are 1,2-dimyristoyl-sn-glycerol, methoxy polyethylene glycol (DMG-PEG). In one embodiment, polyethylene glycol-lipids are 1,2-dimyristoyl-sn-glycerol, methoxy polyethylene glycol-2000 (DMG-PEG2000). DMG-PEGXXXX means 1,2-dimyristoyl-sn-glycerol, methoxypolyethylene glycol-XXXX, wherein XXXX represents the molecular weight of the polyethylene glycol portion, for example, DMG-PEG2000 or DMG-PEG5000.

[0428] In some embodiments, the polyethylene glycol-lipid can be present at a molar ratio of at least 0% (e.g., at least 0.25%, at least 0.5%, at least 0.75%, at least 1%, at least 1.5%, at least 2%, at least 3%, at least 4%, or at least 5%). In some embodiments, the polyethylene glycol-lipid can be present at a molar ratio of 5% or less (e.g., 4% or less, 3% or less, 2% or less, 1% or less, or 0.5% or less).

[0429] Polyethylene glycol-lipid can exist in the molar ratio within the scope of any minimum value mentioned above to any maximum value mentioned above. For example, in some embodiments, Polyethylene glycol-lipid can exist in the molar ratio of 0% to 5% (for example, greater than 0% to 4%, greater than 0% to 3%, greater than 0% to 2%, greater than 0% to 1%, greater than 0% to 0.5%, 1% to 5%, 1% to 4%, 1% to 3%, 1% to 2%, 2% to 5%, 2% to 4%, 2% to 3%, 3% to 5%, 3% to 4% or 4% to 5%). In one embodiment, the molar ratio of Polyethylene glycol-lipid is 0.75%.

[0430] In certain embodiments, the nanoparticles include sterols. Sterols are well known to those skilled in the art and generally refer to compounds having a cyclopentane polyhydrophenanthrene ring system and having one or more OH substituents. Examples of sterols include, but are not limited to, cholesterol, campesterol, ergosterol, sitosterol, etc.

[0431] In some embodiments, sterol is selected from cholesterol-based lipids. In some embodiments, one or more cholesterol-based lipids are selected from cholesterol, pegylated cholesterol, DC-Choi (N, N-dimethyl-N-ethylcarboxamide cholesterol), 1,4-bis(3-N-oleylamino-propyl)piperazine or a combination thereof.

[0432] Sterols can tune the permeability and fluidity of particles based on their function in cell membranes. In one embodiment, the sterol is cholesterol.

[0433] In some embodiments, the molar ratio of sterols is at least 0% (e.g., at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 60%, at least 70%, or at least 80%). In some embodiments, the molar ratio of sterols is 80% or less (e.g., 70% or less, 60% or less, 50% or less, 40% or less, 30% or less, 20% or less, 10% or less, 5% or less, or 1% or less).

[0434] The sterols may be present in a molar ratio ranging from any of the above minimum values ​​to any of the above maximum values. In some embodiments, the sterols may be present in a molar ratio ranging from 0% to 80% (e.g., greater than 0% to 70%, greater than 0% to 60%, greater than 0% to 50%, greater than 0% to 40%, greater than 0% to 30%, greater than 0% to 20%, greater than 0% to 10%, greater than 0% to 5%, greater than 0% to 1%, 5% to 70%, 5% to 60%, 5% to 50%, 5% to 40%, 5% to 30%, 5% to 20%, 5% to 10%, 10% to 70%, 10% to 60%, 10% to 50%, 10% to 40%, 10% to 30%, 10% to 20%, 20% to In some embodiments, the nanoparticles comprise a sterol at a molar ratio of 40%.

[0435] In some embodiments, the nanoparticles may comprise 40% to 60% of a compound described herein; 1% to 2% polyethylene glycol-lipid; 8% to 12% helper lipid; 35% to 40% sterol; and an active agent encapsulated in the nanoparticle.

[0436] In one embodiment, nanoparticles are described that include 50% of a compound described herein; 1.5% polyethylene glycol-lipid; 10% helper lipid (e.g., DSPC); 38.5% sterol (e.g., cholesterol); and an active agent encapsulated in the nanoparticle.

[0437] In one embodiment, the present disclosure provides a nanoparticle comprising:

[0438] A compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof; and

[0439] An active agent is encapsulated within the nanoparticles.

[0440] In some embodiments, nanoparticles are described that comprise:

[0441] A compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof;

[0442] 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE);

[0443] 1,2-Dimyristoyl-sn-glycerol, methoxypolyethylene glycol (DMG-PEG 2000 );

[0444] Cholesterol; and

[0445] An active agent is encapsulated within the nanoparticles.

[0446] In one embodiment, the present disclosure provides a nanoparticle comprising:

[0447] A compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof;

[0448] 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE);

[0449] 1,2-Dimyristoyl-sn-glycerol, methoxypolyethylene glycol (DMG-PEG 2000 );

[0450] Cholesterol; and

[0451] An active agent is encapsulated within the nanoparticles.

[0452] In one embodiment, the present disclosure provides a nanoparticle comprising:

[0453] A compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof;

[0454] 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC);

[0455] 1,2-Dimyristoyl-sn-glycerol, methoxypolyethylene glycol (DMG-PEG 2000 );and

[0456] Cholesterol.

[0457] In one embodiment, the present disclosure provides a nanoparticle comprising:

[0458] A compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof;

[0459] 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE);

[0460] 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000](DSPE-PEG 2000 );

[0461] Cholesterol; and

[0462] An active agent is encapsulated within the nanoparticles.

[0463] In one embodiment, the nanoparticles may further include an active agent. In one embodiment, the nanoparticles may further include a therapeutic agent. In one embodiment, the nanoparticles may further include a diagnostic agent. In one embodiment, the nanoparticles may further include a preventive agent.

[0464] Active Agent

[0465] As used herein, "active agent" refers to a therapeutic agent, a diagnostic agent, or a prophylactic agent. As discussed herein, the therapeutic agent can be released from the disclosed compounds, compositions, and systems in a biologically active form.

[0466] It should also be understood that, as used herein, the term "therapeutic agent" refers to one or more therapeutic agents, active ingredients or substances that can be used to treat medical conditions. Therapeutic agents include any synthetic or naturally occurring biologically active compounds or material compositions that, when applied to an organism (human or non-human animal), induce the desired pharmacological, immunogenic and / or physiological effects through local and / or systemic effects. The term therefore encompasses those compounds or chemicals that are traditionally considered to include drugs, vaccines and biopharmaceuticals such as proteins, peptides, hormones, nucleic acids, genetic constructs and other molecules. Examples of therapeutic agents are described in well-known literature references such as the Merck Index (14th Edition), the Physicians' Desk Reference (64th Edition), and The Pharmacological Basis of Therapeutics (12th Edition), and they include, but are not limited to, pharmaceuticals; vitamins and minerals, such as essential amino acids, calcium, iron, potassium, zinc, vitamin B12, etc.; substances used to treat, prevent, diagnose, cure, or alleviate disease or disorder; substances that affect the structure or function of the body, or prodrugs that become biologically active or more active after being placed in a physiological environment. For example, the term "therapeutic agent" includes compounds or compositions used in all major therapeutic areas, including but not limited to adjuvants; antimicrobial agents (including antibiotics, antivirals, antiparasitics and antifungals), anti-inflammatory agents (including steroids and nonsteroidal anti-inflammatory agents), anticoagulants, ophthalmic agents, gastrointestinal agents, antiplatelet agents, and antibacterial agents, steroidal agents, anti-tumor agents, anti-cancer agents, antigens, antibodies (e.g., cetuximab, anti-CD24 antibodies, panitumumab and bevacizumab), birth control agents, progestational agents, anticholinergic agents, nutritional agents, analgesics and analgesic combinations such as acetaminophen, acetylsalicylic acid, etc.; anesthetics such as lidocaine, xylocaine, etc., anorectics such as dextrothyroxine, phendimetrazine tartrate, etc.; antiepileptics, local and general anesthetics, hypnotics, sedatives, anti Psychotropics, tranquilizers, antidepressants such as isocarboxazid, amoxapine, etc.; anxiolytics, antagonists, neuron blockers, anticholinergics and cholinergics, antimuscarinics and muscarinics, anti-Parkinson's disease agents, anti-Alzheimer's disease agents, antiadrenergics, antiarrhythmics, antihypertensives, hormones such as insulin, progesterone, estrogen, corticosteroids, glucocorticoids, androgens, etc.; and nutrients, antiarthritis drugs such as methylprednisolone, ibuprofen, etc.; antiasthmatics, such as terbutaline sulfate, theophylline, ephedrine, etc.; anticonvulsants, such as phenytoin sodium, diazepam, etc.; antiallergics, antihistamines, such as diphenhydramine hydrochloride, chlorpheniramine maleate, etc.; antinausea drugs, antitumor drugs, antipruritic drugs, antipyretics; antispasmodics, such as belladonna alkaloids, dicyclomine hydrochloride, etc.;Cardiovascular agents such as prazosin hydrochloride, nitroglycerin, propranolol hydrochloride, hydralazine hydrochloride, pancreatic lipase, succinate dehydrogenase, etc.; vasoactive agents, cardiovascular preparations (including calcium channel blockers, beta-blockers, beta-agonists and antiarrhythmic drugs), antihypertensives, diuretics such as furosemide, spironolactone, etc.; vasodilators; central nervous system stimulants; cough and cold preparations; decongestants; diagnostic agents; bone growth stimulants and bone resorption inhibitors; muscle relaxants; psychostimulants; sedatives; tranquilizers such as chlorpromazine, diazepam, chlorpromazine hydrochloride, reserpine, chlordiazepoxide hydrochloride, etc.; anti-ulcer drugs such as ranitidine hydrochloride, cimetidine hydrochloride, etc.; anti-asthma agents, antidiarrheal drugs, anti-obesity agents, antithrombotic agents, antitussive drugs, anti-uric acid drugs, anti-anginal drugs, appetite suppressants preparations, expectorants, hyperglycemic agents, hypoglycemic agents, thyroid drugs and antithyroid drugs, tissue growth agents, uterine relaxants, immunomodulators, including, for example, cytokines, interleukins, interferons, colony stimulating factors, tumor necrosis factors, etc.; immunosuppressants, such as rapamycin, tacrolimus, etc.; immune agents; antigens, factors, growth factors, amino acids, peptides and proteins and fragments thereof (whether naturally occurring, chemically synthesized or recombinantly produced), such as LHRH, somatostatin, calcitonin, growth hormone, glucagon-like peptide, growth releasing factor, angiotensin, FSH, EGF, bone morphogenetic protein (BMP), erythropoietin (EPO), interferon, interleukin, collagen, fibrinogen, insulin, factor VIII, factor IX,; α-Glucosidase, Cerazyme / Vasopressin, ACTH, human serum albumin, gamma globulin, structural proteins, blood product proteins, complex proteins, antigens or antigenic polypeptides, enzymes, antibodies, monoclonal antibodies, etc.; and nucleic acid molecules (polymeric forms of two or more nucleotides, polynucleotides, ribonucleotides (RNA) or deoxyribonucleotides (DNA), including double-stranded and single-stranded molecules, gene constructs, expression mediators, antisense molecules, etc.), small molecules (e.g., doxorubicin) and other biologically active macromolecules, such as proteins and enzymes. The agent can be a bioactive agent for medical (including veterinary) applications and agriculture (such as plants) and other fields. In certain embodiments of the present disclosure, the agent to be delivered can be a mixture of active agents.

[0467] Representative examples of antibiotics include amikacin, amoxicillin, ampicillin, atovaquone, azithromycin, aztreonam, bacitracin, carbenicillin, cefadroxil, cefazolin, cefdinir, cefditoren, cefepime, cefdirox, cefoperazone, cefotetan, cefoxitin, cefotaxime, cefpodoxime, cefprozil, ceftaroline, ceftazidime, ceftibuten, cefazolin, ceftriaxone, chloramphenicol, polymyxin E mesylate, cefuroxime, cephalexin, cephradine, cilastatin, cinoxacin, ciprofloxacin, clarithromycin, clindamycin, dalbavancin, dalfopristin, daptomycin, demeclocycline, dicloxacillin, doripenem, doxycycline, eracycline, ertacycline, , erythromycin, fidaxomicin, fosfomycin, gatifloxacin, gemifloxacin, gentamicin, imipenem, lefamulin, lincomycin, linezolid, lomefloxacin, loracarbef, meropenem, metronidazole, minocycline, moxifloxacin, nafcillin, nalidixic acid, neomycin, norfloxacin, ofloxacin, omadacycline, oritavancin, oxacillin, oxytetracycline, paromomycin, penicillin, pentamidine, piperacillin, prazomicin, quinupristin, rifaximin, sarrecycline, secnidazole, sparfloxacin, spectinomycin, sulfamethoxazole, sulfisoxazole, tedizolid, telavancin, telithromycin, ticarcillin, tigecycline, tobramycin, trimethoprim, trovafloxacin, and vancomycin.

[0468] Representative examples of antiviral agents include, but are not limited to, abacavir, acyclovir, adefovir, amantadine, amprenavir, atazanavir, balavir, baloxavir, boceprevir, cidofovir, cobicistat, daclatasvir, darunavir, delavirdine, didanosine, docasanol, dolutegravir, doravirine, ecoliever, edouridine, efavirenz, elvitegravir, emtricitabine, enfuvirtide, entecavir, etravirine, famciclovir, fomivirsen, fosamprenavir, forscarnet, fosnonet, famciclovir, favipiravir, fomivirsen, foscavir, ganciclovir, ibacitabine, iodine, indinavir, inosine, isoprenavir, type I interferon, type II interferon, type III interferon, lamivudine, lemavir , lemovir, lopinavir, loviramide, maraviroc, methinthiazone, morphine, nelfinavir, nevirapine, nitazoxanide, oseltamivir, peginterferon alfa-2a, peginterferon alfa-2b, penciclovir, peramivir, pleconaril, podophyllotoxin, pyramididine, raltegravir, remdesevir, ribavirin, rilpivirine, rimantadine, retamod, monoprevir, ritonavir, saquinavir, simeprevir, sofosbuvir, stavudine, tarabivirin, telaprevir, telbivudine, tenofovir alafenamide, tenofovir disoproxil, tenofovir, tipranvir, trifluridine, triamterene, tromantanamide, arbidol hydrochloride, valacyclovir, valganciclovir, adenosine, zalcitabine, zanamivir, and zidovudine.

[0469] Representative examples of anticoagulants include, but are not limited to, heparin, warfarin, rivaroxaban, dabigatran, apixaban, edoxaban, enoxaparin, and fondaparinux.

[0470] Representative examples of antiplatelet agents include, but are not limited to, clopidogrel, ticagrelor, prasugrel, dipyridamole, dipyridamole / aspirin, ticlopidine, and eptifibatide.

[0471] Representative examples of antifungal agents include, but are not limited to, voriconazole, itraconazole, posaconazole, fluconazole, ketoconazole, clotrimazole, isavuconazole, miconazole, caspofungin, anidulafungin, micafungin, griseofulvin, terbinafine, flucytosine, terbinafine, nystatin, and amphotericin b.

[0472] Representative examples of steroidal anti-inflammatory agents include, but are not limited to, hydrocortisone, dexamethasone, prednisolone, prednisone, triamcinolone, methylprednisolone, budesonide, betamethasone, cortisone, and deflazacort. Representative examples of nonsteroidal anti-inflammatory drugs include ibuprofen, naproxen, ketoprofen, tolmetin, etodolac, fenoprofen, flurbiprofen, diclofenac, piroxicam, indomethacin, sulindax, meloxicam, nabumetone, oxaprozin, mefenamic acid, and diflunisal.

[0473] Other examples of active agents include chloroquine, hydrochloroquine, pyridoxal phosphate, vitamin D, and vitamin C.

[0474] Representative examples of anti-cytokines or immunomodulators include, but are not limited to, tocilizumab, salinomycin, bevacizumab, fingolimod, imiquimod, and eculizumab.

[0475] Immunotherapeutic agents may include, but are not limited to, anti-CD40 antibodies, anti-PDL1 antibodies (e.g., atezolizumab, durvalumab, or avelumab), anti-PD1 antibodies, anti-CTLA4 antibodies, programmed death protein 1 (PD-1) inhibitors, or programmed death protein ligand 1 or 2 inhibitors including (e.g., nivolumab (BMS), pembrolizumab (Merck), pidilizumab (CureTech / Teva), AMP-244 (Amplimmune / GSK), BMS-936559 (BMS), and MEDI4736 (Roche / Genentech)), or combinations thereof.

[0476] Representative examples of contraceptives include, but are not limited to, progestogens, estrogens, or any combination thereof. For example, suitable progestogens include, but are not limited to, natural and synthetic compounds with progestational activity, such as, for example, progesterone, chlormadinone acetate, norgestrel, cyproterone acetate, norgestrel acetate, desogestrel, levonorgestrel, drospirenone, trimegestone, norgestrel, norgestimate, methylprogesterone, etonogestrel, gestodene, and other natural and / or synthetic progestogens. For example, suitable estrogens include, but are not limited to, natural and synthetic compounds with estrogenic activity, such as, for example, estradiol (17β-estradiol), 17α-estradiol, estriol, estrone and esters thereof, such as acetates, sulfates, valerates or benzoates of these compounds, including, for example, estradiol 17β-cypionate, estradiol 17-propionate, estradiol 3-benzoate and piperazine estrone sulfate; ethinyl estradiol; conjugated estrogens (natural and synthetic); mestranol; agonistic antiestrogens; and selective estrogen receptor modulators. Other examples of contraceptives include gonadotropin-releasing hormone (GnRh) or its analogs, such as deslorelin, avorelin, leuprorelin, triptorelin, nafarelin, goserelin, buserelin and fetirelin.

