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CN-121991153-A - Adjuvant molecule and preparation method and application thereof

CN121991153ACN 121991153 ACN121991153 ACN 121991153ACN-121991153-A

Abstract

The invention provides an adjuvant molecule, a preparation method and application, wherein the adjuvant molecule has a structure shown in a formula 1, has a lymph node targeting function, effectively starts immune response and has wide application prospect.

Inventors

  • LIU JING
  • ZHANG XIAOYU
  • CHEN CHUNYING
  • ZHAO YULIANG

Assignees

  • 国家纳米科学中心

Dates

Publication Date
20260508
Application Date
20241104

Claims (10)

  1. 1. An adjuvanted molecule, wherein the adjuvanted molecule has a structure according to formula 1: Wherein R 1 is a nitrogen-containing six-membered heterocyclic ring or C1-C3 straight-chain alkane, R 2 is a C8-C22 unsaturated chain alkyl, a C8-C22 alkyl, cholesterol hemisuccinate or 1, 2-dioleoyl-SN-glycero-3-phosphorylethanolamine, and R 3 is nitro or amino.
  2. 2. An adjuvanted molecule according to claim 1 wherein R 1 is a nitrogen-containing hexacyclic ring or methylene; preferably, the adjuvanted molecule has any one of the structures shown in the following formulas 2-1 to 2-4: Wherein R 2 is cholesterol hemisuccinate or 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine.
  3. 3. An adjuvanted molecule according to claim 1 or 2 wherein R 1 is a nitrogen-containing six membered heterocyclic ring; preferably, the nitrogen-containing six-membered heterocyclic ring is Preferably, R 3 is amino.
  4. 4. An adjuvanted molecule according to any one of claims 1 to 3 wherein the adjuvanted molecule is any one of the following compounds:
  5. 5. An adjuvanted molecule according to any one of claims 1 to 4 wherein the adjuvanted molecule is a compound of:
  6. 6. An adjuvanted molecule according to any one of claims 1 to 5 wherein the adjuvanted molecule is any one of the following compounds:
  7. 7. A method of preparing an adjuvanted molecule according to any one of claims 1 to 6 comprising: (1) Reacting a compound of formula 2 with N-hydroxysuccinimide to obtain an acyloxy intermediate shown in formula 3; (2) Reacting the acyl oxide intermediate shown in the formula 3 obtained in the step (1) with a nitroimidazole compound containing primary amine shown in the formula 4 to obtain the adjuvant molecule with R3 as nitro shown in the formula 5, wherein the reaction formula is as follows: (3) Reacting an adjuvanted molecule of formula 5 wherein R3 is nitro with a reducing agent to obtain the adjuvanted molecule of formula 6 wherein R3 is amino, the reaction formula is as follows: Wherein R 1 and R 2 are as defined in formula 1.
  8. 8. The preparation method according to claim 7, wherein the compound of formula 2 comprises 1, 2-dioleoyl-SN-glycero-3-phosphorylethanolamine having carboxyl group, which is prepared by reacting 1, 2-dioleoyl-SN-glycero-3-phosphorylethanolamine with succinic anhydride; Preferably, the reaction of 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine and succinic anhydride is carried out in an organic solvent, preferably dichloromethane; Preferably, the molar ratio of the 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine to the succinic anhydride is 1:1-1:1.5; preferably, the reaction of 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine and succinic anhydride is carried out in the presence of a catalyst; Preferably, the catalyst is selected from the group consisting of N, N-diisopropylethylamine; Preferably, the mol ratio of the catalyst to the 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine is (0.5-4): 1; Preferably, the reaction temperature of the 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine (DOPE) and succinic anhydride is 0-37 ℃ and the reaction time is 5-48 hours.
