Techniques useful for oligonucleotide preparation

Abstract

Among other things, the present disclosure provides techniques for the preparation of oligonucleotides, particularly chiral controlled oligonucleotides, which provide significantly improved crude purity and yield, and significantly reduce manufacturing costs.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to U.S. Provisional Patent Application No. 62 / 821,423, filed on 20 March 2019, which is incorporated herein by reference in its entirety. [Background technology]

[0002] background Oligonucleotides can contain various modifications. Certain modifications, such as phosphorothioate internucleotide linkages, can introduce novel chiral centers into oligonucleotides. [Overview of the project] [Problems that the invention aims to solve]

[0003] overview Oligonucleotides are useful for many purposes. However, natural oligonucleotides have been shown to have drawbacks, such as low stability and low activity, which can reduce or even render them ineffective as therapeutic agents. [Means for solving the problem]

[0004] Specific technologies have been developed that can improve the properties and utility of oligonucleotides. For example, specific modifications that can improve the properties and utility of oligonucleotides, such as modifications to nucleic acid bases, sugars, and / or internucleotide bonds, have been described. Furthermore, technologies that enable the preparation of stereochemically and / or chirally controlled oligonucleotide compositions have been demonstrated to provide particularly useful and effective oligonucleotide compositions. Specific useful exemplary technologies are described, for example, in U.S. Patent Application Publication Nos. 20150211006, 20170037399, 20180216107, 20180216108, 20190008986, 20180216107, 20180216108, and 20190008986. These publications are described in one or more of the following: International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2018 / 067973, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 223056, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, etc. (each of which is incorporated herein by reference).

[0005] In particular, given the demonstrated desirability and usefulness of chiral-controlled oligonucleotide compositions, the applicant has understood that there would be great benefit in developing technologies that could improve or accelerate the production of oligonucleotide compositions, especially chiral-controlled oligonucleotide compositions. This disclosure describes specific such developments and provides technologies relating to oligonucleotide compositions, especially chiral-controlled oligonucleotide compositions. The technologies provided may be particularly useful, for example, with respect to therapeutic oligonucleotides.

[0006] In particular, this disclosure encompasses the recognition that certain techniques used in the preparation of oligonucleotides, especially chiral-controlled (e.g., sterically pure) oligonucleotide compositions, may involve the generation of certain impurities and / or the use of certain reagents and / or conditions, including conditions and / or sequence of steps, and that the associated production costs can be further reduced and the associated work can be further improved. In some embodiments, this disclosure therefore identifies the causes of problems and / or challenges related to strategies that have utilized such techniques. In some embodiments, this disclosure provides techniques (e.g., reagents, conditions, reactions, sequence of steps, cycles, methods, etc.) that are described and demonstrated to result in dramatic improvements in crude product purity and yield, significant improvements in work efficiency and / or reductions in production costs.

[0007] For example, in some embodiments, certain techniques provided utilize chiral auxiliary groups that can be readily removed with a base without utilizing HF and / or basic conditions and / or high temperatures, which can cause severe cleavage of oligonucleotide chains and / or undesirable conversion of certain internucleotide bonds (e.g., formation of native phosphate bonds from phosphorothioate internucleotide bonds (or their precursors) and / or neutral internucleotide bonds (or their precursors)). Such techniques can result in novel chemical compatibility and high stereoselectivity, crude purity, and yield as demonstrated herein. In some embodiments, useful compounds are compounds of formula I, formula Ia, formula Ia-1, formula Ia-2, formula Ib, formula Ic, formula Id, formula Ie, formula II, formula II-a, formula II-b, formula III, formula III-a, or formula III-b, or salts thereof. In particular, such compounds can be used, for example, as chiral auxiliary groups in preparing chiral-controlled oligonucleotide compositions as demonstrated herein.

[0008] Oligonucleotide synthesis typically utilizes highly efficient chemical transformations in its steps. However, despite this efficiency, the product of one or more steps may contain one or more reactive functional groups, such as unreacted 5'-OH groups in coupling products and / or newly formed reactive groups (e.g., primary and / or secondary amino groups) when chiral auxiliaries are used in chiral-controlled oligonucleotide synthesis, which can introduce a large amount of impurities if capping is absent. Often, such reactive functional groups are capped during oligonucleotide synthesis to reduce the resulting impurities. In some embodiments, this disclosure encompasses the recognition of the problem that capping steps, such as those typically used in conventional phosphoramidite-based oligonucleotide synthesis, can lead to the generation of large amounts of byproducts, particularly in many chiral-controlled (stereo-controlled, stereoselective) oligonucleotide synthesis processes. In particular, this disclosure provides techniques to address these problems.

[0009] In some embodiments, the disclosure provides a method comprising, for example, a post-modification capping step after a modification step, but before a subsequent deblocking and / or coupling step. In some embodiments, the post-modification capping step is after a modification step that provides a chiral-controlled internucleotide bond, but before a subsequent deblocking and / or coupling step.

[0010] In some embodiments, the Disclosure provides a method comprising a capping step with a different chemical strategy compared to a reference capping step in conventional oligonucleotide synthesis. For example, in some embodiments, the Disclosure provides a method comprising one or more capping steps, each of which selectively caps amino groups rather than hydroxyl groups (compared to, for example, a reference capping reagent system in conventional oligonucleotide synthesis). In some embodiments, the provided capping step is selective to amidation rather than esterification. In some embodiments, the capping reagent system for the capping step contains no or only reduced levels of strongly nucleophiles and / or esterification catalysts (or reagents capable of providing them when in contact with the composition to be capped) (compared to, for example, a reference capping reagent system in conventional oligonucleotide synthesis), and contains no or only reduced levels of, for example, DMAP, NMI, etc. In some embodiments, the Disclosure provides a method comprising a capping step capable of efficiently capping both amino and hydroxyl groups, for example, a capping step equivalent to or identical to a reference capping step in conventional oligonucleotide synthesis.

[0011] In some embodiments, the disclosure provides a method for realizing oligonucleotide synthesis that includes the same or different chemical strategies for a capping step and can provide various advantages, such as improved crude purity and improved yield, particularly for chiral-controlled (stereo-controlled, stereoselective) oligonucleotide synthesis. In some embodiments, the disclosure provides a method for a pre-modification capping step (after a coupling step, but before the next modification step) and a post-modification capping step (after a modification step, but before the next deblocking and / or coupling step). In some embodiments, the pre-modification capping step and the post-modification capping step are different. In some embodiments, the pre-modification capping step and the post-modification capping step have different chemical strategies. In some embodiments, the pre-modification capping step selectively caps amino groups rather than hydroxyl groups (compared to, for example, a reference capping reagent system in conventional oligonucleotide synthesis). In some embodiments, the post-modification capping step can cap both amino groups and hydroxyl groups (compared to, for example, a reference capping reagent system in conventional oligonucleotide synthesis). In some embodiments, the pre-modification capping step selectively caps amino groups rather than hydroxyl groups (compared to, for example, reference capping reagent systems in conventional oligonucleotide synthesis). In some embodiments, the pre-modification capping step can cap both amino and hydroxyl groups (compared to, for example, reference capping reagent systems in conventional oligonucleotide synthesis).

[0012] In some embodiments, the method provided includes two or more capping steps in the oligonucleotide synthesis cycle. In some embodiments, the method provided includes two capping steps in the oligonucleotide synthesis cycle, wherein the two steps are separated by modification steps, such as oxidation or sulfidation. In some embodiments, the method provided includes a step in which a chiral modified internucleotide bond containing a chiral bound phosphorus is formed in a stereoselective manner of at least 80:20, 85:15, 90:10, 91:9, 92:8, 93:7, 94:6, 95:5, 96:4, 97:3, 98:2, or 99:1, favorably in either an Rp configuration or an Sp configuration.

[0013] In some embodiments, this disclosure To provide chiral nucleoside phosphoramidites containing chiral atoms other than phosphorus atoms or sugar carbon atoms; and Capping step immediately following the sulfidation or oxidation step This provides a method that includes [something].

[0014] In some embodiments, this disclosure To provide a chiral nucleoside phosphoramidite containing a chiral atom that is neither a phosphorus atom nor a nucleoside unit atom; and Capping step immediately following the sulfidation or oxidation step This provides a method that includes [something].

[0015] In some embodiments, this disclosure To provide an oligonucleotide intermediate containing a chiral-bonded phosphorus atom, which is bonded to a chiral unit that does not contain a nucleoside unit or a part thereof; and Capping step immediately following the sulfidation or oxidation step This provides a method that includes [something].

[0016] In some embodiments, this disclosure To provide an oligonucleotide intermediate containing a chiral-bonded phosphorus atom that is bonded to a chiral unit that does not contain an atom of the nucleoside unit; and Capping step immediately following the sulfidation or oxidation step This provides a method that includes [something].

[0017] In some embodiments, this disclosure (1) Coupling step; (2) Optionally, a pre-modification capping step; (3) Modification step; (4) Optionally, a post-modification capping step; and (5) Optionally, the deblocking step The present invention provides a method for preparing, for example, an oligonucleotide, which includes [a specific component].

[0018] Exemplary coupling steps, pre-modification capping steps, modification steps, post-modification capping steps, and deblocking steps are described herein. In some embodiments, the cycle includes all optional steps.

[0019] In some embodiments, the present disclosure provides a method for preparing a composition comprising a plurality of oligonucleotides, the method being, (1) A coupling step, Contacting a deblocked composition (deblocked oligonucleotide composition) containing multiple deblocked oligonucleotides or nucleosides, each independently containing a free hydroxyl group, with a coupling reagent system containing a partner compound containing a nucleoside unit; and The coupling of the partner compound with the free hydroxyl groups of multiple declocked oligonucleotides or nucleosides. Includes; A coupling product composition is provided, comprising a plurality of coupling product oligonucleotides, each independently containing an internucleotide bond that links the hydroxyl group of a deblocked oligonucleotide to the nucleoside unit of a partner compound; (2) Optionally, a pre-modification capping step, Contacting the coupling product composition with the pre-modification capping reagent system; and Capping one or more functional groups of a coupling product composition. Includes; A pre-capping step that provides a pre-capping product composition containing multiple pre-capping product oligonucleotides; (3) Modification step Contacting a coupling product composition with a modification reagent system containing a modification reagent, and modifying one or more internucleotide bonds of one or more coupling product oligonucleotides; or The process involves contacting a pre-modification capping product composition with a modification reagent system and modifying one or more bonds of one or more pre-modification capping product oligonucleotides. Includes; A modification step that provides a modified product composition containing multiple modified oligonucleotide products; (4) Optionally, the post-modification capping step is: Contacting the modified product composition with the post-modification capping reagent system; and Capping one or more functional groups of multiple oligonucleotides in a modified product composition. Includes; A post-capping step that provides a post-capping product composition comprising multiple post-capping product oligonucleotides; (5) Optionally, a deblocking step, Contact the modified product composition or the post-modified capping product composition with the deblocking reagent system. Includes; A deblocking step provides a deblocking product composition comprising a plurality of deblocking product oligonucleotides, each independently containing a free hydroxyl group; and (6) Optionally, repeat steps (1) to (5) multiple times (for example, 1 to 100, 1 to 90, 1 to 80, 1 to 70, 1 to 60, 1 to 50, 1 to 40, 1 to 30, 1 to 25, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25 times, etc.; or so that a desired length of oligonucleotide is achieved). Includes.

[0020] In some embodiments, the method provided includes one or more pre-modification capping steps. In some embodiments, the method provided includes one or more post-modification capping steps. In some embodiments, the method provided includes one or more pre-modification and post-modification capping steps. In some embodiments, the method provided includes one or more deblocking steps.

[0021] In some embodiments, this disclosure (1) Coupling step; (2) Optionally, a pre-modification capping step; (3) Modification step; (4) Optionally, a post-modification capping step; and (5) Optionally, the deblocking step The present invention provides a method for preparing, for example, oligonucleotides, comprising one or more cycles, each independently containing a specific compound.

[0022] In some embodiments, the Disclosure provides a method for preparing a composition comprising a plurality of oligonucleotides, the method comprising one or more cycles, each cycle independently (1) A coupling step, Contacting a deblocked composition (deblocked oligonucleotide composition) containing multiple deblocked oligonucleotides or nucleosides, each independently containing a free hydroxyl group, with a coupling reagent system containing a partner compound containing a nucleoside unit; and The coupling of the partner compound with the free hydroxyl groups of multiple declocked oligonucleotides or nucleosides. Includes; A coupling product composition is provided, comprising a plurality of coupling product oligonucleotides, each independently containing an internucleotide bond that links the hydroxyl group of a deblocked oligonucleotide to the nucleoside unit of a partner compound; (2) Optionally, a pre-modification capping step, Contacting the coupling product composition with the pre-modification capping reagent system; and Capping one or more functional groups of a coupling product composition. Includes; A pre-capping step that provides a pre-capping product composition containing multiple pre-capping product oligonucleotides; (3) Modification step, Contacting a coupling product composition with a modification reagent system containing a modification reagent, and modifying one or more internucleotide bonds of one or more coupling product oligonucleotides; or The process involves contacting a pre-modification capping product composition with a modification reagent system and modifying one or more bonds of one or more pre-modification capping product oligonucleotides. Includes; A modification step that provides a modified product composition containing multiple modified oligonucleotide products; (4) Optionally, the post-modification capping step is: Contacting the modified product composition with the post-modification capping reagent system; and Capping one or more functional groups of multiple oligonucleotides in a modified product composition. Includes; A post-capping step that provides a post-capping product composition comprising multiple post-capping product oligonucleotides; (5) Optionally, a deblocking step, Contact the modified product composition or the post-modified capping product composition with the deblocking reagent system. Includes; A deblocking step provides a deblocking product composition comprising a plurality of deblocking product oligonucleotides, each independently containing a free hydroxyl group. Includes.

[0023] In some embodiments, the cycle includes one or more pre-modification capping steps. In some embodiments, the cycle includes one or more post-modification capping steps. In some embodiments, the cycle includes one or more pre-modification and post-modification capping steps. In some embodiments, the cycle includes one or more deblocking steps. In some embodiments, the cycle includes a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step. In some embodiments, the cycle includes a coupling step, a pre-modification capping step, a modification step, and a deblocking step. In some embodiments, the cycle includes a coupling step, a modification step, a post-modification capping step, and a deblocking step. In some embodiments, the cycle includes a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step. In some embodiments, one or more cycles include a coupling step, a pre-modification capping step, a modification step, and a deblocking step. In some embodiments, one or more cycles include a coupling step, a modification step, a post-modification capping step, and a deblocking step.

[0024] In some embodiments, the cycle includes one or more, but not all, optional steps. In some embodiments, the cycle includes a pre-modification step. In some embodiments, the cycle does not include a pre-modification step. In some embodiments, the cycle includes a post-modification step. In some embodiments, the cycle does not include a post-modification step. In some embodiments, the cycle, for example, the final cycle when the desired length of the oligonucleotide is achieved, does not include a deblocking step.

[0025] In some embodiments, each step of the cycle is independently selected from a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step. In some embodiments, the cycle may include two or more identical steps (e.g., two coupling steps), each of which may utilize the same or different reagents, conditions, etc. As those skilled in the art will understand, in some embodiments, for example, if the reaction of a step does not reach completion, it may be beneficial to repeat such a step either immediately after the first such step or after one or more subsequent steps. In many embodiments, the repetition is performed before the next deblocking step is carried out. For example, in some embodiments, a coupling step is repeated one or more times immediately after another coupling step. In some embodiments, a series of steps (e.g., (1)-(2), (1)-(3), (1)-(2)-(3), (1)-(3)-(4), (1)-(2)-(3)-(4)) is repeated one or more times.

[0026] When used in this disclosure, “one or more” means one in some embodiments. In some embodiments, “one or more” means two or more. In some embodiments, "one or more" means 1-200, 1-150, 1-100, 1-90, 1-80, 1-70, 1-60, 1-50, 1-40, 1-30, 1-25, 1-24, 1-23, 1-22, 1-21, 1-20, 5-200, 5-150, 5-100, 5-90, 5-80, 5-70, 5-60, 5-50, 5-40, 5-30, 5-25, 5-24, 5-23, 5-22, 5-21, 5-20, 10-200, 10-150, 10-100, 10-90, 10-80, 10-70, 10-60, 10-50, 10-40, 10-30, 10 ~25, 10~24, 10~23, 10~22, 10~21, 10~20, 15~200, 15~150, 15~100, 15~90, 15~80, 15~70, 15~60, 15~50, 15~40, 15~30, 15~25, 15~24, 15~23, 15~22, 15~21 , 15-20, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 or more.

[0027] When used in this disclosure, “at least one” means one in some embodiments. In some embodiments, “at least one” means two or more. In some embodiments, “at least one” means 1-200, 1-150, 1-100, 1-90, 1-80, 1-70, 1-60, 1-50, 1-40, 1-30, 1-25, 1-24, 1-23, 1-22, 1-21, 1-20, 5-200, 5-150, 5-100, 5-90, 5-80, 5-70, 5-60, 5-50, 5-40, 5-30, 5-25, 5-24, 5-23, 5-22, 5-21, 5-20, 10-200, 10-150, 10-100, 10-90, 10-80, 10-70, 10-60, 10-50, 10-40, 10-30, 1 0-25, 10-24, 10-23, 10-22, 10-21, 10-20, 15-200, 15-150, 15-100, 15-90, 15-80, 15-70, 15-60, 15-50, 15-40, 15-30, 15-25, 15-24, 15-23, 15-22, 15-2 1, 15-20, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 or more.

[0028] In some embodiments, one or more cycles include a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step, and one or more cycles include a coupling step, a pre-modification capping step, a modification step, and a deblocking step. In some embodiments, one or more cycles include a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step, and one or more cycles include a coupling step, a modification step, a post-modification capping step, and a deblocking step.

[0029] In some embodiments, the cycle consists of a coupling step, a pre-modification capping step, a modification step, a post-modification capping step, and a deblocking step. In some embodiments, the cycle consists of a coupling step, a pre-modification capping step, a modification step, and a deblocking step. In some embodiments, the cycle consists of a coupling step, a modification step, a post-modification capping step, and a deblocking step.

[0030] In some embodiments, the coupling step is immediately followed by a pre-modification capping step, for example, within a certain cycle. In some embodiments, the coupling step is immediately followed by a modification step, for example, within a certain cycle. In some embodiments, the coupling step is immediately followed by a modification step, for example, within a certain cycle, which includes an oxidation reaction that converts a P(III) bond to a P(VI) phosphate bond (for example, including the introduction of =O to the P(III) bonded phosphorus). In some embodiments, the pre-modification step is immediately followed by a modification step, for example, within a certain cycle. In some embodiments, the modification step includes the formation of a bond (for example, a single bond, a double bond, etc.) between the bonded phosphorus and a sulfur or nitrogen atom. In some embodiments, the modification step is immediately followed by a post-modification capping step. In some embodiments, the post-modification capping step is immediately followed by a deblocking step.

[0031] As those skilled in the art will understand, the steps described herein may include one or more washes (e.g., washing of oligonucleotides on a support) using a variety of suitable solvents (which in some embodiments may be mixtures of chemicals), which may be carried out before and / or after steps in the various methods and / or cycles described herein. For example, as demonstrated in the examples herein, after carrying out the reaction of a step, the oligonucleotides on the solid support are typically thoroughly washed before carrying out another reaction in the same or immediately following step (e.g., thereby removing excess reagent, removing undesirable products, changing the solvent, and conditioning for the next reaction). In some embodiments, the steps described herein, e.g., the coupling step, the pre-modification capping step, the modification step, the post-modification capping step, or the deblockization step, optionally include one or more washes. In some embodiments, as demonstrated in the examples, the reagents, solvents, and / or reagent systems for the reaction are removed after the reaction has been carried out. In some embodiments, removal is carried out by filtration and / or washing when the product oligonucleotides are on a solid support, for example, when a solid support is used for oligonucleotide synthesis.

[0032] In some embodiments, the disclosure includes the recognition that conventional capping conditions, when used as in conventional oligonucleotide synthesis, can be a significant source of various problems under certain circumstances, and can contribute to the formation of one or more by-products (impurities) and significantly lower oligonucleotide crude purity and yield, particularly in the stereoselective preparation of oligonucleotides containing one or more chiral internucleotide bonds. In particular, the disclosure provides techniques including capping strategies that, when used in combination with other steps in oligonucleotide synthesis, can yield remarkably high crude impurities and yields compared to appropriate reference techniques.

[0033] In some embodiments, the reference technique uses conventional capping conditions, as found in conventional phosphoramidite-based oligonucleotide synthesis, to contact the oligonucleotide and cap the hydroxyl group (e.g., unreacted 5'-OH group), which is typically an esterification condition that acylates the hydroxyl group by using a mixture of, for example, an acylating agent (e.g., acetic anhydride), a base (e.g., 2,6-lutidine), and a catalyst (e.g., N-methylimidazole, DMAP, etc.). Conventional capping conditions typically involve using a considerable amount of acylating agent, base, and catalyst for capping, generally independently of each other, in amounts of about 5% to 15% by volume and / or at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 equivalents relative to the oligonucleotide loading capacity of the support (e.g., the loading capacity of the support used in the preparation of the oligonucleotide can be calculated by multiplying the unit loading capacity of the support (e.g., umol / g) by the amount of support (g)). In some embodiments as used in conventional oligonucleotide synthesis, each synthetic cycle of the reference technique includes a single capping step. In some embodiments, the reference technique comprises only one capping step in each of its synthesis cycles, where the capping is carried out under esterification conditions comprising, for example, an acyling agent (e.g., acetic anhydride), a base (e.g., 2,6-lutidine), and a catalyst (e.g., N-methylimidazole (NMI), DMAP, etc.) (each independently in an amount of about 1 vol%, 2 vol%, 3 vol%, 4 vol%, 5 vol%, 6 vol%, 7 vol%, 8 vol%, 9 vol%, 10 vol%, 11 vol%, 12 vol%, 13 vol%, 14 vol%, or 15 vol% or more of the capping reagent solution and / or the catalyst in an amount of about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.5, or 2 equivalents or more relative to the acyling agent and / or base).

[0034] In some embodiments, the Disclosure provides a technique comprising one or more capping steps, e.g., a pre-modification capping step, a post-modification capping step, etc., each independently equivalent to or identical to a reference capping step in conventional oligonucleotide synthesis based on phosphoramidite chemistry, for example. In some embodiments, the Disclosure reduces byproducts that may be formed in such capping steps by strategically positioning the location (or timing) of the capping steps in the oligonucleotide synthesis method and / or cycle. In some embodiments, such capping steps are positioned either as separate individual capping steps or in combination with other capping steps (e.g., capping steps that selectively cap amino groups, often 5'-OH groups) after amino groups (typically primary and secondary) have been selectively capped (often more selectively than free hydroxyl groups, particularly 5'-OH groups).

[0035] In some embodiments, the Disclosure provides a technique comprising one or more capping steps each independently comprising conditions that are selective or specific to amidation rather than esterification. In some embodiments, the Disclosure provides a technique comprising one or more capping steps using amidation conditions that are not efficient and / or typical esterification conditions. As will be readily apparent to those skilled in the art, esterification and amidation have been extensively studied, and various conditions that are selective or specific to amidation rather than esterification, and various methods for evaluating the selectivity and / or specificity of amidation compared to esterification, are widely known in the art and can be utilized in this Disclosure. For example, typical conditions that are selective or specific to amidation rather than esterification are anhydrous and base (e.g., Ac2O and 2,6-lutidine) without a catalyst, as the corresponding efficient esterification conditions typically require anhydrous, base and catalyst (e.g., Ac2O, 2,6-lutidine and NMI), as are the conventional capping conditions. In some embodiments, the disclosure provides a technique comprising one or more synthetic cycles each comprising independently a coupling step, a modification step (e.g., oxidation, sulfidation, etc.), and one or more capping steps, wherein each capping step after the coupling step and before the modification step includes amidation conditions but does not include esterification conditions. In some embodiments, the amidation conditions include 0.01 vol%, 0.02 vol%, 0.05 vol%, 0.1 vol%, 0.2 vol%, 0.5 vol%, 1 vol%, 2 vol%, 3 vol%, 4 vol% or 5 vol% or less of an esterification catalyst under appropriately corresponding conditions with respect to the acyling agent and / or oligonucleotide loading capacity of the support (having the same acyling agent and base), and / or include about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1 or 1.2 equivalents or less of an esterification catalyst under appropriately corresponding conditions (having the same acyling agent and base). In some embodiments, the acyling agent is an anhydride. In some embodiments, the acylating agent is Ac2O. In some embodiments, the catalyst is NMI. In some embodiments, the catalyst is DMAP. In some embodiments, the catalyst is a nucleophilic nitrogen base.

[0036] Without intending to impose any theoretical limitations, in some embodiments, this disclosure encompasses the recognition of the causes of problems in oligonucleotide synthesis, particularly when nucleophiles are used in capping steps after the coupling step and before the modification step in stereoselective oligonucleotide preparation, which can contribute to the generation of by-products and the overall preparation efficiency and / or crude purity, for example, by degrading the oligonucleotide or reducing the performance of other steps. Accordingly, in some embodiments, this disclosure provides capping techniques that include only significantly reduced levels of a strong nucleophile, such as DMAP or NMI, used in typical capping conditions, or none at all, in contrast to conventional capping conditions that may include a large amount of nucleophilic catalyst (e.g., 5% to 15% by volume of NMI in some cases). In some embodiments, each of one or more capping steps after the coupling step and before the modification step in the oligonucleotide preparation cycle independently includes only significantly reduced levels of a strong nucleophile or none at all. In some embodiments, the reduced level is 0.01%, 0.02%, 0.05%, 0.1%, 0.2%, 0.5%, 1%, 2%, 3%, 4%, or 5% or less by volume of the capping reagent solution. In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, or 1.2 equivalents or less compared to the acylating agent. In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, or 1.2 equivalents or less compared to the oligonucleotide loading capacity of the support.

[0037] In some embodiments, the nucleophile is a nucleophile base. In some embodiments, the nucleophile base is a nitrogen base. In some embodiments, the nucleophile base is a nitrogen base, where the basic nitrogen atom (e.g., =N- or -N(-)-) has no α substituents. In some embodiments, the nucleophile base is a nitrogen base, where the basic nitrogen atom (e.g., =N- or -N(-)-) has no α substituents that are not part of the ring. In some embodiments, the nucleophile base is an optionally substituted 5- to 10-membered ring heteroaryl compound containing a basic nitrogen atom =N-, where the nitrogen atom has fewer than two α substituents or none at all. In some embodiments, the nucleophile base is a nucleophile nitrogen base. In some embodiments, the nucleophile nitrogen base is of formula BI: N(R N )3 BI It is a compound with the structure, in which each R N These are independently R, and the three R groups together with the nitrogen atom form a bicyclic or polycyclic ring which is optionally substituted with respect to the R group (and which group may be R) as described in this disclosure, [ka] The nitrogen (underlined) is a tertiary nitrogen and is not substituted at any position relative to the nitrogen atom at the α-position. In some embodiments, the formed ring is saturated. In some embodiments, the nucleophile is DABCO(1,4-diazabicyclo[2.2.2]octane). In some embodiments, the formed ring contains one or more unsaturated atoms.

[0038] In some embodiments, the nucleophilic nitrogen base is a base containing =N-, where there is no substitution at any position relative to the nitrogen atom at the α-position. In some embodiments, the nucleophilic nitrogen base is a base containing an aromatic moiety containing =N-, where there is no substitution at any position relative to the nitrogen atom at the α-position. In some embodiments, the nucleophilic nitrogen base is of formula B-II: R N -CH=N-CH=CH-R N, B-II It is a compound with the structure, in which each R N These are independently R, and the two R groups together with their intervening atoms form a ring which is optionally substituted as described herein, where the compound contains -CH=N-CH=. In some embodiments, the formed ring is optionally substituted C5- containing 0 to 10 heteroatoms in addition to the nitrogen atom. 30 It is a heteroaryl ring. In some embodiments, the ring formed is an optionally substituted five-membered heteroaryl ring. In some embodiments, the ring formed is a substituted five-membered heteroaryl ring. In some embodiments, the ring formed is a substituted imidazolyl ring. In some embodiments, the nucleophile is a substituted imidazole. In some embodiments, the nucleophile nitrogen base is NMI. In some embodiments, the ring formed is an optionally substituted six-membered heteroaryl ring. In some embodiments, the ring formed is a substituted six-membered heteroaryl ring. In some embodiments, the ring formed is a substituted pyridinyl ring. In some embodiments, the nucleophile is a substituted pyridine. In some embodiments, the nucleophile nitrogen base is DMAP.

[0039] As those skilled in the art will understand, nucleophilicity, such as the nucleophilicity of a basic nitrogen atom in a base, is related to several factors, such as steric hindrance and electron density. Techniques for evaluating nucleophilicity are widely known in the art and can be utilized in this disclosure. In addition or alternatively, bases with various levels of nucleophilicity are well known and can be evaluated and / or utilized in this disclosure. In some embodiments, bases that can efficiently catalyze esterification reactions, such as bases that can be used in combination with anhydrides and 2,6-lutidine in conventional oligonucleotide synthesis for efficient capping of unreacted 5'-OH (e.g., DMAP, NMI, etc.), are strongly nucleophilic bases and should be avoided or used at reduced levels in capping steps that contain only significantly reduced levels or none at all of the strongly nucleophile, such as any capping step after the coupling step and before the modification step. In some embodiments, strongly nucleophilic bases are bases that can effectively replace DMAP or NMI in esterification. In some embodiments, ferrophilic bases are bases that can effectively replace DMAP or NMI in the capping step of conventional oligonucleotide synthesis (which typically uses phosphoramidite chemistry, does not use chiral auxiliaries, and is considered non-stereoselective / uncontrolled).

[0040] In some embodiments, the provided method includes a capping step in which a nucleophile is present in or absent in amounts of 0.1, 0.2, 0.3, 0.4, 0.5, 0.5, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 equivalents or less compared to the loading capacity of the oligonucleotide or support. In some embodiments, such a capping step is immediately before a non-capping step. In some embodiments, such a capping step is immediately after a non-capping step. In some embodiments, such a capping step is immediately before and immediately after a non-capping step. In some embodiments, the non-capping step is a coupling step. In some embodiments, the non-coupling step is a modification step. In some embodiments, the non-capping step immediately before such a capping step is a coupling modification step.

[0041] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain a nucleophile, or, if it contains one or more nucleophiles, the level of each of the one or more nucleophiles is independently reduced compared to appropriate reference capping conditions.

[0042] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain a nucleophile, or, if it contains one or more nucleophiles, the level of each of the one or more nucleophiles is independently about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, or 1 equivalent or less relative to the first nucleoside incorporated by the oligonucleotide.