[0477] The term "steroid" refers to compounds belonging to or related to the following exemplary compound families: corticosteroids, mineral steroids, and sex steroids (including, for example, potential androgens or estrogens or anti-androgens and anti-estrogens). These include, for example, prednisone, prednisolone, methylprednisolone, triamcinolone, fluocinolone, aldosterone, spironolactone, danazol (also known as OPTINA), and others. In some embodiments, the therapeutic agent may include a steroid.

[0478] Exemplary cancer drugs or anticancer agents can include, but are not limited to, antimetabolite anticancer agents and antimitotic anticancer agents and combinations thereof. Various antimetabolite agents and antimitotic anticancer agents (including single such agents or combinations of such agents) can be used in the methods and compositions described herein.

[0479] Antimetabolite anticancer agents are usually similar in structure to natural metabolites, which participate in the normal metabolic processes of cancer cells, such as the synthesis of nucleic acids and proteins. However, antimetabolites are very different from natural metabolites, so that they interfere with the metabolic processes of cancer cells. In cells, antimetabolites are mistaken for their similar metabolites and are processed by cells in a manner similar to normal compounds. The presence of "bait" metabolites prevents cells from performing important functions, and cells cannot grow and survive. For example, antimetabolites can destroy cell division by replacing these fraudulent nucleotides into cell DNA, or by inhibiting key cell enzymes, preventing the replication of DNA, to exert cytotoxic activity.

[0480] Therefore, in one aspect, antimetabolite anticancer agent is nucleotide or nucleotide analog.In certain aspects, for example, antimetabolite can comprise purine (for example, guanine or adenosine) or its analog, or pyrimidine (cytidine or thymidine) or its analog, has or does not have the sugar part of connection.

[0481] Suitable antimetabolite anticancer agents for the present disclosure can generally be classified according to the metabolic processes they affect, and can include, but are not limited to, analogs and derivatives of folic acid, pyrimidine, purine, and cytidine. Thus, in one aspect, antimetabolites are selected from the group consisting of cytidine analogs, folic acid analogs, purine analogs, pyrimidine analogs, and combinations thereof.

[0482] In one particular aspect, for example, the antimetabolite is a cytidine analog. According to this aspect, for example, the cytidine analog can be selected from cytarabine (cytosine arabinoside), azacytidine (5-azacytidine), and salts, analogs, and derivatives thereof.

[0483] In another specific aspect, for example, antimetabolites are folic acid analogs. Folic acid analogs or antifolates generally work by inhibiting dihydrofolate reductase (DHFR), an enzyme involved in nucleotide formation; when this enzyme is blocked, nucleotides are not formed, thereby disrupting DNA replication and cell division. According to certain aspects, for example, folic acid analogs can be selected from the group consisting of normethopterin, methotrexate (methotrexate), pemetrexed, pteropterin, raltitrexed, trimesatide, and salts, analogs, and derivatives thereof.

[0484] In another specific aspect, for example, antimetabolites are purine analogs. Purine-based antimetabolites work by inhibiting DNA synthesis, for example, by interfering with the production of purines containing nucleotides, adenine and guanine, thereby stopping DNA synthesis and thus stopping cell division. Purine analogs can also be incorporated into the DNA molecule itself during DNA synthesis, which can interfere with cell division. According to certain aspects, for example, the purine analog can be selected from the group consisting of acyclovir, allopurinol, 2-aminoadenosine, arabinoadenine (ara-A), azacytidine, thiopurine, 8-aza-adenosine, 8-fluoro-adenosine, 8-methoxy-adenosine, 8-oxo-adenosine, cladribine, deoxycoformycin, fludarabine, ganciclovir, 8-aza-guanosine, 8-fluoro-guanosine, 8-methoxy-guanosine, 8-oxo-guanosine, guanosine diphosphate, guanosine diphosphate-β-L-2-aminofucose, guanosine diphosphate-D-arabinose, guanosine diphosphate-2-fluorofucose, guanosine diphosphate fucose, mercaptopurine (6-MP), pentostatin, thiopurine, thioguanine (6-TG) and salts, analogs and derivatives thereof.

[0485] In another specific aspect, for example, the antimetabolite is a pyrimidine analog. Similar to the purine analogs discussed above, pyrimidine-based antimetabolites block the synthesis of pyrimidine-containing nucleotides (cytosine and thymine in DNA; cytosine and uracil in RNA). By acting as a "bait", pyrimidine-based compounds can prevent the production of nucleotides and / or can be incorporated into the growing DNA chain and cause it to terminate. According to certain aspects, for example, pyrimidine analogs can be selected from ancitabine, azacytidine, 6-azauridine, bromouracil (e.g., 5-bromouracil), capecitabine, carmofur, chlorouracil (e.g., 5-chlorouracil), cytarabine (cytosine arabinoside), cytosine, dideoxyuridine, 3'-azido-3'-deoxythymidine, 3'-dideoxycytidine-2'-ene, 3'-deoxy-3'-deoxythymidine In one aspect, the pyrimidine analog is not 5-fluorouracil. In another aspect, the pyrimidine analog is gemcitabine or a salt thereof.

[0486] In certain aspects, the antimetabolite is selected from the group consisting of 5-fluorouracil, capecitabine, 6-mercaptopurine, methotrexate, gemcitabine, cytarabine, fludarabine, pemetrexed and salts thereof, analogs, derivatives and combinations thereof. In other aspects, the antimetabolite is selected from the group consisting of capecitabine, 6-mercaptopurine, methotrexate, gemcitabine, cytarabine, fludarabine, pemetrexed and salts thereof, analogs, derivatives and combinations thereof. In a specific aspect, the antimetabolite is not 5-fluorouracil. In particularly preferred aspects, the antimetabolite is gemcitabine or a salt thereof (e.g., gemcitabine hydrochloride). ).

[0487] Other antimetabolite anticancer agents may be selected from the group consisting of, but not limited to, acaciferic acid, aminothiadiazole, buquina sodium, Ciba-Geigy CGP-30694, cyclopentylcytosine, cytarabine phosphate stearate, cytarabine conjugate, Lilly DATHF, Merrel Dow DDFC, dezaguanine, dideoxycytidine, dideoxyguanosine, didox, Yoshitomi DMDC, Wellcome EHNA, Merck & Co. EX-015, falazine, fludarabine phosphate, N-(2′-furanyl)-5-fluorouracil, Daiichi Seiyaku FO-152, 5-FU-fibrinogen, isopropylpyrrolizine, Lilly LY-188011; Lilly LY-264618, meprofen, Wellcome MZPES, norspermidine, NCI NSC-127716, NCI NSC-264880, NCI NSC-39661, NCI NSC-612567, Warner-Lambert PALA, pentostatin, pyrimethoxazole, plicamycin, Asahi Chemical PL-AC, Takeda TAC-788, thiazofurine, Erbamont TIF, tyrosine kinase inhibitors, Taiho UFT and uricytin, etc.

[0488] In one aspect, anti-mitotic anticancer agent is a microtubule inhibitor or a microtubule stabilizer. In general, microtubule stabilizers, such as taxanes and epothilones, are combined with the inner surface of β-microtubule chains, and enhance microtubule assembly by promoting the nucleation and extension phase of polymerization and by reducing the critical tubulin subunit concentration required for microtubule assembly. Different from the microtubule inhibitors (such as vinca alkaloids) that hinder microtubule assembly, microtubule stabilizers (such as taxanes) reduce the lag time, and significantly shift the dynamic equilibrium between tubulin dimers and microtubule polymers to the polymerization direction. Therefore, in one aspect, microtubule stabilizers are taxanes or epothilones. In another aspect, microtubule inhibitors are vinca alkaloids.

[0489] In some embodiments, the anticancer agent may include a taxane or a derivative or analog thereof. Taxanes may be naturally derived compounds or related forms, or may be chemically synthesized compounds or derivatives thereof, having antitumor properties. Taxanes are a family of terpenoids, including but not limited to paclitaxel. and docetaxel They are primarily derived from the Pacific yew tree Taxus brevifolia and have activity against certain tumors, particularly breast and ovarian tumors. In one aspect, the taxane is docetaxel or paclitaxel. Paclitaxel is the preferred taxane and is considered an antimitotic agent that promotes assembly of microtubules from tubulin dimers and stabilizes microtubules by preventing depolymerization. This stabilization results in inhibition of the normal dynamic reorganization of the microtubule network, which is essential for interphase and mitotic cell function.

[0490] Also included are various known taxane derivatives, including hydrophilic derivatives and hydrophobic derivatives. Taxane derivatives include, but are not limited to, galactose and mannose derivatives described in International Patent Application No. WO 99 / 18113; piperazine impurities and other derivatives described in WO 99 / 14209; taxane derivatives described in WO 99 / 09021, WO 98 / 22451 and U.S. Pat. No. 5,869,680; 6-thio derivatives described in WO 98 / 28288; sulfonamide derivatives described in U.S. Pat. No. 5,821,263; deoxytaxel compounds, such as those described in U.S. Pat. No. 5,440,056; and paclitaxel derivatives described in U.S. Pat. No. 5,415,869. As described above, it also includes prodrugs of paclitaxel, including, but not limited to, those described in WO 98 / 58927; WO 98 / 13059; and U.S. Pat. No. 5,824,701. The taxane may also be a taxane conjugate, such as, for example, paclitaxel-PEG, paclitaxel-dextran, paclitaxel-xylose, docetaxel-PEG, docetaxel-dextran, docetaxel-xylose, etc. Other derivatives are mentioned in references such as "Synthesis and Anticancer Activity of Taxol Derivatives," DGI Kingston et al., Studies in Organic Chemistry, vol. 26, entitled "New Trends in Natural Products Chemistry" (1986), Atta-ur-Rabman, PWle Quesne, Eds. (Elsevier, Amsterdam 1986). Each of these references is hereby incorporated by reference in its entirety.

[0491] Various taxanes can be readily prepared using techniques known to those skilled in the art (see also WO 94 / 07882, WO 94 / 07881, WO 94 / 07880, WO 94 / 07876, WO 93 / 23555, WO 93 / 10076; U.S. Pat. Nos. 5,294,637; 5,283,253; 5,279,949; 5,274,137; 5,202,448; 5,200,534; 5,229,529; and EP 590,267) (each of which is hereby incorporated by reference in its entirety), or obtained from a variety of commercial sources, including, for example, Sigma-Aldrich Co., St. Louis, Mo.

[0492] Alternatively, the anti-mitotic anticancer agent can be a microtubule inhibitor; in a preferred aspect, the microtubule inhibitor is a vinca alkaloid. Generally speaking, vinca alkaloids are mitotic spindle poisons. Vinca alkaloid agents work during mitosis, when chromosomes divide and begin to migrate along the tubules of the mitotic spindle to one of its poles before cell separation. Under the action of these spindle poisons, the spindle becomes disordered due to the dispersion of chromosomes during mitosis, affecting cell reproduction. According to certain aspects, for example, vinca alkaloids are selected from the group consisting of vinblastine, vincristine, vindesine, vinorelbine and its salts, analogs and derivatives.

[0493] Anti-mitotic anticancer agents can also be epothilones. In general, members of the epothilone class of compounds stabilize microtubule function according to a mechanism similar to that of the taxanes. Epothilones can also cause cell cycle arrest at the G2-M transition, leading to cytotoxicity and eventual apoptosis. Suitable epithiones include epothilone A, epothilone B, epothilone C, epothilone D, epothilone E, and epothilone F and their salts, analogs, and derivatives. A specific epothilone analog is the epothilone B analog ixabepilone (Ixempra TM ).

[0494] In certain aspects, the anti-mitotic anticancer agent is selected from the group consisting of taxanes, epothilones, vinca alkaloids, and salts and combinations thereof. Thus, for example, in one aspect, the anti-mitotic agent is a taxane. In this regard, more preferably, the anti-mitotic agent is paclitaxel or docetaxel, still more preferably paclitaxel. On the other hand, the anti-mitotic agent is an epothilone (e.g., an epothilone B analog). On the other hand, the anti-mitotic agent is a vinca alkaloid.

[0495] Examples of cancer drugs that can be used in the present disclosure include, but are not limited to, thalidomide; platinum coordination complexes, such as cisplatin (cis-DDP), oxaliplatin, and carboplatin; anthracenediones, such as mitoxantrone; substituted ureas, such as hydroxyurea; methylhydrazine derivatives, such as methylbencarbazine (N-methylhydrazine, MIH); adrenocortical suppressants, such as mitotane (o,p′-DDD) and aminoglutethimide; RXR agonists, such as bexarotene; and tyrosine kinase inhibitors, such as sunitinib and imatinib.

[0496] Examples of additional anticancer drugs include alkylating agents, antimetabolites, natural products, hormones and antagonists, and other drugs. Alternative names are indicated in brackets. Examples of alkylating agents include nitrogen mustards, such as dichloromethyldiethylamine, cyclophosphamide, ifosfamide, melphalan (melphalan) and chlorambucil; ethyleneimine and methylmelamine, such as hexamethylmelamine and thiotepa; alkyl sulfonates, such as busulfan; nitrosoureas, such as carmustine (BCNU), semustine (methyl-CCNU), lomustine (CCNU) and streptozotocin (streptozotocin); DNA synthesis antagonists, such as estramustine phosphate; and triazines, such as dacarbazine (DTIC, dimethyl-triazaimidazole carboxamide) and temozolomide. Examples of antimetabolites include folic acid analogs such as methotrexate (methotrexate); pyrimidine analogs such as fluorouracil (5-fluorouracil, 5-FU, SFU), floxuridine (fluorodeoxyuridine, FUdR), cytarabine (cytosine arabinoside) and gemcitabine; purine analogs such as mercaptopurine (6-mercaptopurine, 6-MP), thioguanine (6-thioguanine, TG) and pentostatin (2′-deoxycoformycin, deoxycoformycin), cladribine and fludarabine; and topoisomerase inhibitors such as amsacrine. Examples of natural products include vinca alkaloids, such as vinblastine (VLB) and vincristine; taxanes, such as paclitaxel, protein-bound paclitaxel (Abraxane), and docetaxel (Taxotere); epipodophyllotoxins, e.g., etoposide and teniposide; camptothecins, such as topotecan and irinotecan; antibiotics, such as dactinomycin (actinomycin D), daunomycin (daunomycin, daunomycin), doxorubicin, histrelin, bleomycin, mitomycin (mitomycin C), idarubicin, epirubicin; enzymes, such as L-asparaginase; and biological response modifiers, such as interferon alpha and interleukin 2. Examples of hormones and antagonists include luteinizing hormone-releasing hormone agonists, such as buserelin; adrenocortical steroids, such as prednisone and related preparations; progestins, such as hydroxyprogesterone caproate, medroxyprogesterone acetate, and megestrol acetate; estrogens, such as diethylstilbestrol and ethinyl estradiol and related preparations; estrogen antagonists, such as tamoxifen and anastrozole; androgens, such as testosterone propionate and fluoxymesterone and related preparations; androgen antagonists, such as flutamide and bicalutamide; and gonadotropin-releasing hormone analogs such as leuprolide. Alternative names and trade names for these and other examples of cancer drugs, as well as methods of their use (including dosing and administration regimens) will be known to those skilled in the art.

[0497] In some aspects, anticancer agents may include chemotherapeutic agents. Suitable chemotherapeutic agents include, but are not limited to, alkylating agents, antibiotic agents, antimetabolites, hormone agents, plant-derived agents and synthetic derivatives thereof, anti-angiogenic agents, differentiation inducers, cell growth arrest inducers, apoptosis inducers, cytotoxic agents, agents that affect cell bioenergetics (i.e., agents that affect cell ATP levels and molecules / activities that regulate these levels), biological agents (e.g., monoclonal antibodies, kinase inhibitors, and inhibitors of growth factors and their receptors), gene therapy agents, cell therapies (e.g., stem cells), or any combination thereof.

[0498] According to these aspects, the chemotherapeutic agent is selected from the group consisting of cyclophosphamide, chlorambucil, melphalan, dichloromethyl diethylamine, ifosfamide, busulfan, lomustine, streptozotocin, temozolomide, dacarbazine, cisplatin, carboplatin, oxaliplatin, procarbazine, uramustine, methotrexate, pemetrexed, fludarabine, cytarabine, fluorouracil, floxuridine, gemcitabine, capecitabine, vinblastine, vincristine, vinorelbine, etoposide, paclitaxel, docetaxel, doxorubicin, daunorubicin, epirubicin, idarubicin, mitoxantrone, bleomycin, mitomycin, hydroxyurea, topotecan, irinotecan, amsacrine, teniposide, erlotinib hydrochloride and combinations thereof. Each possibility represents an independent aspect of the present invention.

