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CN-122028931-A - NGF-Fc fusion protein preparation and application thereof

CN122028931ACN 122028931 ACN122028931 ACN 122028931ACN-122028931-A

Abstract

An NGF-Fc fusion protein preparation has strong stability under high temperature, illumination, oscillation, freeze thawing, acceleration and long-term conditions, can ensure that the preparation has good stability during preparation, transportation and storage, and ensures clinical medication safety and quality controllability.

Inventors

  • LIAO CHUAN
  • LI QINGLEI
  • WANG JIA

Assignees

  • 舒泰神（北京）生物制药股份有限公司

Dates

Publication Date
20260512
Application Date
20241012
Priority Date
20231012

Claims (20)

  1. An NGF-Fc fusion protein formulation, comprising an NGF-Fc fusion protein, a stabilizer, a surfactant, an antioxidant, and a buffer, wherein the stabilizer is trehalose or sucrose, the antioxidant is methionine, the surfactant is a poloxamer, and the stabilizer is a histidine buffer.
  2. The formulation of NGF-Fc fusion protein according to claim 1, wherein the fusion protein is at a concentration of 0.05mg/ml to 5.0mg/ml, preferably 0.05mg/ml to 2.0mg/ml, more preferably 0.1mg/ml to 0.5mg/ml.
  3. The NGF-Fc fusion protein formulation according to claim 1 or2, wherein the histidine buffer is at a concentration of 8mM-50mM, preferably 10mM-30mM.
  4. A formulation of NGF-Fc fusion protein according to any one of claims 1-3, wherein the concentration of sucrose or trehalose is 6-12%, preferably 7-10%.
  5. The NGF-Fc fusion protein formulation of any one of claims 1-4, wherein the methionine is present at a concentration of 0.05mM-7mM, preferably 0.1mM-5mM, more preferably 1mM-3mM.
  6. The NGF-Fc fusion protein formulation of any one of claims 1-5, wherein the poloxamer is at a concentration of 0.005-0.3%, preferably 0.01-0.2%, more preferably 0.01-0.03%.
  7. The formulation of NGF-Fc fusion protein according to any one of claims 1-6, wherein the formulation has a pH of 5.0-6.5, preferably 5.0-6.2, more preferably 5.0-5.4.
  8. The NGF-Fc fusion protein formulation according to any one of claims 1-7, further comprising a cyclodextrin, preferably hydroxypropyl betacyclodextrin.
  9. The NGF-Fc fusion protein formulation of claim 8, wherein the cyclodextrin concentration is 0.05mM-8mM, preferably 1mM-6mM, more preferably 5mM.
  10. The formulation of NGF-Fc fusion protein of claim 1, wherein the formulation is any one of the following: (1) The concentration of the fusion protein is 0.05mg/ml, 0.06mg/ml, 0.08mg/ml, 0.09mg/ml, 0.10mg/ml, 0.11mg/ml, 0.12mg/ml, 0.14mg/ml, 0.16mg/ml, 0.18mg/ml or 2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5Mm, the concentration of poloxamer is 0.01% -0.2%, and the pH of the preparation is 5.0-6.2; (2) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer solution is 10mM, 15mM, 20mM, 25mM or 30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5Mm, the concentration of poloxamer is 0.01% -0.2%, and the pH value of the preparation is 5.0-6.2; (3) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7%, 7.5%, 8%, 8.5%, 9%, 9.5% or 10%, the concentration of methionine is 0.1mM-5Mm, the concentration of poloxamer is 0.01% -0.2%, and the pH value of the preparation is 5.0-6.2; (4) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM, 0.5mM, 1mM, 1.5mM, 2mM, 2.5mM, 3mM, 3.5mM, 4mM, 4.5mM or 5mM, the concentration of poloxamer is 0.01% -0.2%, and the pH of the preparation is 5.0-6.2; (5) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5mM, the concentration of poloxamer is 0.01%, 0.02%, 0.03%, 0.04%, 0.06%, 0.08%, 0.1%, 0.12%, 0.14%, 0.16%, 0.18% or 0.2%, and the pH of the preparation is 5.0-6.2; (6) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5mM, the concentration of poloxamer is 0.01% -0.2%, and the pH of the preparation is 5.0, 5.1, 5.2, 5.3, 5.4, 5.6, 5.8, 6.0 or 6.2.