[0043] In some embodiments, the first nucleoside incorporated into the oligonucleotide is the first nucleoside loaded onto the support before the first cycle of internucleotide bonding. In some embodiments, the equivalent of the first nucleoside incorporated into the oligonucleotide relative to the oligonucleotide loading capacity of the support used for oligonucleotide preparation is 1.

[0044] In some embodiments, the nucleophile is a nucleophilic base as described in this disclosure. In some embodiments, the nucleophilic base is a compound of formula BI. In some embodiments, the nucleophilic base is a compound of formula BI and can be used for efficient capping in conventional phosphoramidite-based oligonucleotide synthesis. In some embodiments, the nucleophilic base is a compound of formula B-II. In some embodiments, the nucleophilic base is a compound of formula B-II and can be used for efficient capping in conventional phosphoramidite-based oligonucleotide synthesis. In some embodiments, the nucleophilic base is DMAP. In some embodiments, the nucleophilic base is a nucleophilic base in NMI.

[0045] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain a catalyst that promotes the capping of the 5'-OH group, as in appropriate reference capping conditions, or, if it contains one or more such catalysts, the level of each of the one or more such catalysts is independently reduced compared to appropriate reference capping conditions.

[0046] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain a catalyst that promotes 5'-OH capping, as in appropriate reference capping conditions, or, if it contains one or more such catalysts, the level of each of the one or more such catalysts is independently 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, or 1 equivalent or less relative to the first incorporated nucleoside of the oligonucleotide.

[0047] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain an esterification catalyst, or, if it contains one or more esterification catalysts, the level of each of the one or more such catalysts is independently reduced compared to appropriate reference capping conditions.

[0048] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) either does not contain an esterification catalyst, or, if it contains one or more esterification catalysts, the level of each of the one or more such catalysts is independently 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, or 1 equivalent or less relative to the first nucleoside incorporated into the oligonucleotide.

[0049] In some embodiments, the catalyst, for example for esterification, which promotes 5'-OH capping in oligonucleotide synthesis, is a compound of formula BI. In some embodiments, the catalyst is a compound of formula BI and can be used for efficient capping in conventional phosphoramidite-based oligonucleotide synthesis. In some embodiments, the catalyst is a compound of formula B-II. In some embodiments, the catalyst is a compound of formula B-II and can be used for efficient capping in conventional phosphoramidite-based oligonucleotide synthesis. In some embodiments, the catalyst is DMAP. In some embodiments, the catalyst is a strongly nucleophilic base in NMI.

[0050] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) includes conditions that are more selective to amidation than to esterification.

[0051] In some embodiments, the present disclosure provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles, where, Each cycle independently forms an internucleotide bond; Each cycle independently comprises a coupling step, one or more capping steps, and a modification step, the coupling step forming an internucleotide bond, and the modification step modifying the internucleotide bond formed in the coupling step; Each capping step between the coupling step and the modification step (pre-modification capping step) includes conditions that are more selective to amidation than to esterification, and does not include conditions that are the same as or equivalent to the appropriate reference conditions.

[0052] In some embodiments, conditions that are more selective to amidation than esterification include including only reduced levels or no esterification catalyst at all, e.g., no DMAP, NMI, etc. In some embodiments, capping conditions in conventional phosphoramidite-based oligonucleotide synthesis can be replaced using conditions identical or equivalent to suitable reference conditions without significantly reducing efficiency, crude purity, and / or yield (or with a reduction of 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% or less).

[0053] In some embodiments, the method provided includes a second capping step after a modification step in one or more cycles. In some embodiments, the method provided includes a second capping step after a modification step and before a deblocking step (which deblocks blocked hydroxyl groups) in one or more cycles. In some embodiments, the second capping step includes a nucleophile. In some embodiments, the second capping step includes a nucleophile at a level equivalent to the reference capping conditions. In some embodiments, the second capping step includes an esterification catalyst. In some embodiments, the second capping step includes an esterification catalyst at a level equivalent to the reference capping conditions. In some embodiments, the second capping step includes esterification conditions. In some embodiments, the second capping step includes esterification conditions that are the same as or equivalent to the reference capping conditions in that they cap unreacted 5'-OH in oligonucleotide synthesis, for example. In some embodiments, the nucleophile is DMAP or NMI. In some embodiments, the nucleophile is DMAP. In some embodiments, the nucleophile is NMI. In some embodiments, the esterification catalyst is DMAP or NMI.

[0054] In some embodiments, this disclosure (1) Coupling step; (2) Optionally, the first capping step; (3) Modification step; (4) Optionally, a second capping step; and (5) Optionally, the deblocking step The present invention provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles each independently containing the following elements.

[0055] In some embodiments, the cycle includes a first capping step. In some embodiments, the cycle includes a second capping step. In some embodiments, the cycle includes first and second capping steps. In some embodiments, the cycle includes a deblocking step. In some embodiments, the cycle includes a first capping step and a deblocking step. In some embodiments, the cycle includes a second capping and deblocking step.

[0056] In some embodiments, this disclosure (1) Coupling step; (2) The first capping step; (3) Modification step; (4) Second capping step; and (5) Deblocking step The present invention provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles each independently containing the following elements.

[0057] In some embodiments, this disclosure (1) Coupling step; (2) Pre-modification capping step; (3) Modification step; (4) Post-modification capping; and (5) Deblocking step The present invention provides a method for preparing, for example, an oligonucleotide, comprising one or more cycles each independently containing the following elements.

[0058] In some embodiments, the provided method includes repeating several steps or cycles until a desired length (e.g., a desired length of an oligonucleotide or intermediate) is achieved.

[0059] Some or all steps in the provided method or cycle may be performed in a specific order. In some embodiments, the order is (1)-(2), (1)-(3), (2)-(3), (3)-(4), (3)-(5), (4)-(5), (5)-(1), (1)-(2)-(3), (1)-(3)-(4), (1)-(2)-(3)-(4), or any combination thereof, or including (as those skilled in the art will understand, each of (1)-(5) independently represents the corresponding step in the method or cycle, e.g., (1)-coupling step; (2)-pre-modification capping step or first capping step; (3)-modification step; (4)-post-modification capping or second capping step; (5)-deblocking step). In some embodiments, the order is (1)-(2), or includes therein. In some embodiments, the order is (1)-(3) or includes it. In some embodiments, the order is (2)-(3) or includes it. In some embodiments, the order is (3)-(4) or includes it. In some embodiments, the order is (3)-(5) or includes it. In some embodiments, the order is (4)-(5) or includes it. In some embodiments, the order is (5)-(1) or includes it. In some embodiments, the order is (1)-(2)-(3) or includes it. In some embodiments, the order is (1)-(3)-(4) or includes it. In some embodiments, the order is (1)-(2)-(3)-(4) or includes it. In some embodiments, the order is (1)-(2)-(3)-(5) or includes it. In some embodiments, the order is (1)-(3)-(4)-(5) or includes it. In some embodiments, the order is (1)-(2)-(3)-(4)-(5), or includes that. In some embodiments, the order is (5)-(1)-(2)-(3), or includes that. In some embodiments, the order is (5)-(1)-(3)-(4), or includes that. In some embodiments, the order is (5)-(1)-(2)-(3)-(4), or includes that.

[0060] In some embodiments, a cycle or each cycle independently consists of steps (1) to (5). In some embodiments, a cycle or each cycle independently consists of steps (1) to (5) in the order (1)-(2)-(3)-(4)-(5). In some embodiments, a cycle or each cycle independently consists of steps (1) to (5) in the order (5)-(1)-(2)-(3)-(4). In some embodiments, a cycle or each cycle independently consists of steps (1) to (5) in the order (1)-(2)-(4)-(3)-(5). In some embodiments, a cycle or each cycle independently consists of steps (1) to (5) in the order (5)-(1)-(2)-(4)-(3).

[0061] In some embodiments, the method provided comprises one or more cycles, optionally or in addition, each independently comprising steps (1), (2), (3), and (5) in that order. In some embodiments, the method provided comprises one or more cycles, optionally or in addition, each independently comprising steps (1), (2), (3), and (5) in that order. In some embodiments, such cycles result in a native phosphate bond after, for example, optionally, cleavage, deprotection, etc., of an oligonucleotide. In some embodiments, such cycles result in an uncontrolled chiral phosphorothioate internucleotide bond after, for example, optionally, cleavage, deprotection, etc. In some embodiments, each of such cycles independently results in a native phosphate bond after, for example, optionally, cleavage, deprotection, etc., of an oligonucleotide. In some embodiments, each native phosphate bond in an oligonucleotide is independently formed by such cycles after, optionally, cleavage, deprotection, etc., of an oligonucleotide.

[0062] In some embodiments, the method provided comprises one or more cycles, optionally or in addition, each independently comprising steps (1), (3), (4), and (5) in that order. In some embodiments, the method provided comprises one or more cycles, optionally or in addition, each independently comprising steps (1), (3), (4), and (5) in that order. In some embodiments, such cycles result in a native phosphate bond after, for example, optionally, cleavage, deprotection, etc., of an oligonucleotide. In some embodiments, such cycles result in an uncontrolled chiral phosphorothioate internucleotide bond after, for example, optionally, cleavage, deprotection, etc. In some embodiments, each of such cycles independently results in a native phosphate bond after, for example, optionally, cleavage, deprotection, etc., of an oligonucleotide. In some embodiments, each native phosphate bond in an oligonucleotide is independently formed by such cycles after, optionally, cleavage, deprotection, etc., of an oligonucleotide.

[0063] In some embodiments, the first capping step includes no nucleophile as described in this disclosure, or only a reduced level of it. In some embodiments, the first capping step includes no esterification catalyst, or only a reduced level of it. In some embodiments, the first capping step includes conditions that are more selective to amidation than to esterification. In some embodiments, the first capping step does not include conditions that are the same as or equivalent to suitable reference conditions.

[0064] In some embodiments, the first capping step is a pre-capping step as described in this disclosure. In some embodiments, the first capping step utilizes a capping reagent system that is a pre-capping reagent system. In some embodiments, the second capping step is a post-capping step as described in this disclosure. In some embodiments, the second capping step utilizes a capping reagent system that is a post-capping reagent system.

[0065] In some embodiments, suitable reference capping conditions are those for conventional oligonucleotide synthesis based on phosphoramidite chemistry. An exemplary cycle of conventional phosphoramidite-based oligonucleotide synthesis is described below, where the modification step illustrated herein is an oxidation step introducing P=O: [ka]

[0066] In some embodiments, the cycle is the DPSE cycle as shown below (DPSE chiral auxiliary compound: [ka] For different phosphorus configurations): [ka]

[0067] In some embodiments, the cycle is the PSM cycle as shown below (PSM auxiliary compound: [ka] For different phosphorus configurations): [ka] [ka]

[0068] In some embodiments, capping-1 is pre-modification or the first capping step. In some embodiments, capping-2 is post-modification or the second capping step. In some embodiments, cycle termination is after deblocking but before the next coupling. In some embodiments, cycle termination is before deblocking (for example, to leave the 5'-blocking group such as DMTr attached). In some embodiments, [ka] The support is a nucleoside, nucleotide, or oligonucleotide (optionally bound to the support via a linker). Exemplary steps, reagents, modifications, intermediates, and products are provided. Those skilled in the art will understand that other steps, reagents, modifications, intermediates, and products may also be used in accordance with this disclosure.

[0069] In some embodiments, the first or pre-modification capping step includes only low levels of strongly nucleophilic bases or does not include any strongly nucleophilic bases. In some embodiments, the first or pre-modification capping step includes only low levels of NMI. In some embodiments, the first or pre-modification capping step does not include any NMI. In some embodiments, the first or pre-modification capping step includes only low levels of DMAP. In some embodiments, the first or pre-modification capping step does not include any DMAP. In some embodiments, the second or post-modification capping step includes strongly nucleophilic bases. In some embodiments, the second or post-modification capping step includes NMI. In some embodiments, the second or post-modification capping step includes DMAP.

[0070] In some embodiments, the reduced level of the disclosure is less than or equal to a certain percentage based on the volume of the capping reagent solution, for example, 0.01%, 0.02%, 0.05%, 0.1%, 0.2%, 0.5%, 1%, 2%, 3%, 4%, 5%, etc. In some embodiments, the percentage is 0.01%. In some embodiments, the percentage is 0.02%. In some embodiments, the percentage is 0.05%. In some embodiments, the percentage is 0.1%. In some embodiments, the percentage is 0.2%. In some embodiments, the percentage is 0.5%. In some embodiments, the percentage is 1%. In some embodiments, the percentage is 2%. In some embodiments, the percentage is 3%. In some embodiments, the percentage is 4%. In some embodiments, the percentage is 5%.

[0071] In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, or 1.2 equivalents or less compared to the reference agent. In some embodiments, the reference agent is an acylating agent. In some embodiments, the reference agent is a support (oligonucleotide loading capacity unit). In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, or 1.2 equivalents or less compared to the acylating agent. In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 100 equivalents or less compared to the oligonucleotide. In some embodiments, the reduced level is approximately 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 100 equivalents or less compared to the initial nucleoside incorporated into the oligonucleotide. Often, the equivalent of the initial nucleoside incorporated into the oligonucleotide relative to the oligonucleotide loading capacity of the support used for oligonucleotide preparation is 1. In some embodiments, the reduced level is approximately 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or 100 equivalents or less relative to the oligonucleotide loading capacity of the support. In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 2, 3, 4, 5, or 10 equivalents or less relative to the oligonucleotide loading capacity of the support. In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 2, 3, 4, or 5 equivalents or less relative to the oligonucleotide loading capacity of the support.In some embodiments, the reduced level is about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, or 1 equivalent or less relative to the oligonucleotide loading capacity of the support. In some embodiments, the reduced level is about 0.01 equivalent or less. In some embodiments, the reduced level is about 0.02 equivalent or less. In some embodiments, the reduced level is about 0.05 equivalent or less. In some embodiments, the reduced level is about 0.1 equivalent or less. In some embodiments, the reduced level is about 0.2 equivalent or less. In some embodiments, the reduced level is about 0.5 equivalent or less. In some embodiments, the reduced level is about 1 equivalent or less. In some embodiments, the reduced level is about 1.1 equivalent or less. In some embodiments, the reduced level is about 1.2 equivalent or less.

[0072] In particular, the techniques provided are especially useful for the preparation of chiral-controlled oligonucleotide compositions. In some embodiments, the techniques provided involve the formation of one or more chiral internucleotide bonds each independently comprising a chiral binding phosphorus, where each chiral center of the chiral binding phosphorus is independently formed with stereoselectivity as described herein, for example, at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%.

[0073] In some embodiments, the provided technique involves stereoselectively forming one or more chiral-controlled internucleotide bonds using one or more chiral auxiliaries. In some embodiments, the provided technique involves providing monomeric phosphoramidites with diastereomer purity, for example, at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, as described in this disclosure. In many embodiments, the provided diastereomer-purity phosphoramidites include a chiral auxiliary moiety. In some embodiments, phosphoramidites from conventional oligonucleotide synthesis are used for chiral-uncontrolled and / or non-chiral internucleotide bonds.Suitable chiral auxiliaries and phosphoramidites for chiral-controlled oligonucleotide synthesis that can be used in this disclosure include U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Publication No. 2013,017,8612, 8,470,987, 8,822,671, and U.S. Patent Publication No. 2015,02110. 06, U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2 International Publication No. 017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 201 Examples include those described in International Publication No. 8 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 (each of these chiral auxiliaries and phosphoramidites is incorporated herein by independent reference). In some embodiments, the chiral auxiliaries are those of formulas I, Ia, Ia-1, Ia-2, Ib, Ic, Id, Ie, II, II-a, II-b, III, III-a, or III-b as described herein, or salts thereof.In some embodiments, the phosphoramidite has the structure of formulas IV, IV-a, IV-b, IV-c-1, IV-c-2, IV-d, IV-e, IVa, IVa-a, IVa-b, IVa-c-1, IVa-c-2, IVa-d, IVa-e, V, Va, Vb, Vc-1, Vc-2, Vd, Ve, VI, VI-a, VI-b, VI-c-1, VI-c-2, VI-d, or VI-e, or a salt thereof, as described in this disclosure.

[0074] In some embodiments, the provided technology includes the formation of internucleotide bonds having the structure or salt form thereof of formula VII, formula VII-a-1, formula VII-a-2, formula VII-b, formula VII-c, formula VII-d, formula VII-e, formula NL-n-1, formula NL-n-2, formula NL-n-3, formula NL-n-4, formula NL, formula NL-a-1, formula NL-a-2, formula NL-b-1, formula NL-b-2, formula NL-c-1, formula NL-c-2, formula NL-d-1, formula NL-d-2 as described in this disclosure.

[0075] In some embodiments, the provided technology provides oligonucleotides as intermediates and / or products. In some embodiments, the provided oligonucleotides are of formula OI or salts thereof as described in this disclosure. In some embodiments, the oligonucleotides, e.g., final product, product of reaction, product of step, e.g., U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, 2015,021,1006 U.S. Patent Application Publication No. 0170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / The oligonucleotides described in International Publication No. 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 (each of these oligonucleotides is independently incorporated herein by reference) are or share thereof, or include the base sequence, sugar modification or pattern thereof, internucleotide linkage or pattern thereof, base modification or pattern thereof, and / or skeletal chiral center (bound phosphorus) pattern of such oligonucleotides. In some embodiments, the intermediates and / or products provided are chiral-controlled oligonucleotide compositions.In some embodiments, the intermediate and / or product provided is a chiral-controlled oligonucleotide composition of multiple oligonucleotides of formula OI or a salt thereof. In some embodiments, the intermediate and / or product provided is U.S. Patent No. 9,598,458, U.S. Patent No. 9,744,183, U.S. Patent No. 9,605,019, U.S. Patent No. 9,394,333, U.S. Patent No. 8,859,755, U.S. Patent Application Publication No. 20130178612, U.S. Patent No. 8,470,987, U.S. Patent No. 8,822,671, U.S. Patent Application Publication No. 20150211006, U.S. Patent U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607 These are chiral-controlled oligonucleotide compositions as described in International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 (each of these oligonucleotide compositions is independently incorporated herein by reference).

[0076] In some embodiments, this disclosure Multiple oligonucleotides of the modified product composition; and Post-modification capping reagent system A composition containing the following is provided: The modified capping reagent system is in contact with multiple oligonucleotides.

[0077] In some embodiments, this disclosure A capping reagent system comprising a first compound having the structure of formula BI or B-II, Multiple oligonucleotides, each containing at least one internucleotide bond including a -C(O)-N(-)- moiety or a -PS- moiety. Provide a composition containing; The first compound is present in levels of at least 0.1, 0.2, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, or 100 equivalents relative to the plurality of oligonucleotides.

[0078] In some embodiments, this disclosure A capping reagent system comprising a first compound having the structure of formula BI or B-II, Multiple oligonucleotides, wherein each internucleotide bond of the multiple oligonucleotides is independently an internucleotide bond containing a -C(O)-N(-)- moiety and a tetravalent phosphate group. Provide a composition containing; The first compound is present in levels of at least 0.1, 0.2, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, or 100 equivalents relative to the plurality of oligonucleotides.

[0079] In some embodiments, the plurality of oligonucleotides are plurality of oligonucleotides of the modified product composition.

[0080] In some embodiments, in an oligonucleotide composition comprising multiple oligonucleotides: Multiple oligonucleotides share the same base sequence; Multiple oligonucleotides share the same skeletal bonding pattern; and The oligonucleotides among the multiple include at least one chiral-controlled internucleotide linkage; At least ((DS) of all oligonucleotides that share the same base sequence in the composition Nc×100)% is an oligonucleotide among multiple oligonucleotides, where DS is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, and Nc is the number of chiral-controlled internucleotide bonds.

[0081] Multiple oligonucleotides share the same binding phosphorus configuration (Rp or Sp) in a chiral-controlled internucleotide bond. In some embodiments, the chiral-controlled internucleotide bond is referred to as a "stereodefined" internucleotide bond.

[0082] In some embodiments, the first compound is of formula BI. In some embodiments, the first compound is of formula B-II. In some embodiments, the first compound is a nucleophile as described in this disclosure. In some embodiments, the first compound is an esterification catalyst as described in this disclosure. In some embodiments, the first compound is of formula BI. In some embodiments, the first compound is of formula B-II. In some embodiments, the first compound is a base containing =N-, where there is no substitution at any α-position of the nitrogen of =N-. In some embodiments, the first compound is a base containing a heteroaryl moiety, where the heteroaryl moiety contains =N-, where there is no substitution at any α-position of the nitrogen of =N-.

[0083] In some embodiments, the first compound is NMI. In some embodiments, the first compound is DMAP.

[0084] In some embodiments, a plurality of oligonucleotides are attached to a support, such as a solid support used for the preparation of oligonucleotides. In some embodiments, the molar amount of the plurality of oligonucleotides is equal to the load capacity of the solid support to which they are attached. In some embodiments, one of the plurality of oligonucleotides is attached to the support via a linker. Various linkers are known in the art and may be used in this disclosure; some examples are described herein.

[0085] In some embodiments, multiple oligonucleotides share 1) a common base sequence, 2) a common skeletal binding pattern, and 3) a common skeletal phosphorus modification pattern, where the multiple oligonucleotides share the same binding phosphorus stereochemistry for one or more chiral internucleotide bonds (chiral-controlled or sterically defined internucleotide bonds). In some embodiments, about 1% to 100% of all internucleotide bonds in one of the multiple oligonucleotides (e.g., 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%, 95% to 100%, 50% to 90%, or about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, are chiral-controlled internucleotide bonds. In some embodiments, in addition to one or more chiral-controlled internucleotide bonds, one or more internucleotide bonds are native phosphate bonds. In some embodiments, all chiral internucleotide bonds, including chiral-bound phosphates, are chiral-controlled internucleotide bonds. In some embodiments, one or more chiral internucleotide bonds containing chiral-linked phosphorus are sterically random chiral internucleotide bonds (typically, uncontrolled internucleotide bonds prepared by conventional oligonucleotide synthesis without chiral control, e.g., without the use of chiral auxiliaries or chiral modification (e.g., sulfidation) reagents). In some embodiments, several oligonucleotides share the same configuration. In some embodiments, several oligonucleotides share the same structure (are structurally identical).In some embodiments, multiple oligonucleotides share the same stereochemistry in at least one internucleotide bond containing a -C(O)-N(-)- or -PS- moiety. In some embodiments, approximately 0.1% to 100% (e.g., approximately 1% to 100%, 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%) of all oligonucleotides in a chiral-controlled oligonucleotide composition that share a common base sequence, a common skeletal bonding pattern, and a common skeletal phosphorus modification pattern, or the same configuration. 80-100%, 90-100%, 95-100%, 50-90%, or approximately 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) are oligonucleotides among multiple others.

[0086] In some embodiments, one of the oligonucleotides is the structure of formula OI or each of its salts. In some embodiments, one of the oligonucleotides is U.S. Patent No. 9,598,458, U.S. Patent No. 9,744,183, U.S. Patent No. 9,605,019, U.S. Patent No. 9,394,333, U.S. Patent No. 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, U.S. Patent No. 8,470,987, U.S. Patent No. 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, U.S. National Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 1926 Issue 64, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 20 The oligonucleotides are those of International Publication No. 19 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 (each of these oligonucleotides is independently incorporated herein by reference). In some embodiments, some of the oligonucleotides share the same configuration. In some embodiments, some of the oligonucleotides are identical.

[0087] In some embodiments, the level of the disclosure is at least 0.1 equivalents. In some embodiments, the level is at least 0.2 equivalents. In some embodiments, the level is at least 0.5 equivalents. In some embodiments, the level is at least 1 equivalent. In some embodiments, the level is at least 2 equivalents. In some embodiments, the level is at least 3 equivalents. In some embodiments, the level is at least 4 equivalents. In some embodiments, the level is at least 5 equivalents. In some embodiments, the level is at least 6 equivalents. In some embodiments, the level is at least 7 equivalents. In some embodiments, the level is at least 8 equivalents. In some embodiments, the level is at least 9 equivalents. In some embodiments, the level is at least 10 equivalents. In some embodiments, the level is at least 20 equivalents. In some embodiments, the level is at least 50 equivalents. In some embodiments, the level is at least 100 equivalents.

[0088] In some embodiments, -C(O)-N(-)- is part of the capped amino group in the chiral auxiliary moiety bound to the conjugated phosphorus, where the corresponding chiral auxiliary (replacing the bond of -C(O)-N(-)- to -H and the bond to the conjugated phosphorus to -H) is a compound of formula I, Ia, Ia-1, Ia-2, Ib, Ic, Id, Ie, II, II-a, II-b, III, III-a, III-b or a salt thereof.

[0089] In particular, this disclosure provides high crude oligonucleotide compositions. In some embodiments, this disclosure provides chiral-controlled crude oligonucleotide compositions comprising multiple oligonucleotides, where, Multiple oligonucleotides share the same base sequence; Multiple oligonucleotides share the same skeletal bonding pattern; and The plurality of oligonucleotides comprises at least one chirally controlled internucleotide bond, which is chirally controlled in that the plurality of oligonucleotides share the same stereochemical configuration at the chiral bonding phosphorus of the internucleotide bond; at least ((DS) Nc × 100)% of all oligonucleotides sharing the same base sequence in the crude composition are the plurality of oligonucleotides, where DS is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, and Nc is the number of chirally controlled internucleotide bonds.

[0090] In some embodiments, the disclosure provides a chirally controlled crude oligonucleotide composition comprising a plurality of oligonucleotides, wherein the plurality of oligonucleotides share the same composition; and the plurality of oligonucleotides comprises at least one chirally controlled internucleotide bond, which is chirally controlled in that the plurality of oligonucleotides share the same stereochemical configuration at the chiral bonding phosphorus of the internucleotide bond; at least ((DS) Nc × 100)% of all oligonucleotides sharing the same base sequence in the crude composition are the plurality of oligonucleotides, where DS is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%, and Nc is the number of chirally controlled internucleotide bonds.

[0091] In some embodiments, the provided chiral-controlled crude oligonucleotide composition has a crude purity of 30%-80%, 30%-90%, 30%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, or 75% or higher. In some embodiments, the chiral-controlled crude oligonucleotide composition is cleaved from the support and before any further purification. In some embodiments, the chiral-controlled crude oligonucleotide composition is cleaved after desalting from the support and before any further purification. In some embodiments, the chiral-controlled crude oligonucleotide composition is before any chromatographic or gel purification. In some embodiments, crude purity is % full-length product. In some embodiments, crude purity is % full-length product as evaluated by LC-UV observed at UV260nm.

[0092] In some embodiments, DS is approximately 80% to 100%, 85% to 100%, 87% to 100%, 89% to 100%, 90% to 100%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% or more. In some embodiments, DS is approximately 85% or more. In some embodiments, DS is approximately 86% or more. In some embodiments, DS is approximately 87% or more. In some embodiments, DS is approximately 88% or more. In some embodiments, DS is approximately 89% or more. In some embodiments, DS is approximately 90% or more. In some embodiments, DS is approximately 91% or more. In some embodiments, DS is approximately 92% or more. In some embodiments, DS is approximately 93% or more. In some embodiments, DS is approximately 94% or more. In some embodiments, DS is approximately 95% or more. In some embodiments, DS is approximately 96% or more. In some embodiments, DS is approximately 97% or more. In some embodiments, DS is approximately 98% or more. In some embodiments, DS is approximately 99% or more.

[0093] In some embodiments, the diastereoselectivity and / or diastereopurity of a chiral internucleotide bond in an oligonucleotide at the chiral linking phosphorus can be measured or represented through a model reaction, e.g., the formation of a dimer under essentially the same or equivalent conditions, where the dimer has the same internucleotide bond as the chiral internucleotide bond, the 5'-nucleoside of the dimer is the same as the nucleoside at the 5' end of the chiral internucleotide bond, and the 3'-nucleoside of the dimer is the same as the nucleoside at the 3' end of the chiral internucleotide bond. For example, [ka] The diastereopurity of the underlined bond in can be evaluated from the coupling of the two G moieties under the same or equivalent conditions, for example, monomer, chiral auxiliaries, solvent, activator, temperature, etc. In some embodiments, the dimer [ka] The diastereopurity (and / or diastereoselectivity) of the binding is determined by the oligonucleotide. [ka] This is used as the diastereopurity (and / or diastereoselectivity) of the corresponding bond in a given. In some embodiments, the diastereopurity of a compound containing multiple chiral elements is the product of the diastereomer purities of all of its chiral elements. In some embodiments, the diastereopurity (i.e., diastereomer purity) of a provided oligonucleotide is the product of the diastereomer purities of all of its chiral bound phosphorus in its chiral internucleotide bond.

[0094] In some embodiments, Nc is the number of chiral-controlled internucleotide bonds, and is 1 to 100. In some embodiments, Nc is 1 to 50. In some embodiments, Nc is 1 to 40. In some embodiments, Nc is 1 to 30. In some embodiments, Nc is 1 to 25. In some embodiments, Nc is 1 to 24. In some embodiments, Nc is 1 to 23. In some embodiments, Nc is 1 to 22. In some embodiments, Nc is 1 to 21. In some embodiments, Nc is 1 to 20. In some embodiments, Nc is 1 to 19. In some embodiments, Nc is 1 to 18. In some embodiments, Nc is 1 to 17. In some embodiments, Nc is 1 to 16. In some embodiments, Nc is 1 to 15. In some embodiments, Nc is 1 to 14. In some embodiments, Nc is 1 to 13. In some embodiments, Nc is 1 to 12. In some embodiments, Nc is 1 to 11. In some embodiments, Nc is 1 to 10. In some embodiments, Nc is 1 to 9. In some embodiments, Nc is 1 to 8. In some embodiments, Nc is 1 to 7. In some embodiments, Nc is 1 to 6. In some embodiments, Nc is 1 to 5. In some embodiments, Nc is 1. In some embodiments, Nc is 2. In some embodiments, Nc is 3. In some embodiments, Nc is 4. In some embodiments, Nc is 5. In some embodiments, Nc is 6. In some embodiments, Nc is 7. In some embodiments, Nc is 8. In some embodiments, Nc is 9. In some embodiments, Nc is 10. In some embodiments, Nc is 11. In some embodiments, Nc is 12. In some embodiments, Nc is 13. In some embodiments, Nc is 14. In some embodiments, Nc is 15. In some embodiments, Nc is 16. In some embodiments, Nc is 17.In some embodiments, Nc is 18. In some embodiments, Nc is 19. In some embodiments, Nc is 20. In some embodiments, Nc is 21. In some embodiments, Nc is 22. In some embodiments, Nc is 23. In some embodiments, Nc is 24. In some embodiments, Nc is 25.