[0499] The antineoplastic agent may be selected from the group consisting of abiraterone acetate, Abitrexate (methotrexate), Abraxane (paclitaxel albumin-stabilized nanoparticle formulation), ABVD, ABVE, ABVE-PC, AC, AC-T, Adcetris (brituximab), ADE, Ado-trastuzumab-emtansine, doxorubicin (doxorubicin hydrochloride), Adrucil (fluorouracil), afatinib bismaleate, Afinitor (everolimus), Akynzeo (netupitant and palonosetron hydrochloride), Aldara (imiquimod), aldesleukin, alemtuzumab, Eninda (pemetrexed disodium), Aloxi (palonosetron hydrochloride), ), Ambochlorin (chlorambucil), Amboclorin (chlorambucil), aminolevulinic acid, Anastrozole, Aprepitant, Acorda (pamidronate disodium), Arimidex (anastrozole), Aromasin (exemestane), Arranon (nelarabine), Arsenic trioxide, Arzerra (ofatumumab), Asparaginase Erwinia chrysanthemi, Avastin (bevacizumab), Axitinib, Azacitidine, BEACOPP, Becenum (carmustine), Beleodaq (belinostat), belinostat, bendamustine hydrochloride, BEP, bevacizumab, bexarotene, Bexxar (tositumomab and iodine I 131 tositumomab), bicalutamide, BiCNU (carmustine), bleomycin, blinatumomab, Blincyto (blinatumomab), bortezomib, Bosulif (bosutinib), bosutinib, vebutinib, busulfan, busulfan, cabazitaxel, cabozantinib-S-malate, CAF, Campath (alemtuzumab), Camptosar (irinotecan hydrochloride), capecitabine, CAPOX, carboplatin, carboplatin-paclitaxel, carfilzomib, Carmubris (carmustine), carmustine, carmustine implant, Casodex (bicalutamide), CeeNU (lomustine), ceritinib, Cerubidine (doxorubidine hydrochloride) chlorambucil, chlorambucil-prednisone, CHOP, cisplatin, Clafen (cyclophosphamide), clofarabine, Clofarex (clofarabine), Clolar (clofarabine), CMF, Cometriq (cabozantinib-S-malate), COPP, COPP-ABV, Cosmegen (dactinomycin), crizotinib, CVP, cyclophosphamide, Cyfos (ifosfamide), Cyramza (ramucirumab), cytarabine, cytarabine, liposomal, Cytosar-U (cytarabine), Cytoxan (cyclophosphamide), dabrafenib, dacarbazine,Dacogen (decitabine), dactinomycin, dasatinib, daunomycin hydrochloride, decitabine, degarelix, dasatinib, denileukin, denosumab, DepoCyt (liposomal cytarabine), DepoFoam (liposomal cytarabine), dexrazoxane hydrochloride, denutuximab, docetaxel, doxorubicin (liposomal doxorubicin hydrochloride), doxorubicin hydrochloride, doxorubicin hydrochloride liposomal, Dox-SL (liposomal doxorubicin hydrochloride), doxorubicin hydrochloride, doxorubicin hydrochloride liposomal, Dox-SL (liposomal doxorubicin hydrochloride), DTIC-Dome (dacarbazine), Efudex (fluorouracil), Elitek (rasburicase), Ellence (epirubicin hydrochloride), Eloxatin (oxaliplatin), eltrombopag, Emend (afrapitant), Enzalutamide, epirubicin hydrochloride, EPOCH, Erbitux (cetuximab), Eribulin mesylate, Erivedge (vismodegib), erlotinib hydrochloride, Erwinaze (asparaginase Erwinia chrysanthemi), Etopophos (etoposide phosphate), etoposide, etoposide phosphate, Evacet (liposomal doxorubicin hydrochloride), everolimus, Evista (raloxifene hydrochloride), exemestane, Fareson (toremifene), Farydak (panobinostat), Faslodex (fulvestrant), FEC, Femara (letrozole), Filgrastim, Fludara (fludarabine phosphate), fludarabine phosphate, Fluoroplex (fluorouracil), fluorouracil, Folex (methotrexate), Folex PFS (methotrexate), FOLFIRI, FOLFIRI-bevacizumab, FOLFIRI-cetuximab, FOLFIRINOX, FOLFOX, Folotyn (pralatrexate), FU-LV, fulvestrant, Gardasil (recombinant HPV quadrivalent vaccine), Gardasil 9 (recombinant HPV nine-valent vaccine), Gazyva (otuzumab), gefitinib, gemcitabine hydrochloride, gemcitabine-cisplatin, gemcitabine-oxaliplatin, gemtuzumab ozagamin, Gemzar (gemcitabine hydrochloride), Gilotrif (afatinib bismaleate), Gleevec (imatinib mesylate), Gliadel (carmustine implant), Gliadel wafer (carmustine implant), glutaric acid, goserelin acetate, Halaven (eribulin mesylate), Herceptin (trastuzumab), HPV bivalent vaccine, recombinant, HPV nine-valent vaccine, recombinant, HPV four-valent vaccine, recombinant, Hycamtin (topotecan hydrochloride), Hyper-CVAD, Ibrance (palbociclib), ibrutinib, ibrutinib, ICE, Iclusig (panatinib hydrochloride),Idamycin (idarubicin hydrochloride), idarubicin hydrochloride, idelalisib, Ifex (ifosfamide), ifosfamide, Ifosfamidum (ifosfamide), imatinib mesylate, Imbruvica (ibrutinib), imiquimod, Inlyta (axitinib), interferon alfa-2b, recombinant, intron A (recombinant interferon alfa-2b), iodine I 131 tositumomab and tositumomab, ipilimumab, Iressa (gefitinib), irinotecan hydrochloride, Istodax (romidep), ixabepilone, Ixempra (ixabepilone), Jakafi (ruxolitinib phosphate), Jevtana (cabazitaxel), Kadcyla (Ado-trastuzumab-emtansine), Keoxifene (raloxifene hydrochloride), Kepivance (palifermin), Keytruda (pembrolizumab), Kyp rolis (carfilzomib), lanreotide acetate, lapatinib ditosylate, lenalidomide, lenvatinib mesylate, Lenvima (lenvatinib mesylate), letrozole, folinic acid, Leukeran (chlorambucil), leuprolide acetate, Levulan (aminolevulinic acid), Linfolizin (chlorambucil), LipoDox (liposomal doxorubicin hydrochloride), liposomal cytarabine, lomustine, Lupron (leuprolide acetate), Lupron Depot (leuprolide acetate), Lupron Depot-Ped (leuprolide acetate), Lupron Depot-3Month (leuprolide acetate), Lupron Depot-4 Month (leuprolide acetate), Lynparza (olaparib), Marqibo (liposomal vincristine sulfate), Matulane (procarbazine hydrochloride), Nitrogen mustard hydrochloride, Megace (megestrol acetate), megestrol acetate, Mekinist (trametinib), Mercaptopurine, Mesna, Mesnex (mesna), Methazolastone (temozolomide), Methotrexate, Methotrexate LPF (methotrexate), Mexate (methotrexate), Mexate-AQ (methotrexate), Mitomycin C, Mitoxantrone hydrochloride, Mitozytrex (mitomycin C), MOPP, Mozobil (plerixafor), Mustargen (mechlorethamine hydrochloride), Mutamycin (mitomycin C), Myleran (busulfan), Mylosar (azacitidine), Mylotarg (gemtuzumab ozagamin), Nanoparticulate Paclitaxel (albumin-stabilized nanoparticle formulation of paclitaxel), Navelbine (vinorelbine tartrate), Nelarabine, Neosar (cyclophosphamide), Netupitant and Palonosetron hydrochloride,Neupogen (filgrastim), Nexavar (sorafenib tosylate), nilotinib, nivolumab, Nolvadex (tamoxifen citrate), Nplate (romiplostim), obotuzumab, Odomzo (sonidegi), OEPA, ofatumumab, OFF, olaparib, homoharringtonine, Oncaspar (pegapagastatin), ondansetron hydrochloride, Ontak (denileukin), Opdivo (nivolumab), OPPA, oxaliplatin, paclitaxel, taxane Albumin-stabilized nanoparticle formulation, PAD, palbociclib, palifermin, palonosetron hydrochloride, palonosetron hydrochloride and netupitant, pamidronate disodium, panitumumab, panobinostat, Paraplat (carboplatin), Paraplatin (carboplatin), pazopanib hydrochloride, pegapagastat, peginterferon alfa-2b, PEG-Intron (pegylated interferon alfa-2b), pembrolizumab, pemetrexed disodium, Perjeta (pertuzumab), pertuzumab, Platinol (cisplatin) , Platinol-AQ (cisplatin), Plerixafor, Pomalidomide, Pomalyst (pomalidomide), Ponatinib hydrochloride, Pralatrexate, Prednisone, Procarbazine hydrochloride, Proleukin (aldesleukin), Prolia (denosumab), Promacta (eltrombopag), Provenge (Cipruset-T), Purinethol (mercaptopurine), Purixan (mercaptopurine), Radium 223 dichloride, Raloxifene hydrochloride, Ramucirumab, Rasburicase, R- CHOP, R-CVP, recombinant human papillomavirus (HPV) bivalent vaccine, recombinant human papillomavirus (HPV) nine-valent vaccine, recombinant human papillomavirus (HPV) quadrivalent vaccine, recombinant interferon α-2b, regorafenib, R-EPOCH, Revlimid (lenalidomide), Rheumatrex (methotrexate), Rituxan (rituximab), rituximab, romidepsip, romiplostim, Rubidomycin (daunomycin hydrochloride), ruxolitinib phosphate, Sclerosol Intrapleural Aerosol (talc), Siltuximab, Siprussin-T, Somatuline Depot (lanreotide acetate), Sonidegi, Sorafenib Tosylate, Sprycel (dasatinib), STANFORD V, Sterile Talcum (talc), Steritalc (talc), Stivarga (regorafenib), Sunitinib Malate, Sutent (sunitinib malate), Sylatron (peginterferon alfa-2b), Sylvant (silotuximab), Synovir (thalidomide), Synribo (homoharringtonine), TAC, Tafinlar (dabrafenib), Talc,Tamoxifen citrate, Tarabine PFS (cytarabine), Tarceva (erlotinib hydrochloride), Targretin (bexarotene), Tasigna (nilotinib), Taxol (paclitaxel), Taxotere (docetaxel), Temodar (temozolomide), Temozolomide, Temsirolimus, Thalidomide, Thalomid (thalidomide), Thiotepa, Toposar (etoposide), Topotecan hydrochloride, Toremifene, Torisel (temsirolimus), Tositumomab and Iodine I131 Tositumomab, Totect (dexamfetamine hydrochloride), TPF, Trametinib, Trastuzumab, Treanda (bendamustine hydrochloride), Trisenox (arsenic trioxide), Tykerb (lapatinib ditosylate), Unituxin (denosumab), vandetanib, VAMP, Vectibix (panitumumab), VeIP, Velban (vinblastine sulfate), Velcade (bortezomib), Velsar (vinblastine sulfate), Vemurafenib, VePesid (etoposide), Viadur (leuprorelin acetate), Vidaza (azacitidine), vinblastine sulfate, Vincasar PFS (vincristine sulfate), vincristine sulfate, liposomal vincristine sulfate, vinorelbine tartrate, VIP, vismodegib, Voraxaze (glutaric acid), vorinostat, Votrient (pazopanib hydrochloride), Wellcovorin (leucovorin calcium), Xalkori (crizotinib), Xeloda (capecitabine), XELIRI, XELOX, Xgeva (denosumab), Xofigo (radium 223 dichloride), Xtandi (enzalutamide), Yervo y (ipilimumab), Zaltrap (ziv-aflibercept), Zelboraf (vemurafenib), Zevalin (ibritumomab tiuxetan), Zinecard (dexrazoxane hydrochloride), Ziv-aflibercept, Zofran (ondansetron hydrochloride), Zoladex (goserelin acetate), zoledronic acid, Zolinza (vorinostat), Zometa (zoledronic acid), Zydelig (idelalisib), Zykadia (ceretinib), and Zytiga (abiraterone acetate).

[0500] Growth factors that can be used as therapeutic agents include, but are not limited to, transforming growth factor-α ("TGF-α"), transforming growth factor ("TGF-β"), platelet-derived growth factor ("PDGF"), fibroblast growth factor ("FGF") (including FGF acidic isoforms 1 and 2), FGF basic form 2 and FGF 4, 8, 9 and 10, nerve growth factor ("NGF") (including NGF 2.5s, NGF7.0s and βNGF and neurotrophic factors), brain-derived neurotrophic factor, cartilage-derived factor, bone growth factor (BGF), basic fibroblast growth factor, insulin-like growth factor (IGF), vascular endothelial growth factor (VEGF), granulocyte colony-stimulating factor (G-CSF), insulin-like growth factor (IGF) I and II, hepatocyte growth factor, glial neurotrophic growth factor (GDNF), stem cell factor (SCF), keratinocyte growth factor (KGF), transforming growth factor (TGF) (including TGFα, β, β1, β2, β3) bone growth factor, bone matrix-derived growth factor and bone-derived growth factor, and mixtures thereof.

[0501] Immunoglobulins useful in the present disclosure include, but are not limited to, IgG, IgA, IgM, IgD, IgE, and mixtures thereof. Some preferred growth factors include VEGF (vascular endothelial growth factor), NGF (nerve growth factor), PDGF-AA, PDGF-BB, PDGF-AB, FGFb, FGFa, and BGF.

[0502] Other molecules that can be used as anticancer agents include, but are not limited to, growth hormone, leptin, leukemia inhibitory factor (LIF), tumor necrosis factor alpha and beta, endostatin, thrombospondin, osteogenic protein-1, bone morphogenetic protein 2 and 7, osteonectin, somatomedin-like peptide, osteocalcin.

[0503] Tumor antigens can be based on specific mutations (neo-epitopes) and those mutations expressed by cancer germline genes (antigens common to tumors found in multiple patients, referred to herein as "traditional cancer antigens" or "shared cancer antigens"). In some embodiments, traditional antigens are antigens known to be prevalent in cancer or tumors or in a specific type of cancer or tumor. In some embodiments, traditional cancer antigens are non-mutated tumor antigens. In some embodiments, traditional cancer antigens are mutated tumor antigens.

[0504] Diagnostic agents include gases; metals; commercially available imaging agents for positron emission tomography (PET), computer-assisted tomography (CAT), single photon emission computerized tomography, x-ray, fluoroscopy, and magnetic resonance imaging (MRI); and contrast agents. Examples of suitable materials for use as contrast agents in MRI include gadolinium chelates, as well as iron, magnesium, manganese, copper, and chromium. Examples of materials that can be used for CAT and x-ray imaging include iodine-based materials.

[0505] Vaccines may include isolated proteins or peptides, inactivated organisms and viruses, dead organisms and viruses, genetically altered organisms or viruses, cell extracts, and RNA encoding at least one antigenic polypeptide or its immunogenic fragment (e.g., an immunogenic fragment capable of inducing an immune response to the antigenic polypeptide). Active agents may be combined with interleukins, interferons, cytokines, and adjuvants such as cholera toxin, alum, Freund's adjuvant, etc. Prophylactic agents may include infectious agents such as antigens of bacterial organisms, which are such as Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, Corynebacterium diphtheriae, Listeria monocytogenes, Bacillus anthracis, Clostridium tetani, Clostridium botulinum, Clostridium perfringens, Neisseria meningitidis, Neisseria gonorrhoeae, Streptococcus mutans, Pseudomonas aeruginosa, Salmonella typhi, Haemophilus parainfluenzae, Bordetella pertussis, Francisella tularensis, Yersinia pestis, Vibrio cholerae, Legionella pneumophila, Mycobacterium tuberculosis, Mycobacterium leprae, Treponema pallidum, Leptospira interrogans, Borrelia burgdorferi, Campylobacter jejuni, etc.; Antigens of viruses, such as human metapneumovirus (hMPV), human parainfluenza virus (hPIV) types 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively), respiratory syncytial virus (RSV), measles virus (MeV), coronaviruses (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1), poxviruses (e.g., smallpox, monkeypox), African swine fever virus, influenza A and B, HIV, varicella-zoster, herpes simplex 1 and 2, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps, rabies, rubella, Coxsackie virus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E viruses, etc.; antigens of fungi, protozoa and parasitic organisms, such as Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans, Candida tropicalis, Nocardia asteroides, Rickettsia rickettsii, Rickettsia typhi, Mycoplasma pneumoniae, Chlamydia psittaci, Chlamydia trachomatis, Plasmodium falciparum, Trypanosoma brucei, Entamoeba histolytica, Toxoplasma gondii, Trichomonas vaginalis, Schistosoma mansoni, etc. These antigens can be in the form of whole killed organisms, peptides, proteins, glycoproteins, carbohydrates, or combinations thereof.

[0506] In certain embodiments, the activating agent is a polynucleotide. The polynucleotides or oligonucleotides that can be introduced according to the method herein include all types of DNA, cDNA and RNA sequences. For example, polynucleotides can be double-stranded DNA, single-stranded DNA, composite DNA, encapsulated DNA, naked RNA, encapsulated RNA, messenger RNA (mRNA), tRNA, short interfering RNA (siRNA), double-stranded RNA (dsRNA), microRNA (miRNA), antisense RNA (asRNA), self-amplification mRNA (saRNA), guide RNA (gRNA), cRNA, and combinations thereof. Polynucleotides can also be DNA constructs, such as expression mediators, encoding desired gene products (e.g., homologous or heterologous gene products to be introduced with the subject) expression mediators etc.