  9. 9. The method according to claim 7 or 8, wherein the molar ratio of the compound of formula 2 to N-hydroxysuccinimide in step (1) is (0.5-2): 1; Preferably, the reaction of step (1) is carried out in the presence of a condensing agent; Preferably, the condensing agent is a carbodiimide condensing agent; Preferably, the carbodiimide condensing agent is selected from 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide; preferably, the molar ratio of the compound of formula 2 to the condensing agent in step (1) is 1:1.2-2; preferably, the reaction of step (1) is carried out in the presence of a solvent; Preferably, the solvent comprises dichloromethane Preferably, the temperature of the reaction in the step (1) is 0-25 ℃, and the reaction time is 1-3 hours; preferably, after the reaction structure in step (1), a post-treatment is performed, wherein the post-treatment comprises extraction and drying; preferably, the extracted solvent comprises dichloromethane and water; preferably, the volume ratio of dichloromethane to water in the extracted solvent is 3-5:1; preferably, the number of extractions is 3-5; preferably, the reaction of step (2) is carried out in the presence of a solvent; Preferably, the mixed solvent comprises dichloromethane and methanol; Preferably, the volume ratio of dichloromethane to methanol in the mixed solvent is 5-15:1; preferably, the molar ratio of the acyl oxide intermediate shown in the formula 3 in the step (2) to the nitroimidazole compound containing the primary amine shown in the formula 4 is preferably 1:1-2; preferably, the reaction in step (2) is carried out in the presence of an acid-binding agent; Preferably, the acid binding agent is triethylamine; preferably, the mol ratio of the acid binding agent to the nitroimidazole compound containing primary amine is (0.5-3): 1; Preferably, the reaction time in the step (2) is 12-24 hours; Preferably, after the reaction in the step (2) is finished, the purification treatment is carried out, wherein the purification method comprises the steps of drying a final reaction solution, adding methanol, collecting supernatant, vacuum drying a crude product, dissolving the crude product in a hydrochloric acid aqueous solution, collecting precipitated sediment, washing the pH value back to neutrality by using saturated saline water, and freeze-drying the product; preferably, the concentration of the aqueous hydrochloric acid solution is preferably 1moL/mL; preferably, the purification times with aqueous hydrochloric acid is 3-5 times; preferably, the reducing agent of step (3) is selected from sodium hydrosulfite, a combination of hydrazine hydrate and raney nickel, a combination of iron powder and ammonium chloride or a reducing metal chloride; preferably, the reaction of step (3) is carried out in a solvent comprising methanol and water; preferably, the volume ratio of the methanol to the water in the solvent is 5-20:1; Preferably, in the step (3), the mass ratio of the adjuvant molecule with the R 3 being nitro shown in the formula 5 to the reducing agent is 1-3:1; Preferably, the temperature of the reaction in the step (3) is 0-35 ℃ and the reaction time is 0.5-3 hours; preferably, after the reaction in the step (3) is finished, purifying to obtain a product; preferably, the purification treatment comprises drying the final reaction solution, adding methanol, collecting supernatant, vacuum drying the crude product, dissolving in ultrapure water solution, washing for 3 times, collecting precipitated precipitate, and freeze drying the product.
  10. 10. Use of an adjuvanted molecule according to any one of claims 1-6 in the preparation of a vaccine.