[0095] In some embodiments, the provided technique comprises one or more modification steps, each independently of or including sulfidation (thiolation). In some embodiments, the provided intermediate and / or product comprises one or more chirally controlled phosphorothioate internucleotide bonds or precursors thereof (which can be converted to phosphorothioate internucleotide bonds by deprotection / cleavage). In some embodiments, the provided intermediate and / or product comprises one or more chirally controlled non-negatively charged internucleotide bonds (e.g., neutral internucleotide bonds) or precursors thereof (which can be converted to phosphorothioate internucleotide bonds by deprotection / cleavage). In some embodiments, the provided technique comprises one or more modification steps, each independently of or including oxidation. In some embodiments, the provided intermediate and / or product comprises one or more native phosphate bonds or precursors thereof (which can be converted to native phosphate bonds by deprotection / cleavage). In some embodiments, the provided intermediate and / or product comprises one or more native phosphate bonds and one or more phosphorothioate internucleotide bonds.

[0096] This disclosure covers various supports, for example, U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, and U.S. Patent Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication You may use materials described in International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784, etc. In some embodiments, the support is a polymer. In some embodiments, the support is a solid support. In some embodiments, the solid support is a polymer, such as polystyrene. In some embodiments, the solid support is a Primer Support (e.g., Primer Support 5G, Primer Support 200, etc.). In some embodiments, the solid support is a NittoPhase support (e.g., NittoPhase HL, NittoPhase UnyLinker, etc.). In some embodiments, the solid support is controlled-pore glass (CPG).In some embodiments, the volume of a solid support, such as a certain polystyrene-based solid support, changes during oligonucleotide synthesis, for example, at different synthesis stages and / or in contact with different solvent systems and / or reagents. In some embodiments, the volume of a solid support, such as many CPG supports, changes by less than 25%, 20%, 15%, 10%, or 5% or remains substantially the same during oligonucleotide synthesis. In some embodiments, the disclosure includes the recognition that volume changes of a solid support during synthesis can cause deviations from planned reaction conditions, such as solvent system, reagent concentration, contact time, etc., and can negatively affect synthesis efficiency, crude purity, and / or yield. In some embodiments, a solid support whose volume does not change significantly or remains substantially the same during oligonucleotide synthesis may offer advantages such as smaller deviations from planned reaction conditions, higher crude purity, and higher yield. The support can have several chemical modifications for nucleoside loading and can have various unit loading capacities (e.g., umol / g).

[0097] In oligonucleotide synthesis using a support, typically the oligonucleotide is bound to the support via a linker. This disclosure describes several linkers, for example, U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Publication Nos. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Publication Nos. 2015,021,1006, and U.S. National Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication You may use what is described in Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784, etc. In some embodiments, this disclosure provides a designed linker. Useful supports and / or functionalizations are described herein.

[0098] Oligonucleotide synthesis typically involves a deblocking step, which deblocks blocked hydroxyl groups for the next step, such as a coupling step, while keeping capped hydroxyl groups intact that should not be involved in the next step, such as a coupling step. This disclosure refers to U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, and U.S. Patent Application Publication No. 2017 Patent No. 037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664 International Publication Nos. 2017 / 192679, 2017 / 210647, 2018 / 022473, 2018 / 067973, 2018 / 098264, 2018 / 223056, 2018 / 223073, 2018 / 223081, 2018 / 237194 Various deblocking conditions can be used, including those described in International Publication Nos. 2019 / 032607, 2019 / 032612, 2019 / 032607, 2019 / 055951, 2019 / 075357, 2019 / 200185, or 2019 / 217784. In some embodiments, deblocking removes the DMT group from the DMT-protected hydroxyl (detritylation). In some embodiments, deblocking is carried out by contacting the oligonucleotide with an acid. In some embodiments, the acid is trichloroacetic acid or dichloroacetic acid.In some embodiments, the deblocking conditions are 2% trichloroacetic acid (TCA) or 3% dichloroacetic acid (DCA) in an inert solvent (e.g., dichloromethane, toluene, etc.).

[0099] The coupling step forms an internucleotide bond, which adds a nucleoside unit to an existing oligonucleotide. In some embodiments, the internucleotide bond formed during the coupling step is a phosphite triester bond. In some embodiments, the internucleotide bond can be formed in a chiralally controlled manner, for example, in chiralally controlled oligonucleotide synthesis using diastereomerically pure phosphoramidites, typically containing a chiral auxiliary moiety. Coupling conditions have been widely reported and are referred to in this disclosure in U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, and U.S. Patent Application Publication No. 2015,021,100. 6, U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. Many are available, including those described in International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784. In some embodiments, the coupling reagent system comprises a nucleoside phosphoramidite and an activator.The provided technology includes U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, 2017,003,7399, 2018,021,6107, and U.S. Patent Application Publication No. 20180216108, US Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 International Publication No. / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185 or International Publication No. 2019 / 21778 Various phosphoramidites can be used, including those described in item 4 and those having the structures of formulas IV, IV-a, IV-b, IV-c-1, IV-c-2, IV-d, IV-e, IVa, IVa-a, IVa-b, IVa-c-1, IVa-c-2, IVa-d, IVa-e, V, Va, Vb, Vc-1, Vc-2, Vd, Ve, VI, VI-a, VI-b, VI-c-1, VI-c-2, VI-d, or VI-e, or salts thereof.Exemplary activators include U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, and U.S. Patent Application Publication No. 2017,003. U.S. Patent Application Publication No. 7399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 International Publication No. / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607 Examples include those described in International Publication Nos. 2019 / 032612, 2019 / 032607, 2019 / 055951, 2019 / 075357, 2019 / 200185, 2019 / 217784, and U.S. Patent No. 9403865 (each of these activators is incorporated herein by independent reference). In some embodiments, the activator is CMPT. In some embodiments, the activator is CMIMT. In some embodiments, the activator is ETT. In some embodiments, conditions such as the phosphoramidite concentration, activator concentration, contact time, solvent, etc., can be optimized for each coupling to improve, for example, crude purity, yield, etc.

[0100] In some embodiments, one or more capping steps are performed after the coupling step and before the modification step. In some embodiments, each capping step after the coupling step and before the modification step is performed, as described in this disclosure, for example, with or without a reduced level of nucleophile, with or without a reduced level of esterification catalyst, and / or under conditions that are selective or specific to amidation rather than esterification. In some embodiments, each capping step after the coupling step and before the modification step is performed to cap one or more amino groups, for example, one or more amino groups formed after coupling in a chiral auxiliary moiety attached to a bound phosphorus atom.

[0101] In some embodiments, after one or more capping steps as described in this disclosure, a modification step is performed to modify the internucleotide bond formed after coupling, which in some embodiments includes a trivalent (e.g., as in the phosphite bond) bound phosphorus atom. In some embodiments, the modification step is or includes oxidation, for example, converting the phosphite bond to a tetracoordinate phosphate triester bond (introducing =O to the bound phosphorus). In some embodiments, the modification step is or includes sulfidation. In some embodiments, sulfidation converts the phosphite bond to a tetracoordinate internucleotide bond by introducing =S to the bound phosphorus. In some embodiments, sulfidation is, for example, =N(-LR 5 ), P N By introducing =N- as shown in the example above into the bound phosphorus, the phosphite bond is converted into a four-coordinate internucleotide bond. In some embodiments, as described herein, R 5 It contains a ring containing a nitrogen atom. In some embodiments, the nitrogen atom is in salt form (quaternary) and counterions (e.g., PF6) - It is associated with ). In some embodiments, =N(-LR 5 ) or P NThe bond containing is a precursor to a non-negatively charged internucleotide bond, such as a neutral internucleotide bond. In some embodiments, sulfurization converts the phosphite bond to a tetracoordinate phosphorothioate triester internucleotide bond (e.g., -P(=O)(SL). s -R 5 )-(in the formula, -L s -R 5 Converts to (not hydrogen). Exemplary modifications and related technologies include U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, and U.S. Patent Application Publication No. 2015,021,1006. , U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication Nos. 2017 / 192664, 2017 / 192679, 2017 / 210647, 2018 / 022473, 2018 / 067973, 2018 / 098264, 2018 / 223056, 2018 / 223073, and 2018 / 223081 Examples include those described in International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784.

[0102] In some embodiments, the provided techniques offer greater flexibility in the selection of the type of modification and / or the modification reagents, such as oxidizing reagents, sulfiding reagents, =N- introduction reagents, etc. For example, reagents that tend to yield unsatisfactory results in previously reported chiral-controlled oligonucleotide synthesis can be used in conjunction with the techniques of this disclosure to obtain significantly improved and satisfactory results. In particular, this disclosure provides techniques (e.g., chiral auxiliaries, compounds, methods, etc.) that are particularly effective for the chiral-controlled preparation of oligonucleotides involving specific types of modifications, such as non-negatively charged internucleotide bonds (e.g., n001).

[0103] In some embodiments, a separate capping step is performed after the modification step. In some embodiments, the post-modification capping step is performed using a substantial amount of a nucleophile and / or esterification catalyst (the nucleophile may be the same as the esterification catalyst) under esterification conditions equivalent to or identical to those used for capping in conventional oligonucleotide synthesis. In some embodiments, the post-modification capping step caps free hydroxyl groups, such as hydroxyl groups that remain intact after the modification step as a result of incomplete coupling. After this capping step, the oligonucleotide can be deblocked to expose hydroxyl groups at sites of further chain elongation, allowing for another synthetic cycle.

[0104] After the desired chain length is achieved, the oligonucleotide can be completely deprotected and cleaved from the support for purification and / or further use. This disclosure refers to U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 2013,017,8612, 8,470,987, 8,822,671, U.S. Patent Application Publication No. 2015,021,1006, and U.S. Patent Application Publication No. 2017,000. Patent No. 37399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2017 / 192664, International Publication Nos. 2017 / 192679, 2017 / 210647, 2018 / 022473, 2018 / 067973, 2018 / 098264, 2018 / 223056, 2018 / 223073, 2018 / 223081, 2018 / 237194, Country Various cleavage and / or deprotection techniques can be utilized, including those described in International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784. In some embodiments, cleavage and / or deprotection includes the removal of chiral auxiliary groups. As those skilled in the art will understand, the cleavage and / or deprotection conditions may depend on the chemistry used in oligonucleotide synthesis, such as the properties of the linker that links the oligonucleotide to the support, the properties of the base and / or sugar blocking groups, the properties of the chiral moiety, etc. In some embodiments, the removal of chiral auxiliary, such as DPSE-type chiral auxiliary, includes the use of TEA-HF. In some embodiments, this disclosure unexpectedly demonstrates that the use of TEA-HF in oligonucleotide synthesis using CPG supports can be successful.

[0105] In particular, this disclosure identifies that contact with water for a period of time (e.g., for deprotection and / or cleavage) at optionally high temperatures, for example, under basic conditions, can be a significant cause of impurities / decomposition (e.g., conversion of P=S and / or P=N- to P=O). In some embodiments, this disclosure provides techniques for addressing the causes of such problems. In some embodiments, this disclosure provides chiral auxiliaries that can be readily removed by a base. In some embodiments, such removal does not require strong bases, high temperatures, and / or length of time as previously reported, and can significantly improve product yield and / or purity. In some embodiments, the removal of the chiral auxiliary group preferably involves contacting the oligonucleotide (e.g., a plurality of oligonucleotides) containing the chiral auxiliary group with a base under anhydrous conditions before contacting the oligonucleotide with a large amount of water (e.g., a reagent system containing a base, water, and optionally one or more organic solvents (e.g., useful for deprotection of the base, cleavage of the oligonucleotide from the support, etc.)). In various cases, the applicant has observed that removing chiral auxiliary groups with a base under anhydrous conditions before contact with conditions containing large amounts of water (e.g., deprotection / cleavage conditions containing NH3,H2O) can significantly improve product yield and / or purity. In addition, the removal of chiral auxiliary groups with a base as described herein can simplify the work procedure and reduce manufacturing costs.

[0106] The technologies provided in this disclosure include various sugars and nucleic acid bases, including non-naturally modified sugars and nucleic acid bases, such as U.S. Patent Nos. 9,598,458, 9,744,183, 9,605,019, 9,394,333, 8,859,755, U.S. Patent Application Publication No. 20130178612, 8,470,987, and 8,822,671. U.S. Patent Application Publication No. 20150211006, U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 20 Publication No. 17 / 160741, International Publication No. 2017 / 192664, International Publication No. 2017 / 192679, International Publication No. 2017 / 210647, International Publication No. 2018 / 022473, International Publication No. 2018 / 067973, International Publication No. 2018 / 098264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081 Sugars and nucleic acid bases described in International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 may be used.

[0107] In some embodiments, the provided technology is useful for the large-scale preparation of oligonucleotides. In some embodiments, the scale is 100 g or more. In some embodiments, the scale is 200 g or more. In some embodiments, the scale is 500 g or more. In some embodiments, the scale is at least 1000 g or more. In some embodiments, the composition includes a large amount of oligonucleotide (e.g., the product of a reaction, step, method, etc., or the final product). In particular, this disclosure addresses various challenges associated with the large-scale preparation of oligonucleotide compositions, especially chiral-controlled oligonucleotide compositions.

[0108] In some embodiments, the composition of another step that is brought into contact in one step is the composition of the first step preceding that step. In some embodiments, the composition of another step that is brought into contact in each step is the composition of the first step preceding that step. For example, the deblocked composition brought into contact in the coupling step is the deblocked composition from the first deblocking step preceding the coupling step; the coupling product composition brought into contact in the pre-modification capping step is the coupling product composition from the first coupling step preceding the pre-modification capping step; the coupling product composition brought into contact in the modification step is the coupling product composition from the first coupling step preceding the modification step; the pre-modification capping product composition brought into contact in the modification step is the pre-modification capping product composition from the first pre-modification capping step preceding the modification step; the modified product composition brought into contact in the post-modification capping step is the modified product composition from the first modification step preceding the post-modification capping step; the modified product composition brought into contact in the deblocking step is the modified product composition from the first modification step preceding the deblocking step; and the post-modification capping product composition brought into contact in the deblocking step is the post-modification capping product composition from the first post-modification capping step preceding the deblocking step.

[0109] In some embodiments, the techniques provided can generally be used for the preparation of other oligomeric compounds. In some embodiments, a method for preparing a composition comprising multiple oligomeric compounds is provided. (1) A coupling step, Contacting a deblocked composition containing multiple deblocked compounds, each containing a deblocked monomer unit that is deblocked in such a way that each deblocked monomer unit independently contains a free linking group, with a coupling reagent system containing a partner compound containing monomer units of an oligomeric compound; and To provide a coupling product composition comprising multiple coupling products, each containing an independent bond that connects the linking group of a deblocked monomer unit to the monomer unit of a partner compound, by coupling a partner compound containing monomer units of an oligomeric compound with the free linking groups of multiple deblocked compounds. Steps including; (2) Optionally, a pre-modification capping step, Contacting the coupling product composition with the pre-modification capping reagent system; and To provide a pre-capped product composition containing multiple pre-capped products by capping one or more functional groups of a coupling product composition. Steps including; (3) Modification step, To provide a modification step composition containing multiple modified products by contacting a coupling product composition and modifying one or more bonds of one or more coupling products; or To provide a modified product composition containing multiple modified products by contacting a pre-modified capping product composition and modifying one or more bonds of one or more pre-modified capping products. Steps including; (4) Optionally, the post-modification capping step is: Contacting the modified product composition with the post-modification capping reagent system; and To provide a post-capped product composition containing multiple post-capped products by capping one or more functional groups of one or more compounds in a modified product composition. Steps including; (5) Optionally, a deblocking step, To provide a deblocked composition comprising multiple deblocked products, each independently containing a deblocked monomer unit with a free linking group, by contacting a modified product composition or a post-modified capping product composition with a deblocking reagent system. Steps including Includes.

[0110] In some embodiments, the method optionally includes repeating steps (1) to (5) multiple times, for example, until a desired length is achieved. In some embodiments, the method includes both a pre-modification capping step and a post-modification capping step (where the pre-modification capping reagent system is optionally different from the post-modification reagent system) or a modification step including a pre-modification capping step (where the pre-modification capping reagent system caps multiple uncoupled groups of multiple coupling products) and sulfation (the sulfation provides a modified product composition comprising multiple modified products, each independently containing a P=S moiety) or a post-modification capping step (containing a chiral center and comprising at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the modified product composition having the same composition). The modification includes contacting a modified product composition comprising a plurality of modified products, each independently containing a bond with at least one chiral center, in which % of the oligomer compound is chiral controlled in that it shares the same stereochemical configuration at the chiral center; or a post-modification capping step and a coupling reagent system comprising a chiral partner compound comprising the monomer unit of the oligomer compound (where the chiral partner compound contains a chiral atom not present in the monomer unit); or a coupling step immediately preceding the pre-modification capping step (the pre-modification capping reagent system of the pre-modification capping step does not contain an esterification catalyst or a strong nucleophile).

[0111] As described herein, in some embodiments, the oligomeric compound is an oligonucleotide. In some embodiments, the composition comprising multiple oligomeric compounds is an oligonucleotide composition comprising multiple oligonucleotides. In some embodiments, the coupling step is the coupling step as described herein with respect to oligonucleotide synthesis. In some embodiments, the deblocked compound is a deblocked oligonucleotide, for example, an oligonucleotide after the deblocking step in an oligonucleotide synthesis cycle. In some embodiments, the deblocked monomer unit is a deblocked 5'-terminal nucleoside unit. In some embodiments, the free linking group is a free 5'-hydroxyl group. In some embodiments, the coupling reagent system is the coupling reagent system in an oligonucleotide synthesis cycle. In some embodiments, the partner compound is a phosphoramidite as described herein with respect to oligonucleotide synthesis. In some embodiments, the coupling product is an oligonucleotide formed after coupling in oligonucleotide synthesis. In some embodiments, the bond connecting the linking group of the deblocked monomer unit to the monomer unit of the partner compound is an internucleotide bond formed during the coupling step. In some embodiments, the pre-modification capping step is the capping step in oligonucleotide synthesis as described herein. In some embodiments, the pre-capping reagent system is the pre-capping reagent system in oligonucleotide synthesis as described in this disclosure. In some embodiments, the pre-capping product composition is the composition after the pre-capping step in oligonucleotide synthesis. In some embodiments, the pre-capping product is the product formed after the pre-capping step in oligonucleotide synthesis. In some embodiments, the modification step is the modification step as used in oligonucleotide synthesis.In some embodiments, the modification step, as demonstrated in the oligonucleotide synthesis described herein, modifies internucleotide bonds. In some embodiments, the modified product composition is an oligonucleotide composition provided after the modification step in oligonucleotide synthesis. In some embodiments, the modified product is an oligonucleotide provided after the modification step in oligonucleotide synthesis. In some embodiments, the post-modification capping step is a capping step in oligonucleotide synthesis as described herein. In some embodiments, the post-modification capping reagent system is a post-capping reagent system in oligonucleotide synthesis as described herein. In some embodiments, the post-modification capping product composition is a composition after the post-modification capping step in oligonucleotide synthesis. In some embodiments, the post-modification capping product is a product formed after the post-modification capping step in oligonucleotide synthesis. In some embodiments, the deblocking step is a deblocking step as described herein with respect to oligonucleotide synthesis. In some embodiments, the unlinked group is an amino group. In some embodiments, the chiral-controlled chiral center is a chiral-controlled bounding phosphorus center. In some embodiments, the chiral partner compound containing a chiral atom not present in the monomer unit is a phosphoramidite containing a chiral center that is not present in the nucleoside unit and is not phosphorus. In some embodiments, the reagent system does not contain an esterification catalyst, does not contain DMAP, and does not contain NMI. [Brief explanation of the drawing]

[0112] Brief explanation of the drawing [Figure 1] This is a crude UPLC chromatogram for B6. [Figure 2] This is a crude UPLC chromatogram for B19. [Figure 3] This is a crude UPLC chromatogram for B56. [Figure 4A]This is a crude UPLC chromatogram (after NAP) for B110. [Figure 4B] This is a crude UPLC chromatogram for B110 (before NAP). [Modes for carrying out the invention]

[0113] Detailed description of a specific embodiment 1.Definition As used herein, unless otherwise specified, the following definitions shall apply. For the purposes of this disclosure, chemical elements are identified according to the periodic table, CAS versions, and Handbook of Chemistry and Physics, 75th Ed. In addition, general principles of organic chemistry are described in “Organic Chemistry”, Thomas Sorrell, University Science Books, Sausalito: 1999 and “March's Advanced Organic Chemistry”, 5th Ed., Ed.: Smith, M. b. and March, J., John Wiley & Sons, New York: 2001.

[0114] When used herein, unless otherwise clearly evident from the context, (i) the term “one (a)” or “one (an)” may be understood to mean “at least one”; (ii) the term “or” may be understood to mean “and / or”; (iii) the terms “contains,” “includes,” “incorporates” (whether used with “not limited to these”) and “incorporates” (whether used with “not limited to these”) may be understood to include the components or steps listed, whether presented alone or with one or more additional components or steps; (iv) the term “another” may be understood to mean at least one or more additional / secondary components; (v) the terms “about” and “approximately” may be understood to allow for a standard deviation as a person skilled in the art would understand; and (vi) where a range is provided, the endpoints are included.

[0115] Unless otherwise specified, descriptions of oligonucleotides and their elements (e.g., base sequence, sugar modification, internucleotide bond, bonded phosphorus stereochemistry, etc.) are in 5' to 3'. Unless otherwise specified, oligonucleotides described herein are provided and / or available in salt form, in particular in pharmaceutically acceptable salt form. Unless otherwise indicated, oligonucleotides include various forms of oligonucleotides. As those skilled in the art will understand, in some embodiments, individual oligonucleotides in a composition may be considered to have the same composition and / or structure even if a particular oligonucleotide in such a composition (e.g., a liquid composition) may be in a different form at a particular moment (e.g., one or more different pharmaceutically acceptable salt forms (and may be soluble, and the oligonucleotide chain may exist in anionic form, for example, when in a liquid composition)). For example, those skilled in the art will recognize that, at a given pH, individual internucleotide bonds along an oligonucleotide chain may be in an acidic (H) form or one of several possible salt forms (e.g., a sodium salt, or a salt of another cation depending on which ions may be present in the formulation or composition), and that in their acidic form (e.g., all cations, if present, are H) + You will understand that such individual oligonucleotides can be appropriately considered to have the same composition and / or structure, insofar as they have the same composition and / or structure (replaced by).

[0116] Aliphatic: As used herein, “aliphatic” means a straight (i.e., unbranched) or branched, substituted or unsubstituted hydrocarbon chain that is fully saturated or contains one or more unsaturated units, or a substituted or unsubstituted monocyclic, dicyclic or polycyclic hydrocarbon ring that is fully saturated or contains one or more unsaturated units (but is not aromatic), or a combination thereof. In some embodiments, the aliphatic group contains 1 to 50 aliphatic carbon atoms. In some embodiments, the aliphatic group contains 1 to 20 aliphatic carbon atoms. In other embodiments, the aliphatic group contains 1 to 10 aliphatic carbon atoms. In other embodiments, the aliphatic group contains 1 to 9 aliphatic carbon atoms. In other embodiments, the aliphatic group contains 1 to 8 aliphatic carbon atoms. In other embodiments, the aliphatic group contains 1 to 7 aliphatic carbon atoms. In other embodiments, the aliphatic group contains 1 to 6 aliphatic carbon atoms. In yet another embodiment, the aliphatic group comprises 1 to 5 aliphatic carbon atoms, and in yet another embodiment, the aliphatic group comprises 1, 2, 3, or 4 aliphatic carbon atoms. Suitable aliphatic groups include, but are not limited to, linear or branched, substituted or unsubstituted alkyl, alkenyl, alkynyl groups and their hybrids, such as (cycloalkyl)alkyl, (cycloalkenyl)alkyl, or (cycloalkyl)alkenyl.

[0117] Alkenyl: As used herein, the term “alkenyl” means an aliphatic group having one or more double bonds as defined herein.

[0118] Alkyl: As used herein, the term "alkyl" is given in the ordinary sense of the art and includes saturated aliphatic groups, including linear alkyl groups, branched alkyl groups, cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, and cycloalkyl-substituted alkyl groups. In some embodiments, alkyl groups have 1 to 100 carbon atoms. In some embodiments, linear or branched alkyl groups have about 1 to 20 carbon atoms in their skeleton (for example, C1 to C1 in the case of a linear group). 20 In the case of a branched chain, C2~C 20) or having about 1 to 10. In some embodiments, the cycloalkyl ring has about 3 to 10 carbon atoms in its ring structure (in which case such a ring is monocyclic, dicyclic, or polycyclic) or about 5, 6, or 7 carbon atoms in its ring structure. In some embodiments, the alkyl group may be a lower alkyl group, which contains 1 to 4 carbon atoms (for example, C1 to C4 in the case of a linear lower alkyl group).

[0119] Alkynyl: As used herein, the term “alkynyl” refers to an aliphatic group having one or more triple bonds as defined herein.

[0120] Analogue: The term "analogue" includes any chemical moiety that is structurally different from a reference chemical moiety or subclass but is capable of performing at least one function of such reference chemical moiety or subclass. Non-limiting examples include nucleotide analogues that are structurally different from nucleotides but perform at least one function of nucleotides; and nucleic acid base analogues that are structurally different from nucleic acid bases but perform at least one function of nucleic acid bases, and so on.

[0121] The term "aryl," used alone or as part of a larger term such as "aralkyl," "aralkoxy," or "aryloxyalkyl," refers to a monocyclic, dicyclic, or polycyclic system having a total of 5 to 30 ring members, wherein at least one ring in the system is an aromatic ring. In some embodiments, the aryl group refers to a monocyclic, dicyclic, or polycyclic system having a total of 5 to 14 ring members, wherein at least one ring in the system is aromatic, and each ring in the system contains 3 to 7 ring members. In some embodiments, the aryl group is a biaryl group. The term "aryl" may be used interchangeably with the term "aryl ring." In some embodiments of the present invention, "aryl" refers to an aromatic ring system, including, but not limited to, phenyl, biphenyl, naphthyl, binaphthyl, and anthracyl, which may contain one or more substituents. In some embodiments, when the term “aryl” is used herein, its scope also includes groups in which an aromatic ring is condensed with one or more non-aromatic rings, such as indanyl, phthalimidyl, naphthoimidyl, phenantridinyl, or tetrahydronaphthyl, where the radical or bond site is on the aryl ring.

[0122] Chiral Control: As used herein, “chiral control” refers to the control of the stereochemical designation of the chiral bounding phosphorus in a chiral internucleotide bond within an oligonucleotide. As used herein, a chiral internucleotide bond is an internucleotide bond whose bounding phosphorus is chiral. In some embodiments, control is achieved through a chiral element not present in the sugar and base portions of the oligonucleotide, for example, in some embodiments, control is achieved by using one or more chiral auxiliaries during the preparation of the oligonucleotide, as described herein, which are often part of a chiral phosphoramidite used during the preparation of the oligonucleotide. Those skilled in the art will understand that, in contrast to chiral control, when chiral internucleotide bonds are formed using conventional oligonucleotide synthesis without chiral auxiliaries, such conventional oligonucleotide synthesis cannot control the stereochemistry of the chiral internucleotide bonds. In some embodiments, the stereochemical designation of each chiral bounding phosphorus in each chiral internucleotide bond within the oligonucleotide is controlled.

[0123] Chiral-controlled oligonucleotide composition: The terms “chiral-controlled oligonucleotide composition,” “chiral-controlled nucleic acid composition,” etc., as used herein, refer to a composition comprising multiple oligonucleotides (or nucleic acids) that share 1) a common base sequence, 2) a common skeletal linkage pattern, and 3) a common skeletal phosphorus modification pattern, wherein the multiple oligonucleotides (or nucleic acids) share the same linkage phosphorus stereochemistry in one or more chiral internucleotide bonds (the chiral linkage phosphorus in the composition is Rp or Sp ("stereodefined"), and is not a random mixture of Rp and Sp like an unchiral-controlled internucleotide bond, but a chiral-controlled or stereodefined internucleotide bond). The levels of the multiple oligonucleotides (or nucleic acids) in a chiral-controlled oligonucleotide composition are predetermined / controlled (e.g., through chiral-controlled oligonucleotide preparations that stereoselectively form one or more chiral internucleotide bonds). In some embodiments, about 0.1% to 100% of all oligonucleotides in the chiral-controlled oligonucleotide composition (e.g., about 1% to 100%, 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%, 95% to 100%, 50% to 90%, or about 5%, 10%, 2%). 0%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% (or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) are multiple oligonucleotides.In some embodiments, approximately 1% to 100% of all oligonucleotides in a chiral-controlled oligonucleotide composition sharing a common base sequence, common skeletal bonding pattern, and common skeletal phosphorus modification pattern (e.g., approximately 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%, 95% to 100%, 50%) ~90% or approximately 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) are multiple oligonucleotides. In some embodiments, the level is about 1% to 100% of all oligonucleotides in the composition, or all oligonucleotides in the composition that share a common base sequence (e.g., multiple oligonucleotides or oligonucleotide types), or all oligonucleotides in the composition that share a common base sequence, a common skeletal binding pattern and a common skeletal phosphorus modification pattern, or all oligonucleotides in the composition that share a common base sequence, a common nucleotide modification pattern, a common sugar modification pattern, a common internucleotide binding type pattern and / or a common internucleotide binding modification pattern (e.g., about 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%, 95% to 100%, 50% to 90%, or about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%. (%), 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%).In some embodiments, multiple oligonucleotides share the same stereochemistry in about 1 to 50 (e.g., about 1 to 10, 1 to 20, 5 to 10, 5 to 20, 10 to 15, 10 to 20, 10 to 25, 10 to 30 or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 or at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) chiral internucleotide bonds. In some embodiments, multiple oligonucleotides are present in concentrations of approximately 1% to 100% (e.g., approximately 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%, 95% to 100%, 50% to 90%, approximately 5%, 10%, 15%, 20%, 25%, 30%, 35%, 4%). They share the same stereochemistry in chiral internucleotide bonds at 0%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% (or at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%). In some embodiments, multiple oligonucleotides (or nucleic acids) have the same configuration.In some embodiments, the level of one of the oligonucleotides (or nucleic acids) is approximately 1% to 100% of all oligonucleotides (or nucleic acids) in the composition that share the same composition as one of the oligonucleotides (or nucleic acids), (for example, approximately 5% to 100%, 10% to 100%, 20% to 100%, 30% to 100%, 40% to 100%, 50% to 100%, 60% to 100%, 70% to 100%, 80% to 100%, 90% to 100%) The percentages are 95-100%, 50-90%, or approximately 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%). In some embodiments, each chiral internucleotide bond is a chiral-controlled internucleotide bond, and the composition is a completely chiral-controlled oligonucleotide composition. In some embodiments, several oligonucleotides (or nucleic acids) are structurally identical. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5%, typically at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5%. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 95%. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 96%. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 97%. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 98%. In some embodiments, chiral-controlled internucleotide bonds have a diastereopurity of at least 99%. In some embodiments, the percentage of the level is (DS).nc is or at least (DS) nc The formula is where DS is the diastereopurity as described in this disclosure (e.g., 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% or more), and nc is the number of chiral-controlled internucleotide bonds as described in this disclosure (e.g., 1-50, 1-40, 1-30, 1-25, 1-20, 5-50, 5-40, 5-30, 5-25, 5-20, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more). In some embodiments, the percentage of the level is (DS) nc is or at least (DS) nc In this formula, DS is between 95% and 100%. For example, when DS is 99% and nc is 10, the percentage is 90%, or at least 90% ((99%)). 10≈0.90 = 90%). In some embodiments, the level of multiple oligonucleotides in a composition is expressed as the product of the diastereopurities of each chiral-controlled internucleotide bond in the oligonucleotide. In some embodiments, the diastereopurity of an internucleotide bond connecting two nucleosides in an oligonucleotide (or nucleic acid) is expressed by the diastereopurity of a dimeric internucleotide bond connecting the same two nucleosides, where the dimer is prepared under equivalent conditions, and in some examples, under the same synthetic cycle conditions (e.g., for a bond between Nx and Ny in oligonucleotide...NxNy..., the dimer is NxNy). In some embodiments, not all chiral internucleotide bonds are chiral-controlled internucleotide bonds, and the composition is a partially chiral-controlled oligonucleotide composition. In some embodiments, the internucleotide bonds that are not chiralized have diastereoplegies of about 80%, 75%, 70%, 65%, 60%, less than 55%, or about 50%, as typically observed in sterically random oligonucleotide compositions (e.g., conventional oligonucleotide synthesis, e.g., by phosphoramidite methods, as those skilled in the art will understand). In some embodiments, several oligonucleotides (or nucleic acids) are of the same type. In some embodiments, a chiralized oligonucleotide composition contains individual oligonucleotide or nucleic acid types at non-random or controlled levels. For example, in some embodiments, a chiralized oligonucleotide composition contains one and only one oligonucleotide type. In some embodiments, a chiralized oligonucleotide composition contains two or more oligonucleotide types. In some embodiments, a chiralized oligonucleotide composition contains multiple oligonucleotide types. In some embodiments, a chiralized oligonucleotide composition is a composition of oligonucleotides of a certain oligonucleotide type, and this composition contains multiple oligonucleotides of that oligonucleotide type at non-random or controlled levels.