[0507] In some embodiments, RNA (e.g., mRNA) can be used to induce a balanced immune response against an infectious agent. In some embodiments, RNA (e.g., mRNA) can be used to induce a balanced immune response against a virus, such as a metapneumovirus such as human metapneumovirus (hMPV), a parainfluenza virus such as human parainfluenza virus (hPIV) 1, 2, and 3 (hPIV1, hPIV2, and hPIV3, respectively), respiratory syncytial virus (RSV), measles virus (MeV), coronaviruses (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-29E, HCoV-390, HCoV-406, HCoV-414, HCoV-429E, HCoV-434, HCoV-435, HCoV-436, HCoV-437, HCoV-438, HCoV-439 ... V-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1), poxvirus (e.g., smallpox, monkeypox), African swine fever virus, influenza A and B, HIV, varicella-zoster, herpes simplex 1 and 2, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps, rabies, rubella, coxsackievirus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E virus, etc. In some embodiments, RNA (e.g., mRNA) can be used to induce a balanced immune response. In some embodiments, RNA (e.g., mRNA) can be used to induce a balanced immune response against respiratory viruses. The term "respiratory virus" refers to a virus that causes a respiratory disease in this article. For example, negative sense single stranded RNA viruses of the family Paramyxoviridae, such as human metapneumovirus (hMPV), human parainfluenza virus (hPIV) types 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively), RSV and measles virus (MeV). Another example of a respiratory virus is a coronavirus. Metapneumovirus is an enveloped virus with a positive sense single stranded RNA genome and a helical symmetrical nucleocapsid. Coronavirus is a virus species belonging to the subfamily Coronavirus of the family Coronaviridae of the order Nidovirales.

[0508] Representative examples of betacoronaviruses include, but are not limited to, Erbecon coronavirus 1 (e.g., betacoronavirus 1, human coronavirus OC43, Chinese mouse coronavirus HKU24, human coronavirus HKU1, mouse coronavirus), Haibei virus (e.g., bat Hp-betacoronavirus Zhejiang 2013), Meibei virus (e.g., hedgehog coronavirus 1, Middle East respiratory syndrome-related coronavirus (MERS-CoV), scutellaria bat coronavirus HKU5, flat-headed bat coronavirus HKU4), Norbe virus (e.g., fruit bat coronavirus GCCDC1, fruit bat coronavirus HKU9), Shell coronavirus (e.g., severe acute respiratory syndrome coronavirus (SARS-CoV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

[0509] Representative examples of gammacoronaviruses include, but are not limited to, cegacovirus (eg, beluga coronavirus SQ1) and Igacovirus (eg, avian coronavirus (IBV)).

[0510] Representative examples of delta coronaviruses include, but are not limited to, andecovirus (e.g., wild duck coronavirus HKU20), buldecovirus (e.g., nightingale coronavirus HKU11, porcine coronavirus HKU15 (PorCoV HKU15), munia subcoronavirus HKU13, white-eye coronavirus HKU16), herdecovirus (e.g., night heron coronavirus HKU19), and moordecovirus (e.g., common swamphen coronavirus HKU21).

[0511] In some embodiments, the coronavirus is a human coronavirus. Representative examples of human coronaviruses include, but are not limited to, human coronavirus 229E (HCoV-229E), human coronavirus OC43 (HCoV-OC43), human coronavirus HKU1 (HCoV-HKU1), human coronavirus NL63 (HCoV-NL63), severe acute respiratory syndrome coronavirus (SARS-CoV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and Middle East respiratory syndrome-related coronavirus (MERS-CoV).

[0512] In some embodiments, the RNA (e.g., mRNA) polynucleotide having an open reading frame encodes at least one (e.g., at least 2, 3, 4, or 5) human metapneumovirus (hMPV) antigenic polypeptide, human parainfluenza virus (hPIV) types 1, 2, and 3 (hPIV1, hPIV2, and hPIV3, respectively) antigenic polypeptide, respiratory syncytial virus (RSV) antigenic polypeptide, measles virus (MeV) antigenic polypeptide, varicella-zoster virus antigenic polypeptide, influenza virus antigenic polypeptide, herpes simplex virus 1 (HSV1) antigenic polypeptide, herpes simplex virus 2 (HSV2) antigenic polypeptide, poxvirus (e.g., smallpox, monkeypox) antigenic polypeptide, African swine fever virus antigenic polypeptide, cytomegalovirus antigenic polypeptide, Epste in-Barr virus antigen polypeptides, rotavirus antigen polypeptides, rhinovirus antigen polypeptides, adenovirus antigen polypeptides, papillomavirus antigen polypeptides, poliovirus antigen polypeptides, mumps antigen polypeptides, rabies antigen polypeptides, rubella antigen polypeptides, coxsackievirus antigen polypeptides, equine encephalitis antigen polypeptides, Japanese encephalitis antigen polypeptides, yellow fever antigen polypeptides, Rift Valley fever antigen polypeptides, hepatitis A, B, C, D and E virus antigen polypeptides, or coronavirus (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1) antigen polypeptides. As used herein, the term "antigenic polypeptide" encompasses immunogenic fragments of an antigenic polypeptide that induce (or are capable of inducing) an immune response to the following viruses: human metapneumovirus (hMPV), human parainfluenza virus (hPIV) types 1, 2, and 3 (hPIV1, hPIV2, and hPIV3, respectively), respiratory syncytial virus (RSV), measles virus (MeV), varicella-zoster, influenza virus, herpes simplex virus type 1 (HSV1), herpes simplex virus type 2 (HSV2), poxviruses (e.g., smallpox, monkeypox), African swine fever virus, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps, rabies, rubella, coxsackievirus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E viruses or coronaviruses (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1), or any combination thereof.

[0513] In some embodiments, for example, the agent is RNA (e.g., mRNA) that can induce a balanced immune response to hMPV, PIV, RSV, MeV and / or coronavirus (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH and / or HCoV-HKU1), or any combination of two or more of the foregoing viruses, including both cellular and humoral immunity, without the risk of the possibility of insertional mutations.

[0514] In some embodiments, the nucleic acids disclosed herein may include at least one chemically modified nucleotide. In some embodiments, at least one chemically modified nucleotide includes a chemically modified nucleobase, a chemically modified ribose, a chemically modified phosphodiester bond, or a combination thereof.

[0515] In one embodiment, the at least one chemically modified nucleotide is a chemically modified nucleobase.

[0516] In one embodiment, the chemically modified nucleobase is selected from 5-formylcytidine (5fC), 5-methylcytidine (5meC), 5-methoxycytidine (5moC), 5-hydroxycytidine (5hoC), 5-hydroxymethylcytidine (5hmC), 5-formyluridine (5fU), 5-methyluridine (5-meU), 5-methoxyuridine (5moU), 5-carboxymethyl ester uridine (5camU), pseudouridine (Ψ), N 1 -methyl pseudouridine (me 1 Ψ), N 6 -methyladenosine (me 6 A) or tenoguanosine ( th G).

[0517] In some embodiments, the chemically modified nucleobase is 5-methoxyuridine (5moU). In some embodiments, the chemically modified nucleobase is pseudouridine (Ψ). In some embodiments, the chemically modified nucleobase is N 1 -methyl pseudouridine (me 1 Ψ).

[0518] The structures of these modified nucleobases are shown below:

[0519]

[0520] In one embodiment, at least one chemically modified nucleotide is a chemically modified ribose.

[0521] In one embodiment, the chemically modified ribose is selected from 2'-O-methyl (2'-O-Me), 2'-fluoro (2'-F), 2'-deoxy-2'-fluoro-β-D-arabinoic acid (2'F-ANA), 4'-S, 4'-SFANA, 2'-azido, UNA, 2'-O-methoxy-ethyl (2'-O-ME), 2'-O-allyl, 2'-O-ethylamine, 2'-O-cyanoethyl, locked nucleic acid (LAN), methylene-cLAN, N-MeO-amino BNA or N-MeO-aminooxy BNA. In one embodiment, the chemically modified ribose is 2'-O-methyl (2'-O-Me). In one embodiment, the chemically modified ribose is 2'-fluoro (2'-F).

[0522] The structures of these modified ribose sugars are shown below:

[0523]

[0524] In one embodiment, at least one chemically modified nucleotide is a chemically modified phosphodiester bond.

[0525] In one embodiment, the chemically modified phosphodiester bond is selected from phosphorothioate (PS), borophosphate, phosphorodithioate (PS2), 3',5'-amide, N3'-phosphoramide (NP), phosphodiester (PO) or 2',5'-phosphodiester (2',5'-PO). In one embodiment, the chemically modified phosphodiester bond is phosphorothioate.

[0526] The structures of these modified phosphodiester bonds are shown below:

[0527]

[0528] In some embodiments, the mRNA may include a heterologous 5' untranslated region (5'UTR). In some embodiments, the mRNA may include a heterologous 3' untranslated region (3'UTR).

[0529] method

[0530] The composition of the present invention can be used as a composition for diagnosis, prevention and treatment of diseases or disorders (e.g., infection, cancer, pain, depression, inflammatory diseases, intestinal diseases, brain diseases, allergic diseases, arrhythmias, hypertension, eye diseases, endocrine diseases or cardiovascular diseases), depending on the type of active agent.

[0531] Described herein are methods for delivering an active agent (e.g., a polynucleotide) to an organ, tissue, and / or cell, the method comprising: introducing into the organ, tissue, and / or cell a composition or nanoparticle described herein, the composition or nanoparticle comprising: a compound of formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0532] Described herein are methods for diagnosing, treating, or preventing a disease, the method comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle described herein, the composition or nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0533] Described herein are methods for treating or preventing an infection, the method comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle described herein, the composition or nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0534] Described herein are methods for treating or preventing respiratory infections, comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle described herein, the composition or nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0535] Described herein are methods of inducing an immune response against a virus, the method comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle described herein, the composition or nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0536] Described herein are methods of inducing an immune response against a respiratory virus, the method comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle comprising a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0537] In some embodiments, provided herein are methods for delivering polynucleotides. In some embodiments, provided herein are methods for delivering polynucleotides (e.g., mRNA) to provide expression (and translation to produce protein) of polynucleotides in cells. In some embodiments, provided herein are methods for delivering polynucleotides (e.g., mRNA) to induce an immune response in a subject.

[0538] In some embodiments, the method for delivering an active agent can introduce a polynucleotide into a cell or tissue to express a protein encoded by the polynucleotide in vivo. In some embodiments, the delivery of an active agent (e.g., a polynucleotide) can correct a defect caused by a defect in the polynucleotide in a cell or tissue.

[0539] In some embodiments, the method for delivering an active agent can introduce a polynucleotide into a cell or tissue to regulate (e.g., turn off or reduce / increase) specific gene expression. For example, delivery of antisense mRNA or shRNA to tumor cells can regulate protein expression encoded by mRNA or shRNA.

[0540] In some embodiments, methods for delivering active agents may introduce polynucleotides into cells or tissues for cellular reprogramming, where the polynucleotides may be used to modulate cellular behavior by expressing transcription factors or growth factors.

[0541] In some embodiments, methods for delivering active agents can introduce polynucleotides into cells or tissues in vivo, such as introducing a "suicide gene" or drug sensitivity gene into a tumor cell, allowing the cell to express a prodrug-activating enzyme, such as herpes simplex virus thymidine kinase, which can kill the tumor cell when exposed to a prodrug (e.g., acyclovir, ganciclovir, valganciclovir, or famciclovir).

[0542] In some embodiments, the method for delivering an active agent can introduce a polynucleotide into a cell or tissue to encode a polypeptide that causes a specific immune response to a target cell. In some embodiments, the target cell can be a cancer cell, an infected cell, or any combination thereof. In some embodiments, the polynucleotide can encode an infection antigen polypeptide, a tumor antigen, a cytokine (e.g., interferon, interleukin, colony stimulating factor), a monoclonal antibody or an antibody fragment (single-chain fragment variable (scFv), fragment antigen binding ((Fab')2), intracellular antibody, nanobody), or any combination thereof.

[0543] In some aspects, disclosed herein is a method of treating cancer, comprising administering to a subject in need thereof an effective amount of a composition or nanoparticle described herein, wherein the composition or nanoparticle comprises a compound of Formula Y, I, Ia, II, III, IV, V, or any combination thereof; and an active agent.

[0544] In some embodiments, the method for delivering an active agent may include introducing a polynucleotide into a specific cell or tissue in vitro to express a polypeptide encoded by the polynucleotide on the specific cell or tissue, thereby producing a chimeric cell or tissue; and administering the chimeric cell or tissue to a subject for tumor treatment. In some embodiments, the specific cell may include a CAR T cell, a CAR-NK cell, or other cells derived from a subject that can be used for immunotherapy.

[0545] In some embodiments, the subject is a mammal. In some embodiments, the mammal is a human. In some embodiments, the subject is a veterinary patient.

[0546] In some embodiments, the compositions herein are used to treat both localized and metastatic tumors.

[0547] In some embodiments, the compositions and methods described herein can be used to treat or prevent the metastasis or recurrence of cancer. In some embodiments, the compositions and methods described herein can be used to prevent the recurrence of resected solid tumors. In some embodiments, the compositions and methods described herein can be used to prevent the metastasis of resected solid tumors.

[0548] In one aspect, the methods described herein are used to treat cancer, such as melanoma, lung cancer (including lung adenocarcinoma, basal cell carcinoma, squamous cell carcinoma, large cell carcinoma, bronchioloalveolar carcinoma, bronchial carcinoma, non-small cell carcinoma, small cell carcinoma, mesothelioma); breast cancer (including ductal carcinoma, lobular carcinoma, inflammatory breast cancer, clear cell carcinoma, mucinous carcinoma, serous cavity breast cancer); colorectal cancer (colon cancer, rectal cancer, colorectal adenocarcinoma); anal cancer; pancreatic cancer (including pancreatic adenocarcinoma, islet cell carcinoma, neuroendocrine tumors); prostate cancer; prostate adenocarcinoma; ovarian cancer (ovarian epithelial carcinoma or surface epithelial stromal tumors, including serous tumors, endometrioid tumors and mucinous cystadenocarcinomas, sex cord stromal tumors); =Includes ovarian cancer (including endometrial adenocarcinoma, eye cancer, uterine papillary serous carcinoma, uterine clear cell carcinoma, uterine sarcoma, leiomyosarcoma, mixed Müllerian duct tumor); glioma, glioblastoma, medulloblastoma and other brain tumors; kidney cancer (including renal cell carcinoma, clear cell carcinoma, Wilms' tumor); head and neck cancer (including squamous cell carcinoma); stomach cancer (gastric cancer, gastric adenocarcinoma, gastrointestinal stromal tumor); testicular cancer; germ cell tumors; neuroendocrine tumors carcinomas; cervical cancer; carcinoids of the gastrointestinal tract, breast, and other organs; signet ring cell carcinoma; mesenchymal tumors, including sarcomas, fibrosarcomas, hemangiomas, angiomatosis, hemangiopericytoma, pseudoangiomatous stromal hyperplasia, myofibroblastic tumors, fibromatosis, inflammatory myofibroblastic tumors, lipomas, angiolipomas, granular cell tumors, neurofibromas, neurotheca tumors, angiosarcomas, liposarcoma, rhabdomyosarcomas, osteosarcomas, leiomyomas, leiomyosarcomas, skin cancers, including melanomas, cervical cancer, retinoblastomas, head and neck cancers, pancreatic cancer, brain cancer, thyroid cancer, testicular cancer, kidney cancer, bladder cancer, soft tissue cancers, adenocarcinomas, urethral cancer, penile cancer, myxosarcoma, chondrosarcoma , osteosarcoma, chordoma, malignant fibrous histiocytoma, lymphangiosarcoma, mesothelioma, squamous cell carcinoma; epidermoid carcinoma, malignant skin adnexal tumor, adenocarcinoma, hepatocellular carcinoma, renal cell carcinoma, adrenal tumor, bile duct carcinoma, transitional cell carcinoma, choriocarcinoma, seminoma, embryonal cell carcinoma, anaplastic glioma; glioblastoma multiforme, neuroblastoma, medulloblastoma, malignant meningioma, malignant neurilemmoma, neurofibrosarcoma, parathyroid carcinoma, medullary thyroid carcinoma, bronchial carcinoid, pheochromocytoma, islet cell carcinoma, malignant carcinoid, malignant paraganglioma, melanoma, Merkel cell tumor, phyllodes cystosarcoma, salivary gland carcinoma, thymic carcinoma and vaginal cancer, etc.

[0549] In some embodiments, the compositions and methods described herein can be used to treat or prevent cancer. In some cases, the cancer is a circulating cancer cell (circulating tumor cell). In some cases, the cancer is a metastatic cancer cell.

[0550] In some embodiments, the method further comprises administering an additional therapeutic agent. In some embodiments, the additional therapeutic agent comprises an additional immunotherapeutic agent or an anti-tumor agent.

[0551] Also disclosed herein are methods of treating a disease or condition such as an inflammatory disorder (including an autoimmune disease) or a lymphoproliferative disorder, comprising administering to a subject in need thereof an effective amount of a compound, combination of compounds, or composition provided herein, or a pharmaceutically acceptable form thereof, or a pharmaceutical composition provided herein.

[0552] Further disclosed herein are methods of treating a disease or condition such as an inflammatory disorder (including an autoimmune disease) or a lymphoproliferative disorder, the method comprising administering to a subject in need thereof an effective amount of a compound, combination of compounds, or composition provided herein, or a pharmaceutically acceptable form thereof, or a pharmaceutical composition provided herein.