  11. The formulation of NGF-Fc fusion protein of claim 1, wherein the formulation is any one of the following: (1) The concentration of the fusion protein is 0.1mg/ml, the concentration of histidine buffer is 10mM, the concentration of sucrose or trehalose is 7%, the concentration of methionine is 1.5mM, the concentration of poloxamer is 0.01%, and the pH of the preparation is 5.4; (2) The concentration of the fusion protein is 0.1mg/ml, the concentration of histidine buffer solution is 20mM, the concentration of sucrose or trehalose is 9%, the concentration of methionine is 1.0mM, the concentration of poloxamer is 0.03%, and the pH of the preparation is 5.2; (3) The concentration of the fusion protein is 0.5mg/ml, the concentration of histidine buffer solution is 30mM, the concentration of sucrose or trehalose is 10%, the concentration of methionine is 3mM, the concentration of poloxamer is 0.02%, and the pH of the preparation is 5.0; (4) The concentration of the fusion protein is 2.0mg/ml, the concentration of histidine buffer is 20mM, the concentration of sucrose or trehalose is 8%, the concentration of methionine is 2.0mM, the concentration of poloxamer is 0.1%, and the pH of the preparation is 5.8; (5) The concentration of the fusion protein is 0.5mg/ml, the concentration of histidine buffer is 20mM, the concentration of sucrose or trehalose is 10%, the concentration of methionine is 0.1mM, the concentration of poloxamer is 0.2%, and the pH of the preparation is 6.2; (6) The fusion protein concentration was 0.05mg/ml, histidine buffer concentration was 20mM, sucrose or trehalose concentration was 8.5%, methionine concentration was 5.0mM, poloxamer concentration was 0.04%, and pH of the formulation was 5.2.
  12. The NGF-Fc fusion protein formulation of any one of claims 1-11, wherein the cyclodextrin concentration is 1mM, 2mM, 3mM, 4mM, 5mM or 6mM.
  13. The NGF-Fc fusion protein formulation of any one of claims 1-12, wherein the formulation is a liquid formulation, preferably the formulation is an injection formulation or an eye drop formulation.
  14. The NGF-Fc fusion protein formulation of claim 13, wherein the formulation is an eye drop formulation, further comprising a viscosity enhancing agent;
  15. The NGF-Fc fusion protein formulation of claim 14, wherein the viscosity enhancing agent is hypromellose, more preferably wherein the viscosity enhancing agent is hypromellose K4M.
  16. The NGF-Fc fusion protein formulation of claim 15, wherein the concentration of the viscosity enhancing agent is 0.4% -0.5%.
  17. The formulation of NGF-Fc fusion protein according to claim 15, wherein the formulation of eye drops comprises a fusion protein concentration of 0.1mg/ml, a histidine buffer concentration of 20mM, a trehalose concentration of 8.5%, a methionine concentration of 1.0mM, a poloxamer concentration of 0.03%, a cyclodextrin concentration of 5mM, a hypromellose concentration of 0.4%, 0.45% or 0.5%, and a pH of 5.2.
  18. The NGF-Fc fusion protein formulation of any one of claims 1-17, wherein the NGF-Fc fusion protein comprises an NGF moiety and an Fc moiety from N-terminus to C-terminus, the Fc moiety being from an Fc moiety of an IgG, or a mutant of an Fc moiety of an IgG, the mutant of an Fc moiety of an IgG comprising a site mutation associated with ADCC/CDC activity.
  19. The NGF-Fc fusion protein formulation of claim 18, wherein the NGF moiety is a human nerve growth factor, preferably a human nerve growth factor mutant.
  20. The NGF-Fc fusion protein formulation of claim 19, wherein the human nerve growth factor mutant comprises Phe12Glu at a mutation site relative to the amino acid position of a wild-type human nerve growth factor, preferably wherein the human nerve growth factor comprises the amino acid sequence of SEQ ID No. 2.