[0124] Equivalent: The term “equivalent” is used herein to mean two (or more) sets of conditions or circumstances that are similar enough to each other to allow for a comparison of the results obtained or the phenomena observed. In some embodiments, equivalent sets of conditions or circumstances are characterized by several substantially identical features and one or a few variable features. Those skilled in the art will understand that sets of conditions are equivalent when they are characterized by a sufficient number and variety of substantially identical features to guarantee a reasonable conclusion that the differences in the results obtained or the phenomena observed under different sets of conditions or circumstances are caused by or exhibit differences in such variable features.

[0125] Alicyclic Group: The terms “alicyclic group,” “carbocyclic,” “carbocykyl,” “carbocyclic group,” and “carbocyclic ring” are used synonymously and, when used herein, refer to saturated or partially unsaturated, but non-aromatic, cyclic aliphatic monocyclic, bicyclic, or polycyclic ring systems having 3 to 30 ring members, as described herein, unless otherwise specified. Alicyclic groups include, without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cycloheptenyl, cyclooctyl, cyclooctenyl, norbornyl, adamantyl, and cyclooctadienyl. In some embodiments, alicyclic groups have 3 to 6 carbons. In some embodiments, alicyclic groups are saturated and cycloalkyl. The term “alicyclic group” may also include aliphatic rings condensed to one or more aromatic or non-aromatic rings, such as decahydronaphthyl or tetrahydronaphthyl. In some embodiments, the alicyclic group is bicyclic. In some embodiments, the alicyclic group is tricyclic. In some embodiments, the alicyclic group is polycyclic. In some embodiments, "alicyclic" refers to a C3-C6 monocyclic hydrocarbon that is fully saturated or contains one or more unsaturated units, but is not aromatic, and has a single bond with the rest of the molecule, or a C8-C6 monocyclic hydrocarbon. 10Bicyclic or polycyclic hydrocarbons, or C9-C hydrocarbons that are fully saturated or contain one or more unsaturated units but are not aromatic, and have a single bond with the rest of the molecule. 16 This refers to polycyclic hydrocarbons.

[0126] Halogen: The term "halogen" refers to F, Cl, Br, or I.

[0127] Heteroaliphatic: As used herein, the term “heteroaliphatic” is given its common meaning in the art and refers to an aliphatic group as described herein, in which one or more carbon atoms are independently replaced by one or more heteroatoms (e.g., oxygen, nitrogen, sulfur, silicon, phosphorus, etc.). In some embodiments, one or more units selected from C, CH, CH2, and CH3 are independently replaced by one or more heteroatoms (including their oxidized and / or substituted forms). In some embodiments, the heteroaliphatic group is heteroalkyl. In some embodiments, the heteroaliphatic group is heteroalkenyl.

[0128] Heteroalkyl: As used herein, the term “heteroalkyl” is given its ordinary meaning in the art and refers to the alkyl groups described herein in which one or more carbon atoms are independently substituted with one or more heteroatoms (e.g., oxygen, nitrogen, sulfur, silicon, phosphorus, etc.). Examples of heteroalkyl groups include, but are not limited to, alkoxy, poly(ethylene glycol)-, alkyl-substituted amino, tetrahydrofuranyl, piperidinyl, and morpholinyl.

[0129] Heteroaryl: As used herein, the terms “heteroaryl” and “hetero-” are used alone or as part of a larger term (e.g., “heteroaralkyl” or “heteroaralkoxy”) to refer to a monocyclic, dicyclic, or polycyclic system having a total of 5 to 30 ring members, wherein at least one ring in the system is aromatic, and at least one aromatic ring atom is a heteroatom. In some embodiments, the heteroaryl group has 5 to 10 ring atoms (i.e., monocyclic, dicyclic, or polycyclic), and in some embodiments, it has 5, 6, 9, or 10 ring atoms. In some embodiments, the heteroaryl group has 6, 10, or 14π electrons shared within the cyclic array; in addition to carbon atoms, it has 1 to 5 heteroatoms. It possesses. Examples of heteroaryl groups, though not limited to them, include thienyl, furanyl, pyronyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl, thiadiazolyl, pyridyl, pyridadinyl, pyrimidinyl, pyrazinyl, indolidinyl, purinyl, naphthilidinyl, and pteridinyl. In some embodiments, the heteroaryl is a heterobiaryl group such as bipyridyl. The terms "heteroaryl" and "hetero-" as used herein also include groups in which the heteroaromatic ring is fused with one or more aryl, alicyclic, or heterocyclyl rings, and the binding group or bond site is located on the heteroaromatic ring. Non-limiting examples include indolyl, isoindolyl, benzothienyl, benzofuranyl, dibenzofuranyl, indazolyl, benzimidazolyl, benzthiazolyl, quinolyl, isoquinolyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinolidinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, and pyrido[2,3-b]-1,4-oxazine-3(4H)-one. Heteroaryl groups can be monocyclic, bicyclic, or polycyclic. The term "heteroaryl" may be interchangeable with the terms "heteroaryl ring," "heteroaryl group," or "heteroaromatic," all of which include optionally substituted rings.The term "heteroaralkyl" refers to an alkyl group substituted with a heteroaryl group, where the alkyl and heteroaryl portions are substituted independently and at will.

[0130] Heteroatom: The term "heteroatom," as used herein, means an atom that is neither carbon nor hydrogen. In some embodiments, the heteroatom is boron, oxygen, sulfur, nitrogen, phosphorus, or silicon (including nitrogen, sulfur, phosphorus, or silicon in oxidized forms; and charged forms of nitrogen (e.g., quaternized forms, forms such as those found in iminium groups), phosphorus, sulfur, oxygen, etc.). In some embodiments, the heteroatom is oxygen, sulfur, or nitrogen.

[0131] Heterocyclic: As used herein, the terms “heterocyclic,” “heterocyclyl,” “heterocyclic group,” and “heterocyclic” are interchangeable as used herein and refer to monocyclic, dicyclic, or polycyclic ring moieties (e.g., 3 to 30-membered) that are saturated or partially unsaturated and have one or more heteroatom ring atoms. In some embodiments, a heterocyclyl group is a stable 5 to 7-membered monocyclic or 7 to 10-membered bicyclic heterocyclic moiety that is either saturated or partially unsaturated and has one or more, preferably 1 to 4, heteroatoms (as defined above) in addition to carbon atoms. When used with respect to the ring atoms of a heterocyclic ring, the term “nitrogen” includes substituted nitrogen. For example, in the case of a saturated or partially unsaturated ring having 0 to 3 heteroatoms selected from oxygen, sulfur, and nitrogen, nitrogen is N (e.g., in the case of 3,4-dihydro-2H-pyrrolyl), NH (e.g., in the case of pyrrolidinyl), or +It can be NR (for example, in the case of N-substituted pyrrolidinyl). The heterocycle can be bonded to its pendant group at any heteroatom or carbon atom, thereby obtaining a stable structure, and any of the ring atoms can be optionally substituted. Examples of such saturated or partially unsaturated heterocyclic radicals include, but are not limited to, tetrahydrofuranyl, tetrahydrothienyl, pyrrolidinyl, piperidinyl, pyrrolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepinyl, oxazepinyl, thiazepinyl, morpholinyl, and quinuclidinyl. The terms “heterocyclic,” “heterocyclyl,” “heterocyclyl ring,” “heterocyclic group,” “heterocyclic moiety,” and “heterocyclic radical” are used synonymously herein and include groups in which a heterocyclyl ring is fused to one or more aryl, heteroaryl, or alicyclic rings, such as indolinyl, 3H-indolyl, chromanyl, phenantridinyl, or tetrahydroquinolinyl. The heterocyclyl group may be monocyclic, dicyclic, or polycyclic. The term “heterocyclylalkyl” refers to an alkyl group substituted with a heterocyclyl, where the alkyl and heterocyclyl moieties are independently and optionally substituted.

[0132] Internucleotide bond: As used herein, the term “internucleotide bond” generally refers to a bond that connects nucleoside units of an oligonucleotide or nucleic acid. In some embodiments, the internucleotide bond is a phosphate diester bond (natural phosphate bond (-OP(=O)(OH)O-)), as is widely found in naturally occurring DNA and RNA molecules, which may exist in salt form, as will be understood by those skilled in the art. In some embodiments, the internucleotide bond is a modified internucleotide bond (not a natural phosphate bond). In some embodiments, the internucleotide bond is a “modified internucleotide bond,” where at least one oxygen atom or -OH of the phosphate diester bond is replaced by a different organic or inorganic moiety. In some embodiments, such organic or inorganic moiety is selected from =S, =Se, =NR', -SR', -SeR', -N(R')2, B(R')3, -S-, -Se-, and -N(R')-, where each R' is independently as defined and described herein. In some embodiments, the internucleotide bond is a phosphate triester bond, a phosphorothioate bond (or a phosphorothioate diester bond, -OP(=O)(SH)O-, which, as those skilled in the art will understand, may exist in salt form), or a phosphorothioate triester bond. In some embodiments, the modified internucleotide bond is a phosphorothioate bond. In some embodiments, the internucleotide bond is, for example, a PNA (peptide nucleic acid) or a PMO (phosphodiamidate morpholino oligomer) bond. In some embodiments, the modified internucleotide bond is a non-negatively charged internucleotide bond. In some embodiments, the modified internucleotide bond is a neutral internucleotide bond (e.g., n001 in a particular provided oligonucleotide). As those skilled in the art will understand, the internucleotide bond may exist as an anion or a cation at a given pH due to the presence of an acid or base moiety in the bond.In some embodiments, the modified internucleotide linkage is a modified internucleotide linkage designated as s, s1, s2, s3, s4, s5, s6, s7, s8, s9, s10, s11, s12, s13, s14, s15, s16, s17, and s18 as described in International Publication No. 2017 / 210647.

[0133] Bound phosphorus: The term “bound phosphorus” as defined herein is used to mean that the particular phosphorus atom referred to is a phosphorus atom present in an internucleotide bond, which corresponds to the phosphorus atom in a phosphodiester internucleotide bond, as found in naturally occurring DNA and RNA. In some embodiments, the bound phosphorus atom is in a modified internucleotide bond, where each oxygen atom of the phosphodiester bond is optionally and independently replaced by an organic or inorganic moiety. In some embodiments, the bound phosphorus atom is, for example, P in formula VII as defined herein. In some embodiments, the bound phosphorus atom is chiral. In some embodiments, the bound phosphorus atom is chiral (for example, as in a natural phosphate bond).

[0134] Modified nucleic acid bases: The terms "modified nucleic acid base" and "modified base" refer to a chemical portion that is chemically different from a nucleic acid base but has the ability to perform at least one function of a nucleic acid base. In some embodiments, a modified nucleic acid base is a nucleic acid base that includes modifications. In some embodiments, a modified nucleic acid base has the ability to perform at least one function of a nucleic acid base, for example, the ability to form a portion in a polymer that has the ability to base pair with nucleic acids containing at least a complementary base sequence. In some embodiments, a modified nucleic acid base is a substituted A, T, C, G, or U or a substituted tautomer of A, T, C, G, or U. In some embodiments, a modified nucleic acid base in the context of oligonucleotides refers to a nucleic acid base that is not A, T, C, G, or U.

[0135] Modified Nucleoside: The term "modified nucleoside" refers to a portion of a nucleoside that is derived from or chemically similar to a natural nucleoside but contains chemical modifications that distinguish it from the natural nucleoside. Non-limiting examples of modified nucleosides include those with modifications to bases and / or sugars. Non-limiting examples of modified nucleosides include those with 2' modifications to sugars. Non-limiting examples of modified nucleosides include debasic nucleosides (which lack nucleic acid bases). In some embodiments, modified nucleosides have the ability to form a portion of a polymer that has the ability to perform at least one function of a nucleoside, for example, the ability to base pair with nucleic acids containing at least a complementary base sequence.

[0136] Modified nucleotides: The term "modified nucleotide" includes any chemical portion that is structurally different from a natural nucleotide but has the ability to perform at least one function of a natural nucleotide. In some embodiments, modified nucleotides include modifications to sugars, bases, and / or internucleotide bonds. In some embodiments, modified nucleotides include modified sugars, modified nucleic acid bases, and / or modified internucleotide bonds. In some embodiments, modified nucleotides have the ability to perform at least one function of a nucleotide, for example, the ability to form subunits in polymers that have the ability to base pair with nucleic acids containing at least a complementary base sequence.

[0137] Modified sugar: The term “modified sugar” refers to a portion of a sugar that can be replaced. Modified sugars mimic the spatial arrangement, electronic properties, or any other physicochemical properties of a sugar. In some embodiments, when described herein, the modified sugar is a substituted ribose or deoxyribose. In some embodiments, the modified sugar includes 2'-modifications. Useful examples of 2'-modifications are widely available in the art and are described herein. In some embodiments, the 2'-modification is 2'-OR, where R is optionally substituted C. 1~10It is aliphatic. In some embodiments, the 2'-modification is 2'-OMe. In some embodiments, the 2'-modification is 2'-MOE. In some embodiments, the modified sugar is a bicyclic sugar (e.g., a sugar used in LNA, BNA, etc.). In some embodiments, in the context of oligonucleotides, the modified sugar is a sugar other than ribose or deoxyribose, as is typically found in natural RNA or DNA.

[0138] Nucleic acid bases: The term "nucleic acid base" refers to the parts of nucleic acids involved in hydrogen bonding, which sequence-specifically connects one nucleic acid chain to another complementary chain. The most common naturally occurring nucleic acid bases are adenine (A), guanine (G), uracil (U), cytosine (C), and thymine (T). In some embodiments, the naturally occurring nucleic acid bases are modified adenine, guanine, uracil, cytosine, or thymine. In some embodiments, the naturally occurring nucleic acid bases are methylated adenine, guanine, uracil, cytosine, or thymine. In some embodiments, the nucleic acid bases include heteroaryl rings in which the ring atom is nitrogen and, when in a nucleoside, that nitrogen is bonded to a sugar moiety. In some embodiments, the nucleic acid bases include heterocyclic rings in which the ring atom is nitrogen and, when in a nucleoside, that nitrogen is bonded to a sugar moiety. In some embodiments, the nucleic acid base is a “modified nucleic acid base,” i.e., a nucleic acid base other than adenine (A), guanine (G), uracil (U), cytosine (C), and thymine (T). In some embodiments, the modified nucleic acid base is a substituted A, T, C, G, or U. In some embodiments, the modified nucleic acid base is a substituted tautomer of A, T, C, G, or U. In some embodiments, the modified nucleic acid base is a methylated adenine, guanine, uracil, cytosine, or thymine. In some embodiments, the modified nucleic acid base mimics the spatial arrangement, electronic properties, or any other physicochemical properties of the nucleic acid base and retains the properties of hydrogen bonding, which sequence-specifically binds one nucleic acid chain to another. In some embodiments, the modified nucleic acid base can pair with all five naturally occurring bases (uracil, thymine, adenine, cytosine, or guanine) without substantially affecting melting behavior, recognition by intracellular enzymes, or the activity of the oligonucleotide double helix. As used herein, the term “nucleic acid base” also includes structural analogues used in place of natural or naturally occurring nucleotides, such as modified nucleic acid bases and nucleic acid base analogues. In some embodiments, the nucleic acid base is A, T, C, G, or U, which is optionally substituted, or a tautomer which is optionally substituted with A, T, C, G, or U.In some embodiments, “nucleic acid base” refers to a nucleic acid base unit in an oligonucleotide or nucleic acid (for example, A, T, C, G, or U, as found in an oligonucleotide or nucleic acid).

[0139] Nucleoside: The term "nucleoside" refers to a portion of a nucleic acid base or modified nucleic acid base that is covalently bonded to a sugar or modified sugar. In some embodiments, the nucleoside is a natural nucleoside, e.g., adenosine, deoxyadenosine, guanosine, deoxyguanosine, thymidine, uridine, cytidine, or deoxycytidine. In some embodiments, the nucleoside is a substituted natural nucleoside selected from a modified nucleoside, e.g., adenosine, deoxyadenosine, guanosine, deoxyguanosine, thymidine, uridine, cytidine, and deoxycytidine. In some embodiments, the nucleoside is a substituted tautomer of a natural nucleoside selected from a modified nucleoside, e.g., adenosine, deoxyadenosine, guanosine, deoxyguanosine, thymidine, uridine, cytidine, and deoxycytidine. In some embodiments, "nucleoside" refers to a nucleoside unit in an oligonucleotide or nucleic acid.

[0140] Nucleoside analogs: The term "nucleoside analog" refers to a chemical moiety that is chemically different from a natural nucleoside but has the ability to perform at least one function of a nucleoside. In some embodiments, nucleoside analogs include sugar analogs and / or nucleic acid base analogs. In some embodiments, modified nucleosides have the ability to perform at least one function of a nucleoside, for example, the ability to form a moiety in a polymer that has the ability to base pair with nucleic acids containing complementary base sequences.

[0141] Nucleotide: As used herein, the term “nucleotide” refers to the monomeric unit of a polynucleotide, which consists of a nucleic acid base, a sugar, and one or more internucleotide links (e.g., a phosphate linkage in natural DNA and RNA). Naturally occurring bases [guanine, (G), adenine, (A), cytosine, (C), thymine, (T), and uracil (U)] are derivatives of purines or pyrimidines, but it should be understood that this also includes naturally occurring and non-naturally occurring base analogs. Naturally occurring sugars are pentose (five-carbon sugar) deoxyribose (which forms DNA) or ribose (which forms RNA), but it should be understood that this also includes naturally occurring and non-naturally occurring sugar analogs. Nucleotides are joined by internucleotide links to form nucleic acids or polynucleotides. Numerous internucleotide links are known in the art (but are not limited to phosphates, phosphorothioates, boranophosphates, etc.). Artificial nucleic acids include PNA (peptide nucleic acid), phosphate triesters, phosphorothionic acid, H-phosphonic acid, phosphoramidic acid, boranophosphate, methylphosphonic acid, phosphonoacetic acid, thiophosphonoacetic acid, and other variants of the phosphate skeleton of natural nucleic acids, as described herein. In some embodiments, natural nucleotides include naturally occurring bases, sugars, and internucleotide bonds. As used herein, the term “nucleotide” also includes structural analogues used in place of natural or naturally occurring nucleotides, such as modified nucleotides and nucleotide analogues. In some embodiments, “nucleotide” refers to an oligonucleotide or a nucleotide unit in a nucleic acid.

[0142] Oligonucleotides: The term "oligonucleotide" refers to polymers or oligomers of nucleotides, and may encompass any combination of natural and unnatural nucleic acid bases, sugars, and internucleotide bonds.

[0143] Oligonucleotides can be single-stranded or double-stranded. Single-stranded oligonucleotides may have a double-stranded region (formed by two parts of a single-stranded oligonucleotide), and double-stranded oligonucleotides containing two oligonucleotide strands may have a single-stranded region, for example, in a region where the two oligonucleotide strands are not complementary to each other. Exemplary oligonucleotides include, but are not limited to, structural genes, genes containing regulatory and termination regions, self-renewal systems such as viral DNA or plasmid DNA, single-stranded and double-stranded RNAi agents and other RNA interference reagents (RNAi agents or iRNA agents), shRNA, antisense oligonucleotides, ribozymes, microRNAs, microRNA mimetic compounds, supermirs, aptamers, antimirs, antagonistmirs, Ul adapters, triple-stranded oligonucleotides, G quadruple-stranded oligonucleotides, RNA activators, immunostimulatory oligonucleotides, and decoy oligonucleotides.

[0144] The oligonucleotides of this disclosure may vary in length. In detailed embodiments, oligonucleotides may range in length from about 2 to about 200 nucleosides. In various related embodiments, single-stranded, double-stranded, or triple-stranded oligonucleotides may range in length from about 4 to about 10 nucleosides, about 10 to about 50 nucleosides, about 20 to about 50 nucleosides, about 15 to about 30 nucleosides, and about 20 to about 30 nucleosides. In some embodiments, oligonucleotides are about 9 to about 39 nucleosides long. In some embodiments, oligonucleotides are at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleosides long. In some embodiments, oligonucleotides are at least 4 nucleosides long. In some embodiments, oligonucleotides are at least 5 nucleosides long. In some embodiments, the oligonucleotide has a length of at least 6 nucleosides. In some embodiments, the oligonucleotide has a length of at least 7 nucleosides. In some embodiments, the oligonucleotide has a length of at least 8 nucleosides. In some embodiments, the oligonucleotide has a length of at least 9 nucleosides. In some embodiments, the oligonucleotide has a length of at least 10 nucleosides. In some embodiments, the oligonucleotide has a length of at least 11 nucleosides. In some embodiments, the oligonucleotide has a length of at least 12 nucleosides. In some embodiments, the oligonucleotide has a length of at least 15 nucleosides. In some embodiments, the oligonucleotide has a length of at least 15 nucleosides. In some embodiments, the oligonucleotide has a length of at least 16 nucleosides. In some embodiments, the oligonucleotide has a length of at least 17 nucleosides. In some embodiments, the oligonucleotide has a length of at least 18 nucleosides. In some embodiments, the oligonucleotide has a length of at least 19 nucleosides. In some embodiments, the oligonucleotide has a length of at least 20 nucleosides.In some embodiments, the oligonucleotide is at least 25 nucleosides long. In some embodiments, the oligonucleotide is at least 30 nucleosides long. In some embodiments, the oligonucleotide is a double helix of complementary chains at least 18 nucleosides long. In some embodiments, the oligonucleotide is a double helix of complementary chains at least 21 nucleosides long. In some embodiments, each nucleoside counted in the oligonucleotide length independently comprises A, T, C, G, or U, or A, T, C, G, or U substituted by choice, or a tautomer substituted by choice of A, T, C, G, or U.

[0145] Oligonucleotide type: As used herein, the term “oligonucleotide type” refers to a specific base sequence, skeletal bonding pattern (i.e., internucleotide bonding type, e.g., phosphate, phosphorothioate, etc. pattern), skeletal chiral center pattern (i.e., bonding phosphorus stereochemistry (Rp / Sp) pattern), and skeletal phosphorus modification pattern (e.g., “-XL” in formula VII). s -R 5 Oligonucleotides having a "group pattern" are defined and used. In some embodiments, oligonucleotides of a common designated "type" are structurally identical, including being stereochemically identical to one another.

[0146] Those skilled in the art will understand that the synthesis methods of the present disclosure provide a degree of control in oligonucleotide chain synthesis such that each nucleotide unit of the oligonucleotide chain can be pre-designed and / or selected to have a specific stereochemistry and / or specific modification of the bound phosphorus, and / or a specific base and / or a specific sugar. In some embodiments, the oligonucleotide chain is pre-designed and / or selected to have a specific combination of stereocenters in the bound phosphorus. In some embodiments, the oligonucleotide chain is designed and / or determined to have a specific combination of modifications in the bound phosphorus. In some embodiments, the oligonucleotide chain is designed and / or selected to have a specific combination of bases. In some embodiments, the oligonucleotide chain is designed and / or selected to have one or more specific combinations of the above structural properties. In some embodiments, the present disclosure provides a composition comprising or consisting of a plurality of oligonucleotide molecules (e.g., a chiral-controlled oligonucleotide composition). In some embodiments, all such molecules are of the same type (i.e., structurally identical to one another). However, in some embodiments, the composition provided comprises a plurality of different types of oligonucleotides, typically in predetermined relative amounts.

[0147] Optionally Substituted: As described herein, the compounds of this disclosure, e.g., oligonucleotides, may include optionally substituted and / or substituted moieties. Generally, the term “substituted,” whether preceded by the term “optionally,” means that one or more hydrogens of the indicated moiety are replaced with preferred substituents. Unless otherwise specified, an “optionally substituted” group may have preferred substituents at each of its substituted positions, and when two or more positions in any given structure can be replaced with two or more substituents selected from a specified group, the substituents may be the same or different at each position. In some embodiments, the optionally substituted group is unsubstituted. The substituent combinations envisioned in this disclosure preferably result in the formation of stable or chemically feasible compounds. As used herein, “stable” means a compound that is substantially unchanged when subjected to conditions that enable its generation, detection, and, in certain embodiments, its recovery, purification, and use for one or more of the purposes disclosed herein. Specific substituents are described below.

[0148] Substitutable atoms, for example, preferred monovalent substituents on preferred carbon atoms, are independently halogens;-(CH2) 0~4 R 〇 ;-(CH2) 0~4 Ure 〇 ;-O(CH2) 0~4 R o -O-(CH2) 0~4 C(O)OR 〇 ;-(CH2) 0~4 CH(OR 〇 )2;R 〇 -(CH2) can be substituted with 0~4 Ph;R 〇 -(CH2) can be substituted with 0~4 O(CH2) 0~1 Ph;R 〇 -CH=CHPh;R can be substituted. 〇 -(CH2) can be substituted with 0~4 O(CH2) 0~1-ピリジル;-NO2;-CN;-N3;-(CH2) 0~4 N(R 〇 )2;-(CH2) 0~4 N(R 〇 )C(O)R 〇 ;-N(R 〇 )C(S)R 〇 ;-(CH2) 0~4 N(R 〇 )C(O)NR 〇 2;-N(R 〇 )C(S)NR 〇 2;-(CH2) 0~4 N(R 〇 )C(O)OR 〇 ;-N(R 〇 )N(R 〇 )C(O)R 〇 ;-N(R 〇 )N(R 〇 )C(O)NR 〇 2;-N(R 〇 )N(R 〇 )C(O)OR 〇 ;-(CH2) 0~4 C(O)R 〇 ;-C(S)R 〇 ;-(CH2) 0~4 C(O)OR 〇 ;-(CH2) 0~4 C(O)SR 〇 ;-(CH2) 0~4 C(O)OSiR 〇 3;-(CH2) 0~4 OC(O)R 〇 ;-OC(O)(CH2) 0~4 SR 〇 、-SC(S)SR 〇 ;-(CH2) 0~4 SC(O)R 〇 ;-(CH2) 0~4 C(O)NR 〇 2;-C(S)NR 〇 2;-C(S)SR 〇 ;-(CH2) 0~4 OC(O)NR 〇 2;-C(O)N(OR 〇 )R 〇 ;-C(O)C(O)R 〇 ;-C(O)CH2C(O)R 〇;-C(NOR 〇 )R 〇 ;-(CH2) 0~4 SSR 〇 ;-(CH2) 0~4 S(O)2R 〇 ;-(CH2) 0~4 S(O)2OR 〇 ;-(CH2) 0~4 OS(O)2R 〇 ;-S(O)2NR 〇 2;-(CH2) 0~4 S(O)R 〇 ;-N(R 〇 )S(O)2NR 〇 2;-N(R 〇 )S(O)2R 〇 ;-N(OR 〇 )R 〇 ;-C(NH)NR 〇 2;-Si(R 〇 )3;-OSi(R 〇 )3;-B(R 〇 )2;-OB(R 〇 )2;-OB(OR 〇 )2;-P(R 〇 )2;-P(OR 〇 )2;-P(R 〇 )(OR 〇 );-OP(R 〇 )2;-OP(OR 〇 )2;-OP(R 〇 )(OR 〇 );-P(O)(R 〇 )2;-P(O)(OR 〇 )2;-OP(O)(R 〇 )2;-OP(O)(OR 〇 )2;-OP(O)(OR 〇 )(SR 〇 );-SP(O)(R 〇 )2;-SP(O)(OR 〇 )2;-N(R 〇 )P(O)(R 〇 )2;-N(R 〇 )P(O)(OR 〇 )2;-P(R 〇 )2[B(R 〇 )3];-P(OR 〇 )2[B(R 〇)3];-OP(R 〇 )2[B(R 〇 )3];-OP(OR 〇 )2[B(R 〇 )3];-(C 1~4 Linear or branched alkylenes) ON(R 〇 )2; or -(C 1~4 Linear or branched alkylene)C(O)ON(R 〇 )2, and in the formula, each R 〇 These may be substituted as defined herein, independently of hydrogen, C 1~20 Aliphatic carbon atoms having 1 to 5 heteroatoms independently selected from nitrogen, oxygen, sulfur, silicon, and phosphorus. 1~20 Heteroliphatic, -CH2-(C 6~14 aryl), -O(CH2) 0~1 (C 6~14 aryl), -CH2- (5-14 member heteroaryl ring), a 5-20 member monocyclic, bicyclic, or polycyclic saturated, partially unsaturated or aryl ring having 0-5 heteroatoms independently selected from nitrogen, oxygen, sulfur, silicon, and phosphorus, or notwithstanding the above definition, R 〇 Two independent entities, together with one or more intervening atoms, form a 5-20 member monocyclic, bicyclic, or polycyclic saturated, partially unsaturated, or aryl ring having 0-5 heteroatoms independently selected from nitrogen, oxygen, sulfur, silicon, and phosphorus, which may be substituted as defined below.