[0553] In one embodiment, a method for treating an inflammatory condition (including an autoimmune disease) of a subject is provided herein. The method includes administering to the subject a therapeutically effective amount of a compound provided herein, a combination of compounds, or a composition, or a pharmaceutically acceptable form thereof, or a pharmaceutical composition provided herein. Examples of autoimmune diseases include, but are not limited to, acute disseminated encephalomyelitis (ADEM), Addison's disease, antiphospholipid antibody syndrome (APS), aplastic anemia, autoimmune hepatitis, autoimmune skin disease, celiac disease, Crohn's disease, diabetes (type 1), Goodpasture's syndrome, Graves' disease, Guillain-Barre syndrome (GBS), Hashimoto's disease, lupus erythematosus, multiple sclerosis, myasthenia gravis, opsoclonus-myoclonus syndrome (OMS), optic neuritis, Odd's thyroiditis, o The diseases that may be considered include inflammatory bowel disease, ...

[0554] Inflammation takes many forms and includes, but is not limited to, acute, adhesive, atrophic, catarrhal, chronic, cirrhotic, diffuse, disseminated, exudative, fibrinous, fibrotic, focal, granulomatous, proliferative, hypertrophic, interstitial, metastatic, necrotic, occlusive, parenchymal, plastic, prolific, proliferative, pseudomembranous, purulent, sclerotic, serofibrinous, serous, simple, idiopathic, subacute, purulent, toxic, traumatic, and / or ulcerative inflammation.

[0555] Exemplary inflammatory conditions include, but are not limited to, inflammation associated with acne, anemia (e.g., aplastic anemia, hemolytic autoimmune anemia), asthma, arteritis (e.g., polyarteritis, temporal arteritis, periarteritis nodosa, polyarteritis), arthritis (e.g., crystalline arthritis, osteoarthritis, psoriatic arthritis, gout flares, gouty arthritis, reactive arthritis, rheumatoid arthritis, and Reiter's arthritis), ankylosing spondylitis, amyloidosis, amyotrophic lateral sclerosis, autoimmune diseases, allergies or allergic reactions, atherosclerosis, bronchitis, bursitis, chronic prostatitis , conjunctivitis, Chagas disease, chronic obstructive pulmonary disease, dermatomyositis, diverticulitis, diabetes (e.g., type 1 diabetes, type 2 diabetes), skin diseases (e.g., psoriasis, eczema, burns, dermatitis, pruritus (itching)), endometriosis, Guillain-Barré syndrome, infection, ischemic heart disease, Kawasaki disease, glomerulonephritis, gingivitis, allergies, headaches (e.g., migraines, tension headaches), intestinal obstruction (e.g., postoperative intestinal obstruction and intestinal obstruction during sepsis), idiopathic thrombocytopenic purpura, interstitial cystitis (bladder pain syndrome), gastrointestinal disorders (e.g., selected from peptic ulcer, localized intestinal eosinophilic gastrointestinal disorders (e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilic gastroenteritis, eosinophilic colitis), gastritis, diarrhea, gastroesophageal reflux disease (GORD or its synonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis, collagenous colitis, lymphocytic colitis, ischemic colitis, diversion colitis, Behcet's syndrome, indeterminate colitis) and inflammatory bowel syndrome (IBS)), lupus, multiple sclerosis, morphea, myasthenia gravis, myocardial ischemia, nephrotic syndrome, common Pemphigus, pernicious anemia, peptic ulcer, polymyositis, primary biliary cirrhosis, neuroinflammation associated with brain disorders (e.g., Parkinson's disease, Huntington's disease, and Alzheimer's disease), prostatitis, chronic inflammation associated with skull radiation injury, pelvic inflammatory disease, polymyalgia rheumatica, reperfusion injury, regional enteritis, rheumatic fever, systemic lupus erythematosus, scleroderma, sclerosis, sarcoidosis, spondyloarthropathies, Sjögren's syndrome, thyroiditis, transplant rejection, tendinitis, trauma or injury (e.g., frostbite, chemical irritants, toxins, scars, burns, physical injuries), vasculitis, vitiligo, and Wegener's granulomatosis. In certain embodiments, the inflammatory disorder is selected from arthritis (e.g., rheumatoid arthritis), inflammatory bowel disease, inflammatory bowel syndrome, asthma, psoriasis, endometriosis, interstitial cystitis, and prostatitis. In certain embodiments, the inflammatory condition is an acute inflammatory condition (e.g., inflammation caused by infection). In certain embodiments, the inflammatory condition is a chronic inflammatory condition (eg, conditions resulting from asthma, arthritis, and inflammatory bowel disease).The compounds may also be useful in treating inflammation associated with trauma and non-inflammatory myalgia.

[0556] Immune disorders such as autoimmune disorders include, but are not limited to, arthritis (including rheumatoid arthritis, spondyloarthropathies, gouty arthritis, degenerative joint diseases such as osteoarthritis, systemic lupus erythematosus, Sjögren's syndrome, ankylosing spondylitis, undifferentiated spondylitis, Behcet's disease, hemolytic autoimmune anemia, multiple sclerosis, amyotrophic lateral sclerosis, amyloidosis, acute shoulder pain, psoriasis, and juvenile arthritis), asthma, atherosclerosis, osteoporosis, bronchitis, tendinitis, bursitis, skin diseases (e.g., psoriasis, eczema, burns, dermatitis, pruritus (itching)), enuresis, eosinophilic diseases, gastrointestinal disorders (e.g., selected from peptic ulcer, regional enteritis, diverticulitis, gastrointestinal bleeding, eosinophilic gastrointestinal diseases (e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilic gastroenteritis, eosinophilic colon enteritis), gastritis, diarrhea, gastroesophageal reflux disease (GORD or its synonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis, collagenous colitis, lymphocytic colitis, ischemic colitis, diversion colitis, Behcet's syndrome, indeterminate colitis, and inflammatory bowel syndrome (IBS)), relapsing polychondritis (e.g., atrophic polychondritis and systemic polychondromalacia), and conditions improved by prokinetic agents (e.g., ileus, postoperative ileus, and ileus during sepsis; gastroesophageal reflux disease (GORD, or its synonym GERD); eosinophilic esophagitis, gastroparesis such as diabetic gastroparesis; food intolerances and food allergies and other functional bowel disorders such as non-ulcer dyspepsia (NUD) and non-cardiac chest pain (NCCP, including costochondritis)).

[0557] In one embodiment, a method for treating an infection in a subject is provided herein, and the infection is caused by an infectious agent. The method includes administering to the subject a therapeutically effective amount of a compound, a combination of compounds, or a composition, or a pharmaceutically acceptable form thereof, or a pharmaceutical composition provided herein. Infectious agents may include, but are not limited to, bacterial organisms such as Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, Corynebacterium diphtheriae, Listeria monocytogenes, Bacillus anthracis, Clostridium tetani, Clostridium botulinum, Clostridium perfringens, Neisseria meningitidis, Neisseria gonorrhoeae, Streptococcus mutans, Pseudomonas aeruginosa, Salmonella typhi, Haemophilus parainfluenzae, Bordetella pertussis, Francisella tularensis, Yersinia pestis, Vibrio cholerae, Legionella pneumophila, Mycobacterium tuberculosis, Leprosy Mycobacterium, Treponema pallidum, Leptospira interrogans, Borrelia burgdorferi, Campylobacter jejuni, etc.; viruses, such as metapneumoviruses such as human metapneumovirus (hMPV), parainfluenza viruses such as human parainfluenza virus (hPIV) types 1, 2, and 3 (hPIV1, hPIV2, and hPIV3, respectively), respiratory syncytial virus (RSV), measles virus (MeV), coronaviruses (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV- OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1), poxviruses (e.g., smallpox, monkeypox), African swine fever virus, influenza A and B, human immunodeficiency virus (HIV), varicella-zoster, herpes simplex 1 and 2, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps, rabies, Rubella, Coxsackievirus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E viruses, etc.; fungi, protozoa and parasitic organisms, such as Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans, Candida tropicalis, Nocardia asteroides, Rickettsia rickettsii, Rickettsia typhi, Mycoplasma pneumoniae, Chlamydia psittaci, Chlamydia trachomatis, Plasmodium falciparum, Trypanosoma brucei, Entamoeba histolytica, Toxoplasma gondii, Trichomonas vaginalis, Schistosoma mansoni, etc.

[0558] In some embodiments, the infection can be a coronavirus infection. In one embodiment, the coronavirus infection is an infection of the upper respiratory tract and / or the lower respiratory tract. The "upper respiratory tract" includes the mouth, nose, sinuses, middle ear, throat, larynx and trachea. The "lower respiratory tract" includes the bronchi (bronchial tubes) and lungs (bronchi, bronchioles and alveoli) and interstitial lung tissue.

[0559] In another embodiment, the coronavirus infection is a gastrointestinal infection."Gastrointestinal tract" can include any area of ​​the tube from mouth to anus, including the mouth, esophagus, stomach, and intestines.

[0560] In yet another embodiment, the coronavirus infection is a kidney infection.

[0561] It is understood and contemplated herein that the coronavirus infection disclosed herein may cause a pathological state associated with a coronavirus infection, referred to herein as a "coronavirus disease". In some embodiments, the coronavirus disease is selected from the group consisting of the common cold, pneumonia, pneumonia, bronchitis, severe acute respiratory syndrome (SARS), coronavirus disease 2019 (COVID-2019), Middle East respiratory syndrome (MERS), sinusitis, swine diarrhea, porcine epidemic diarrhea, avian infectious bronchitis, otitis, and pharyngitis. In some embodiments, the coronavirus infection is the common cold. In some embodiments, the coronavirus infection is selected from the group consisting of SARS, COVID-19, and MERS. In a specific embodiment, the coronavirus infection is COVID-19. In another specific embodiment, the coronavirus infection is IBV, PorCoV HKU15, or PEDV.

[0562] Most SARS patients have been previously healthy adults between the ages of 25 and 70. Some suspected SARS cases have been reported in children under the age of 15. The case fatality rate for patients who meet the current World Health Organization definition of probable and suspected SARS cases is approximately 3%.

[0563] Other indications associated with coronavirus infection are described in Gralinski and Baric, 2015, J. Pathol. 235: 185-195 and Cavanagh, 2005, “Coronaviridae: a review of coronavirus and toroviruses”, Coronaviruses with Special Emphasis on First Insights Concerning SARS1, A. Schmidt, MH Wolff and O. Weber, eds., Birkhauser Verlag Baser, Switzerland, which are incorporated herein by reference in their entirety.

[0564] The coronavirus causing the infection may be selected from an alphacoronavirus, a betacoronavirus, a gammacoronavirus or a deltacoronavirus.

[0565] Ocular conditions that can be treated according to the compositions and methods disclosed herein include amebic keratitis, fungal keratitis, bacterial keratitis, viral keratitis, onchocercal keratitis, bacterial keratoconjunctivitis, viral keratoconjunctivitis, corneal dystrophic diseases, Fuchs endothelial dystrophy, Sjögren's syndrome, Stevens-Johnson syndrome, autoimmune dry eye disease, environmental dry eye disease, corneal neovascular disease, prevention and treatment of rejection after corneal transplantation, autoimmune uveitis, infectious uveitis, anterior uveitis, posterior uveitis (including toxoplasmosis), panuveitis, vitreous or retinal inflammatory diseases, prevention and treatment of endophthalmitis, macular edema, macular degeneration, age-related macular degeneration, proliferative and non-proliferative diabetic retinopathy, hypertensive retinopathy, retinal autoimmune diseases, primary and metastatic intraocular melanoma, other intraocular metastatic tumors, open-angle glaucoma, angle-closure glaucoma, pigmentary glaucoma, and combinations thereof.

[0566] Endocrine disorders that can be treated according to the compositions and methods disclosed herein include thyroid disorders (e.g., hyperthyroidism, hypothyroidism, thyroiditis, goiter, diabetes, hypoglycemia, glucagonoma, calcium homeostasis disorders (e.g., parathyroid, osteoporosis, osteomalacia, rickets, sex hormone disorders (e.g., disorders of sexual development, hypogonadism, puberty disorders, menstrual function or fertility disorders).

[0567] Examples

[0568] The following examples are set forth below to illustrate compounds, compositions, methods and results according to the disclosed subject matter. These examples are not intended to include all aspects of the subject matter disclosed herein, but are intended to illustrate representative methods and results. These examples are not intended to exclude equivalents and variations of the present invention that are apparent to those skilled in the art.

[0569] Example 1: Synthesis of ionizable lipids (Compound Target 1)

[0570] The experimental procedures used to synthesize compound target 1 are described. The synthetic route of compound 1 is shown in Figure 1 middle.

[0571]

[0572] Preparation procedure of compound 2: Mg (48.5 g, 1.99 mol, 3.08 eq) and I2 (20.0 mg, 78.8 μmol) were loaded into R1 (3.00 L three-necked round-bottom flask) at 30 to 40 ° C under N2. Compound 1 (125 g, 647 mmol, 113 mL, 1.00 eq) in THF (1.20 L) was loaded into R2 (2.00 L flask) at 30 to 40 ° C. 5.00 mL of R2 was loaded into R1 at 30 to 40 ° C. An exothermic reaction was noted, and immediate decolorization was observed and the solution began to reflux. The remaining R2 was loaded into R1 at 30 to 40 ° C. R1 was stirred at 35 ° C under N2 for 1 hour. An exothermic reaction was noted and the color became dark brown. The mixture was used for the next step without further purification. Compound 2 (140 g, 644 mmol, 99.5% yield) in THF (1.20 L) was used in the next step as a dark brown liquid.

[0573]

[0574] Preparation procedure of compound 3: Compound 2 (141 g, 647 mmol, 146 mL, 1.40 eq) in THF (500 mL) was charged into R1 (2.00 L three-necked flask). R1 ​​was cooled to 0°C. THF (400 mL) was charged into R2 (1.00 L bucket). Compound 2a (52.8 g, 462 mmol, 64.5 mL, 1.00 eq) was charged into R2. The solution of R2 was added dropwise to R1 at 0°C. R2 was stirred at 25°C for 16 hours. H2O (200 mL) was charged into R2 (2.00 L bucket) at 25°C. The reaction was extracted with ethyl acetate (800 mL*3) at 25°C. The reaction was dried over anhydrous Na2SO4 at 25°C. The organic phase was concentrated below 40°C to 50°C to obtain a residue. The residue was purified by column chromatography to obtain compound 3 (129 g, 565 mmol, crude) as a white solid ( 1 H NMR: 400MHz, CDCl3δ3.61-3.57(m,1H),1.47-1.43(m,6H),1.29-1.25(m,18H),0.91-0.87(m,6H)).

[0575]

[0576] Preparation procedure of compound 4: At 25°C, EDCI (49.6g, 259mmol, 1.30eq), DIEA (103g, 796mmol, 139mL, 4.00eq) and DMAP (4.86g, 39.8mmol, 0.20eq) were charged into a solution of compound 3 (50.0g, 219mmol, 1.10eq) and compound 3a (41.6g, 199mmol, 1.00eq) in 500mL dichloromethane. The mixture was stirred at 25°C for 16 hours. 500mL H2O was charged into the mixture, and the reaction was extracted with dichloromethane (400mL*3) at 25°C. The reaction was washed with saturated brine (300mL*3) at 25°C. The reaction was dried with anhydrous Na2SO4 at 25°C. The organic phase was concentrated below 40°C to 50°C to obtain a residue. The residue was purified by column chromatography to give Compound 4 (32.0 g, 78.7 mmol, 39.5% yield) as a colorless liquid ( 1 H NMR: 400MHz, CDCl3δ4.90-4.84(m,1H),3.40(t,J=6.8Hz,2H),2.30(t,J=7.2Hz,2H),1.90-1.82(m,2H) ,1.68-1.61(m,2H),1.52-1.45(m,5H),1.43-1.34(m,3H),1.30-1.25(m,20H),0.882(t,J=6.4Hz,6H)).

[0577]

[0578] Preparation procedure of compound 5: At 25 ° C, to a solution of compound 4 (32.0 g, 78.7 mmol, 1.00 eq) in 33 mL EtOH, compound 4a (210 g, 2.36 mol, 219 mL, 30.0 eq) was loaded into the solution. The mixture was heated to 65 ° C and stirred at 65 ° C for 16 hours. Cooled to 25 ° C, and 100 mL H2O was loaded into the mixture. The reaction was extracted with ethyl acetate (60.0 mL*3) at 25 ° C, and the reaction was washed with saturated brine (50.0 mL*3). The reaction was dried with anhydrous Na2SO4 at 25 ° C. The organic phase was concentrated below 40 ° C to 50 ° C to obtain a residue. The residue was purified by column chromatography to give Compound 5 (16.4 g, 65.5 mmol, 83.2% yield) as a red oil (1H NMR: 400 MHz, CDCl3 δ 4.89-4.83 (m, 1H), 3.58-3.56 (m, 2H), 2.66-2.62 (m, 2H), 2.55-2.52 (m, 2H), 2.25-2.19 (m, 2H), 1.64-1.53 ​​(m, 6H), 1.44-1.42 (m, 6H), 1.28-1.25 (m, 7H), 1.19 (s, 17H), 0.89-0.86 (m, 6H)).