Description

Adjuvant molecule and preparation method and application thereof Technical Field The invention belongs to the technical field of chemistry, and particularly relates to an adjuvanted molecule, a preparation method and application thereof. Background Vaccines can be divided into attenuated live vaccines, inactivated vaccines and subunit vaccines made with certain components of the natural microorganism, and nucleic acid vaccines. The vaccine mainly comprises antigen, adjuvant and carrier, and further comprises antiseptic, emulsifier or stabilizer components, which do not affect immunogenicity of the vaccine. Adjuvants are classified into a variety of forms, including small molecule adjuvants (e.g., TLR receptor agonists), biological adjuvants (e.g., cytokine GM-CSF), inorganic compounds (e.g., aluminum hydroxide [ AL (OH) 3 ]), and artificial compositions (e.g., double-stranded polyinosinic-cytidylic acid (poly I: C) that mimics double-stranded RNA), and the like. Toll-like receptors (TLRs) are one of the earliest PRRs found in the innate immune system and play an important role in the inflammatory response. Researchers have utilized TLR agonists to initiate responses of the innate immune system as candidate adjuvants to increase the immunogenicity of vaccines. Compared with other adjuvants, TOll-like agonists have definite action sites and obvious action effects. TLR7 and TLR8 are located on endosomal membranes of immune cells. TLR7/8 agonists are mostly imidazole quinoline structures. Imidazoquinoline derivatives are mostly TLR7/8 dual agonists, wherein the free amine group at C-4 and the n-butyl group at C-2 are the groups that optimally activate TLR7/8 activity, respectively. Vaccine adjuvants often require immune cells in the lymph nodes to exert the greatest immune activating function. The immune adjuvant small molecules are not only difficult to enter lymph nodes after traditional interstitial injection, but also can be distributed to the whole body to cause systemic inflammatory response. Disclosure of Invention Aiming at the defects of the prior art, the invention aims to provide an adjuvant molecule, a preparation method and application. The adjuvanted molecules of the invention have lymph node targeting and are effective in initiating immune responses. In order to achieve the aim of the invention, the invention adopts the following technical scheme: In one aspect, the invention provides an adjuvanted molecule having a structure according to formula 1: Wherein R 1 is a nitrogen-containing six-membered heterocyclic ring or a C1-C3 linear alkane, R 2 is a C8-C22 (e.g., C8, C10, C12, C14, C16, C18, C20, or C22, etc.) unsaturated chain hydrocarbon group, a C8-C22 (e.g., C8, C10, C12, C14, C16, C18, C20, or C22, etc.) alkane group, cholesterol Hemisuccinate (CHOL) or 1, 2-dioleoyl-SN-glycerol-3-phosphorylethanolamine (DOPE), and R 3 is nitro or amino. The invention selects a clinically common radiotherapy sensitization drug-Pimonidazole (PMND), and the amino imidazole structure obtained by reduction under the condition of hypoxia has similarity with the main structure of a commercially available TLR7/8 agonist. According to the invention, through modifying the pimonidazole structure and the reduced form (aminoimidazole) thereof, a compound structure with a lymph node targeting function is introduced into the structure, so that the construction of a novel immunoadjuvant molecular library with a lymph node targeting effect and an immune activation effect is realized. The adjuvanted molecules of the invention have lymph node targeting and are effective in initiating immune responses. The adjuvant molecule based on cholesterol structure in the adjuvant molecule can be used as a lipid component to be doped in liposome or lipid nano-particles to prepare a novel nano-delivery platform, so that the novel nano-delivery platform can be applied to different microenvironments to enhance the immunocompetence of the nano-particles, and has wider application prospect. Preferably, R 1 is a nitrogen-containing hexacyclic ring or methylene. Preferably, the adjuvanted molecule has any one of the structures shown in the following formulas 2-1 to 2-4: Wherein R 2 is Cholesterol Hemisuccinate (CHOL) or 1, 2-dioleoyl-SN-glycero-3-phosphoethanolamine (DOPE). Preferably, the R 1 is a nitrogen-containing six-membered heterocyclic ring; preferably, the nitrogen-containing six-membered heterocyclic ring is Preferably, R 3 is amino. Preferably, the adjuvanting molecule is any one of the following compounds: Further preferably, the adjuvanting molecule is a compound as follows: preferably, the adjuvanting molecule is any one of the following compounds: in a second aspect, the present invention provides a process for the preparation of an adjuvanted molecule as described above, comprising (1) reacting a compound of formula 2 with N-hydroxysuccinimide to give an acyloxy intermediate of formula 3; (2) Reacting the acyl oxide intermediat