[0149] R 〇 Up (or R 〇 The preferred monovalent substituents on the ring formed by the two independent entities of -(CH2) together with their intervening atoms are, independently, halogens, -(CH2) 0~2 R ● ,-(HaroR ● ), -(CH2) 0~2 OH, -(CH2) 0~2 Ure ● ,-(CH2) 0~2 CH(OR ● )2;-O(HaroR ● ), -CN, -N3, -(CH2) 0~2 C(O)R ● ,-(CH2)0~2 C(O)OH, -(CH2) 0~2 C(O)OR ● ,-(CH2) 0~2 SR ● ,-(CH2) 0~2 SH, -(CH2) 0~2 NH2, -(CH2) 0~2 NHR ● ,-(CH2) 0~2 NR ● 2, -NO2, -SiR ● 3. -OSiR ● 3. -C(O)SR ● ,-(C 1~4 Linear or branched alkylene)C(O)OR ● , or -SSR ● And in the formula, each R ● It is either unsubstituted, or if preceded by "halo", it is substituted by only one or more halogens, independently, C 1~4 Aliphatic, -CH2Ph, -O(CH2) 0~1 The pH is independently selected from nitrogen, oxygen, and sulfur, and the 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms. 〇 Suitable divalent substituents on the saturated carbon atom include =O and =S.

[0150] For example, suitable divalent substituents on suitable carbon atoms are independently: =O, =S, =NNR * 2. =NNHC(O)R * ,=NNHC(O)OR * ,=NNHS(O)2R * ,=NR * 、=NOR * , -O(C(R * 2)) 2~3 O- or -S(C(R * 2)) 2~3 S-, and in the formula, R * Each independent entity is a hydrogen atom, which can be substituted as defined below. 1~6Selected from aliphatic and unsubstituted 5-6 member saturated, partially unsaturated, or aryl rings having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur. Suitable divalent substituents bonded to the vicinal substituted carbon of the "optionally substituted" group include -O(CR * 2) 2~3 O- is mentioned, and in the formula, R * Each independent entity is a hydrogen atom, which can be substituted as defined below. 1~6 The compounds are selected from aliphatic and unsubstituted 5-6 member saturated, partially unsaturated, and aryl rings having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0151] R * Preferred substituents on the aliphatic group are, independently, halogens, -R ● ,-(HaroR ● ), -OH, -OR ● ,-O(HaroR ● ), -CN, -C(O)OH, -C(O)OR ● -NH2, -NHR ● , -NR ● 2, or -NO2, where each R ● It is either unsubstituted, or if preceded by "halo", it is substituted by only one or more halogens, independently, C 1~4 Aliphatic, -CH2Ph, -O(CH2) 0~1 The pH is a 5-6 member saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0152] In some embodiments, suitable substituents on the replaceable nitrogen are independently -R † , -NR † 2, -C(O)R † , -C(O)OR † ,-C(O)C(O)R † -C(O)CH2C(O)R † -S(O)2R † -S(O)2NR † 2, -C(S)NR † 2, -C(NH)NR † 2, or -N(R† )S(O)2R † And; in the formula, each R † These are, independently, hydrogen, and C which can be substituted as defined below. 1~6 Aliphatic, unsubstituted-OPh, or an unsubstituted 5-6 member saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur, or notwithstanding the above definition, R † Two independent entities, together with one or more intervening atoms, form an unsubstituted 3-12 member saturated, partially unsaturated, or aryl monocyclic or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0153] R † Preferred substituents on the aliphatic group are, independently, halogens, -R ● ,-(HaroR ● ), -OH, -OR ● ,-O(HaroR ● ), -CN, -C(O)OH, -C(O)OR ● -NH2, -NHR ● , -NR ● 2, or -NO2, where each R ● It is either unsubstituted, or if preceded by "halo", it is substituted by only one or more halogens, independently, C 1~4 Aliphatic, -CH2Ph, -O(CH2) 0~1 The pH is a 5-6 member saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, and sulfur.

[0154] Partially unsaturated: As used herein, the term “partially unsaturated” refers to a ring moiety containing at least one double or triple bond. The term “partially unsaturated” is intended to encompass rings having multiple unsaturated moies, as defined herein, but not intended to encompass aryl or heteroaryl moies.

[0155] Pharmaceutical Composition: As used herein, the term “pharmaceutical composition” refers to an active agent formulated with one or more pharmaceutically acceptable carriers. In some embodiments, the active agent is present in a unit dose appropriate for administration in a therapeutic regimen that demonstrates a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population. In some embodiments, the pharmaceutical composition may be specifically formulated for administration in solid or liquid form, including: oral administration, e.g., oral tablets (aqueous or nonaqueous solutions or suspensions), tablets, e.g., those targeted for buccal, sublingual and systemic absorption, boluses, powders, granules, pastes for application to the tongue; parenteral administration, e.g., subcutaneous, intramuscular, intravenous or epidural injection, as sterile solutions or suspensions or sustained-release formulations; topical application, e.g., creams, ointments or controlled-release patches or sprays applied to the skin, lungs or oral cavity; e.g., pessaries, creams or foams for vaginal or rectal administration; sublingual; intraocular; transdermal; or adapted for the nasal cavity, lungs and other mucosal surfaces.

[0156] Pharmacopoeia-acceptable: As used herein, the term "pharmacopoeia-acceptable" means, within reasonable medical judgment, a compound, material, composition and / or dosage form suitable for use in contact with human and animal tissues without excessive toxicity, irritation, allergic reaction or other problems or complications, in proportion to a reasonable risk-benefit ratio.

[0157] Pharmaceutically acceptable carrier: As used herein, the term “pharmaceutically acceptable carrier” means a pharmaceutically acceptable material, composition, or medium, such as a liquid or solid filler, diluent, excipient, or solvent, that encapsulates a material and is involved in the transport or delivery of a compound of interest from one organ or part of the body to another organ or part of the body. Each carrier must be “acceptable” in the sense that it is compatible with the other components of the formulation and is not harmful to the patient. Some examples of materials that can serve as pharmaceutically acceptable carriers include sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; celluloses and their derivatives such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; tragacanth powder; malt; gelatin; talc; excipients such as cocoa butter and suppository wax; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols such as propylene glycol; polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; pH buffer solutions; polyesters, polycarbonates, and / or polyanhydrides; and other non-toxic, suitable substances used in pharmaceutical formulations.

[0158] pharmaceutically acceptable salts: When used herein, the term “pharmaceutically acceptable salt” refers to a salt of such compound that is appropriate for use in a pharmaceutical context, i.e., a salt that, within reasonable medical judgment, does not impose excessive toxicity, irritation, or allergic reactions, is suitable for use in contact with human and lower animal tissues, and has a reasonable risk-benefit ratio. pharmaceutically acceptable salts are well known in the art. For example, S.M. Berge, et al. described pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 66:1-19 (1977). In some embodiments, pharmaceutically acceptable salts include, but are not limited to, non-toxic acid addition salts, which are salts of amino groups formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid, or with organic acids such as acetic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid, or by other methods used in the art, such as ion exchange. In some embodiments, pharmaceutically acceptable salts include, but are not limited to, adipine, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, and 2-hydroxy- Examples include ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, and valerate.In some embodiments, the compounds provided comprise one or more acidic groups, for example, oligonucleotides, and pharmaceutically acceptable salts are alkali salts, alkaline earth metal salts, or ammonium salts (e.g., ammonium salts of N(R)3 (wherein each R is independently defined and described herein)). Typical alkali salts or alkaline earth metal salts include sodium, lithium, potassium, calcium, and magnesium. In some embodiments, the pharmaceutically acceptable salt is a sodium salt. In some embodiments, the pharmaceutically acceptable salt is a potassium salt. In some embodiments, the pharmaceutically acceptable salt is a calcium salt. In some embodiments, pharmaceutically acceptable salts include, where appropriate, non-toxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, alkyls having 1 to 6 carbon atoms, sulfonates, and arylsulfonates. In some embodiments, the compounds provided comprise two or more acidic groups, for example, oligonucleotides may comprise two or more acidic groups (e.g., in a natural phosphate bond and / or a modified internucleotide bond). In some embodiments, a pharmaceutically acceptable salt of such compound, or a salt in general, contains two or more cations that may be the same or different. In some embodiments, in a pharmaceutically acceptable salt (or a salt in general), all ionizable hydrogens in the acidic group (e.g., in aqueous solutions with pKas of about 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2 or less; in some embodiments, about 7 or less; in some embodiments, about 6 or less; in some embodiments, about 5 or less; in some embodiments, about 4 or less; in some embodiments, about 3 or less) are replaced by cations. In some embodiments, each phosphorothioate and phosphate group exists independently in their salt form (e.g., in the case of the sodium salt, -OP(O)(SNa)-O- and -OP(O)(ONa)-O-, respectively).In some embodiments, each phosphorothioate and phosphate internucleotide bond exists independently in its salt form (e.g., in the case of the sodium salt, -OP(O)(SNa)-O- and -OP(O)(ONa)-O-, respectively). In some embodiments, the pharmaceutically acceptable salt is the sodium salt of the oligonucleotide. In some embodiments, the pharmaceutically acceptable salt is the sodium salt of the oligonucleotide, where each acidic phosphate group and modified phosphate group (e.g., phosphorothioate, phosphate, etc.) exists, if present, in salt form (all sodium salts).

[0159] Predetermined: Predetermined (or pre-determined) means intentionally selected, not random, or controlled, as opposed to something that happens randomly, is random, or is realized without control. Those skilled in the art will understand, upon reading this specification, that this disclosure provides techniques that enable the selection of specific chemical and / or stereochemical features into oligonucleotide compositions and further enable the controlled preparation of oligonucleotide compositions having such chemical and / or stereochemical features. Such provided compositions are “predetermined” as described herein. A composition that may contain a particular oligonucleotide because it occurs accidentally through a process that is not controlled in such a way as to intentionally produce the particular chemical and / or stereochemical features is not a “predetermined” composition. In some embodiments, a predetermined composition is intentionally reproducible (e.g., through the repetition of a controlled process). In some embodiments, a predetermined level of multiple oligonucleotides in a composition means that the absolute and / or relative amounts (ratio, percentage, etc.) of the multiple oligonucleotides in the composition are controlled. In some embodiments, a predetermined level of multiple oligonucleotides in a composition is achieved through chiral-controlled oligonucleotide preparation.

[0160] Protecting group: When the term "protecting group" is used herein, it refers to a term well known in the art, as seen in Protecting Groups in Organic Synthesis, TW Greene and PGM Wuts, 3 rdExamples include those described in detail in edition, John Wiley & Sons, 1999 (the entire work is incorporated herein by reference). Also included are protecting groups specifically adapted to nucleoside and nucleotide chemistry, as described in Current Protocols in Nucleic Acid Chemistry, edited by Serge L. Beaucage et al. 06 / 2012 (the entire Chapter 2 is incorporated herein by reference). Suitable amino protecting groups include methyl carbamate, ethyl carbamate, 9-fluorenyl methyl carbamate (Fmoc), 9-(2-sulfo)fluorenyl methyl carbamate, 9-(2,7-dibromo)fluorenyl methyl carbamate, 2,7-di-t-butyl-[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxantyl)] methyl carbamate (DBD-Tmoc), 4-methoxyphenacyl carbamate (Phenoc), 2,2,2-trichloroethyl carbamate (Troc), 2-trimethylsilylethyl carbamate (Teoc), 2- Phenylethylcarbamate (hZ), 1-(1-adamantyl)-1-methylethylcarbamate (Adpoc), 1,1-dimethyl-2-haloethylcarbamate, 1,1-dimethyl-2,2-dibromoethylcarbamate (DB-t-BOC), 1,1-dimethyl-2,2,2-trichloroethylcarbamate (TCBOC), 1-methyl-1-(4-biphenylyl)ethylcarbamate (Bpoc), 1-(3,5-di-t-butylphenyl)-1-methylethylcarbamate (t-Bumeoc), 2-(2'-and 4'-pyridyl)ethylcarbamate (Pyoc), 2-(N,N-Dicyclohexylcarboxamide) ethylcarbamate, t-butylcarbamate (BOC), 1-adamantylcarbamate (Adoc), vinylcarbamate (Voc), allylcarbamate (Alloc), 1-isopropylallylcarbamate (Ipaoc), cinnamylcarbamate (Coc), 4-nitrocinnamylcarbamate (Noc), 8-quinolylcarbamate, N-hydroxypiperidinylcarbamate, alkyldithiocarbamate, benzylcarbamate (Cbz), p-methoxybenzylcarbamate (Moz), p- Trobenzylcarbamate, p-bromobenzylcarbamate, p-chlorobenzylcarbamate, 2,4-dichlorobenzylcarbamate, 4-methylsulfinylbenzylcarbamate (Msz), 9-anthrylmethylcarbamate, diphenylmethylcarbamate, 2-methylthioethylcarbamate, 2-methylsulfonylethylcarbamate, 2-(p-toluenesulfonyl)ethylcarbamate, [2-(1,3-dithianyl)]methylcarbamate (Dmoc), 4-methylthiophenylcarbamate (Mtpc), 2,4-dimethylthio Phenylcarbamate (Bmpc), 2-phosphonioethylcarbamate (Peoc), 2-triphenylphosphonioisopropylcarbamate (Ppoc), 1,1-dimethyl-2-cyanoethylcarbamate, m-chloro-p-acyloxybenzylcarbamate, p-(dihydroxyboryl)benzylcarbamate, 5-benzisoxazolylmethylcarbamate, 2-(trifluoromethyl)-6-chromonylmethylcarbamate (Tcroc), m-nitrophenylcarbamate, 3,5-dimethoxybenzylcarbamate, o-nitrobenzyl Carbamates, 3,4-dimethoxy-6-nitrobenzylcarbamate, phenyl(o-nitrophenyl)methylcarbamate, phenothiazinyl-(10)-carbonyl derivatives, N'-p-toluenesulfonylaminocarbonyl derivatives, N'-phenylaminothiocarbonyl derivatives, t-amylcarbamate, S-benzylthiocarbamate, p-cyanobenzylcarbamate, cyclobutylcarbamate, cyclohexylcarbamate, cyclopentylcarbamate, cyclopropylmethylcarbamate, p-decyloxybenzylcarbamate, 2,2-Dimethoxycarbonylvinylcarbamate, o-(N,N-dimethylcarboxamide)benzylcarbamate, 1,1-dimethyl-3-(N,N-dimethylcarboxamide)propylcarbamate, 1,1-dimethylpropynylcarbamate, di(2-pyridyl)methylcarbamate, 2-furanylmethylcarbamate, 2-iodoethylcarbamate, isobornylcarbamate, isobutylcarbamate, isonicotinylcarbamate, p-(p'-methoxyphenylazo)benzylcarbamate, 1-methylcyclobutylcarbamate, 1- Methylcyclohexylcarbamate, 1-methyl-1-cyclopropylmethylcarbamate, 1-methyl-1-(3,5-dimethoxyphenyl)ethylcarbamate, 1-methyl-1-(p-phenylazophenyl)ethylcarbamate, 1-methyl-1-phenylethylcarbamate, 1-methyl-1-(4-pyridyl)ethylcarbamate, phenylcarbamate, p-(phenylazo)benzylcarbamate, 2,4,6-tri-t-butylphenyl, 4-(trimethylammonium)benzylcarbamate, 2,4,6-trimethylbenzylcarbamate Rubamate, formamide, acetamide, chloroacetamide, trichloroacetamide, trifluoroacetamide, phenylacetamide, 3-phenylpropanamide, picolinamide, 3-pyridylcarboxamide, N-benzoylphenylalanyl derivatives, benzamide, p-phenylbenzamide, o-nitrophenylacetamide, o-nitrophenoxyacetamide, acetacetamide, (N'-dithiobenzyloxycarbonylamino)acetamide, 3-(p-hydroxyphenyl)propanamide, 3-(o-nitrophenyl ) Propanamide, 2-methyl-2-(o-nitrophenoxy)propanamide, 2-methyl-2-(o-phenylazofenoxy)propanamide, 4-chlorobutanamide, 3-methyl-3-nitrobutanamide, o-nitrosinamide, N-acetylmethionine derivative, o-nitrobenzamide, o-(benzoyloxymethyl)benzamide, 4,5-diphenyl-3-oxazolin-2-one, N-phthalimide, N-dithiasuccinimide (Dts), N-2,3-diphenylmaleimide, N-2,5-dimethylpyrrole, N-1,1,4,4-Tetramethyldisilylazacyclopentane adduct (STABASE), 5-substituted 1,3-dimethyl-1,3,5-triazacyclohexane-2-one, 5-substituted 1,3-dibenzyl-1,3,5-triazacyclohexane-2-one, 1-substituted 3,5-dinitro-4-pyridone, N-methylamine, N-allylamine, N-[2-(trimethylsilyl)ethoxy]methylamine (SEM), N-3-acetoxypropylamine, N-(1-isopropyl-4-nitro-2-oxo-3-pyrroline-3-yl)amine, 4 Ammonium salts, N-benzylamine, N-di(4-methoxyphenyl)methylamine, N-5-dibenzosberylamine, N-triphenylmethylamine (Tr), N-[(4-methoxyphenyl)diphenylmethyl]amine (MMTr), N-9-phenylfluorenylamine (PhF), N-2,7-dichloro-9-fluorenylmethyleneamine, N-ferrocenylmethylamino (Fcm), N-2-picolylamino N'-oxide, N-1,1-dimethylthiomethyleneamine, N-benzylideneamine, Np-meth Xybenzylideneamine, N-diphenylmethyleneamine, N-[(2-pyridyl)mesityl]methyleneamine, N-(N',N'-dimethylaminomethylene)amine, N,N'-isopropylidenediamine, Np-nitrobenzylideneamine, N-salicylideneamine, N-5-chlorosalicylideneamine, N-(5-chloro-2-hydroxyphenyl)phenylmethyleneamine, N-cyclohexylideneamine, N-(5,5-dimethyl-3-oxo-1-cyclohexenyl)amine, N-borane derivatives, N-diphenyl Poric acid derivatives, N-[phenyl(pentacarbonylchromium- or tungsten)carbonyl]amine, N-copper chelate, N-zinc chelate, N-nitroamine, N-nitrosamine, amine N-oxide, diphenylphosphinamide (Dpp), dimethylthiophosphinamide (Mpt), diphenylthiophosphinamide (Ppt), dialkylphosphoramides, dibenzylphosphoramide, diphenylphosphoramide, benzenesulfenamide, o-nitrobenzenesulfenamide (Nps), 2,4-Dinitrobenzenesulfenamide, Pentachlorobenzenesulfenamide, 2-Nitro-4-methoxybenzenesulfenamide, Triphenylmethylsulfenamide, 3-Nitropyridinesulfenamide (Npys), p-Toluenesulfonamide (Ts), Benzenesulfonamide, 2,3,6-Trimethyl-4-methoxybenzenesulfonamide (Mtr), 2,4,6-Trimethoxybenzenesulfonamide (Mtb), 2,6-Dimethyl-4-methoxybenzenesulfonamide (Pme), 2,3,5,6-Tetramethyl-4-methoxybenzenesulfonamide (Mte) Examples include 4-methoxybenzenesulfonamide (Mbs), 2,4,6-trimethylbenzenesulfonamide (Mts), 2,6-dimethoxy-4-methylbenzenesulfonamide (iMds), 2,2,5,7,8-pentamethylchroman-6-sulfonamide (Pmc), methanesulfonamide (Ms), β-trimethylsilylethanesulfonamide (SES), 9-anthracene sulfonamide, 4-(4',8'-dimethoxynaphthylmethyl)benzenesulfonamide (DNMBS), benzylsulfonamide, trifluoromethylsulfonamide, and phenacylsulfonamide.

[0161] Suitable protected carboxylic acids include, but are not limited to, silyl-protected, alkyl-protected, alkenyl-protected, aryl-protected, and arylalkyl-protected carboxylic acids. Examples of suitable silyl groups include trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, and triisopropylsilyl. Examples of suitable alkyl groups include methyl, benzyl, p-methoxybenzyl, 3,4-dimethoxybenzyl, trityl, t-butyl, and tetrahydropyran-2-yl. An example of a suitable alkenyl group is allyl. Examples of suitable aryl groups include optionally substituted phenyl, biphenyl, or naphthyl. Examples of suitable arylalkyl groups include optionally substituted benzyl (e.g., p-methoxybenzyl (MPM), 3,4-dimethoxybenzyl, O-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl) and 2- and 4-picolyl.

[0162] Suitable hydroxyl protecting groups include methyl, methoxymethyl (MOM), methylthiomethyl (MTM), t-butylthiomethyl, (phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM), p-methoxybenzyloxymethyl (PMBM), (4-methoxyphenoxy)methyl (p-AOM), guaiacolmethyl (GUM), t-butoxymethyl, 4-pentenyloxymethyl (POM), siloxymethyl, and 2-methoxyethoxymethyl (M EM), 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl, 2-(trimethylsilyl)ethoxymethyl (SEMOR), tetrahydropyranyl (THP), 3-bromotetrahydropyranyl, tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl (MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxide, 1-[(2-chloro-4-methyl)phenyl] -4-methoxypiperidine-4-yl (CTMP), 1,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl, 2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methanobenzofuran-2-yl, 1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 1-methyl-1-methoxyethyl, 1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-fluoroethyl, 2,2,2-trichloro Roethyl, 2-trimethylsilylethyl, 2-(phenylselenenyl)ethyl, t-butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl, benzyl, p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2-picolyl, 4-picolyl, 3-methyl-2-picolyl N-oxide, diphenylmethyl, p,p'-Dinitrobenzhydryl, 5-Dibenzosberyl, Triphenylmethyl, α-Naphthyldiphenylmethyl, p-Methoxyphenyldiphenylmethyl, Di(p-Methoxyphenyl)phenylmethyl, Tri(p-Methoxyphenyl)methyl, 4-(4'-Bromophenacyloxyphenyl)diphenylmethyl, 4,4',4”-Tris(4,5-Dichlorophthalimidophenyl)methyl, 4,4',4”-Tris(Lebrinoyloxyphenyl)methyl, 4,4',4”-Tris(Benzoyloxyphenyl)methyl, 3-(imidazole-1-yl)bis(4',4"-dimethoxyphenyl)methyl, 1,1-bis(4-methoxyphenyl)-1'-pyrenylmethyl, 9-anthryl, 9-(9-phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl, 1,3-benzodithiolan-2-yl, benzisothiazolyl S,S-dioxide, trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), Dimethyltexylsilyl, t-Butyldimethylsilyl (TBDMS), t-Butyldiphenylsilyl (TBDPS), Tribenzylsilyl, Tri-p-Xylylsilyl, Triphenylsilyl, Diphenylmethylsilyl (DPMS), t-Butylmethoxyphenylsilyl (TBMPS), Formate, Benzoylformate, Acetate, Chloroacetate, Dichloroacetate, Trichloroacetate, Trifluoroacetate, Methoxyacetate, Triphenylmethoxyacetate, Phenoxyacetate Tate, p-chlorophenoxyacetate, 3-phenylpropionate, 4-oxopentanoate (rebrinate), 4,4-(ethylenedithio)pentanoate (rebrinoyldithioacetal), pivaloate, adamantoate, crotonate, 4-methoxycrotonate, benzoate, p-phenylbenzoate, 2,4,6-trimethylbenzoate (mesitoate), alkylmethyl carbonate, 9-fluorenylmethyl carbonate (Fmoc), alkylethyl carbonate, alkyl 2,22-Trichloroethyl carbonate (Troc), 2-(trimethylsilyl)ethyl carbonate (TMSEC), 2-(phenylsulfonyl)ethyl carbonate (Psec), 2-(triphenylphosphonio)ethyl carbonate (Peoc), alkyl isobutyl carbonate, alkyl vinyl carbonate Alkylallyl carbonate, alkyl p-nitrophenyl carbonate, alkyl benzyl carbonate, alkyl p-methoxybenzyl carbonate, alkyl 3,4-dimethoxybenzyl carbonate, alkyl o-nitrobenzyl carbonate, alkyl p-nitrobenzyl carbonate, alkyl S-benzylthiocarbonate, 4-ethoxy-1-naphthyl carbonate, methyl dithiocarbonate, 2-iodobenzoate, 4-azidobutyrate, 4-nitro-4-methylpentanoate, o-(dibromomethyl)benzoate, 2-formylbenzenesulfonate, 2-(methylthiomethoxy)ethyl, 4-(methylthiomethoxy)butyrate, 2-(methylthiomethoxymethyl)benzoate, 2,6 Examples include dichloro-4-methylphenoxyacetate, 2,6-dichloro-4-(1,1,3,3-tetramethylbutyl)phenoxyacetate, 2,4-bis(1,1-dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate, monosuccinoate, (E)-2-methyl-2-butenoate, o-(methoxycarbonyl)benzoate, α-naphthoate, nitrate, alkyl N,N,N',N'-tetramethylphosphodiamide, alkyl N-phenylcarbamate, borate, dimethylphosphinthioyl, alkyl 2,4-dinitrophenyl sulfenate, sulfate, methanesulfonate (mesylate), benzyl sulfonate, and tosylate (Ts). For the protection of 1,2- or 1,3-diols, the protecting groups include methylene acetal, ethylidene acetal, 1-t-butylethylidene ketal, 1-phenylethylidene ketal, (4-methoxyphenyl)ethylidene acetal, 2,2,2-trichloroethylidene acetal, acetonide, cyclopentylidene ketal, cyclohexylidene ketal, cycloheptylidene ketal, benzylidene acetal, p-methoxybenzylidene acetal, 2,4-Dimethoxybenzylidene ketal, 3,4-Dimethoxybenzylidene acetal, 2-Nitrobenzylidene acetal, Methoxymethylene acetal, Ethoxymethylene acetal, Dimethoxymethylene orthoester, 1-Methoxyethylidene orthoester, 1-Ethoxyethylidine orthoester, 1,2-Dimethoxyethylidene orthoester, α-Methoxybenzylidene orthoester, 1-(N,N-dimethylamino) Examples include chilidene derivatives, α-(N,N'-dimethylamino)benzylidene derivatives, 2-oxacyclopentylidene orthoesters, di-t-butylsilylene groups (DTBS), 1,3-(1,1,3,3-tetraisopropyldisiloxanylidene) derivatives (TIPDS), tetra-t-butoxydisiloxane-1,3-diylidene derivatives (TBDS), cyclic carbonates, cyclic boronates, ethyl boronates, and phenyl boronates.

[0163] In some embodiments, the hydroxyl protecting group is acetyl, t-butyl, t-butoxymethyl, methoxymethyl, tetrahydropyranyl, 1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 2-trimethylsilylethyl, p-chlorophenyl, 2,4-dinitrophenyl, benzyl, benzoyl, p-phenylbenzoyl, 2,6-dichlorobenzyl, diphenylmethyl, p-nitrobenzyl, triphenylmethyl(trityl), 4,4'-dimethoxytrityl, trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, triphenylsilyl, triisopropylsilyl, benzoylformate, chloroacetyl, trichloroacetyl, trifluoroacetyl, pivaloyl, 9-fluorenylmethyl carbonate, mesylate, tosylate, triflate, trityl, monomethoxytrityl (MMTr), 4,4'-dimethoxytri Trimethyl (DMTr) and 4,4',4”-trimethoxytrityl (TMTr), 2-cyanoethyl (CE or Cne), 2-(trimethylsilyl)ethyl (TSE), 2-(2-nitrophenyl)ethyl, 2-(4-cyanophenyl)ethyl, 2-(4-nitrophenyl)ethyl (NPE), 2-(4-nitrophenylsulfonyl)ethyl, 3,5-dichlorophenyl, 2,4-dimethylphenyl, 2-nitrophenyl, 4-nitrophenyl, 2 These include 4,6-trimethylphenyl, 2-(2-nitrophenyl)ethyl, butylthiocarbonyl, 4,4',4”-tris(benzoyloxy)trityl, diphenylcarbamoyl, levlinyl, 2-(dibromomethyl)benzoyl (Dbmb), 2-(isopropylthiomethoxymethyl)benzoyl (Ptmt), 9-phenylxanthene-9-yl (Pixyl), or 9-(p-methoxyphenyl)xanthin-9-yl (MOX). In some embodiments, each of the hydroxyl protecting groups is independently selected from acetyl, benzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, and 4,4'-dimethoxytrityl. In some embodiments, the hydroxyl protecting group is selected from the group consisting of trityl, monomethoxytrityl, and 4,4'-dimethoxytrityl.In some embodiments, the phosphite-protecting group is a group that is attached to the phosphite bond (e.g., the internucleotide bond) throughout the entire oligonucleotide synthesis. In some embodiments, the protecting group is attached to the sulfur atom of the phosphorothioate group. In some embodiments, the protecting group is attached to the oxygen atom of the internucleotide phosphorothioate bond. In some embodiments, the protecting group is attached to the oxygen atom of the internucleotide phosphate bond. In some embodiments, the protecting group is 2-cyanoethyl (CE or Cne), 2-trimethylsilylethyl, 2-nitroethyl, 2-sulfonylethyl, methyl, benzyl, o-nitrobenzyl, 2-(p-nitrophenyl)ethyl (NPE or Npe), 2-phenylethyl, 3-(N-tert-butylcarboxamide)-1-propyl, 4-oxopentyl, 4-methylthio-l-butyl, 2-cyano-1,1-dimethylethyl, 4-N-methylaminobutyl, 3-(2-pyridyl)-1-propyl, 2-[N-methyl-N-(2-pyridyl)]aminoethyl, 2-(N-formyl,N-methyl)aminoethyl, or 4-[N-methyl-N-(2,2,2-trifluoroacetyl)amino]butyl.

[0164] Substantially: As used herein, the term “substantially” refers to a qualitative condition that exhibits the entire or nearly entire range or degree of the desired feature or characteristic. A nucleotide sequence substantially complementary to a second sequence is not identical to the second sequence, but is approximately or nearly identical. In addition, those skilled in the art of biology and / or chemistry will understand that it is rare, if any, for biological and chemical phenomena to go to completion and / or progress to completeness or to achieve or avoid absolute results. Thus, the term “substantially” is used herein to capture the potential lack of completeness inherent in many biological and / or chemical phenomena.