[0579]

[0580] Preparation procedure of compound 5a: At 25°C, compound 5c (20.0g, 51.7mmol, 13.51mL, 1.10eq) was added to a solution of 400mL DCM. Then, EDCI (11.7g, 61.1mmol, 1.30eq), DMAP (1.15g, 9.40mmol, 0.200eq) and DIEA (24.3g, 188mmol, 32.8mL, 4.00eq) were added and the mixture was stirred at 25°C for 16 hours. 300mL H2O was added to the mixture, and the mixture was extracted with DCM (100mL*3) at 25°C. The reaction was dried with anhydrous Na2SO4, and the organic phase was concentrated at 40°C to 50°C to obtain a residue. The residue was purified by column chromatography to give compound 5a (12.1 g, 21.5 mmol, 45.6% yield) as a white solid ( 1 H NMR:EC3947-13-P1A1,400MHz,CDCl3

[0581] δ5.38(d,J=4.8Hz,1H),4.67–4.59(m,1H),3.44–3.38(m,2H),2.34–2.29(m,4H),2.03–2.00(m,2H),1.66–1.51(m,4H),1.50 -1.48(m,6H),1.35–1.33(m,5H),1.27-1.13(m,7H),1.03(s,3H),0.92(d,J=6.4Hz,3H),0.089–0.087(m,6H),0.07(s,3H)).

[0582]

[0583] Preparation procedure of lipid target 1: At 25 ° C, compound 6 (6.40 g, 14.9 mmol, 1.00 eq) was added to a solution of compound 6 (6.40 g, 14.9 mmol, 1.00 eq) in 140 mL ACN. K2CO3 (8.27 g, 59.9 mmol, 4.00 eq), KI (2.73 g, 16.5 mmol, 1.10 eq) and CPME (35.0 mL) were added to the solution. The mixture was heated to 90 ° C and stirred at 90 ° C for 16 hours. After cooling to 25 ° C, the residue was dissolved in dichloromethane (100 mL) and filtered. The residue was purified by column chromatography to obtain compound target 1 (MS: m / z=910.7 (M+H) as a yellow oil. + , 1 H NMR: 400MHz, CDCl3δ5.38(d,J=4.0Hz,1H),4.89-4.79(m,1H),4.64-4.58(m,1H),3 .55(d,J=8.0Hz,2H),2.45-2.35(m,6H),2.25-2.19(m,6H),1.98-1.88(m,2H),1.80 -1.72(m,3H),1.57-1.52(m,8H),1.47-1.34(m,13H),1.27-1.14(m,31H),1.10-1. 01(m,8H),0.95-0.90(m,6H),0.89-0.82(m,5H),0.81-0.78(m,10H),0.68(s,3H)). FIG. 2A to FIG. 2C The 1H NMR spectrum (2A), mass spectrum (2B), and HPLC spectrum (2C) of compound target 1 are shown.

[0584] Example 2: Synthesis of ionizable lipids (Compound Target 11)

[0585]

[0586] To a solution of compound 4a (10.0 g, 31.6 mmol, 1.00 eq) in DCM (270 mL) was added (COCl) 2 (27.2 g, 214 mmol, 18.8 mL, 6.80 eq) and DMF (231 mg, 3.16 mmol, 243 μL, 0.01 eq). The mixture was stirred at 25 ° C for 16 hours. One drop of the reaction mixture was diluted with 3 drops of methanol to obtain a mixture, and the mixture was then monitored by TLC. TLC (DCM: methanol = 10: 1) showed compound 4a (R f = 0.30) was completely consumed, and new spots were detected (R f =0.80). The reaction mixture was then concentrated in vacuo to give compound 4b (9.00 g, crude) as a brown oil.

[0587]

[0588] To a solution of compound 1 (4.00 g, 30.4 mmol, 1.00 eq) in DCM (200 mL) was added (COCl) 2 (11.6 g, 91.4 mmol, 8.01 mL, 3.00 eq) and DMF (22.2 mg, 304 μmol, 23.4 μL, 0.01 eq). The mixture was stirred at 25 ° C for 16 hours. One drop of the reaction mixture was diluted with 3 drops of methanol to obtain a mixture, and the mixture was then monitored by TLC. TLC (DCM: methanol = 10: 1) showed compound 1 (R f = 0.00) is completely consumed, and new spots are detected (R f =0.50). The reaction mixture was then concentrated in vacuo to give compound 1b (4.50 g, crude) as a brown solid.

[0589]

[0590] To a solution of compound b1 (20.0 g, 153 mmol, 1.00 eq) in THF (400 mL) was added LiAlH4 (2.50 M, 61.4 mL, 1.00 eq) at 0°C under N2. The mixture was stirred at 25°C under N2 for 1 hour. TLC (petroleum ether: EtOAc = 1:1) showed compound b1 (R f = 0.80) is consumed and a new spot is detected (R f=0.20). The reaction mixture was quenched with Na2SO4·10H2O until a large amount of white solid was produced and no gas was produced at 0°C. The solid was filtered and the filter cake was washed with DCM (3 x 100 mL). The combined filtrate was dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated in vacuo to give compound b2 (18.0 g, 136 mmol, 88.6% yield) as a colorless oil. 1 H NMR: (400MHz, DMSO-d6) δ4.94-4.91(m,1H),3.80-3.74(m,2H),3.52-3.44(m,3H),1.33(s,3H),1.27(s,3H).

[0591]

[0592] Compound 4b (8.99 g, 30.2 mmol, 1.00 eq) was added to a solution of compound b2 (4.00 g, 30.2 mmol, 1.00 eq) and pyridine (4.79 g, 60.5 mmol, 4.89 mL, 2.00 eq) in DCM (200 mL). The mixture was stirred at 25 °C for 2 hours. TLC (petroleum ether: EtOAc = 1:1) showed compound b2 (R f = 0.20) is consumed and a new spot is detected (R f =0.80). The reaction mixture was poured into 400 mL of water and then extracted with DCM (3 x 100 mL). The combined organic layers were washed with brine (100 mL), dried over Na2SO4 and filtered. The filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether: EtOAc = 100: 1 to 5: 1, R f =0.40 (petroleum ether:EtOAc=5:1)) to give compound b3 (10.0 g, 23.1 mmol, 76.5% yield, purity in LCMS under ELSD was 91.0%) as a light yellow oil. 1 H NMR: (400MHz, CDCl3) δ5.45-5.28(m,6H),4.74-4.69(m,1H),4.10(dd,J=13.2,3.6Hz,2H),3.81(dd,J=12.8,3.6Hz,2H),2.86-2 .75(m,4H),2.38(t,J=7.6Hz,2H),2.11-2.03(m,4H),1.68-1.60(m,2H),1.45(s,6H),1.36-1.30(m,8H),0.98(t,J=7.6Hz,3H).

[0593]

[0594] A solution of compound b3 (10.0 g, 23.1 mmol, 1.00 eq) in AcOH (80.0 mL) and H2O (20.0 mL) was stirred at 50°C for 1 hour. TLC (petroleum ether: EtOAc = 1:1) showed that compound b3 (R f = 0.80) is consumed and a new spot is detected (R f =0.20). The reaction mixture was concentrated in vacuo and then diluted with EtOAc (100 mL). The organic layer was washed successively with saturated NaHCO solution (50.0 mL), water (50.0 mL) and brine (50.0 mL). The organic layer was then dried over NaSO and filtered. The filtrate was concentrated in vacuo to obtain compound b4 (7.00 g, 18.9 mmol, 81.7% yield, 95.4% purity in LCMS under ELSD) as a yellow oil. 1 H NMR: (400MHz, DMSO-d6) δ5.40-5.24(m,6H),4.73-4.67(m,2H),4.02(dd,J=14.4,7.2Hz,1H),3.53-3.39(m,4H),2. 81-2.71(m,4H),2.30-2.24(m,2H),2.08-2.00(m,4H),1.55-1.47(m,2H),1.30-1.24(m,8H),0.92(t,J=7.2Hz,3H).

[0595]

[0596] To compound b4 (7.00g, 18.9mmol, 1.00eq) and pyridine (3.00g, 37.8mmol, 3.06mL, 2.00eq) solution in DCM (140mL) add compound 1b (2.27g, 15.1mmol, 0.80eq) solution in DCM (20.0mL). The mixture was stirred at 25 ° C for 2 hours. LCMS (EC15884-22-P1B1) shows that most of compound b4 (Rt=2.118 minutes) is consumed, and the desired mass (Rt=1.845 minutes) is detected. The reaction mixture is poured into 200mL water and extracted with DCM (3x 100mL). The combined organic layer is washed with brine (100mL), dried over Na2SO4 and filtered. The filtrate is concentrated in vacuo to obtain a residue. The precipitate was purified by column chromatography (SiO 2 , petroleum ether:EtOAc=100:1 to 5:1, R f=0.40 (petroleum ether:EtOAc=5:1)) to give compound 6 (3.50 g, 7.34 mmol, 38.7% yield, purity in LCMS under ELSD was 97.7%) as a yellow oil. 1 H NMR: (400MHz, CDCl3) δ5.41-5.28(m,6H),5.11-5.04(m,1H),4.37-4.30(m,1H) ,4.25-4.18(m,1H),4.16-4.06(m,2H),2.84-2.74(m,4H),2.61-2.51(m,2H),2 .47-2.42(m,2H),2.40(d,J=4.4Hz,6H),2.36-2.30(m,2H),2.11-2.01(m,4H), 1.97-1.86(m,2H),1.65-1.57(m,2H),1.34-1.28(m,8H),0.96(t,J=7.6Hz,3H).

[0597]

[0598] Compound SM_1 (1.63 g, 8.06 mmol, 1.20 eq), EDCI (1.93 g, 10.0 mmol, 1.50 eq), DIEA (3.47 g, 26.8 mmol, 4.68 mL, 4.00 eq) and DMAP (164 mg, 1.34 mmol, 0.20 eq) were added to a solution of compound 6 (3.20 g, 6.71 mmol, 1.00 eq) in DCM (60.0 mL). The mixture was stirred at 25 ° C for 12 hours. LCMS (EC15884-24-P1A1) showed that compound 6 was completely consumed and the desired mass (Rt = 1.617 minutes) was detected. The reaction mixture was concentrated in vacuo to obtain a residue. The residue was diluted with 50.0 mL EtOAc. The organic layer was washed with H2O (50.0 mL), brine (50.0 mL), dried over Na2SO4 and filtered. The filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography (SiO2, DCM:methanol = 100:1 to 10:1, R f =0.50 (DCM:methanol=10:1)) to give compound 6d (3.00 g, 4.14 mmol, 61.6% yield, purity in LCMS under ELSD was 89.6%) as a yellow oil.

[0599]

[0600] HCl / dioxane (4.00M, 30.0mL, 29.0eq) was added to a solution of compound 6d (3.00g, 4.14mmol, 1.00eq) in DCM (10.0mL). The mixture was stirred at 25°C for 2 hours. LCMS (EC15884-25-P1A1) showed that compound 6d was completely consumed, and the desired mass (Rt=0.334 minutes) was detected. The reaction mixture was concentrated in vacuo to obtain compound 6c (2.46g, crude product) in a yellow oily state.

[0601]

[0602] Compound Int-7 (2.40 g, 6.21 mmol, 1.62 mL, 1.50 eq), EDCI (1.03 g, 5.39 mmol, 1.30 eq), DIEA (2.68 g, 20.7 mmol, 3.61 mL, 5.00 eq) and DMAP (101 mg, 828 μmol, 0.20 eq) were added to a solution of compound 6c (2.46 g, 4.14 mmol, 1.00 eq) in DCM (25.0 mL). The mixture was stirred at 25 ° C for 12 hours. LCMS (EC15884-26-P1A2) showed that compound 6c was completely consumed, and the desired mass (Rt=2.090 minutes) was detected. The reaction mixture was concentrated in vacuo to obtain a residue. The residue was diluted with 50.0 mL EtOAc. The organic layer was washed with H2O (50.0 mL), brine (50.0 mL), dried over Na2SO4 and filtered. The filtrate was concentrated in vacuo to give the target compound 11 (4.00 g, crude) as a yellow oil.

[0603] The crude target 11 (4.00 g, crude) was purified by preparative HPLC (column: Welch Ultimate XB-CN 250*50*10 um; mobile phase: [EtOH+MeOH (4:1, neutral)]; gradient: 12%-100% B over 16 minutes), and then by column chromatography (SiO2, DCM:methanol=100:0 to 50:1, R f =0.35 (DCM:methanol=10:1)) to obtain the target 11 (511.07 mg, 0.522 mmol, 98.3% purity, 12.5% ​​yield) as a yellow oil. 1H NMR: (400MHz, CDCl3) δ5.45-5.23(m,8H),4.67-4.57(m,1H),4.35-4.27(m,2H),4.19-4 .11(m,2H),2.86-2.75(m,4H),2.41-2.27(m,18H),2.13-1.94(m,7H),1.89-1.78(m,6H ),1.70-1.41(m,16H),1.39-1.33(m,5H),1.29-1.22(m,2H),1.21-1.04(m,8H),1.02(s ,3H),1.01-0.94(m,5H),0.92(d,J=6.4Hz,3H),0.87(d,J=6.8,1.6Hz,6H),0.68(s,3H).

[0604] Example 3: Synthesis of ionizable lipids (Compound Target 12)

[0605]

[0606] Compound 1a (5.04 g, 25.8 mmol, 1.00 eq), EDCI (6.45 g, 33.6 mmol, 1.30 eq), DMAP (631 mg, 5.17 mmol, 0.20 eq) and DIEA (13.3 g, 103 mmol, 18.0 mL, 4.00 eq) were added to a solution of compound Int_7 (11.0 g, 28.4 mmol, 7.43 mL, 1.10 eq) in DCM (220 mL) at 25°C. The mixture was stirred at 25°C for 16 hours. TLC (petroleum ether: ethyl acetate = 10: 1) showed that most of compound Int_7 (R f = 0.10) is consumed and a new spot is formed (R f =0.50). The reaction mixture was concentrated in vacuo to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether:ethyl acetate = 100:1 to 10:1, R f =0.50 (petroleum ether:ethyl acetate=10:1)) to give Compound 1 (6.80 g, 12.0 mmol, 46.6% yield) as a white solid. 1H NMR: (400MHz, CDCl3) δ5.38 (d, J=4.0Hz, 1H), 4.67-4.56 (m, 1H), 3.41 (t, J=6.8 Hz,2H),2.36-2.27(m,4H),2.05-1.94(m,2H),1.93-1.81(m,5H),1.70-1.58(m, 4H),1.55-1.46(m,6H),1.45-1.27(m,5H),1.23-1.05(m,8H),1.02(s,3H),1.0 1-0.95(m,2H),0.92(d,J=6.4Hz,3H),0.87(dd,J=6.4,1.6Hz,6H),0.68(s,3H).

[0607]

[0608] Compound 1 (4.00 g, 7.10 mmol, 1.00 eq), K2CO3 (3.92 g, 28.3 mmol, 4.00 eq), KI (1.30 g, 7.81 mmol, 1.10 eq) and CPME (15.0 mL) were added to a solution of compound 2a (1.14 g, 7.10 mmol, 1.00 eq) in ACN (60.0 mL). The mixture was stirred at 90 ° C for 12 hours. LCMS (EC15884-12-P1B1) showed that compound 1 was completely consumed, and the desired mass (Rt = 1.825 minutes) was detected. The reaction mixture was diluted with 100 mL DCM and filtered. The filtrate was concentrated in vacuo to obtain a residue. By column chromatography (SiO2, DCM: methanol = 100: 1 to 5: 1, R f =0.35 (DCM:methanol=10:1)) to give Compound 2 (3.00 g, 4.41 mmol, 62.1% yield, 94.7% purity in LCMS under ELSD) as a yellow oil. 1HNMR: (400MHz, CDCl3) δ5.38 (d, J=4.0Hz, 1H), 4.66-4.57 (m, 1H), 3.61 (t, J=5.6Hz, 4H), 2 .55-2.46(m,6H),2.35-2.26(m,4H),2.06-1.93(m,3H),1.90-1.79(m,4H),1.72-1.66(m, 3H),1.62-1.59(m,2H),1.58-1.40(m,10H),1.36-1.23(m,7H),1.22-1.04(m,9H),1.02(s ,3H),1.01-0.95(m,2H),0.92(d,J=6.4Hz,3H),0.87(dd,J=6.8,2.0Hz,6H),0.68(s,3H).

[0609]

[0610] Compound 3a (900 mg, 3.24 mmol, 1.10 eq), EDCI (732 mg, 3.82 mmol, 1.30 eq), DIEA (1.90 g, 14.7 mmol, 2.56 mL, 5.00 eq) and DMAP (71.8 mg, 588 μmol, 0.20 eq) were added to a solution of compound 2 (2.00 g, 2.94 mmol, 1.00 eq) in DCM (20.0 mL). The mixture was stirred at 25 ° C for 12 hours. LCMS (EC15884-14-P1A2) showed that most of compound 2 (Rt = 1.814 minutes) was consumed, and the desired mass (Rt = 2.052 minutes) was detected. The reaction mixture was concentrated in vacuo to obtain a residue. The residue was purified by preparative HPLC (column: Welch Ultimate XB-NH2 250*50*10um; mobile phase: [EtOH+MeOH (4:1, neutral)]; B%: 1%, isocratic elution mode) to give target 12 (504.60 mg, 0.530 mmol, 18.0% yield, 95.0% purity in HPLC under ELSD) as off-white oil. 1H NMR: (400MHz, CDCl3) δ5.44-5.28(m,7H),4.66-4.57(m,1H),4.07(t,J=6.4Hz,2H),3.6 0-3.54(m,2H),2.85-2.75(m,4H),2.55-2.43(m,6H),2.34-2.26(m,7H),2.13-1.94(m,7 H),1.90-1.79(m,4H),1.69-1.43(m,28H),1.30-1.20(m,4H),1.19-1.05(m,8H),1.02(s ,3H),1.01-0.94(m,5H),0.92(d,J=6.4Hz,3H),0.87(d,J=6.4,1.6Hz,6H),0.68(s,3H).