Description

NGF-Fc fusion protein preparation and application thereof Technical Field The invention belongs to the technical field of biological medicines, and particularly relates to an NGF-Fc fusion protein preparation and application thereof. Background Nerve Growth Factor (NGF) was the first member of the neurotrophic factor family to be discovered in murine sarcoma cells by Italian scientist Levi-Montlcini in 1953. NGF plays an important regulatory role in the development, differentiation, growth, regeneration and functional expression of central and peripheral neurons. NGF has been used to treat nervous system dysplasia, including amblyopia, neuroma, various nerve injuries, and nervous system diseases. However, it causes pain, has a short half-life in vivo, limits the widespread use of NGF to avoid low dose limitations of nociception and frequent dosing and other side effects. Fusion of protein drugs to longer half-life and/or higher molecular weight moieties is a strategy to make certain protein drugs active for long duration. However, there are few Fc fusion protein pharmaceutical formulations currently on the market, and no NGF-Fc fusion protein formulations have been disclosed. The NGF-Fc fusion protein preparation with high stability is finally obtained through reasonable design, screening and detection of the preparation prescription, the fusion protein preparation can be used for preparing injection and eye drops, and the fusion protein preparation can ensure that the preparation has good stability in the preparation, transportation and storage processes and ensures quality controllability and clinical medication safety. Disclosure of Invention In one aspect, the invention provides an NGF-Fc fusion protein formulation comprising an NGF-Fc fusion protein, a stabilizer, a surfactant, an antioxidant, and a buffer, wherein the stabilizer is trehalose or sucrose, the antioxidant is methionine, the surfactant is a poloxamer, and the stabilizer is a histidine buffer. In some embodiments, the fusion protein concentration is 0.05mg/ml to 5.0mg/ml, preferably 0.05mg/ml to 2.0mg/ml, more preferably 0.1mg/ml to 0.5mg/ml. In some embodiments, the fusion protein is at a concentration of 0.05mg/ml, 0.06mg/ml, 0.08mg/ml, 0.09mg/ml, 0.10mg/ml, 0.11mg/ml, 0.12mg/ml, 0.14mg/ml, 0.16mg/ml, 0.18mg/ml, or 2.0mg/ml. In some embodiments, the histidine buffer is at a concentration of 8mM-50mM, preferably 10mM-30mM. In some embodiments, the histidine buffer is at a concentration of 10mM, 15mM, 20mM, 25mM or 30mM. In some embodiments, the sucrose or trehalose is at a concentration of 6% to 12%, preferably 7% to 10%. In some embodiments, the sucrose or trehalose is at a concentration of 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10%. In some embodiments, the methionine is present at a concentration of 0.05mM-7mM, preferably 0.1mM-5mM, more preferably 1mM-3mM. In some embodiments, the methionine is present at a concentration of 0.1mM, 0.5mM, 1mM, 1.5mM, 2mM, 2.5mM, 3mM, 3.5mM, 4mM, 4.5mM, or 5mM. In some embodiments, the poloxamer is at a concentration of 0.005% to 0.3%, preferably 0.01% to 0.2%, more preferably 0.01% to 0.03%. In some embodiments, the poloxamer is at a concentration of 0.01%, 0.02%, 0.03%, 0.04%, 0.06%, 0.08%, 0.1%, 0.12%, 0.14%, 0.16%, 0.18%, or 0.2%. In some embodiments, the pH of the formulation is from 5.0 to 6.5, preferably from 5.0 to 6.2, more preferably from 5.0 to 5.4. In some embodiments, the pH of the formulation is 5.0, 5.1, 5.2, 5.3, 5.4, 5.6, 5.8, 6.0, or 6.2. In some embodiments, the fusion protein is at a concentration of 0.05mg/ml to 2.0mg/ml, the histidine buffer is at a concentration of 10mM to 30mM, the sucrose or trehalose is at a concentration of 7% to 10%, the methionine is at a concentration of 0.1mM to 5mM, the poloxamer is at a concentration of 0.01% to 0.2%, and the pH of the formulation is between 5.0 and 6.2. In some embodiments, the formulation further comprises a cyclodextrin, preferably hydroxypropyl betacyclodextrin. In some embodiments, the cyclodextrin concentration is 0.05mM-8mM, preferably 1mM-6mM, more preferably 5mM. In some embodiments, the cyclodextrin is at a concentration of 1mM, 2mM, 3mM, 4mM, 5mM, or 6mM. In some embodiments, the formulation is any one of the following formulations: (1) The concentration of the fusion protein is 0.05mg/ml, 0.06mg/ml, 0.08mg/ml, 0.09mg/ml, 0.10mg/ml, 0.11mg/ml, 0.12mg/ml, 0.14mg/ml, 0.16mg/ml, 0.18mg/ml or 2.0mg/ml, the concentration of histidine buffer is 10mM-30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5Mm, the concentration of poloxamer is 0.01% -0.2%, and the pH of the preparation is 5.0-6.2; (2) The concentration of the fusion protein is 0.05mg/ml-2.0mg/ml, the concentration of histidine buffer solution is 10mM, 15mM, 20mM, 25mM or 30mM, the concentration of sucrose or trehalose is 7% -10%, the concentration of methionine is 0.1mM-5Mm, the concentration of poloxamer is 0.01% -0.2%, and the