[0165] Sugars: The term "sugar" refers to closed-chain and / or open-chain monosaccharides or polysaccharides. In some embodiments, the sugar is a monosaccharide. In some embodiments, the sugar is a polysaccharide. Examples of sugars, but not limited to, include ribose, deoxyribose, pentofuranose, pentopyranose, and hexopyranose moieties. As used herein, the term "sugar" also includes structural analogs used in place of conventional sugar molecules, such as glycols, whose polymers form the backbone of nucleic acid analogs, glycol nucleic acids ("GNAs"), etc. As used herein, the term "sugar" also includes structural analogs used in place of natural or naturally occurring nucleotides, such as modified sugars and nucleotide sugars. In some embodiments, the sugar is an RNA or DNA sugar (ribose or deoxyribose). In some embodiments, the sugar is a modified ribose or deoxyribose sugar, e.g., 2'-modified, 5'-modified, etc. When used in oligonucleotides and / or nucleic acids, as described herein, in some embodiments, modified sugars may provide one or more desired properties, activities, etc. In some embodiments, the sugar is optionally substituted ribose or deoxyribose. In some embodiments, "sugar" refers to a sugar unit in an oligonucleotide or nucleic acid.

[0166] Susceptible to: An individual "susceptible" to a disease, disorder, and / or condition is an individual with a higher risk of developing the disease, disorder, and / or condition compared to a member of the general public. In some embodiments, an individual susceptible to a disease, disorder, and / or condition has a predisposition to that disease, disorder, and / or condition. In some embodiments, an individual susceptible to a disease, disorder, and / or condition may not have been diagnosed with that disease, disorder, and / or condition. In some embodiments, an individual susceptible to a disease, disorder, and / or condition may exhibit symptoms of that disease, disorder, and / or condition. In some embodiments, an individual susceptible to a disease, disorder, and / or condition may not exhibit symptoms of that disease, disorder, and / or condition. In some embodiments, an individual susceptible to a disease, disorder, and / or condition will develop that disease, disorder, and / or condition. In some embodiments, individuals susceptible to disease, disorder, and / or pathology will not develop that disease, disorder, and / or pathology.

[0167] Therapeutic Agent: As used herein, the term “therapeutic agent” generally refers to any agent that, when administered to a subject, produces a desired effect (e.g., a desired biological, clinical, or pharmacological effect). In some embodiments, an agent is considered a therapeutic agent if it demonstrates a statistically significant effect in an appropriate population. In some embodiments, an appropriate population is a population of subjects suffering from and / or susceptible to a disease, disorder, or condition. In some embodiments, an appropriate population is a population of model organisms. In some embodiments, an appropriate population may be defined by one or more criteria, such as age group, sex, genetic background, pre-existing clinical condition, or history of exposure to a therapy. In some embodiments, a therapeutic agent is a substance that, when administered to a subject in an effective dose, alleviates, improves, reduces, inhibits, prevents, delays the onset of, reduces the severity of, and / or decreases the incidence of one or more symptoms or characteristics of the disease, disorder, and / or condition in question. In some embodiments, the “therapeutic agent” is a drug that has received or is required to receive approval from a government agency before it can be placed on the market for administration to humans. In some embodiments, the “therapeutic agent” is a drug that requires a physician’s prescription for administration to humans. In some embodiments, the therapeutic agent is a compound provided, for example, a oligonucleotide provided.

[0168] Therapeutic dose: As used herein, the term “therapeutic dose” means the amount of a substance (e.g., a therapeutic agent, composition, and / or formulation) administered as part of a therapeutic regimen to elicit a desired biological response. In some embodiments, the therapeutic dose of a substance is an amount sufficient to treat, diagnose, prevent, and / or delay the onset of a disease, disorder, and / or condition when administered to a subject suffering from or susceptible to that disease, disorder, and / or condition. As those skilled in the art will understand, the effective dose of a substance may vary depending on factors such as the desired biological endpoint, the substance to be delivered, the target cells or tissues, etc. For example, the effective dose of a compound in a formulation for treating a disease, disorder, and / or condition is an amount that alleviates, improves, reduces, inhibits, prevents, delays the onset, reduces the severity, and / or decreases the incidence of one or more symptoms or characteristics of that disease, disorder, and / or condition. In some embodiments, the therapeutic dose is administered in a single dose; in some embodiments, multiple unit doses are required to deliver the therapeutic dose.

[0169] To treat: As used herein, the terms “to treat,” “treatment,” or “to treat” mean any method used to partially or completely alleviate, improve, reduce, inhibit, prevent, delay the onset, reduce the severity, and / or decrease the incidence of one or more symptoms or features of a disease, disorder, and / or condition. Treatment may be administered to subjects who are not exhibiting signs of a disease, disorder, and / or condition. In some embodiments, treatment may be administered to subjects exhibiting only the initial signs of a disease, disorder, and / or condition, for example, to reduce the risk of developing lesions associated with that disease, disorder, and / or condition.

[0170] Unsaturated: As used herein, the term “unsaturated” means that a portion of a material has one or more unsaturated units.

[0171] 2. Detailed description of specific embodiments In particular, this disclosure provides techniques for preparing oligonucleotide compositions, especially chiral-controlled oligonucleotide compositions, with unexpectedly improved crude purity and yield. In some embodiments, the techniques provided can dramatically reduce commercial costs, and in some embodiments, enable the mass production of therapeutic oligonucleotides under commercially acceptable conditions for clinical use and commercialization, such as cost, purity, and yield. As those skilled in the art will understand, the techniques provided enable the production of various oligonucleotide compositions independent of nucleotide sequence, chemical / stereochemical modifications, modes of activity, chiral adjuvants, etc. Exemplary embodiments of the techniques provided are described herein.

[0172] Oligonucleotides and oligonucleotide compositions In some embodiments, the oligonucleotide compositions of the provided technology, such as product oligonucleotide compositions of various steps, the final oligonucleotide composition, etc., are chiral-controlled oligonucleotide compositions. In some embodiments, the oligonucleotides of the provided technology, such as product oligonucleotides of various steps, the final product oligonucleotide, etc., are oligonucleotides of formula OI or a salt thereof. In some embodiments, each oligonucleotide of a plurality of oligonucleotides is independently an oligonucleotide of formula OI or a salt thereof.

[0173] In some embodiments, the oligonucleotide is of formula OI or a salt thereof: [ka] (In the formula, R 5s These are independently R' or -OR'; Each L s These are independently, covalently, or C 1~30 C having an aliphatic group and 1 to 10 heteroatoms 1~30A linear or branched group that is substituted by optional divalent selection from heteroaliphatic groups, wherein one or more methylene units are optionally and independently C 1~6 Alkylene, C 1~6 Alkenylene, -C≡C-, divalent C1-C6 heteroaliphatic group having 1-5 heteroatoms, -C(R')2-, -Cy-, -O-, -S-, -SS-, -N(R')-, -C(O)-, -C(S)-, -C(NR')-, -C(O)N(R')-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -S(O)-, -S(O)2-, -S(O)2N(R')-, -C(O)S-, -C(O)O-, -P(O)(OR')-, -P(O)(SR')-, -P(O)(R')-, -P(O)(NR')-, -P(S)(OR')-, -P(S)(SR') -, -P(S)(R')-, -P(S)(NR')-, -P(R')-, -P(OR')-, -P(SR')-, -P(NR')-, -P(OR')[B(R')3]-, -OP(O)(OR')O-, -OP(O)(SR')O-, -OP(O)(R')O-, -OP(O)(NR')O-, -OP(OR')O-, -OP(SR')O-, -OP(NR')O-, -OP(R')O- or -OP(OR')[B(R')3]O- are replaced by optionally substituted groups, and one or more carbon atoms are optionally and independently replaced with Cy L Replaced by; Each -Cy- is independent of C 3~20 alicyclic ring, C 6~20 A divalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms; Each Cy L Independently, C 3~20 alicyclic ring, C 6~20 A tetravalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms; Each ring A sThese are independently substituted 3-20 member monocyclic, bicyclic, or polycyclic rings having 0-10 heteroatoms; Each R s These are independently -H, halogen, -CN, -N3, -NO, -NO2, and -L s -R', -L s -Si(R')3, -L s -OR', -L s -SR', -L s -N(R')2, -OL s -R', -OL s -Si(R)3, -OL s -OR', -OL s -SR' or -OL s -N(R')² is; Each t is independently between 0 and 20; Each BA is independent of C 3~30 alicyclic, C 6~30 Aryl, C having 1 to 10 heteroatoms 5~30 Heteroaryls, C having 1 to 10 heteroatoms 3~30 A group that is optionally substituted, selected from heterocyclyl, native nucleic acid base moieties, and modified nucleic acid base moieties; Each L P These are independently internucleotide bonds; z is between 1 and 1000; L 3E is, -L s - or -L s -L s -and; R 3E -R', -L s -R', -OR', or support; Each R' is independently -R, -C(O)R, -C(O)OR, or -S(O)2R; Each R is independently -H or C 1~30 Aliphatic carbon atoms containing 1 to 10 heteroatoms 1~30 Heteroliphatic, C 6~30 Ariel, C 6~30 Aryl aliphatic, C having 1 to 10 heteroatoms 6~30The group is optionally substituted, selected from aryl heteroaliphatic groups, 5-30 membered heteroaryls having 1-10 heteroatoms, and 3-30 membered heterocyclines having 1-10 heteroatoms, or Two R groups, by arbitrary and independent choice, come together to form a covalent bond, or; Two or more R groups on the same atom, by arbitrary choice and independently, combine with that atom to form an optionally substituted 3-30 member monocyclic, bicyclic, or polycyclic ring having 0-10 heteroatoms in addition to that atom; or Two or more R groups on two or more atoms, by arbitrary choice and independently, combine with their intercalating atoms to form an optionally substituted 3- to 30-membered monocyclic, bicyclic, or polycyclic ring having 0 to 10 heteroatoms in addition to the intercalating atoms.

[0174] In some embodiments, the oligonucleotide composition is a chiral-controlled oligonucleotide composition.

[0175] In some mechanisms, chiral controlled oligonucleotide compositions are 1) Common base sequence; 2) Common skeletal connection patterns; 3) One or more independently, for example, about 1 to 50 (e.g., about 5 to 50, about 10 to 50, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45 or 50, etc.) common stereochemistry in chiral internucleotide bonds ("chiral-controlled internucleotide bonds") An oligonucleotide composition comprising multiple oligonucleotides sharing; This composition is chiral-controlled in that the levels of several oligonucleotides in the composition are predetermined.

[0176] In some embodiments, the chiral-controlled oligonucleotide composition is an oligonucleotide composition comprising a plurality of oligonucleotides, where the plurality of oligonucleotides are 1) Base sequence; 2) Skeletal connection patterns; 3) Chiral center patterns of the skeleton; and 4) Skeletal phosphorus modification patterns A specific oligonucleotide type defined by; This composition is chiral-controlled in that the levels of several oligonucleotides in the composition are predetermined.

[0177] In some mechanisms, chiral controlled oligonucleotide compositions are 1) Common base sequence; 2) Common skeletal connection patterns; and 3) Common skeletal chiral center pattern The oligonucleotide composition comprises multiple oligonucleotides sharing a common base sequence and length, a common skeletal bonding pattern, and a common skeletal chiral center pattern, and is substantially a pure formulation of a single oligonucleotide in that a predetermined level of oligonucleotides in the composition have a common base sequence and length, a common skeletal bonding pattern, and a common skeletal chiral center pattern.

[0178] In some embodiments, the chiral-controlled oligonucleotide composition is an oligonucleotide composition comprising multiple oligonucleotides, where, Multiple oligonucleotides share the same base sequence; Multiple oligonucleotides share the same skeletal bonding pattern; and Multiple oligonucleotides contain at least one chiral-controlled internucleotide bond, which is chiral-controlled in that the multiple oligonucleotides share the same stereochemical configuration with respect to the chiral linkage phosphorus of the internucleotide bond; At least ((DS)Nc×100)% of all oligonucleotides sharing the same base sequence in the composition are multiple oligonucleotides, where DS is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, and Nc is the number of chiral-controlled internucleotide bonds.

[0179] In some embodiments, the chiral-controlled oligonucleotide composition is an oligonucleotide composition comprising multiple oligonucleotides, where, Multiple oligonucleotides share the same composition; and Multiple oligonucleotides contain at least one chiral-controlled internucleotide bond, which is chiral-controlled in such a way that the multiple oligonucleotides share the same stereochemical configuration with respect to the chiral linkage phosphorus of the internucleotide bond; and ((DS) Nc (×100)% or more are multiple oligonucleotides, where DS is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, and Nc is the number of chiral-controlled internucleotide bonds.

[0180] In some embodiments, the oligonucleotide provided contains 1 to 30 non-natural internucleotide bonds (not in the form of -OP(O)(OH)-O- or its salts). In some embodiments, the oligonucleotide provided contains 2 to 30 non-natural internucleotide bonds. In some embodiments, the oligonucleotide provided contains 5 to 30 non-natural internucleotide bonds. In some embodiments, the oligonucleotide provided contains 10 to 30 non-natural internucleotide bonds.

[0181] In some embodiments, the provided oligonucleotide contains 1 to 30 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 2 to 30 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 5 to 30 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 10 to 30 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 1 chiral-controlled internucleotide bond. In some embodiments, the provided oligonucleotide contains 2 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 3 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 4 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 5 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 6 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 7 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains 8 chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains nine chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains ten chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains eleven chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains twelve chiral-controlled internucleotide bonds. In some embodiments, the provided oligonucleotide contains thirteen chiral-controlled internucleotide bonds.In some embodiments, the oligonucleotide provided contains 14 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 15 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 16 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 17 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 18 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 19 chiral-controlled internucleotide bonds. In some embodiments, the oligonucleotide provided has 20 chiral-controlled internucleotide bonds. In some embodiments, about 1-100% of all internucleotide bonds are chiral-controlled internucleotide bonds. In some embodiments, about 1-100% of all chiral internucleotide bonds (including chiral linkage phosphates) are chiral-controlled internucleotide bonds. In some embodiments, the percentage is about 5-100%. In some embodiments, the percentage is at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. In some embodiments, the percentage is about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. In some embodiments, each chiral internucleotide bond is chiral-controlled. In some embodiments, some or all of the chiral-controlled internucleotide bonds in the provided oligonucleotide or one or more segments thereof (e.g., 5'-wing region, core wing, 3'-wing region) are continuous.In some embodiments, all chiral-controlled internucleotide bonds in the provided oligonucleotide or one or more segments thereof (e.g., 5'-wing region, core wing, 3'-wing region) are contiguous.

[0182] In some embodiments, the oligonucleotide provided contains 1 to 30 natural phosphate bonds (not in the form of -OP(O)(OH)-O- or its salts). In some embodiments, the oligonucleotide provided further contains 1 to 30 natural phosphate bonds (not in the form of -OP(O)(OH)-O- or its salts) in addition to natural phosphate bonds or chiral internucleotide bonds or chiral-controlled internucleotide bonds as described herein. In some embodiments, the oligonucleotide provided contains 2 to 30 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 5 to 30 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 10 to 30 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 1 chiral-controlled internucleotide bond. In some embodiments, the oligonucleotide provided contains 2 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 3 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 4 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 5 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 6 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 7 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 8 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 9 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 10 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 11 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 12 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 13 natural phosphate bonds. In some embodiments, the oligonucleotide provided contains 14 natural phosphate bonds. In some embodiments, the oligonucleotide provided has 15 natural phosphate bonds.In some embodiments, the oligonucleotide provided has 16 natural phosphate bonds. In some embodiments, the oligonucleotide provided has 17 natural phosphate bonds. In some embodiments, the oligonucleotide provided has 18 natural phosphate bonds. In some embodiments, the oligonucleotide provided has 19 natural phosphate bonds. In some embodiments, the oligonucleotide provided has 20 natural phosphate bonds. In some embodiments, about 1-100% of all internucleotide bonds are natural phosphate bonds. In some embodiments, approximately 1–99% of all internucleotide bonds are native phosphate bonds, and approximately 1–99% of all internucleotide bonds are non-native internucleotide bonds (e.g., internucleotide bonds of formulas VII, VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1 or NL-d-2 or their salt forms and non-native internucleotide bonds). In some embodiments, approximately 1–99% of all internucleotide bonds are native phosphate bonds, and approximately 1–99% of all internucleotide bonds are chiral internucleotide bonds (e.g., internucleotide bonds of formulas VII, VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1, or NL-d-2, or their salt forms and being chiral).In some embodiments, approximately 1-99% of all internucleotide bonds are natural phosphate bonds, and approximately 1-99% of all internucleotide bonds are chiral-controlled oligonucleotide composition internucleotide bonds (e.g., internucleotide bonds of formulas VII, VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1, or NL-d-2 or their salt forms and chiral-controlled). In some embodiments, the percentage is approximately 5-100%. In some embodiments, the percentage is at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. In some embodiments, the percentage is about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. In some embodiments, each non-natural internucleotide bond independently has the structure of formula VII, formula VII-a-1, formula VII-a-2, formula VII-b, formula VII-c, formula VII-d, formula VII-e, formula NL-n-1, formula NL-n-2, formula NL-n-3, formula NL-n-4, formula NL, formula NL-a-1, formula NL-a-2, formula NL-b-1, formula NL-b-2, formula NL-c-1, formula NL-c-2, formula NL-d-1 or formula NL-d-2, or a salt form thereof, where formula V II. The structures or salt forms of formulas VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1, or NL-d-2 are non-natural internucleotide bonds (not -OP(O)(OH)-O- or their salt forms).In some embodiments, each chiral bond independently has the structure of formula VII, formula VII-a-1, formula VII-a-2, formula VII-b, formula VII-c, formula VII-d, formula VII-e, formula NL-n-1, formula NL-n-2, formula NL-n-3, formula NL-n-4, formula NL, formula NL-a-1, formula NL-a-2, formula NL-b-1, formula NL-b-2, formula NL-c-1, formula NL-c-2, formula NL-d-1, or formula NL-d-2, or a salt form thereof, where formula VI I, the structure of formulas VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1, or NL-d-2, or their salt forms, are not natural phosphate bonds (-OP(O)(OH)-O- or their salt forms). In some embodiments, each chiral controlled phosphate bond independently has the structure of formula VII, formula VII-a-1, formula VII-a-2, formula VII-b, formula VII-c, formula VII-d, formula VII-e, formula NL-n-1, formula NL-n-2, formula NL-n-3, formula NL-n-4, formula NL, formula NL-a-1, formula NL-a-2, formula NL-b-1, formula NL-b-2, formula NL-c-1, formula NL-c-2, formula NL-d-1 or formula NL-d-2, or a salt form thereof, where formula VII The structures or salt forms thereof of formulas VII-a-1, VII-a-2, VII-b, VII-c, VII-d, VII-e, NL-n-1, NL-n-2, NL-n-3, NL-n-4, NL, NL-a-1, NL-a-2, NL-b-1, NL-b-2, NL-c-1, NL-c-2, NL-d-1, or NL-d-2 or VII-e are not natural phosphate bonds (-OP(O)(OH)-O- or their salt forms). In some embodiments, some or all of the natural phosphate bonds in the provided oligonucleotide or one or more segments thereof (e.g., 5'-wing region, core wing, 3'-wing region) are continuous. In some embodiments, all native phosphate bonds in the provided oligonucleotide or one or more segments thereof (e.g., 5'-wing region, core wing, 3'-wing region) are contiguous.

[0183] In some embodiments, the non-natural internucleotide bond is a phosphorothioate bond or a salt thereof (-OP(O)(SH)-O- or a salt thereof). In some embodiments, the chiral internucleotide bond is a phosphorothioate bond or a salt thereof (-OP(O)(SH)-O- or a salt thereof). In some embodiments, the chiral-controlled internucleotide bond is a phosphorothioate bond or a salt thereof (-OP(O)(SH)-O- or a salt thereof).

[0184] In some embodiments, the provided oligonucleotides contain 5-200, 5-150, 5-100, 5-50, 5-40, 5-35, 5-30, 5-25, 10-200, 10-150, 10-100, 10-50, 10-40, 10-35, 10-30, 10-25, 15-200, 15-150, 15-100, 15-50, 15-40, 15-35, 15-30, or 15-25 nucleic acid bases. In some embodiments, the oligonucleotides provided contain at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, or 50 nucleic acid bases. In some embodiments, the oligonucleotides provided contain at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, or 50 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, or 50 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 35, 40, 45, or 50 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 15 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 16 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 17 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 18 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 19 nucleic acid bases. In some embodiments, the provided oligonucleotide contains at least 20 nucleic acid bases. In some embodiments, the provided oligonucleotide contains at least 21 nucleic acid bases.In some embodiments, the oligonucleotide provided contains at least 22 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 23 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 24 nucleic acid bases. In some embodiments, the oligonucleotide provided contains at least 25 nucleic acid bases. In some embodiments, the nucleic acid bases are optionally substituted adenine, cytosine, guanosine, thymine, or uracil, or tautomers thereof.

[0185] In some embodiments, each chiral bound phosphorus independently has the diastereomer purity as described in this disclosure. In some embodiments, the provided compound has the purity, diastereomer purity and / or enantiomer purity as described in this disclosure. In some embodiments, the provided compound has the purity as described in this disclosure. In some embodiments, the provided compound has the diastereomer purity as described in this disclosure. In some embodiments, the provided compound has the enantiomer purity as described in this disclosure. In some embodiments, the provided compound has the diastereomer purity and enantiomer purity as described in this disclosure.

[0186] In some embodiments, the oligonucleotide provided comprises or has a 5'-wing region-core region-3'-wing region structure. In some embodiments, the oligonucleotide provided comprises or has a 5'-wing region-core region structure. In some embodiments, the oligonucleotide provided comprises or has a core region-3'-wing region structure. In some embodiments, the oligonucleotide provided comprises a 5'-wing region-core region-3'-wing region structure. In some embodiments, the oligonucleotide provided comprises a 5'-wing region-core region structure. In some embodiments, the oligonucleotide provided comprises a core region-3'-wing region structure. In some embodiments, the oligonucleotide provided has a 5'-wing region-core region-3'-wing region structure. In some embodiments, the oligonucleotide provided has a 5'-wing region-core region structure. In some embodiments, the oligonucleotide provided has a core region-3'-wing region structure. In some embodiments, the wing-core-wing (i.e., XYX) motif is represented by the numbers, for example, 5-10-4, meaning that the 5'-wing region is 5 nucleotides long, the core region is 10 nucleotides long, and the 3'-wing region is 4 nucleotides long. In some embodiments, the wing-core-wing motif is one of the following, for example: 2-16-2, 3-14-3, 4-12-4, 5-10-5, 2-9-6, 3-9-3, 3-9-4, 3-9-5, 4-7-4, 4-9-3, 4-9-4, 4-9-5, 4-10-5, 4-11-4, 4-11-5, 5-7-5, 5-8-6, 8-7-5, 7-7-6, 5-9-3, 5-9-5, 5-10-4, 5-10-5, 6-7-6, 6-8-5, and 6-9-2. In a particular embodiment, the wing-core-wing motif is 5-10-5. In a particular embodiment, the wing-core-wing motif is 7-7-6. In certain embodiments, the wing-core-wing motif is 8-7-5. In some embodiments, the wing-core motifs are 5-15, 6-14, 7-13, 8-12, 9-12, etc.In some embodiments, the core-wing motifs are 5-15, 6-14, 7-13, 8-12, 9-12, etc.

[0187] In some embodiments, the wing region contains sugar modifications not present in the core region. In some embodiments, the wing region contains 2'-modifications. In some embodiments, each nucleotide unit of the wing region independently contains a 2'-modification. In some embodiments, each nucleotide unit of the wing region independently contains the same 2'-modification. In some embodiments, each nucleotide unit of the 5'-wing region independently contains the same 2'-modification. In some embodiments, each nucleotide unit of the 3'-wing region independently contains the same 2'-modification. In some embodiments, the 2'-modifications of the 5'-wing region are the same. In some embodiments, the 2'-modifications of the 5'-wing region are different. In some embodiments, the 2'-modification is 2'-OR, where R' is not hydrogen. In some embodiments, the 2'-modification is 2'-OR, where R' is optionally substituted C. 1~6 It is aliphatic. In some embodiments, the 2'-modification is 2'-OR, where R' is optionally substituted with C. 1~6 It is alkyl. In some embodiments, the 2'-modification is 2'-OMe. In some embodiments, the 2'-modification is 2'-OCH2CH2OMe. In some embodiments, the wing region comprises one or more native phosphate bonds as described in this disclosure. In addition or alternatively, in some embodiments, the wing region comprises one or more non-native internucleotide bonds, e.g., phosphorothioate internucleotide bonds. In some embodiments, the core region comprises one or more native phosphate bonds. In some embodiments, the core region comprises one or more consecutive native phosphate bonds. In some embodiments, the core region comprises one or more chiral phosphate bonds. In some embodiments, the core region comprises one or more consecutive chiral phosphate bonds. In some embodiments, the chiral phosphate bond is a phosphorothioate bond. In some embodiments, the chiral phosphate bond is chiral controlled.

[0188] The oligonucleotides of this disclosure may contain a skeletal chiral center pattern. In some embodiments, the skeletal chiral center pattern of the oligonucleotide or its segment, for example, the core region, results in increased stability. In some embodiments, the skeletal chiral center pattern unexpectedly results in increased activity. In some embodiments, the skeletal chiral center pattern results in increased stability and activity. In some embodiments, the skeletal chiral center pattern unexpectedly results in increased binding to specific proteins. In some embodiments, the skeletal chiral center pattern unexpectedly results in enhanced delivery. In some embodiments, the skeletal chiral center pattern includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, (Sp)p(Rp)n(Sp)m, (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m (from 5' to 3' unless otherwise specified), where n is 1 to 10 and each of p and m is independently 0 to 50. In some embodiments, the skeletal chiral center pattern includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, (Sp)p(Rp)n(Sp)m, (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m (from 5' to 3' unless otherwise specified), where n is 1 to 10 and each of p and m is independently 1 to 50. In some embodiments, the skeletal chiral center pattern includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m (from 5' to 3' unless otherwise specified), where n is 1 to 10 and each of p and m is independently 1 to 50. In some embodiments, the skeletal chiral center pattern includes (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m.In some embodiments, the skeletal chiral center pattern is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is a repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is a repeating pattern of (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m (from 5' to 3'). In some embodiments, the skeletal chiral center pattern includes or is a repeating pattern of (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is a repeating pattern of (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m (from 5' to 3').

[0189] In some embodiments, the skeletal chiral center pattern includes or is (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, where m>2. In some embodiments, the skeletal chiral center pattern includes or is (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, where n is 1, p>1, and m>2. In some embodiments, m>3. In some embodiments, m>4. In some embodiments, the skeletal chiral center pattern includes or is (Op)n(Sp)m, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m, where m>2. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m, where n is 1, p>1, and m>2. In some embodiments, m>3. In some embodiments, m>4. In some embodiments, the skeletal chiral center pattern of an oligonucleotide or a region thereof includes or comprises two or more units independently selected from (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, (Sp)p(Rp)n(Sp)m, (Op)n(Sp)m, (Np)p(Op)n(Sp)m, and (Sp)p(Op)n(Sp)m, where each variable element is independently as described in this disclosure. In some embodiments, n is 1. In some embodiments, n is 1, and m in each unit is independently 2 or more as described in this disclosure. In some embodiments, at least two m in two units are different. In some embodiments, the skeletal chiral center pattern includes 2, 3, 4, 5, 6, 7, 8, 9, or 10 such units. In some embodiments, the skeletal chiral center pattern includes 2 such units, but not more than 2. In some embodiments, the skeletal chiral center pattern includes 3 such units, but not more than 3. In some embodiments, the skeletal chiral center pattern includes 4 such units, but not more than 4.In some embodiments, the skeletal chiral center pattern contains five such units, but not more than five. In some embodiments, a region of an oligonucleotide contains such skeletal chiral center pattern. In some embodiments, such region does not contain a 2'-substitution (two 2'-H) in its sugar portion. In some embodiments, such region is adjacent to a 5'-region containing a sugar modification as described in this disclosure (e.g., a 2'-modification as described in this disclosure, e.g., 2'-OMe, 2'-MOE, 2'-F, etc.) and / or a 5'-region containing a sugar modification as described in this disclosure (e.g., a 2'-modification as described in this disclosure, e.g., 2'-OMe, 2'-MOE, 2'-F, etc.).

[0190] In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m, (Sp)p(Sp)m, or (Sp)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, and the oligonucleotide includes one or more 2'-modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, and the oligonucleotide includes one or more 2'-F modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, and the oligonucleotide includes one or more 2'-OR modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m, and the oligonucleotide contains one or more 2'-OR modifications, where R is not -H. In some embodiments, the skeletal chiral center pattern includes or comprises (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m, and each nucleoside unit between internucleotide bonds having the pattern of (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m does not contain 2'-modifications.In some embodiments, the skeletal chiral center pattern includes or is (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m, and each nucleoside unit between internucleotide bonds having the pattern (Rp)n(Sp)m, (Sp)p(Rp)n, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m does not contain a 2'-substitution (-CH2- at the 2'-position). In some embodiments, the skeletal chiral center pattern includes or is (Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is (Sp)p(Rp)n. In some embodiments, the skeletal chiral center pattern includes or is (Np)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes (Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes (Sp)p(Rp)n. In some embodiments, the skeletal chiral center pattern includes (Np)(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern is (Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern is (Sp)p(Rp)n. In some embodiments, the skeletal chiral center pattern is (Np)p(Rp)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is (Sp)p(Sp)m, and optionally there are n achiral phosphodiester internucleotide bonds and / or stereorandom (un-chiral controlled) chiral internucleotide bonds between the section having (Sp)p and the section having (Sp)m. In some embodiments, there are n achiral phosphodiester internucleotide bonds between them. In some embodiments, there are n stereorandom chiral internucleotide bonds between them. In some embodiments, the skeletal chiral center pattern contains or is (Sp)p(Rp)n(Sp)m.

[0191] In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m, (Sp)p(Sp)m, or (Sp)p(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m, and the oligonucleotide includes one or more 2'-modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m, and the oligonucleotide includes one or more 2'-F modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m, and the oligonucleotide includes one or more 2'-OR modifications as described herein. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m, and the oligonucleotide contains one or more 2'-OR modifications, where R is not -H. In some embodiments, the skeletal chiral center pattern includes or comprises (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m, and each nucleoside unit between internucleotide bonds having the pattern of (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m does not contain 2'-modifications.In some embodiments, the skeletal chiral center pattern includes or is (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m, and each nucleoside unit between internucleotide bonds having the pattern (Op)n(Sp)m, (Sp)p(Op)n, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m does not contain a 2'-substitution (-CH2- at the 2'-position). In some embodiments, the skeletal chiral center pattern includes or is (Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is (Sp)p(Op)n. In some embodiments, the skeletal chiral center pattern includes or is (Np)p(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes (Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes (Sp)p(Op)n. In some embodiments, the skeletal chiral center pattern includes (Np)(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern is (Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern is (Sp)p(Op)n. In some embodiments, the skeletal chiral center pattern is (Np)p(Op)n(Sp)m. In some embodiments, the skeletal chiral center pattern includes or is (Sp)p(Sp)m, and optionally there are n achiral phosphodiester internucleotide bonds and / or stereorandom (un-chiral controlled) chiral internucleotide bonds between the section having (Sp)p and the section having (Sp)m. In some embodiments, there are n achiral phosphodiester internucleotide bonds between them. In some embodiments, there are n stereorandom chiral internucleotide bonds between them. In some embodiments, the skeletal chiral center pattern contains or is (Sp)p(Op)n(Sp)m.