[0611] Example 4: Synthesis of ionizable lipids (Compound Target 13)

[0612]

[0613] Compound 1 (30.0 g, 131 mmol, 1.00 eq) was charged into R1 (1.00 L triple bottom flask) at 20°C. DCM (350 mL) was charged into R1 at 20°C. Compound 1a (27.4 g, 131 mmol, 1.00 eq) was charged into R1 at 20°C. DIEA (67.9 g, 525 mmol, 91.5 mL, 4.00 eq), DMAP (3.21 g, 26.2 mmol, 0.20 eq), and EDCI (32.7 g, 170 mmol, 1.30 eq) were charged into R1 at 20°C. The mixture was stirred at 20°C for 16 hours. TLC (petroleum ether: ethyl acetate = 20:1) showed that most of compound 1 (R f =0.60) is consumed, and a new main spot (R f =0.50). H2O (300mL) was loaded into R2 (1.00L bucket) at 20°C. The reaction was loaded into R2 at 20°C. The reaction was extracted with dichloromethane (300mL*3) at 20°C. The reaction was washed with saturated brine (200mL*2) at 20°C. The reaction was dried over anhydrous Na2SO4 at 20°C. The organic phase was concentrated at 40°C to 50°C to obtain a residue. The product was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 50:1 to 10:1, R f =0.50) to purify the residue.

[0614] Compound 2 (30.0 g, 71.5 mmol, 54.4% yield) was obtained as a colorless liquid.

[0615]

[0616] Compound 2 (30.0 g, 71.5 mmol, 1.00 eq) was charged into R1 (250 mL three-necked bottom flask) at 20°C. EtOH (30.0 mL) was added into R1 at 20°C. Compound 2a (191 g, 2.15 mol, 199 mL, 30.0 eq) was charged into R1 at 20°C. The mixture was heated to 65°C. The mixture was stirred at 65°C for 16 hours. TLC (dichloromethane: methanol = 5: 1) showed that most of compound 2 (R f =0.95) is consumed, and a new main spot (R f =0.20). H2O (100 mL) was loaded into R2 (250 mL bucket) at 20°C. The reaction was loaded into R2 at 20°C. The reaction was extracted with ethyl acetate (60.0 mL*3) at 20°C. The reaction was washed with saturated brine (50.0 mL*3) at 20°C. The reaction was dried over anhydrous Na2SO4 at 20°C. The organic phase was concentrated at 40°C to 50°C to obtain a residue. The product was purified by column chromatography (SiO2, dichloromethane:methanol=50:1 to 10:1, R f =0.20) to purify the residue. A pale yellow oily compound 3 (17.0 g, 39.7 mmol, 55.5% yield) was obtained.

[0617]

[0618] Compound 3-1 (5.00 g, 12.1 mmol, 1.10 eq) was charged into R1 (2.00 L triple bottom flask) at 20°C. DCM (100 mL) was charged into R1 at 20°C. Compound 3-2 (2.15 g, 11.0 mmol, 1.00 eq) was charged into R1 at 20°C. EDCI (2.74 g, 14.3 mmol, 1.30 eq), DIEA (5.69 g, 44.0 mmol, 7.67 mL, 4.00 eq), and DMAP (269 mg, 2.20 mmol, 0.20 eq) were charged into R1 at 20°C. The mixture was stirred at 20°C for 20 hours. TLC (petroleum ether: ethyl acetate = 5:1) showed that most of compound 3-1 (R f =0.30) is consumed, and a new main spot (R f=0.60). H2O (200 mL) was loaded into R2 (1.00 L bucket) at 20°C. The reaction was loaded into R2 at 20°C. The reaction was extracted with dichloromethane (200 mL*3) at 20°C. The reaction was dried over anhydrous Na2SO4 at 20°C. The organic phase was concentrated at 40°C to 50°C to obtain a residue. The product was purified by column chromatography (SiO2, petroleum ether: ethyl acetate = 20:1 to 1:3, R f =0.60) to purify the residue to obtain compound 3-3 (3.30 g, 5.60 mmol, 50.8% yield) as a white solid.

[0619]

[0620] Compound 3-3 (1.50 g, 2.54 mmol, 1.00 eq) was charged into R1 (500 mL three-necked flask) at 20°C. ACN (33.0 mL) was charged into R1 at 20°C. Compound 3 (1.09 g, 2.54 mmol, 1.00 eq) was charged into R1 at 20°C. K2CO3 (1.41 g, 10.1 mmol, 4 eq), KI (464 mg, 2.80 mmol, 1.10 eq), and CMPE (7.05 g, 70.4 mmol, 8.20 mL, 27.6 eq) were charged into R1 at 20°C. The mixture was heated to 90°C. The mixture was stirred at 90°C for 16 hours. TLC (dichloromethane: methanol = 5: 1) showed that most of the reactant 1 (R f =0.20) is consumed, and a new main spot (R f =0.50). H2O (100 mL) was loaded into R2 (1.00 L bucket) at 20°C. The reaction was loaded into R2 at 20°C. The reaction was extracted with dichloromethane (100 mL*3) at 20°C. The reaction was dried with anhydrous Na2SO4 at 20°C. The organic phase was concentrated at 40°C to 50°C to obtain a residue. The residue was purified by column chromatography (SiO2, dichloromethane:methanol=50:1 to 10:1, R f =0.50) to purify the residue to obtain target 13 (1.04 g, 1.11 mmol, 43.5% yield) as a colorless oil.

[0621] Example 5: Lipid Nanoparticles for Nucleic Acid Delivery

[0622] Lipid nanoparticles (LNPs) play a key role in effectively protecting nucleic acids and delivering them to cells for preventive and therapeutic applications. Although data from ongoing clinical trials are promising, clinical applications of genetic medicine require the discovery and development of more effective delivery systems. This article describes nanoparticles for gene and drug delivery applications.

[0623] EGFP mRNA, self-amplifying mRNA, SARS-CoV-2 mRNA lipid nanoparticles (LNP).

[0624] Target 1 (ARV-T1), target 11 (ARV-T11), target 12 (ARV-T12), target 13 (ARV-T13) or commercially available SM-102 lipids are used herein to prepare lipid nanoparticle (LNP) formulations for comparison. Lipids dissolved in ethanol are used to prepare LNP formulations at a molar ratio of 50:10:38.5:1.5 (ionizable lipids: DSPC: cholesterol: PEG-lipids). NanoAssemblr Ignite is used to combine lipid mixtures with 100mM sodium citrate buffer (pH 4.0) containing mRNA at a volume ratio of 3:1 (water: ethanol). The formulation is dialyzed for at least 16 hours in a Slide-A-Lyzer dialysis cassette for 10mMTris (pH 7.4) containing 8% sucrose, and concentrated using an Amicon ultracentrifugal filter, and then passed through a 0.22 μm filter and stored at 4°C or -20°C until use. Size, PDI and charge were measured by dynamic light scattering and Ribogreen spectrometry was used and Measuring mRNA packaging efficiency.

[0625] Figure 3A Particle diameter and polydispersity index (PDI) of LNPs are shown. Figure 3B Shown are the surface charge (Z potential) and mRNA encapsulation efficiency of SM-102 and ARV-T1 LNPs.

[0626] Plasmid DNA lipid nanoparticles (LNPs).

[0627] Lipid nanoparticle (LNP) formulations were prepared using target 1 (ARV-T1), target 11 (ARV-T11), target 12 (ARV-T12), or target 13 (ARV-T13) lipids for comparison. LNP formulations were prepared using lipids dissolved in ethanol and supplemented with DOTAP (10% molar ratio) lipids. The lipid mixture was combined with 100mM sodium citrate buffer (pH 4.0) containing mRNA in a volume ratio of 3:1 (water:ethanol) using NanoAssemblr Ignite. The formulation was dialyzed against 10mM Tris (pH 7.4) containing 8% sucrose in a Slide-A-Lyzer dialysis cassette for at least 16 hours and concentrated using an Amicon ultracentrifugal filter, and then passed through a 0.22μm filter and stored at 4°C or -20°C until use. Size, PDI, and charge were measured by dynamic light scattering, while Ribogreen spectrometry was used and Measuring mRNA packaging efficiency.

[0628] Figure 3A Particle diameter and polydispersity index (PDI) of LNPs are shown. Figure 3B Shown are the surface charge (Z potential) and mRNA encapsulation efficiency of SM-102 and ARV-T1 LNPs.

[0629] In vitro studies

[0630] 293T cells were transfected with mRNA encoding the SARS-CoV-2 spike protein. Figure 4 Shown is the change in in vitro expression of spike protein over time after cells were transfected with 0.5 and 1.0 μg / mL mRNA.

[0631] The transfection efficiency was studied by delivering GFP mRNA (1 μg / mL) into BHK cells using LNP delivery. After 24 hours, the transfection efficiency was determined by imaging the transfected BHK cells and analyzing GFP expression using flow cytometry. The results are shown in Figure 5A and Figure 5B middle.

[0632] In vivo studies

[0633] LNPs were formulated with indicated ionizable lipids and 1 μg of mRNA expressing luciferase was injected intramuscularly. After administration, luciferase expression was determined by whole body bioluminescence imaging using the IVIS spectrum in vivo imaging system at 6 hours, 24 hours, 48 ​​hours and 72 hours, respectively. The results of the in vivo transfection efficiency study of mRNA expressing luciferase are shown in FIG6 .

[0634] 1 μg of the vaccine formulated with target 1 lipid (ARV-T1) and mRNA encoding the full-length spike glycoprotein (delta variant) of SARS-CoV-2 was injected intramuscularly as planned. A formulation containing the commercially available lipid SM-102 and mRNA encoding the full-length spike glycoprotein (delta variant) of SARS-CoV-2 was prepared and used as a comparison. Total spike-specific total IgG was assessed on days 14 and 35 after the first immunization. The results are shown in Figure 7B and Figure 7C Neutralizing antibodies in serum were evaluated by pseudotyped virus. The results are shown in Fig.7D The results of evaluating antigen-specific T cell responses by Elispot are shown in Fig. 7E Data are expressed as mean ± SD. One-way ANOVA and Tukey's multiple comparison test were used for statistical comparison. *p<0.05, **p<0.01. The results demonstrate effective induction of immunity in vivo.

[0635] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosed invention belongs.Publications cited herein and the materials for which they are cited are specifically incorporated herein by reference.

[0636] Those skilled in the art will appreciate that various changes and modifications may be made to the preferred embodiments of the present invention, and such changes and modifications may be made without departing from the spirit of the present invention. Therefore, the following claims are intended to cover all such changes and modifications that fall within the true spirit and scope of the present invention.

Claims

1. A compound of formula Y: or a pharmaceutically acceptable salt thereof, wherein: R 1 is H or -CH3; represents a carbon-carbon bond or a carbon-carbon double bond; R 7 Is H or OR 16 ; R 8a and R 8b Each independently is H or OR 16 , or R 8a and R 8b Together with the atoms to which they are attached, they form a cycloalkyl, aryl, heterocycloalkyl or heteroaryl group; R 9 is H, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl; W is CR 4a or CR 4a R 4b , where if there is a double bond between W and the adjacent carbon, then W is CR 4a ; and if there is a single bond between W and the adjacent carbon, then W is CR 4a R 4b ; R 4a and R 4b Each of is independently H, halogen or C1-C6 alkyl; A is X is O or S; L 1 is -O(C=O)-, -C(=O)-, -S(O) x -, -C(=O)R a C(=O)-, -NR a C(=O)-, -NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-; L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -, -S-S-, -NR a C(=O)- or -C(=O)R a N-; L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-; R a yes wherein n is an integer from 0 to 12; x is 0, 1, or 2; M is CH, G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl; G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl; R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl; R 3 yes R 6 is H, OR b , CN, -C(=O)OR b , -NC(=O)R b , -C(=O)NR b , R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; p and q are independently 0, 1, 2, 3 or 4; R 35 yes L 4 Does not exist, yes R 10 Not present, or C1-C6 alkyl; L 5 does not exist, or or m is an integer 1, 2 or 3; L 6 Does not exist, yes R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl or C2-C9 heteroaryl; L 7 is C1-C6 alkylene; R 13a , R 13b and R 13c Each is independently C1-C6 alkyl or C6-C 10 Aryl; R 14 , R 16 , R 19 , R 21 , R 22 , R 24 , R 28 , R 29 and R 31 Each independently is H or C1-C6 alkyl; R 15 Yes-OR 16 、-NR 17 R 17 or R 17 Each independently is H, -OR 16 、C6-C 10 Aryl or C1-C6 alkyl; R 18 Each is independently halogen or C1-C6 alkyl; o1 is an integer from 0 to 8; p1 and p2 are each independently an integer from 0 to 2; Z is CH2, O, S or NR 16 ; s and t are each independently 0 or 1; R 23 is halo, hydroxy, C1-C6 alkyl or C1-C6 heteroalkyl; R 20 , R 25a and R 25b , R 30a , R 30b , R 30c , R 32a , R 32b and R 34 Each is independently a C1-C6 alkyl group; R 26a and R 26b are each independently H, C1-C6 alkyl, or R 26a and R 26b Combined with the atoms to which they are connected, Where R 26c and R 26d Each is independently H or substituted C1-C6 alkyl; R 27a and R 26b Each is independently H, hydroxyl or C1-C6 alkyl; u is 1, 2, or 3; R 33a and R 33b Each is independently C1-C6 alkyl, C1-C6 heteroalkyl, halogen or hydroxy; Q is O, S or NR 16 ;as well as or a pharmaceutically acceptable salt thereof.

2. The compound according to claim 1, wherein R 35 yes in L 4 Does not exist, yes R 10 Not present, or C1-C6 alkyl; L 5 does not exist, or or m is an integer 1, 2 or 3; L 6 Does not exist, yes and R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl or C2-C9 heteroaryl; or a pharmaceutically acceptable salt thereof.

3. A compound of formula I: or a pharmaceutically acceptable salt thereof, wherein: R 1 is H or -CH3; represents a carbon-carbon bond or a carbon-carbon double bond; L 4 Does not exist, yes R 10 Not present, or C1-C6 alkyl; L 5 does not exist, or or m is an integer 1, 2 or 3; L 6 Does not exist, yes R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl, C2-C9 heteroaryl; R 7 Is H or OR 16 ; R 8a and R 8b Each independently is H or OR 16 , or R 8a and R 8b Together with the atoms to which they are attached, they form a cycloalkyl, aryl, heterocycloalkyl or heteroaryl group; R 16 is H or C1-C6 alkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl; W is CR 4a or CR 4a R 4b , where if there is a double bond between W and the adjacent carbon, then W is CR 4a ; and If there is a single bond between W and the adjacent carbon, then W is CR 4a R 4b ; R 4a and R 4b Each of is independently H, halogen or C1-C6 alkyl; A is X is O or S; L 1 is -O(C=O)-, -C(=O)-, -S(O) x -, -C(=O)R a C(=O)-, -NR a C(=O)-, -NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-; L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -, -S-S-, -NR a C(=O)- or -C(=O)R a N-; L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-; R a yes wherein n is an integer from 0 to 12; x is 0, 1, or 2; M is CH, G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl; G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl; R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl; R 3 yes R 6 is H, OR b , CN, -C(=O)OR b , -NC(=O)R b , -C(=O)NR b , R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and p and q are independently 0, 1, 2, 3 or 4; or a pharmaceutically acceptable salt thereof.

4. A compound of formula Ia: or a pharmaceutically acceptable salt thereof, wherein: R 1 is H or -CH3; represents a carbon-carbon bond or a carbon-carbon double bond; L 4 Does not exist, yes L 5 does not exist, or or Where m is an integer 1, 2 or 3; L 6 Does not exist, yes R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl, C3-C 10 Aryl, C2-C9 heterocyclyl, C2-C9 heteroaryl; R 10 Not present, or C1-C6 alkyl; A is X is O or S; L 1 is -O(C=O)-, -C(=O)-, -S(O) x -, -C(=O)R a C(=O)-, -NR a C(=O)-, -NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-; L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -, -S-S-, -NR a C(=O)- or -C(=O)R a N-; L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-; R a yes wherein n is an integer from 0 to 12; x is 0, 1, or 2; M is CH, G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl; G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl; R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl; R 3 yes R 6 is H, OR b , CN, -C(=O)OR b , -NC(=O)R b , -C(=O)NR b , R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and p and q are independently 0, 1, 2, 3 or 4; or a pharmaceutically acceptable salt thereof.

5. The compound according to claim 1, wherein the compound is selected from: or a pharmaceutically acceptable salt thereof; Among them, A, R 1 , R 7 , R 8a , R 8b , R 9 , W is the same as formula Y.

6. The compound according to any one of claims 1 to 5, wherein the compound is selected from: or a pharmaceutically acceptable salt thereof, or any combination thereof; Wherein A is the same as formula Y.

7. The compound according to any one of claims 1 to 6, wherein the compound is selected from: or a pharmaceutically acceptable salt thereof; Wherein A is the same as formula Y.

8. A compound of formula II or III: or a pharmaceutically acceptable salt thereof, wherein: R 1 is H or -CH3; X is O or S; represents a carbon-carbon bond or a carbon-carbon double bond; L 4 Does not exist, yes R 10 Not present, or C1-C6 alkyl; L 5 does not exist, or or m is an integer 1, 2 or 3; L 6 Does not exist, yes R 4 It is C3-C 10 Alkyl, C3-C 10 Cycloalkyl, C3-C 10 Alkenyl, C3-C 10 Cycloalkenyl, C3-C 10 Alkynyl or its derivatives; L 1 is -O(C=O)-, -C(=O)-, -S(O) x -, -C(=O)R a C(=O)-, -NR a C(=O)-, -NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-; L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -, -S-S-, -NR a C(=O)- or -C(=O)R a N-; L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-; R a yes wherein n is an integer from 0 to 12; x is 0, 1, or 2; M is CH, G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl; G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl; R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl; R 3 yes R 6 is H, OR b , CN, -C(=O)OR b , -NC(=O)R b , -C(=O)NR b , R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and p and q are independently 0, 1, 2, 3 or 4; or a pharmaceutically acceptable salt thereof.

9. A compound according to any one of claims 1 to 8, wherein R 1 It is -CH3.

10. The compound according to any one of claims 1 to 9, wherein X is O.

11. The compound according to any one of claims 1 to 10, wherein m is 3.

12. A compound according to any one of claims 1 to 11, wherein L 4 yes 13. A compound according to any one of claims 1 to 12, wherein L 5 yes 14. A compound according to any one of claims 1 to 13, wherein L 6 Does not exist.

15. A compound according to any one of claims 1 to 14, wherein R 4 It is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, hexyl, heptyl, octyl, nonyl or decyl.

16. A compound according to any one of claims 1 to 15, wherein R 4 It is isopropyl.

17. A compound according to any one of claims 1 to 16, wherein M is CH.

18. A compound according to any one of claims 1 to 17, wherein R 3 yes 19. A compound according to any one of claims 1 to 17, wherein R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b , 20. A compound of formula IV or formula V: or a pharmaceutically acceptable salt thereof, wherein: L 1 is -O(C=O)-, -C(=O)-, -S(O) x -, -C(=O)R a C(=O)-, -NR a C(=O)-, -NC(=O)R a C(=O)- or -C(=O)NR a C(=O)-; L 2 is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -S(O) x -, -S-S-, -NR a C(=O)- or -C(=O)R a N-; L 3 does not exist, and is -O(C=O)-, -(C=O)O-, -O(C=O)O-, -C(=O)-, -O-, -NR a C(=O)- or -C(=O)R a N-; R a yes wherein n is an integer from 0 to 12; x is 0, 1, or 2; M is CH, G 1 , G 2 and G 3 Each independently selected from C1-C 12 Alkyl or C1-C 12 alkenyl; G 4 Not present or selected from C1-C 12 Alkyl or C1-C 12 alkenyl; R 2 Yes-CR b R c 、C6-C 24 Alkyl or C6-C 24 alkenyl; R 3 yes R 6 is H, OR b , CN, -C(=O)OR b , -NC(=O)R b , -C(=O)NR b , R b and R c independently H, C1-C 12 Alkyl or C1-C 12 alkenyl; and p and q are independently 0, 1, 2, 3 or 4.

21. The compound according to claim 20, wherein L 1 It is -C(=O)-.

22. A compound according to any one of claims 20 to 21, wherein L 2 It is -(C=O)O-.

23. A compound according to any one of claims 20 to 22, wherein L 3 Does not exist.

24. A compound according to any one of claims 20 to 23, wherein G 1 It is a C5 alkyl group.

25. A compound according to any one of claims 20 to 24, wherein G 2 It is a C6 alkyl group.

26. A compound according to any one of claims 20 to 25, wherein G 3 It is a C2 alkyl group.

27. A compound according to any one of claims 20 to 26, wherein G 4 Does not exist.

28. A compound according to any one of claims 20 to 27, wherein R 2 Yes-CR b R c .

29. A compound according to any one of claims 20 to 28, wherein R 3 yes 30. A compound according to any one of claims 20 to 29, wherein R b and R c It is a C8 alkyl group.

31. A compound according to any one of claims 20 to 30, wherein p and q are 0.

32. A compound according to any one of claims 20 to 31, wherein M is CH.

33. A compound according to any one of claims 20 to 32, wherein R 3 yes 34. A compound according to any one of claims 20 to 32, wherein R 6 Yes H, OR b 、CN、-C(=O)OR b 、-NC(=O)R b 、-C(=O)NR b , 35. A compound according to any one of claims 1 to 34, wherein the compound of formula Y, I, Ia or II to V can be selected from: or a pharmaceutically acceptable salt thereof.

36. A compound according to any one of claims 1 to 39, wherein the compound is selected from: or a pharmaceutically acceptable salt thereof, or any combination thereof.

37. A composition comprising: A compound according to any one of claims 1 to 36; and Active agent.

38. The composition of claim 37, wherein the active agent is a diagnostic agent, a prophylactic agent, or a therapeutic agent.

39. The composition of any one of claims 37 to 38, wherein the active agent is a nucleic acid, a protein / peptide, a small molecule, or any combination thereof.

40. The composition of 39, wherein the agent is a nucleic acid.

41. The composition of any one of claims 37 to 40, wherein the nucleic acid comprises double-stranded DNA, single-stranded DNA, complexed DNA, encapsulated DNA, naked RNA, encapsulated RNA, messenger RNA (mRNA), tRNA, short interfering RNA (siRNA), double-stranded RNA (dsRNA), micro RNA (miRNA), antisense RNA (asRNA), self-amplifying mRNA (saRNA), guide RNA (gRNA), cRNA, or any combination thereof.

42. The composition of claims 37 to 41, wherein the nucleic acid comprises mRNA.

43. A composition according to claim 42, wherein the mRNA encodes at least one hMPV antigenic polypeptide, parainfluenza virus (hPIV) type 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively) antigenic polypeptide, respiratory syncytial virus (RSV) antigenic polypeptide, measles virus (MeV) antigenic polypeptide, varicella-zoster virus antigenic polypeptide, influenza virus antigenic polypeptide, herpes simplex virus 1 (HSV1) antigenic polypeptide, herpes simplex virus 2 (HSV2) antigenic polypeptide, poxvirus (e.g., smallpox, monkeypox) antigenic polypeptide, African swine fever virus antigenic polypeptide, cytomegalovirus antigenic polypeptide, Epstein-Barr virus antigenic polypeptide, Rotavirus antigen polypeptides, rhinovirus antigen polypeptides, adenovirus antigen polypeptides, papillomavirus antigen polypeptides, poliovirus antigen polypeptides, mumps antigen polypeptides, rabies antigen polypeptides, rubella antigen polypeptides, coxsackievirus antigen polypeptides, equine encephalitis antigen polypeptides, Japanese encephalitis antigen polypeptides, yellow fever antigen polypeptides, Rift Valley fever antigen polypeptides, hepatitis A, B, C, D and E virus antigen polypeptides or coronavirus (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1) antigen polypeptides, or any combination thereof.

44. The composition of claim 43, wherein the coronavirus is a human coronavirus.

45. The composition of claim 44, wherein the human coronavirus is selected from HCoV-229E, HCoV-OC43, HCoV-HKU1, HCoV-NL, HCoV-NH, HCoV-NL63, SARS-CoV, SARS-CoV-2, and MERS-CoV.

46. ​​A composition according to any one of claims 44 to 45, wherein the human coronavirus is SARS-CoV-2.

47. A composition according to claim 42, wherein the mRNA encodes a tumor antigen, a cytokine (e.g., interferon, interleukin, colony stimulating factor), a monoclonal antibody or antibody fragment (single-chain fragment variable (scFv), fragment antigen binding ((Fab')2), intracellular antibody, nanobody), or any combination thereof.

48. The composition of any one of claims 36 to 47, wherein the composition comprises a lipid nanoparticle dispersion, a liposomal formulation, a lipid emulsion, or any combination thereof.

49. A lipid nanoparticle comprising: 20% to 80% of a compound according to any one of claims 1 to 36; 0% to 5% PEGylated lipids; 0% to 40% helper lipids; 0% to 80% sterols; as well as An active agent is encapsulated within the nanoparticles.

50. The nanoparticle of claim 49, wherein the active agent comprises a nucleic acid, a protein / peptide, a small molecule, or any combination thereof.

51. The nanoparticle of claim 50, wherein the active agent is a nucleic acid.

52. The nanoparticle of claim 51, wherein the nucleic acid comprises double-stranded DNA, single-stranded DNA, complexed DNA, encapsulated DNA, naked RNA, encapsulated RNA, messenger RNA (mRNA), tRNA, short interfering RNA (siRNA), double-stranded RNA (dsRNA), micro RNA (miRNA), antisense RNA (asRNA), self-amplifying mRNA (saRNA), guide RNA (gRNA), cRNA, or any combination thereof.

53. The nanoparticle of any one of claims 50 to 52, wherein the nucleic acid comprises mRNA.

54. A nanoparticle according to claim 53, wherein the mRNA encodes a tumor antigen, a cytokine (e.g., interferon, interleukin, colony stimulating factor), a monoclonal antibody or an antibody fragment (single-chain fragment variable (scFv), fragment antigen binding ((Fab')2), intracellular antibody, nanobody), or any combination thereof.

55. A nanoparticle according to claim 53, wherein the mRNA encodes at least one metapneumovirus (hMPV) antigenic polypeptide, parainfluenza virus (hPIV) type 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively) antigenic polypeptide, respiratory syncytial virus (RSV) antigenic polypeptide, measles virus (MeV) antigenic polypeptide, varicella-zoster virus antigenic polypeptide, influenza virus antigenic polypeptide, herpes simplex virus 1 (HSV1) antigenic polypeptide, herpes simplex virus 2 (HSV2) antigenic polypeptide, poxvirus (e.g., smallpox, monkeypox) antigenic polypeptide, African swine fever virus antigenic polypeptide, cytomegalovirus antigenic polypeptide, Epstein-Barr virus antigenic polypeptide, Rotavirus antigen polypeptides, rhinovirus antigen polypeptides, adenovirus antigen polypeptides, papillomavirus antigen polypeptides, poliovirus antigen polypeptides, mumps antigen polypeptides, rabies antigen polypeptides, rubella antigen polypeptides, coxsackievirus antigen polypeptides, equine encephalitis antigen polypeptides, Japanese encephalitis antigen polypeptides, yellow fever antigen polypeptides, Rift Valley fever antigen polypeptides, hepatitis A, B, C, D and E virus antigen polypeptides or coronavirus (e.g., MERS-CoV, SARS-CoV, SARS-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1) antigen polypeptides, or any combination thereof.

56. The nanoparticle of claim 55, wherein the coronavirus is a human coronavirus.

57. The nanoparticle of claim 56, wherein the human coronavirus is selected from the group consisting of HCoV-229E, HCoV-OC43, HCoV-HKU1, HCoV-NL, HCoV-NH, HCoV-NL63, SARS-CoV, SARS-CoV-2, and MERS-CoV.

58. The nanoparticle of any one of claims 55 to 57, wherein the human coronavirus is SARS-CoV-2.

59. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and an effective amount of the lipid nanoparticles according to any one of claims 49 to 58.

60. A method for delivering an active agent into a cell, the method comprising: The composition of any one of claims 37 to 48 or the nanoparticle of any one of claims 49 to 58 is introduced into the cell.

61. A method for treating or preventing an infection caused by an infectious agent, the method comprising administering to a subject in need thereof a composition according to any one of claims 37 to 48, a nanoparticle according to any one of claims 49 to 58.

62. A method of inducing an immune response to infection caused by an infectious agent, the method comprising administering to a subject in need thereof a composition according to any one of claims 37 to 48, a nanoparticle according to any one of claims 49 to 58.

63. The method of any one of claims 61 to 62, wherein the infectious agent comprises a bacterial organism such as Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, Corynebacterium diphtheriae, Listeria monocytogenes, Bacillus anthracis, Clostridium tetani, Clostridium botulinum, Clostridium perfringens, Neisseria meningitidis, Neisseria gonorrhoeae, Streptococcus mutans, Pseudomonas aeruginosa, Salmonella typhi, Haemophilus parainfluenzae, Bordetella pertussis, Francisella tularensis, Yersinia pestis, Cholera Vibrio, Legionella pneumophila, Mycobacterium tuberculosis, Mycobacterium leprae, Treponema pallidum, Leptospira interrogans, Borrelia burgdorferi, Campylobacter jejuni, etc.; viruses, such as metapneumoviruses such as human metapneumovirus (hMPV), parainfluenza viruses such as human parainfluenza virus (hPIV) types 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively), respiratory syncytial virus (RSV), measles virus (MeV), coronaviruses (e.g., MERS-CoV, SARS-CoV, SAR S-CoV2, HCoV-OC43, HCoV-229E, HCoV-NL63, HCoV-NL, HCoV-NH, HCoV-HKU1), poxviruses (e.g., smallpox, monkeypox), African swine fever virus, influenza A and B, human immunodeficiency virus (HIV), varicella-zoster, herpes simplex 1 and 2, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps Inflammation, rabies, rubella, coxsackievirus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E viruses, etc.; fungi, protozoa and parasitic organisms, such as Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans, Candida tropicalis, Nocardia asteroides, Rickettsia rickettsii, Rickettsia typhi, Mycoplasma pneumoniae, Chlamydia psittaci, Chlamydia trachomatis, Plasmodium falciparum, Trypanosoma brucei, Entamoeba histolytica, Toxoplasma gondii, Trichomonas vaginalis, Schistosoma mansoni, etc.

64. A method for treating or preventing a respiratory tract infection, the method comprising administering to a subject in need thereof a composition according to any one of claims 37 to 48, a nanoparticle according to any one of claims 49 to 58.

65. A method of inducing an immune response against a respiratory virus, the method comprising administering to a subject in need thereof a composition according to any one of claims 37 to 48, a nanoparticle according to any one of claims 49 to 58.

66. A method according to claim 65, wherein the respiratory tract infection is caused by metapneumovirus (MPV), human parainfluenza virus (hPIV) types 1, 2 and 3 (hPIV1, hPIV2 and hPIV3, respectively), RSV, measles virus (MeV) or coronavirus infection.

67. The method of claim 66, wherein the coronavirus is a human coronavirus.

68. The method of claim 67, wherein the human coronavirus is selected from HCoV-229E, HCoV-OC43, HCoV-HKU1, HCoV-NL63, SARS-CoV, SARS-CoV-2, and MERS-CoV.

69. The method of any one of claims 67 to 68, wherein the human coronavirus is SARS-CoV-2.

70. The method of any one of claims 61 to 69, wherein the subject is a veterinary patient.

71. The method of any one of claims 61 to 70, wherein the subject is a mammal.

72. The method of claim 71, wherein the mammal is a human.

73. The method of any one of claims 60 to 72, which can be administered orally, topically, transdermally, intra-articularly, intra-arteriole, intradermally, intraventricularly, intralesionally, intranasally, rectally, vaginally, by inhalation, via an implanted reservoir, subcutaneously, intravenously, intramuscularly, intra-articularly, intrasynovially, intrasternally, intrathecally, intraperitoneally, intrahepatically, intralesionally, by intracranial injection or infusion techniques.

74. A method for delivering an active agent into a cell, the method comprising: The composition of any one of claims 37 to 48 or the nanoparticle of any one of claims 49 to 58 is introduced into the cell.

75. The method of claim 74, wherein the composition or nanoparticle comprises an active agent comprising a nucleic acid, a protein / peptide, a small molecule, or any combination thereof.

76. The method of claim 75, wherein the active agent comprises a nucleic acid.

77. according to the method described in any one of claims 74 to 76, it comprises introducing the composition or nanoparticle comprising nucleic acid into specific cells or tissues to express the protein encoded by the nucleic acid to correct the defect caused by the lack of the nucleic acid in the cell or tissue.

78. according to the method described in any one of claims 74 to 76, it comprises the composition or nanoparticle that comprises nucleic acid is introduced into specific cells or tissue in vivo, and the protein encoded by the nucleic acid is expressed on the specific cells or tissue to induce the immune response of the experimenter.

79. A method according to any one of claims 74 to 76, comprising Introducing a composition or nanoparticle comprising a nucleic acid into specific cells or tissues ex vivo to express a protein encoded by the nucleic acid on the specific cells or tissues, thereby generating chimeric cells or tissues; and The chimeric cell or tissue is administered to a subject.

80. The method of any one of claims 74 to 76, wherein the specific cells are T cells (e.g., CAR T cells) derived from a subject.

81. The method of any one of claims 74 to 76, wherein the method introduces a polynucleotide to modulate gene expression, protein expression, or any combination thereof.

82. The method of any one of claims 74 to 76, wherein the method introduces a polynucleotide for cell reprogramming by regulating cell behavior.

83. The method of any one of claims 74 to 76, wherein the polynucleotide encodes an intracellular antibody.

84. according to the method described in any one of claims 74 to 76, it comprises introducing the composition or nanoparticle comprising nucleic acid into specific cells or tissues so that the expression of the protein encoded by the nucleic acid is silenced to correct the defect caused by the overexpression of the nucleic acid in the cell or tissue.

85. according to the method described in any one of claims 74 to 76, it comprises introducing the composition or nanoparticle comprising nucleic acid into specific cells or tissues, so that the expression of the protein encoded by nucleic acid on the specific cells or tissues is silenced to induce the immune response of the subject.

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