[0192] In some embodiments, an oligonucleotide or a region thereof contains a skeletal chiral center pattern or repeating pattern of (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m (structurally, each starting from the first internucleotide bond of an internucleotide bond having the pattern or repeating pattern of (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m, and ending at the last internucleotide bond; depending on whether it is repeating or not, these are respectively the "(repeating) (Sp)m(Rp)n region," the "(repeating) (Rp)n(Sp)m region," the "(repeating) (Np)p(Rp)n(Sp)m region," or the "(repeating) (Sp)p(Rp)n(Sp)m region"). In some embodiments, an oligonucleotide or a region thereof contains a (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m skeletal chiral center pattern or repeating pattern (structurally, each starting from the first internucleotide bond of an internucleotide bond having the pattern or repeating pattern of (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m, and ending at the last internucleotide bond; depending on whether it is repeating or not, these are respectively the "(repeating) (Sp)m(Op)n region", the "(repeating) (Op)n(Sp)m region", the "(repeating) (Np)p(Op)n(Sp)m region", or the "(repeating) (Sp)p(Op)n(Sp)m region"). For example, the (Sp)p(Rp)n(Sp)m region ((Sp)7(Rp)1(Sp)3 in WV-2555): [ka] ) does not contain 2'-OR sugar modifications. In some embodiments, each sugar moiety in this region has -CH2- at the 2' position. In some embodiments, each sugar moiety in this region is an unmodified native 2'-deoxyribose moiety of DNA. In some embodiments, a region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m is adjacent to a 5'-wing region which structurally terminates with a nucleoside moiety (this nucleoside moiety is linked at its 3' end to the first internucleotide bond of the region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m or (Sp)p(Rp)n(Sp)m). For example, the adjacent 5'-wing region in WV-2555: [ka] In some embodiments, a region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m is adjacent to a 5'-wing region which structurally terminates with a nucleoside moiety (this nucleoside moiety is linked at its 3' end to the first internucleotide bond of the region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m). In some embodiments, a region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m is adjacent to a 3'-wing region which structurally begins with a nucleoside moiety (this nucleoside moiety is linked at its 5' end to the last internucleotide bond of the region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m). For example, adjacent 3'-wing regions in WV-2555: [ka] In some embodiments, a 3'-wing region is adjacent to a region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m, which structurally begins with a nucleoside moiety (this nucleoside moiety is linked at its 5' end to the last internucleotide bond of the region containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m or (Sp)p(Op)n(Sp)m). In some embodiments, regions containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m are flanked by a 5' end and a 3'-wing region. In some embodiments, regions containing a skeletal chiral center pattern or repeating pattern that includes or is (Sp)m(Op)n, (Op)n(Sp)m, (Np)p(Op)n(Sp)m, or (Sp)p(Op)n(Sp)m are flanked by a 5' end and a 3'-wing region. In some embodiments, the flanked 5'-wing region and / or 3'-wing region contain a non-natural internucleotide bond. In some embodiments, the flanked 5'-wing region and / or 3'-wing region contain a chiral internucleotide bond. In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions include chiral-controlled internucleotide bonds. In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions include modified internucleotide bonds containing Sp-linked phosphate. In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions include Sp-phosphorothioate bonds. In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions include one or more native phosphate bonds. In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions include one or more consecutive native phosphate bonds.In some embodiments, the adjacent 5' end comprises a single modified internucleotide bond, which is a 5' terminal internucleotide bond, and one or more consecutive native phosphate bonds (for example, in WV-2555). [ka] ). In some embodiments, the adjacent 3' end includes a single modified internucleotide bond which is a 3' terminal internucleotide bond and one or more consecutive native phosphate bonds (for example, in WV-2555, [ka] ). In some embodiments, adjacent 5'-wing regions and / or 3'-wing regions contain 2'-modified sugar units. In some embodiments, each sugar unit in the 5'-wing region and / or 3'-wing region is independently modified. In some embodiments, each sugar unit in the 5'-wing region and / or 3'-wing region independently contains 2'-modification (e.g., m, 2'-OMe:mA in WV-2555). * SmGmCmUmU * SC * ST * ST * SG * ST * SC * SC * RA * SG * SC * SmUmUmUmA * SmU). In some embodiments, each sugar unit in the 5'-wing region and / or 3'-wing region contains the same 2'-modification. In some embodiments, the 2'-modification is 2'-OR, where R is optionally substituted with C. 1~6 It is aliphatic. In some embodiments, the 2'-modification is 2'-OMe. In some embodiments, the 2'-modification is 2'-MOE. In some embodiments, the 2'-modification is LNA modification (which includes a type of C2-C4 crosslink).

[0193] In some embodiments, the oligonucleotide comprises a 2'-F modified sugar and includes one or more wings and one or more cores (e.g., wing-core-wing, core-wing, wing-core, etc.). In some embodiments, the wings, e.g., 5'-wing, 3'-wing, etc., contain one or more 2'-F modified sugars. In some embodiments, most of the sugars in the wings contain 2'-F modification. In some embodiments, each sugar in the wings contains 2'-F modification. In some embodiments, the internucleotide bond bound to two 2'-F modified sugars is a chiral modified internucleotide bond. In some embodiments, each internucleotide bond bound to two 2'-F modified sugars is independently a chiral modified internucleotide bond. In some embodiments, each chiral modified internucleotide bond is independently a phosphorothioate internucleotide bond or a non-negatively charged internucleotide bond. In some embodiments, each chiral modified internucleotide bond is independently a phosphorothioate internucleotide bond or a neutral internucleotide bond. In some embodiments, the chiralized internucleotide bond is a phosphorothioate internucleotide bond. In some embodiments, each chiralized internucleotide bond is a phosphorothioate internucleotide bond. In some embodiments, the chiralized internucleotide bond is chiralized. In some embodiments, the chiralized internucleotide bond is chiralized and Sp at the bound phosphorus. In some embodiments, the chiralized internucleotide bond is a chiralized phosphorothioate internucleotide bond and Sp at the bound phosphorus. In some embodiments, the chiralized internucleotide bond is a non-negatively charged internucleotide bond and Rp at the bound phosphorus. In some embodiments, each chiralized phosphorothioate internucleotide bond is Sp. In some embodiments, each chiralized phosphorothioate internucleotide bond attached to two 2'-F modified sugars is Sp.In some embodiments, each internucleotide bond attached to the two 2'-F modified sugars is independently a chiral-controlled phosphorothioate internucleotide bond, which is Sp.

[0194] In some embodiments, the core comprises one or more 2'-F modified sugars and one or more 2'-OR modified sugars (wherein R is as described herein). In some embodiments, R is optionally substituted with C 1~4It is alkyl. In some embodiments, R is -OMe. In some embodiments, the core comprises alternating 2'-F modified sugars and 2'-OR modified sugars. In some embodiments, one or more 2'-OR modified sugars are consecutive. In some embodiments, the proportion of one or more 2'-F modified sugars in the core is lower than the proportion for a wing or for each wing. In some embodiments, the core contains one or more native phosphate bonds. In some embodiments, the internucleotide bond attached to two 2'-OR modified sugars is a native phosphate bond. In some embodiments, each internucleotide bond attached to two 2'-OR modified sugars is a native phosphate bond. In some embodiments, the internucleotide bonds attached to the 2'-F modified sugar and the 2'-OR modified sugar are native phosphate bonds. In some embodiments, the internucleotide bond was attached to the 2'-OR modified sugar at its 3' position. In some embodiments, each native phosphate bond is independently attached to the 2'-OR modified sugar at its 3' position. In some embodiments, the internucleotide bonds attached to the 2'-F modified sugar and the 2'-OR modified sugar are modified internucleotide bonds (e.g., phosphorothioate internucleotide bonds), which are optionally chiralized (and optionally sp at their phosphate group). In some embodiments, each modified internucleotide bond is independently a phosphorothioate internucleotide bond. In some embodiments, each modified internucleotide bond is chiralized. In some embodiments, each modified internucleotide bond is a chiralized sp phosphorothioate internucleotide bond. In some embodiments, each internucleotide bond in the core is independently a native phosphate bond or a phosphorothioate internucleotide bond. In some embodiments, each internucleotide bond in the core is independently a native phosphate bond or a chiralized phosphorothioate internucleotide bond. In some embodiments, each internucleotide bond in the core is independently a native phosphate bond or a chiralized sp internucleotide bond.In some embodiments, each internucleotide bond in the core is independently either a native phosphate bond or a chiral-controlled Sp phosphorothioate internucleotide bond.

[0195] Examples of oligonucleotides including the wing and core regions described herein include WV-1714, WV-2444, WV-2445, WV-2526, WV-2527, WV-2528, WV-2530, WV-2531, WV-2578, WV-2580, WV-2587, WV-3047, WV-3152, WV-3472, WV-3473, WV-3507, WV-3508, and WV-35 09, WV-3510, WV-3511, WV-3512, WV-3513, WV-3514, WV-3515, WV-3545, WV-3546, WV-9517, WV-12555, WV-1255 6, WV-12558, WV-12876, WV-12877, WV-12878, WV-12880, WV-13826, WV-13835, WV-13864, or WV-14344.

[0196] In some embodiments, n is 1 to 10. In some embodiments, n is 1, 2, 3, 4, 5, 6, 7, or 8. In some embodiments, n is 1. In some embodiments, n is 2, 3, 4, 5, 6, 7, or 8. In some embodiments, n is 3, 4, 5, 6, 7, or 8. In some embodiments, n is 4, 5, 6, 7, or 8. In some embodiments, n is 5, 6, 7, or 8. In some embodiments, n is 6, 7, or 8. In some embodiments, n is 7 or 8. In some embodiments, n is 1. In some embodiments, n is 2. In some embodiments, n is 3. In some embodiments, n is 4. In some embodiments, n is 5. In some embodiments, n is 6. In some embodiments, n is 7. In some embodiments, n is 8. In some embodiments, n is 9. In some embodiments, n is 10.

[0197] In some embodiments, n is 1. In some embodiments, m is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, m is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, p is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, m is at least 2. In some embodiments, p is at least 2. In some embodiments, n is 1. In some embodiments, m is at least 2, p is at least 2, and n is 1. In some embodiments, p is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, each of m and p is independently at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, each of m and p is independently 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, at least one of m and p is at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, m is at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15. In some embodiments, p is at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15.

[0198] In some embodiments, p is 0 to 50. In some embodiments, p is 1 to 50. In some embodiments, p is 1. In some embodiments, p is 2 to 50. In some embodiments, p is 2, 3, 4, 5, 6, 7, or 8. In some embodiments, p is 3, 4, 5, 6, 7, or 8. In some embodiments, p is 4, 5, 6, 7, or 8. In some embodiments, p is 5, 6, 7, or 8. In some embodiments, p is 6, 7, or 8. In some embodiments, p is 7 or 8. In some embodiments, p is 2. In some embodiments, p is 3. In some embodiments, p is 4. In some embodiments, p is 5. In some embodiments, p is 6. In some embodiments, p is 7. In some embodiments, p is 8. In some embodiments, p is 9. In some embodiments, p is 10. In some embodiments, p is 11. In some embodiments, p is 12. In some embodiments, p is 13. In some embodiments, p is 14. In some embodiments, p is 15. In some embodiments, p is 16. In some embodiments, p is 17. In some embodiments, p is 18. In some embodiments, p is 19. In some embodiments, p is 20. In some embodiments, p is 21. In some embodiments, p is 22. In some embodiments, p is 23. In some embodiments, p is 24. In some embodiments, p is 25. In some embodiments, p is at least 2. In some embodiments, p is at least 3. In some embodiments, p is at least 4. In some embodiments, p is at least 5. In some embodiments, p is at least 6. In some embodiments, p is at least 7. In some embodiments, p is at least 8. In some embodiments, p is at least 9. In some embodiments, p is at least 10.In some embodiments, p is at least 11. In some embodiments, p is at least 12. In some embodiments, p is at least 13. In some embodiments, p is at least 14. In some embodiments, p is at least 15. In some embodiments, p is at least 16. In some embodiments, p is at least 17. In some embodiments, p is at least 18. In some embodiments, p is at least 19. In some embodiments, p is at least 20. In some embodiments, p is at least 21. In some embodiments, p is at least 22. In some embodiments, p is at least 23. In some embodiments, p is at least 24. In some embodiments, p is at least 25.

[0199] In some embodiments, m is 0 to 50. In some embodiments, m is 1 to 50. In some embodiments, m is 1. In some embodiments, m is 2 to 50. In some embodiments, m is 2, 3, 4, 5, 6, 7, or 8. In some embodiments, m is 3, 4, 5, 6, 7, or 8. In some embodiments, m is 4, 5, 6, 7, or 8. In some embodiments, m is 5, 6, 7, or 8. In some embodiments, m is 6, 7, or 8. In some embodiments, m is 7 or 8. In some embodiments, m is 0. In some embodiments, m is 1. In some embodiments, m is 2. In some embodiments, m is 3. In some embodiments, m is 4. In some embodiments, m is 5. In some embodiments, m is 6. In some embodiments, m is 7. In some embodiments, m is 8. In some embodiments, m is 9. In some embodiments, m is 10. In some embodiments, m is 11. In some embodiments, m is 12. In some embodiments, m is 13. In some embodiments, m is 14. In some embodiments, m is 15. In some embodiments, m is 16. In some embodiments, m is 17. In some embodiments, m is 18. In some embodiments, m is 19. In some embodiments, m is 20. In some embodiments, m is 21. In some embodiments, m is 22. In some embodiments, m is 23. In some embodiments, m is 24. In some embodiments, m is 25. In some embodiments, m is at least 2. In some embodiments, m is at least 3. In some embodiments, m is at least 4. In some embodiments, m is at least 5. In some embodiments, m is at least 6. In some embodiments, m is at least 7. In some embodiments, m is at least 8. In some embodiments, m is at least 9.In some embodiments, m is at least 10. In some embodiments, m is at least 11. In some embodiments, m is at least 12. In some embodiments, m is at least 13. In some embodiments, m is at least 14. In some embodiments, m is at least 15. In some embodiments, m is at least 16. In some embodiments, m is at least 17. In some embodiments, m is at least 18. In some embodiments, m is at least 19. In some embodiments, m is at least 20. In some embodiments, m is at least 21. In some embodiments, m is at least 22. In some embodiments, m is at least 23. In some embodiments, m is at least 24. In some embodiments, m is at least 25. In some embodiments, m is at least greater than 25.

[0200] In some embodiments, at least one of m and p is greater than 2. In some embodiments, at least one of m and p is greater than 3. In some embodiments, at least one of m and p is greater than 4. In some embodiments, at least one of m and p is greater than 5. In some embodiments, at least one of m and p is greater than 6. In some embodiments, at least one of m and p is greater than 7. In some embodiments, at least one of m and p is greater than 8. In some embodiments, at least one of m and p is greater than 9. In some embodiments, at least one of m and p is greater than 10. In some embodiments, at least one of m and p is greater than 11. In some embodiments, at least one of m and p is greater than 12. In some embodiments, at least one of m and p is greater than 13. In some embodiments, at least one of m and p is greater than 14. In some embodiments, at least one of m and p is greater than 15. In some embodiments, at least one of m and p is greater than 16. In some embodiments, at least one of m and p is greater than 17. In some embodiments, at least one of m and p is greater than 18. In some embodiments, at least one of m and p is greater than 19. In some embodiments, at least one of m and p is greater than 20. In some embodiments, at least one of m and p is greater than 21. In some embodiments, at least one of m and p is greater than 22. In some embodiments, at least one of m and p is greater than 23. In some embodiments, at least one of m and p is greater than 24. In some embodiments, at least one of m and p is greater than 25.

[0201] In some embodiments, each of m and p is greater than 2. In some embodiments, each of m and p is greater than 3. In some embodiments, each of m and p is greater than 4. In some embodiments, each of m and p is greater than 5. In some embodiments, each of m and p is greater than 6. In some embodiments, each of m and p is greater than 7. In some embodiments, each of m and p is greater than 8. In some embodiments, each of m and p is greater than 9. In some embodiments, each of m and p is greater than 10. In some embodiments, each of m and p is greater than 11. In some embodiments, each of m and p is greater than 12. In some embodiments, each of m and p is greater than 13. In some embodiments, each of m and p is greater than 14. In some embodiments, each of m and p is greater than 15. In some embodiments, each of m and p is greater than 16. In some embodiments, each of m and p is greater than 17. In some embodiments, each of m and p is greater than 18. In some embodiments, each of m and p is greater than 19. In some embodiments, each of m and p is greater than 20.

[0202] In some embodiments, the sum of m and p is greater than 3. In some embodiments, the sum of m and p is greater than 4. In some embodiments, the sum of m and p is greater than 5. In some embodiments, the sum of m and p is greater than 6. In some embodiments, the sum of m and p is greater than 7. In some embodiments, the sum of m and p is greater than 8. In some embodiments, the sum of m and p is greater than 9. In some embodiments, the sum of m and p is greater than 10. In some embodiments, the sum of m and p is greater than 11. In some embodiments, the sum of m and p is greater than 12. In some embodiments, the sum of m and p is greater than 13. In some embodiments, the sum of m and p is greater than 14. In some embodiments, the sum of m and p is greater than 15. In some embodiments, the sum of m and p is greater than 16. In some embodiments, the sum of m and p is greater than 17. In some embodiments, the sum of m and p is greater than 18. In some embodiments, the sum of m and p is greater than 19. In some embodiments, the sum of m and p is greater than 20. In some embodiments, the sum of m and p is greater than 21. In some embodiments, the sum of m and p is greater than 22. In some embodiments, the sum of m and p is greater than 23. In some embodiments, the sum of m and p is greater than 24. In some embodiments, the sum of m and p is greater than 25.

[0203] In some embodiments, n is 1 and at least one of m and p is greater than 1. In some embodiments, n is 1 and each of m and p is independently greater than 1. In some embodiments, m > n and p > n. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)2Rp(Sp)2. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)2Rp(Sp)2. In some embodiments, (Sp)p(Rp)n(Sp)m is SpRp(Sp)2. In some embodiments, (Np)p(Rp)n(Sp)m is (Np)tRp(Sp)m. In some embodiments, (Np)p(Rp)n(Sp)m is (Np)2Rp(Sp)m. In some embodiments, (Np)p(Rp)n(Sp)m is (Rp)2Rp(Sp)m. In some embodiments, (Np)p(Rp)n(Sp)m is (Sp)2Rp(Sp)m. In some embodiments, (Np)p(Rp)n(Sp)m is RpSpRp(Sp)m. In some embodiments, (Np)p(Rp)n(Sp)m is SpRpRp(Sp)m.

[0204] In some embodiments, (Sp)p(Rp)n(Sp)m is SpRpSpSp. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)2Rp(Sp)2. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)3Rp(Sp)3. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)4Rp(Sp)4. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)tRp(Sp)5. In some embodiments, (Sp)p(Rp)n(Sp)m is SpRp(Sp)5. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)2Rp(Sp)5. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)3Rp(Sp)5. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)4Rp(Sp)5. In some embodiments, (Sp)p(Rp)n(Sp)m is (Sp)5Rp(Sp)5.

[0205] In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)2Rp(Sp)2. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)3Rp(Sp)3. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)4Rp(Sp)4. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)mRp(Sp)5. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)2Rp(Sp)5. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)3Rp(Sp)5. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)4Rp(Sp)5. In some embodiments, (Sp)m(Rp)n(Sp)p is (Sp)5Rp(Sp)5.

[0206] In some embodiments, the oligonucleotides provided are blockmers. In some embodiments, the oligonucleotides provided are altomers. In some embodiments, the oligonucleotides provided are altomers comprising alternating blocks. In some embodiments, blockmers or altomers can be defined by chemical modifications (including or omitting them), such as base modifications, sugar modifications, internucleotide bond modifications, stereochemistry, etc., or patterns thereof. Exemplary chemical modifications, stereochemistry, and patterns thereof of blocks and / or alternating units include, but are not limited to, those described for oligonucleotides and others described in this disclosure. In some embodiments, blockmers include the pattern ..SS..RR..SS..RR.. In some embodiments, altomers include the pattern SRSRSRSR.

[0207] In some embodiments, the provided skeletal chiral center pattern includes repeating (Sp)m(Rp)n, (Rp)n(Sp)m, (Np)p(Rp)n(Sp)m, or (Sp)p(Rp)n(Sp)m units. In some embodiments, the repeating unit is (Sp)m(Rp)n. In some embodiments, the repeating unit is SpRp. In some embodiments, the repeating unit is SpSpRp. In some embodiments, the repeating unit is SpRpRp. In some embodiments, the repeating unit is RpRpSp. In some embodiments, the repeating unit is (Rp)n(Sp)m. In some embodiments, the repeating unit is (Np)p(Rp)n(Sp)m. In some embodiments, the repeating unit is (Sp)p(Rp)n(Sp)m.

[0208] In some embodiments, the oligonucleotides of this disclosure are U.S. Patent Publication No. 20150211006, U.S. Patent Publication No. 20170037399, U.S. Patent Publication No. 20180216107, U.S. Patent Publication No. 20180216108, U.S. Patent Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication Nos. 2017 / 015575, 2017 / 062862, 2017 / 160741, 2017 / 192664, 2017 / 192679, 2017 / 210647, 2018 / 022473, 2018 / 067973, 2018 / 09 Includes nucleotide sequences, nucleotide modifications, sugar modifications, skeletal linkage patterns (internucleotide links) and / or skeletal chiral center patterns (e.g., of linked phosphorus atoms) as described in International Publication No. 8264, International Publication No. 2018 / 223056, International Publication No. 2018 / 223073, International Publication No. 2018 / 223081, International Publication No. 2018 / 237194, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185 or International Publication No. 2019 / 217784 (each of which is independently incorporated herein by reference).

[0209] In some embodiments, the provided technique includes labeling oligonucleotides using, for example, isotopes. In some embodiments, the provided oligonucleotides contain increased levels of one or more isotopes. In some embodiments, the provided oligonucleotides are labeled with, for example, one or more isotopes of one or more elements, such as hydrogen, carbon, nitrogen. In some embodiments, the provided oligonucleotides in a provided composition, for example, a first plurality of oligonucleotides, include one or more base modifications, sugar modifications and / or internucleotide bond modifications, where the oligonucleotides contain increased levels of isotopes. In some embodiments, the isotope is deuterium. In some embodiments, the hydrogen of a sugar is replaced with deuterium (for example, at the 2' position of 2'-deoxy). In some embodiments, the hydrogen of a base is replaced with deuterium. In some embodiments, the hydrogen of an internucleotide bond is replaced with deuterium. In some embodiments, the provided oligonucleotides are labeled with deuterium at one or more positions (- 1 H 2 Labeled (by replacing hydrogen with deuterium). In some embodiments, replacing hydrogen with deuterium can improve the stability, activity, bioavailability, ease of use, convenience, efficacy and / or systemic exposure of the oligonucleotide. In some embodiments, one or more of the oligonucleotide or any portion conjugated to the oligonucleotide (e.g., targeting portion, lipid, etc.) are labeled. 1 H 2 It is substituted with H. Such oligonucleotides can be used in any composition or method described herein. In some embodiments, the oligonucleotide targeting HTT comprises one or more isotopes. In some embodiments, the oligonucleotide targeting dystrophin comprises one or more isotopes.

[0210] Chiral enhancer In some embodiments, the techniques provided are particularly useful for preparing chiral-controlled oligonucleotide compositions with high crude purity and / or yield. In some embodiments, chiral-controlled (stereo-controlled / stereoselective) oligonucleotide synthesis typically utilizes chiral auxiliaries to control the stereochemistry of the bounding line chiral centers formed. In some embodiments, the disclosure provides compounds that can be used as chiral auxiliaries for oligonucleotide synthesis, such as compounds of formulas I, Ia, Ia-1, Ia-2, Ib, Ic, Id, Ie, II, II-a, II-b, III, III-a, or III-b, or salts thereof. In some embodiments, the chiral auxiliary group is as specified in U.S. Patent Nos. 9598458, 9744183, 9605019, 9394333, 9598458, 8859755, U.S. Patent Application Publication No. 20130178612, 8470987, 8822671, and U.S. Patent Application Publication No. 20150211. U.S. Patent Application Publication No. 006, U.S. Patent Application Publication No. 20170037399, U.S. Patent Application Publication No. 20180216107, U.S. Patent Application Publication No. 20180216108, U.S. Patent Application Publication No. 20190008986, International Publication No. 2017 / 015555, International Publication No. 2017 / 015575, International Publication No. 2017 / 062862, International Publication No. 2017 / 160741, International Publication No. 2 International Publication Nos. 017 / 192664, 2017 / 192679, 2017 / 210647, 2018 / 022473, 2018 / 067973, 2018 / 098264, 2018 / 223056, 2018 / 223073, 2018 / 223081, and 2018 / 23719 These are described in International Publication No. 4, International Publication No. 2019 / 032607, International Publication No. 2019 / 032612, International Publication No. 2019 / 032607, International Publication No. 2019 / 055951, International Publication No. 2019 / 075357, International Publication No. 2019 / 200185, or International Publication No. 2019 / 217784 (each of these chiral auxiliary groups is incorporated herein by reference).

[0211] In some embodiments, this disclosure relates to formula I: [ka] The present invention provides a compound or salt thereof having the structure, in which, L is replaced by C by covalent bonds or optional substitution. 1~6 It is an alkylene, where one or more methylene units are optionally and independently replaced with -L'-; or L is L s and; Each L' is independently substituted with a divalent C by covalent bond, of arbitrary choice. 1~3 Alkylene, -C(R 3 )(R 4 )-,-C(R 3 )(R 4 )-C(R 3 )(R 4 )-, -Cy- or -C(R 3 )[C(R 4 )3]- and; R 1 , R 2 , R 3 , R 4 and R 5 Each of them independently corresponds to -H and -L s -R', halogen, -CN, -NO2, -L s -Si(R')3, -OR', -SR', or -N(R')2; Each L s These are, independently, covalently bonded, or C 1~30 A carbon atom having an aliphatic group and 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. 1~30 A linear or branched group that is optionally substituted with a divalent heteroaliphatic group, wherein one or more methylene units are optionally and independently substituted with C 1~6 Alkylene, C 1~6Alkenylene, -C≡C-, divalent C1-C6 heteroaliphatic group having 1-5 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus and silicon, -C(R')2-, -Cy-, -O-, -S-, -SS-, -N(R')-, -C(O)-, -C(S)-, -C(NR')-, -C(O)N(R')-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -S(O)-, -S(O)2-, -S(O)2N(R')-, -C(O)S-, -C(O)O-, -P(O)(OR')-, -P(O)(SR')-, -P(O)(R')-, -P(O)(NR')-, -P(S)(O The carbon atoms are replaced by groups that are optionally substituted from R')-, -P(S)(SR')-, -P(S)(R')-, -P(S)(NR')-, -P(R')-, -P(OR')-, -P(SR')-, -P(NR')-, -P(OR')[B(R')3]-, -OP(O)(OR')O-, -OP(O)(SR')O-, -OP(O)(R')O-, -OP(O)(NR')O-, -OP(OR')O-, -OP(SR')O-, -OP(NR')O-, -OP(R')O- or -OP(OR')[B(R')3]O-, and one or more carbon atoms are optionally and independently substituted with Cy L Replaced with; Each -Cy- is independent of C 3~20 Alicyclic rings and C 6~20 A divalent group that is optionally substituted with an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each Cy L Independently, C 3~20 Alicyclic rings and C 6~20 A tetravalent group that is optionally substituted with an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each R' is independently -R, -C(O)R, -CO2R, or -SO2R; R 6 is -L-R'; R 7 is -OH or -SH; R 1 , R 2 , R 3 and R 4 At least one of them is not -H; Each R is independently either -H or C 1~30 Aliphatic C atoms and C atoms having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. 1~30 Heteroliphatic and C 6~30 Aryl and C 6~30 A C atom having aryl aliphatic elements and 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. 6~30 The group is optionally substituted from an aryl heteroaliphatic, a 5-30 membered ring heteroaryl having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-30 membered ring heterocyclyl having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, or The two R groups may, by choice and independently, come together to form a covalent bond, or Two or more R groups on the same atom may, by choice and independently, combine with that atom to form a optionally substituted monocyclic, bicyclic, or polycyclic ring of 3 to 30 members, having in addition to that atom 0 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; or Two or more R groups on two or more atoms, optionally and independently, together with their intercalating atoms, form optionally substituted monocyclic, bicyclic, or polycyclic rings of 3 to 30 members, each containing 0 to 10 heteroatoms, independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, in addition to the intercalating atoms.

[0212] In some embodiments, L is replaced by C by covalent bonding or optional means. 1~6It is an alkylene, where one or more methylene units are optionally and independently replaced by -L'-. In some embodiments, L is a covalent bond. In some embodiments, the provided compound is [ka] It has the structure or a salt thereof. In some embodiments, R 5 And, R 1 and R 2 One or both of these, together with their intervening atoms, form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 1 to 5 heteroatoms. In some embodiments, R 1 and R 2 One of them is R 5 Together with these intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 1 to 5 heteroatoms. As described extensively in this disclosure, the formed rings can be monocyclic, bicyclic, or polycyclic of various sizes and can contain various numbers of heteroatoms. In some embodiments, the ring is a 3-membered ring. In some embodiments, the ring is a 4-membered ring. In some embodiments, the ring is a 5-membered ring. In some embodiments, the ring is a 6-membered ring. In some embodiments, the ring is a monocyclic ring. In some embodiments, the ring contains additional ring heteroatoms other than the intervening heteroatoms. In some embodiments, the ring is a 3-membered ring containing one ring heteroatom. In some embodiments, the ring is a 3-membered ring containing two ring heteroatoms. In some embodiments, the ring is a 3-membered ring containing one carbon, one nitrogen, and one oxygen ring atom.

[0213] In some embodiments, L is -C(R 3 )(R 4 )-. In some embodiments, the compound provided is of formula Ia: [ka] The compound has the structure of or a salt thereof, where each variable element is independently as described herein. In some embodiments, the compound of formula I has the structure of formula Ia. In some embodiments, the compound provided is [ka] Having the structure or a salt thereof, where each variable element is independently as described herein, where R 4 and R 5 It is not hydrogen.

[0214] In some embodiments, the provided compound is of formula (Ia-1): [ka] Having the structure or a salt thereof, where each variable element is independently as described herein, where R 4 and R 5 It is not hydrogen, and R 2 R 1 It has a larger size than [formula]. In some embodiments, the compound of formula Ia has the structure of formula Ia-1.

[0215] In some embodiments, the provided compound is of formula (Ia-2): [ka] Having the structure or a salt thereof, where each variable element is independently as described herein, where R 4 and R 5 It is not hydrogen, and R 2 R 1 It has a larger size than [formula]. In some embodiments, the compound of formula Ia has the structure of formula Ia-2.

[0216] In some embodiments, R 6 is -H. In some embodiments, R 6 is -H, and R4 and R 5 These intercalating atoms, together with the intercalating atoms, form an optionally substituted 3-20 membered heterocyclyl ring having 1-5 heteroatoms. In some embodiments, R 4 and R 5 These intervening atoms, together with the intervening atoms, form a 4- to 10-membered heterocyclyl ring that is optionally substituted, including the intervening nitrogen atom as the sole ring heteroatom. In some embodiments, the formed ring is a 3-membered ring. In some embodiments, the formed ring is a 4-membered ring. In some embodiments, the formed ring is a 5-membered ring. In some embodiments, the formed ring is a 6-membered ring. In some embodiments, the formed ring is a 7-membered ring. In some embodiments, the formed ring is substituted. In some embodiments, the formed ring is unsubstituted. In some embodiments, the formed ring is a monoring. In some embodiments, the formed ring is a diring. In some embodiments, the formed ring is polyring. In some embodiments, the formed ring is saturated. In some embodiments, the formed ring is R 5 It does not have any ring heteroatoms other than the nitrogen atom to which it is bonded.

[0217] In some embodiments, R 1 and R 2 At least one of them is not hydrogen. In some embodiments, R 1 is hydrogen, and R 2 is not hydrogen. In some embodiments, R 1 It is not hydrogen, and R 2 is hydrogen. In some embodiments, R 1 and R 2 None of them are hydrogen.

[0218] In some embodiments, R 1 and R 2 One of the elements is -H, and the other is R, where R is not hydrogen. In some embodiments, R 1 and R 2One of them is -H, and the other is C which is substituted by choice. 1~6 It is aliphatic. In some embodiments, R 1 and R 2 One of them is -H, and the other is C which is substituted by choice. 1~4 It is aliphatic. In some embodiments, R 1 and R 2 One of them is -H, and the other is C which is substituted by choice. 1~3 It is aliphatic. In some embodiments, R 1 and R 2 One of them is -H, and the other is C which is substituted by choice. 1~2 It is aliphatic. In some embodiments, R 1 and R 2 One of them is -H, and the other is C which is substituted by choice. 1~6 It is an alkenyl. In some embodiments, R 1 and R 2 One of them is -H, and the other is vinyl. In some embodiments, R 1 and R 2 One of them is -H, and the other is C which is substituted by choice. 1~6 It is an alkynyl. In some embodiments, R 1 and R 2 One of them is -H, and the other is ethynyl. In some embodiments, R 1 and R 2 One of them is -H, and the other is benzyl which is optionally substituted. In some embodiments, R 1 and R 2 One of them is -H, and the other is benzyl, where the phenyl group of benzyl is optionally substituted. In some embodiments, R 1 is -H, and R 2 is benzyl. In some embodiments, R 1 is -H, and R 2 is -R, where R is as described in this disclosure and is not hydrogen. In some embodiments, R 2 C is replaced by an optional substitution. 1~6It is aliphatic. In some embodiments, R 2 R is optionally substituted with -CH2-CPh2Me. In some embodiments, R 2 is -CH2-CPh2Me. In some embodiments, R 2 R is a phenyl compound that is optionally substituted. In some embodiments, R 2 is benzyl which is optionally substituted. In some embodiments, the provided compound is [ka] Or its salt.

[0219] In some embodiments, R 1 is not -H and R 2 is not -H. In some embodiments, R 1 and R 2 R is independent of R, where R is not -H. In some embodiments, R 1 C is replaced by an optional substitution. 1~6 It is aliphatic, R 2 R is a phenyl compound that is optionally substituted. In some embodiments, R 1 It is methyl, and R 2 It is phenyl.

[0220] In some embodiments, R 1 and R 2 One of the elements is R, where R includes a ring portion. In some embodiments, R is C 3~20 alicyclic, C 6~20 The group is optionally substituted from aryl, 5-20 membered heteroaryl rings having 1-5 heteroatoms, and 3-20 membered heterocyclyl rings having 1-5 heteroatoms. In some embodiments, R is optionally substituted with C 3~20 It is an alicyclic group. In some embodiments, R is optionally substituted with C. 3~10 It is an alicyclic group. In some embodiments, R is optionally substituted with C.3~10 It is a cycloalkyl group. In some embodiments, R is optionally substituted with C 4~10 It is a cycloalkyl group. In some embodiments, R is optionally substituted with cyclopropyl. In some embodiments, R is optionally substituted with cyclobutyl. In some embodiments, R is optionally substituted with cyclopentyl. In some embodiments, R is optionally substituted with cyclohexyl. In some embodiments, R is optionally substituted with cycloheptyl. In some embodiments, R is cyclopropyl. In some embodiments, R is cyclobutyl. In some embodiments, R is cyclopentyl. In some embodiments, R is cyclohexyl. In some embodiments, R is cycloheptyl. In some embodiments, R is optionally substituted with C 6~20 It is an aryl. In some embodiments, R is optionally substituted with phenyl. In some embodiments, R is a phenyl. In some embodiments, R is an optionally substituted 5-20 membered ring heteroaryl having 1-5 heteroatoms. In some embodiments, R is an optionally substituted 5-membered ring heteroaryl having 1-5 heteroatoms. In some embodiments, R is an optionally substituted 6-membered ring heteroaryl having 1-5 heteroatoms. In some embodiments, R is an optionally substituted 3-20 membered ring heterocyclyl having 1-5 heteroatoms. In some embodiments, R 1 and R 2 The other is R, where R is not hydrogen. In some embodiments, R is optionally substituted with C. 1~6 It is aliphatic. In some embodiments, R is optionally replaced by C 1~6 It is alkyl. In some embodiments, R is C 1~6It is alkyl. In some embodiments, R is methyl. In some embodiments, R is substituted methyl. In some embodiments, R is ethyl. In some embodiments, R is substituted ethyl. In some embodiments, R 1 and R 2 One of them is R, which includes a cyclic portion as described in this disclosure, and the other is an alkyl group as described in this disclosure.

[0221] In some embodiments, R 1 and R 2 Each of these is independently R, where R is optionally substituted with C. 1~20 It is aliphatic. In some embodiments, R is an unsubstituted C 1~20 It is aliphatic. In some embodiments, R is optionally replaced by C 1~20 It is alkyl. In some embodiments, R is optionally substituted with C 1~6 It is alkyl. In some embodiments, R is linear C 1~6 It is alkyl. In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~6 It is an alkyl group, and the other is a C group that is optionally substituted. 1~6 It is alkyl. In some embodiments, R 1 and R 2 This is the same as. In some embodiments, R 1 and R 2 That is different.

[0222] In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~6 It is an alkyl group, and the other is a C group that is optionally substituted. 1~6 It is an alkenyl. In some embodiments, R 1 and R 2 One of them is optionally substituted with methyl or ethyl, and the other is vinyl. In some embodiments, R 1 and R2 One of them is methyl, and the other is vinyl.

[0223] In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~6 It is an alkyl group, and the other is a C group that is optionally substituted. 1~6 It is an alkynyl. In some embodiments, R 1 and R 2 One of them is optionally substituted with methyl or ethyl, and the other is ethynyl. In some embodiments, R 1 and R 2 One of them is methyl, and the other is ethynyl.

[0224] In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~6 It is an alkyl group, and the other is a C group that is optionally substituted. 1~6 It is alkyl. In some embodiments, R 1 and R 2 C is replaced by the same arbitrary choice. 1~6 It is alkyl. In some embodiments, R 1 and R 2 C is replaced by the same arbitrary choice. 1~2 It is alkyl, R 1 and R 2 It contains two or fewer carbon atoms. In some embodiments, R 1 and R 1 Both are methyl. In some embodiments, R 1 and R 1 Both are ethyl. In some embodiments, R 1 and R 1 Both are isopropyl. In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~3 It is a linear alkyl group, and the other side is optionally substituted with C 3~10It is cycloalkyl. In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~3 It is a linear alkyl group, and the other side is optionally substituted with C 5~6 It is cycloalkyl. In some embodiments, R 1 is methyl. In some embodiments, R 2 is cyclopentyl. In some embodiments, R 2 is cyclohexyl. In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~3 One is a linear alkyl group, and the other is a benzyl group that is optionally substituted. In some embodiments, R 1 It is methyl, and R 2 is benzyl which is optionally substituted. In some embodiments, R 2 is benzyl. In some embodiments, R 2 It is p-CH3O-C6H4-CH2-. In some embodiments, R 1 R is selected from methyl, ethyl, cyclohexyl, and optionally phenyl-substituted benzyl. In some embodiments, 2 R is selected from methyl, ethyl, cyclohexyl, and optionally phenyl-substituted benzyl. In some embodiments, 1 and R 2 Each of these is independently selected from methyl, ethyl, cyclohexyl, and optionally phenyl-substituted benzyl. In some embodiments, the provided compounds are [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] Or its salt.

[0225] In some embodiments, R 1 and R 2 One of them is C which is replaced by arbitrary choice. 1~6 One is alkyl, and the other is optionally substituted with phenyl. In some embodiments, R 1 It is methyl, and R 2 R is a phenyl compound that is optionally substituted. In some embodiments, R 1 It is methyl, and R 2 R is phenyl. In some embodiments, R 1 It is methyl, and R 2 teeth, [ka] In some embodiments, the provided compound is [ka] or selected from salts thereof. In some embodiments, the provided compound is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] Or its salt.

[0226] In some embodiments, R 1 and R 2 R is independently R, where R is an optionally substituted aryl group. In some embodiments, R 1 and R 2 R is independently and optionally substituted phenyl. In some embodiments, R 1 and R 2 is phenyl. In some embodiments, the provided compound is [ka] Or its salt.

[0227] In some embodiments, R 1 and R 2 These, together with the carbon atoms to which they are bonded, form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 0 to 5 heteroatoms. In some embodiments, R 1 and R 2These, together with the carbon atoms to which they are bonded, form a monocyclic ring of 3 to 7 members that is optionally substituted and does not contain heteroatoms. In some embodiments, the monocyclic ring formed is a 3-membered ring; in some embodiments, a 4-membered ring; in some embodiments, a 5-membered ring; in some embodiments, a 6-membered ring; in some embodiments, a 7-membered ring; in some embodiments, an 8-membered ring; in some embodiments, a 9-membered ring; and in some embodiments, a 10-membered ring. In some embodiments, the ring formed is monocyclic. In some embodiments, the ring formed is dicyclic. In some embodiments, the ring formed is polycyclic. In some embodiments, the ring formed is aliphatic. In some embodiments, the ring formed does not contain unsaturation. In some embodiments, the ring formed is saturated, partially unsaturated and / or partially aromatic, for example, a bicyclic or polycyclic ring containing a condensed saturated, partially unsaturated and / or aromatic moiety. In some embodiments, the ring formed is substituted. In some embodiments, the ring formed is unsubstituted. In some embodiments, R 1 and R 2 The carbon atom to which it is bonded is not chiral. In some embodiments, R 1 and R 2 These are the same, and the carbon atoms to which they are bonded are not chiral. In some embodiments, R 1 and R 2 The ring formed by them together with the carbon atoms to which they are bonded does not introduce chirality, R 1 and R 2 The carbon atom to which it is bonded is not chiral. In some embodiments, R 1 and R 2 Unlike the carbon atoms to which they are bonded, the carbon atoms to which they are bonded are chiral. In some embodiments, R 1 and R 2 The ring formed by them together with the carbon atoms to which they are bonded introduces chirality, R 1 and R 2 The carbon atom to which it is bonded is not chiral. In some embodiments, the provided compound is [ka] and selected from salts thereof. In some embodiments, the provided compound is [ka] and selected from salts thereof. In some embodiments, the provided compound is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] Or its salt.

[0228] In some embodiments, R 4 and R 5 These interposing atoms, together with the interposing atoms, form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 1 to 5 heteroatoms. In some embodiments, the formed ring is a 3-membered ring. In some embodiments, the formed ring is a 4-membered ring. In some embodiments, the formed ring is a 5-membered ring. In some embodiments, the formed ring is a 6-membered ring. In some embodiments, the formed ring is a 7-membered ring. In some embodiments, the formed ring is an 8-membered ring. In some embodiments, the formed ring is a 9-membered ring. In some embodiments, the formed ring is a 10-membered ring. In some embodiments, R3 is -H, and R 4 and R 5 These intervening atoms, together with the intervening atoms, form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 1 to 5 heteroatoms. In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 Forms a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members that is optionally substituted with (those with ). 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 It forms a monocyclic ring of 4 to 7 members that is optionally substituted with (those with ). In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 Forms a monocyclic ring of a four-membered ring that is optionally substituted with (those with ). In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 It forms a monocyclic ring of a five-membered ring that is optionally substituted with (those with ). In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 Forms a monocyclic ring of a 6-membered ring that is optionally substituted with (those with ). In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 It forms a monocyclic ring of a 7-membered ring that is optionally substituted with (those with R). In some embodiments,3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 Forms an optionally substituted monocyclic ring of an 8-membered ring having (with) R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 It forms a monocyclic ring of a 9-membered ring that is optionally substituted with (those with ). In some embodiments, R 3 is -H, and R 4 and R 5 Together with those intervening atoms, it forms a nitrogen atom (R 5 Forms a monocyclic ring of a 10-membered ring that is optionally substituted with (those with ). In some embodiments, R 4 and R 5 The ring that is formed by these intervening atoms is substituted. In some embodiments, R 4 and R 5 The rings that are formed by these intervening atoms are not substituted. In some embodiments, R 4 and R 5 The ring formed by these intervening atoms together is a monoring. In some embodiments, R 4 and R 5 The ring formed by these intervening atoms together is a diring ring. In some embodiments, R 1 and R 2 One of the two, and R 3 and R 4One of these atoms, together with the intervening atoms, forms a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 0 to 5 heteroatoms. In some embodiments, the formed ring is a 3-membered ring. In some embodiments, the formed ring is a 4-membered ring. In some embodiments, the formed ring is a 5-membered ring. In some embodiments, the formed ring is a 6-membered ring. In some embodiments, the formed ring is a 7-membered ring. In some embodiments, the formed ring is substituted. In some embodiments, the formed ring is not substituted. In some embodiments, the formed ring is a monocyclic ring. In some embodiments, the formed ring is a bicyclic ring. In some embodiments, the formed ring is polycyclic. In some embodiments, the formed ring has no additional heteroatoms in addition to the intervening atoms. In some embodiments, the formed ring has additional ring heteroatoms in addition to the intervening atoms. Exemplary rings formed are described extensively in this disclosure. In some embodiments, the provided compound is [ka] and selected from salts thereof. In some embodiments, the provided compound is [ka] and selected from salts thereof. In some embodiments, the provided compound is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] or a salt thereof. In some embodiments, the compound provided is [ka] Or its salt.

[0229] In some embodiments, R 1 and R 2 One or two of them are R 3 , R 4 and R 5Together with one or more of the intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 0 to 5 heteroatoms. In some embodiments, R 1 and R 2 One or two of them are R 3 and R 4 Together with one or two of the intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 0 to 5 heteroatoms. In some embodiments, R 1 and R 2 One or two of them are R 5 Together with intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members having 1 to 5 heteroatoms and optionally substituted. In some embodiments, R 1 and R 2 One or two of them are R 5 , R 3 and R 4 Together with one or two of the intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members, optionally substituted with 1 to 5 heteroatoms. In some embodiments, R 1 and R 2 One or two of them are R 5 , R 3 and R 4 Together with one or two of the intervening atoms, they form a bicyclic or polycyclic ring of 6 to 20 members, optionally substituted with 1 to 5 heteroatoms. In some embodiments, R 1 and R 2 One or two of them are R 5 , R 3 and R 4 Together with one or two of the intervening atoms, they form an optionally substituted 8-20 membered bicyclic or polycyclic ring having 1-5 heteroatoms. In some embodiments, R 1 and R 2 One of them is R 5 , R 3 and R 4Together with one of the intervening atoms, it forms an optionally substituted 8-20 membered bicyclic or polycyclic ring having 1-5 heteroatoms. In some embodiments, R 1 and R 2 One of them is R 5 , R 3 and R 4 Together with one of the intervening atoms, it forms an optionally substituted, 8- to 20-membered bicyclic ring having 1 to 5 heteroatoms. In some embodiments, the formed ring is an 8-membered ring. In some embodiments, the formed ring is a 9-membered ring.

[0230] In some embodiments, R 5 R 1 and R 2 Together with one of them and their intervening atoms, they form a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members having 1 to 5 heteroatoms and being optionally substituted. In some embodiments, R 5 R 3 and R 4 Together with one of them and their i...

Claims

1. The following steps: (1) Coupling step; (2) Pre-modification capping step; (3) Modification step; (4) Post-modification capping step; and (5) Deblockization step A method for preparing oligonucleotides comprising two or more cycles, each independently containing, In at least one cycle, the coupling step independently includes reacting a free hydroxyl group of an oligonucleotide or nucleoside with a coupling partner compound containing a chiral auxiliary group, The modification step includes sulfidation, which includes converting a -P(-)- bonded phosphorus atom to a -P(=S)(-)- bonded phosphorus atom. The modification step involves modifying the -P(-)- bonded phosphorus atom. ＊Ｎ S and ＊Ｎ This includes converting to a -P(=N-)(-)- bonded phosphorus atom, as in R, Each partner compound containing chiral auxiliary groups in two or more rings can be independently, 【Chemistry 1】 Having the structure or a salt thereof, ＊Ｎ S is independent of the formula 【Chemistry 2】 It is a substance or a salt form thereof, ＊Ｎ R is independent of the formula 【Transformation 3】 It is a substance or a salt form thereof, (In the formula, BA is a group that is optionally substituted, selected from the native nucleic acid base moiety and the modified nucleic acid base moiety; R ２ｓ and R ４ｓ each independently is -H, halogen, -CN, -N ３ , -NO, -NO ２ , -L ｓ -R', -L ｓ -Si(R) ３ , -L ｓ -OR', -L ｓ -SR', -L ｓ -N(R')[[]] ２ , -O-L ｓ -R', -O-L ｓ -Si(R) ３ , -O-L ｓ -OR', -O-L ｓ -SR' or -O-L ｓ -N(R')[[]] ２ ; R ２ is, -CH ２ SO ２ R' is C １～３０ Aliphatic carbon atoms containing 1 to 10 heteroatoms １～３０ Heteroliphatic, C ６～３０ Ariel, C ６～３０ Aryl aliphatic carbon atoms containing 1 to 10 heteroatoms ６～３０ The group is optionally substituted, selected from aryl heteroaliphatic groups, 5-30 membered heteroaryl groups having 1-10 heteroatoms, and 3-30 membered heterocyclines having 1-10 heteroatoms. P = W Ｎ P Ｎ And, -X-L ｓ -R ５ teeth, 【Chemistry 4】 And, P Ｎ P(=N-L-R) ５ ), 【Transformation 5】 And; Q － is an anion; Each R ｓ These are independently -H, halogen, -CN, and -N ３ , -NO, -NO ２ , -L ｓ -R', -L ｓ -Si(R') ３ , -L ｓ -OR', -L ｓ -SR', -L ｓ -N(R') ２ , -O-L ｓ -R', -O-L ｓ -Si(R) ３ , -O-L ｓ -OR', -OL-L ｓ -SR' or -OL-L ｓ -N(R') ２ And; g is between 0 and 20; Ring A Ｌ This is a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members that is optionally substituted with 0 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; P Ｎ In R １ and R ５ Each of them independently corresponds to -H and -L ｓ -R', halogen, -CN, -NO ２ , -L ｓ -Si(R') ３ , -OR', -SR', or -N(R') ２ And; R ６ is -C(O)R', where R' is independently -R, -C(O)R, -C(O)OR, or -S(O) ２ It is R, L and L ｂ Each of them independently, L ｓ And; Each L ｓ These are independently, covalently, or C １～３０ A carbon atom having an aliphatic group and 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. １～３０ A linear or branched group substituted by optional divalent selection from heteroaliphatic groups, wherein one or more methylene units are optionally and independently C １～６ Alkylene, C １～６ Alkenylene, -C≡C-, divalent C １ ~C ６ A heteroaliphatic group having 1 to 5 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a divalent carbon １ ~C ６ Heteroaliphatic group, -C(R') ２ -, -Cy-, -O-, -S-, -S-S-, -N(R')-, -C(O)-, -C(S)-, -C(NR')-, -C(O)N(R')-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -S(O)-, -S(O) ２ -, -S(O) ２ N(R')-, -C(O)S-, -C(O)O-, -P(O)(OR')-, -P(O)(SR')-, -P(O)(R')-, -P(O)(NR')-, -P(S)(OR')-, -P(S)(SR')-, -P(S)(R')-, -P(S)(NR')-, -P(R')-, -P(OR')-, -P(SR')-, -P(NR')-, -P(OR') [B(R') ３ ]-, -OP(O)(OR')O-, -OP(O)(SR')O-, -OP(O)(R')O-, -OP(O)(NR')O-, -OP(OR')O-, -OP(SR')O-, -OP(NR')O-, -OP(R')O- or -OP(OR') [B(R') ３ Replaced by a group selected from ]O-, which is optionally substituted, and one or more carbon atoms are optionally and independently substituted with Cy Ｌ Replaced by; Each -Cy- is independent of C ３～２０ Alicyclic rings and C ６～２０ A divalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each Cy Ｌ Independently, C ３～２０ Alicyclic rings and C ６～２０ A tetravalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each R' is independently -R, -C(O)R, -C(O)OR, or -S(O) ２ It is R; Each R is independently -H or C １～３０ aliphatic, C having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus and silicon １～３０ heteroaliphatic, C ６～３０ aryl, C ６～３０ aryl aliphatic, C having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus and silicon ６～３０ aryl heteroaliphatic, 5- to 30-membered heteroaryl having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus and silicon, 3- to 30-membered heterocyclyl having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus and silicon, which is an optionally substituted group, or The two R groups may, by choice and independently, come together to form a covalent bond, or; Two or more R groups on the same atom may, by choice and independently, combine with the atom to form a optionally substituted 3-30 membered monocyclic, bicyclic, or polycyclic ring having, in addition to the atom, 0-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; or Two or more R groups on two or more atoms can be optionally and independently combined with the intervening atoms to form an optionally substituted 3-30 member monocyclic, bicyclic, or polycyclic ring, which has 0-10 heteroatoms, independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, in addition to the intervening atoms. ,method.

2. In the partner compound, R ４ｓ The method according to claim 1, wherein is -H.

3. In the partner compound, R ２ｓ The method according to claim 2, wherein is -H.

4. In the partner compound, R ２ｓ is -OR, wherein R is optionally substituted C １～６ The method according to claim 2, which is aliphatic.

5. In the partner compound, R ２ｓ is -O-Si(R) ３ The method according to claim 2, wherein each R is independently not -H.

6. R ２ is, -CH ２ SO ２ The method according to any one of claims 1 to 5, wherein R' is a phenyl compound optionally substituted.

7. R ２ is, -CH ２ SO ２ The method according to any one of claims 1 to 5, wherein R' is phenyl.

8. R ２ is, -CH ２ SO ２ The method according to any one of claims 1 to 5, wherein R' is tert-butyl.

9. The method according to any one of claims 1 to 8, comprising removing the chiral auxiliary by contacting the oligonucleotide containing the chiral auxiliary with a base under anhydrous conditions.

10. The method according to claim 9, wherein the base is diethylamine.

11. The method according to any one of claims 1 to 10, wherein the product oligonucleotide comprises a sugar containing a 2'-OH group.

12. The method according to claim 11, wherein the product oligonucleotide further comprises a non-negatively charged internucleotide bond.

13. The method according to claim 12, wherein the non-negatively charged internucleotide bond includes a guanidine moiety.

14. The method according to any one of claims 1 to 13, wherein the product oligonucleotide comprises a modified internucleotide bond containing a cyclic guanidine moiety that is optionally substituted.

15. The aforementioned non-negatively charged internucleotide bond is 【Transformation 6】 The method according to claim 12.

16. The method according to any one of claims 1 to 15, wherein the product oligonucleotide further comprises a native phosphate bond.

17. P Ｎ teeth, 【Transformation 7】 The method according to claim 9.

18. R ２ is, -CH ２ SO ２ R' is phenyl, and Q － PF ６ － The method according to claim 17.

19. The process includes the step of contacting an oligonucleotide with a base under anhydrous conditions to remove the chiral auxiliary group, The oligonucleotide is ＊Ｐ S or ＊Ｐ Includes R internucleotide linkage, ＊Ｐ S is independent of the formula 【Transformation 8】 It is a substance or a salt form thereof, ＊Ｐ R is independent of the formula 【Chemistry 9】 It is a substance or a salt form thereof, ＊Ｎ S or ＊Ｎ Further containing R internucleotide bonds, ＊Ｎ S is independent of the formula 【Chemistry 10】 It is either a substance or a salt thereof, in the formula P = W Ｎ P Ｎ And, ＊Ｎ R is independent of the formula 【Chemistry 11】 It is either a substance or a salt thereof, in the formula P = W Ｎ P Ｎ And, In the formula, -XL ｓ -R ５ teeth, 【Chemistry 12】 And, (In the formula, R ６ is -C(O)R', where R' is independently -R, -C(O)R, -C(O)OR, or -S(O) ２ It is R, R ２ is, -CH ２ SO ２ R' is C １～３０ Aliphatic carbon atoms containing 1 to 10 heteroatoms １～３０ Heteroliphatic, C ６～３０ Ariel, C ６～３０ Aryl aliphatic carbon atoms containing 1 to 10 heteroatoms ６～３０ The group is optionally substituted, selected from aryl heteroaliphatic groups, 5-30 membered heteroaryl groups having 1-10 heteroatoms, and 3-30 membered heterocyclines having 1-10 heteroatoms. P Ｎ P(=N-L-R) ５ ), 【Chemistry 13】 And; Q － is an anion; Each R ｓ These are independently -H, halogen, -CN, and -N ３ , -NO, -NO ２ , -L ｓ -R', -L ｓ -Si(R') ３ , -L ｓ -OR', -L ｓ -SR', -L ｓ -N(R') ２ , -O-L ｓ -R', -O-L ｓ -Si(R) ３ , -O-L ｓ -OR', -OL-L ｓ -SR' or -OL-L ｓ -N(R') ２ And; g is between 0 and 20; Ring A Ｌ This is a monocyclic, bicyclic, or polycyclic ring of 3 to 20 members that is optionally substituted with 0 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; P Ｎ In R １ and R ５ Each of them independently corresponds to -H and -L ｓ -R', halogen, -CN, -NO ２ , -L ｓ -Si(R') ３ , -OR', -SR', or -N(R') ２ And; L and L ｂ Each of them independently, L ｓ And; Each L ｓ These are independently, covalently, or C １～３０ A carbon atom having an aliphatic group and 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. １～３０ A linear or branched group substituted by optional divalent selection from heteroaliphatic groups, wherein one or more methylene units are optionally and independently C １～６ Alkylene, C １～６ Alkenylene, -C≡C-, divalent C １ ~C ６ A heteroaliphatic group having 1 to 5 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a divalent carbon １ ~C ６ Heteroaliphatic group, -C(R') ２ -, -Cy-, -O-, -S-, -S-S-, -N(R')-, -C(O)-, -C(S)-, -C(NR')-, -C(O)N(R')-, -N(R')C(O)N(R')-, -N(R')C(O)O-, -S(O)-, -S(O) ２ -, -S(O) ２ N(R')-, -C(O)S-, -C(O)O-, -P(O)(OR')-, -P(O)(SR')-, -P(O)(R')-, -P(O)(NR')-, -P(S)(OR')-, -P(S)(SR')-, -P(S)(R')-, -P(S)(NR')-, -P(R')-, -P(OR')-, -P(SR')-, -P(NR')-, -P(OR') [B(R') ３ ]-, -OP(O)(OR')O-, -OP(O)(SR')O-, -OP(O)(R')O-, -OP(O)(NR')O-, -OP(OR')O-, -OP(SR')O-, -OP(NR')O-, -OP(R')O- or -OP(OR') [B(R') ３ Replaced by a group selected from ]O-, which is optionally substituted, and one or more carbon atoms are optionally and independently substituted with Cy Ｌ Replaced by; Each -Cy- is independent of C ３～２０ Alicyclic rings and C ６～２０ A divalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each Cy Ｌ Independently, C ３～２０ Alicyclic rings and C ６～２０ A tetravalent group that is optionally substituted, selected from an aryl ring, a 5-20 membered heteroaryl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-20 membered heterocyclyl ring having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon; Each R' is independently -R, -C(O)R, -C(O)OR, or -S(O) unless otherwise specified. ２ It is R; Each R is independently -H or C １～３０ Aliphatic C12C having 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. １～３０ Heteroliphatic and C ６～３０ Aryl and C ６～３０ A carbon atom having an aryl aliphatic and 1 to 10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon. ６～３０ The group is optionally substituted, selected from an aryl heteroaliphatic, a 5-30 membered heteroaryl having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, and a 3-30 membered heterocyclyl having 1-10 heteroatoms independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, or Two R groups, by arbitrary and independent choice, come together to form a covalent bond, or; Two or more R groups on the same atom, by arbitrary and independent choice, together with that atom to form an optionally substituted 3-30 member monocyclic, bicyclic, or polycyclic ring having 0-10 heteroatoms, independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, in addition to that atom; or (Two or more R groups on two or more atoms, by arbitrary and independent choice, together with their intervening atoms, form an optionally substituted 3-30 member monocyclic, bicyclic, or polycyclic ring, which has 0-10 heteroatoms, independently selected from oxygen, nitrogen, sulfur, phosphorus, and silicon, in addition to the intervening atoms.) A method for producing oligonucleotides.

20. The method according to claim 19, wherein the base is diethylamine.

21. R ２ is, -CH ２ SO ２ The method according to claim 19 or 20, wherein R' is a phenyl compound that is optionally substituted.

22. R ２ is, -CH ２ SO ２ The method according to claim 19 or 20, wherein R' is phenyl.

23. R ２ is, -CH ２ SO ２ The method according to claim 19 or 20, wherein R' is tert-butyl.

24. P Ｎ teeth, 【Chemistry 14】 The method according to any one of claims 19 to 23.

25. R ２ is, -CH ２ SO ２ R' is phenyl, and Q － PF ６ － The method according to claim 24.

Images