# US 2021/0340481 A1 · Alginate hollow microfiber

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- Number: US 2021/0340481 A1
- Document ID: US20210340481A1
- Status: Published application
- Kind: A1
- Application number: 17/267,320
- Filed: 2019-08-09
- Published: 2021-11-04
- Earliest claimed: 2018-08-10
- Assignees: Mochida Pharmaceutical Co., Ltd.; The University of Tokyo
- Applicants: Mochida Pharmaceutical Co., Ltd.; The University of Tokyo
- Inventors: Shoji FURUSAKO; Shoji TAKEUCHI
- Web page: https://patentgrep.com/patent/US20210340481A1

## CPC

- C12M 25/12: {the culture medium flowing outside the fiber or tube}
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C07K 16/2866: {against receptors for cytokines, lymphokines, interferons}
- C12M 25/14: {Scaffolds; Matrices (in general C12N5/0068)}
- C12M 25/16: {Particles; Beads; Granular material; Encapsulation (chemical or physical processes conducted in the presence of fluids and solid particles B01J8/00)}
- C12N 5/0068: {General culture methods using substrates (for specific animal cell type C12N5/06)}
- C07K 2317/14: Specific host cells or culture conditions, e.g. components, pH or temperature
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C12N 2533/74: Alginate
- C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS}
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Provided are an alginate gel fiber for antibody production wherein an antibody-producing cell (for example, an antibody-producing CHO cell) is comprised in a core layer, and an antibody production method using the alginate gel fiber. An antibody production method is thereby additionally provided.

## Claims

### Claim 1

An alginate gel fiber formed by covering a core layer comprising antibody-producing cells and a base material with a shell layer comprising an alginate gel having greater mechanical strength than the core layer.

### Claim 2 (depends on 1)

The alginate gel fiber according to claim 1, wherein the antibody-producing cells comprised in the core layer are cells selected from the group consisting of the CHO cells, CHO cell substrains, COS cells, Sp2/0 cells, NSO cells, SP2 cells, PERC6 cells, YB2/0 cells, YE2/0 cells, 1R983F cells, Namalwa cells, Wil-2 cells, Jurkat cells, Vera cells, Molt-4 cells, 293-HEK cells, BHK cells, KGH6 cells, P3X63Ag8.653 cells, C127 cells, JC cells, LA7 cells, ZR-45-30 cells, hTERT cells, NM2C5 cells and UACC-812 cells.

### Claim 3 (depends on 1)

The alginate gel fiber according to claim 1, wherein the base material comprised in the core layer is a base material comprising at least one material selected from the group consisting of a collagen solution, a collagen gel, a medium, a culture solution, an alginic acid solution an alginate gel and mixtures thereof.

### Claim 4 (depends on 1)

The alginate gel fiber according to claim 1, wherein the raw material of the alginate gel comprised in the shell layer is sodium alginate, and the M/G ratio of the sodium alginate is in the range of from 0.4 to 1.8 or in the range of from 0.1 to 0.4.

### Claim 5 (depends on 1)

The alginate gel fiber according to claim 1, wherein the raw material of the alginate gel comprised in the shell layer is sodium alginate, and the molecular weight (GPC) of the sodium alginate is in the range of from 700,000 to 1,000,000 or in the range of from 800,000 to 1,000,000.

### Claim 6 (depends on 1)

The alginate gel fiber according to claim 1, wherein the raw material of the alginate gel comprised in the shell layer is sodium alginate, and the 1 w/w % viscosity of the sodium alginate is in the range of from 50 to 150 (mPa·s) or in the range of from 70 to 150 (mPa·s).

### Claim 7 (depends on 1)

The alginate gel fiber according to claim 1, wherein the outer diameter of the alginate gel fiber is from 0.2 μm to 2,000 μm, while the core layer of the alginate gel fiber has a diameter of from 0.1 μm to 1,000 μm.

### Claim 8

A method for manufacturing an alginate gel fiber formed by covering a core layer comprising antibody-producing cells and a base material with a shell layer comprising an alginate gel having greater mechanical strength than the core layer, wherein
the method for manufacturing an alginate gel fiber uses a microfluidic device 10 comprising an introduction pipe 40, an inlet 1 of the introduction pipe 40, an inlet 2 of the introduction pipe 40 located downstream from the inlet 1, an inlet 3 of the introduction pipe 40 located downstream from the inlet 2, and an outlet 50 of the introduction pipe 40 located downstream from the inlet 2, and comprises
(1) a step of forming a first laminar flow of the antibody-producing cells 6 and the base material in the introduction pipe 40 by introducing antibody-producing cells 6 and a base material from the inlet 1 and injecting the same,
(2) a step of forming a second laminar flow of the sodium alginate solution covering the outer circumference of the first laminar flow by introducing a sodium alginate solution from the inlet 2 and injecting the same,
(3) a step of forming a third laminar flow of the solution comprising the divalent metal ion covering the outer circumference of the second laminar flow by introducing a solution comprising a divalent metal ion from the inlet 3 and injecting the same, and
(4) a step of obtaining an alginate gel fiber formed by covering a core layer comprising antibody-producing cells and a base material that are ejected from the outlet 50 with a shell layer comprising an alginate gel.

### Claim 9 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the antibody-producing cells are selected from the group consisting of the CHO cells, CHO cell substrains, COS cells, Sp2/0 cells, NSO cells, SP2 cells, PERC6 cells, YB2/0 cells, YE2/0 cells, 1R983F cells, Namalwa cells, Wil-2 cells, Jurkat cells, Vera cells, Molt-4 cells, 293-HEK cells, BHK cells, KGH6 cells, P3X63Ag8.653 cells, C127 cells, JC cells, LA7 cells, ZR-45-30 cells, hTERT cells, NM2C5 cells and UACC-812 cells.

### Claim 10 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the base material comprised in the core layer is a base material comprising at least one material selected from the group consisting of a collagen solution, a collagen gel, a medium or culture solution, an alginic acid solution, an alginate gel and mixtures thereof.

### Claim 11 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the divalent metal ion is an ion selected from the group consisting of calcium ions, magnesium ions, barium ions, strontium ions and zinc ions.

### Claim 12 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the solution comprising the divalent metal ion is an aqueous solution selected from the group consisting of a calcium chloride aqueous solution, a calcium carbonate aqueous solution and a calcium gluconate aqueous solution.

### Claim 13 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the temperature during manufacture of the alginate gel fiber is in the range of from 4° C. to 25° C.

### Claim 14 (depends on 1)

A method for manufacturing an antibody using an alginate gel fiber formed by covering a core layer comprising antibody-producing cells and a base material with a shell layer comprising an alginate gel, wherein an alginate gel fiber according to claim 1, is placed in a culture vessel, medium is added to impregnate the alginate gel fiber, and shaking culture is performed to produce the antibody.

### Claim 15 (depends on 1)

The alginate gel fiber according to claim 1, wherein the antibody-producing cells comprised in the core layer are cells selected from the group consisting of the muromonab-CD3-producing CHO cells, trastuzumab-producing CHO cells, rituximab-producing CHO cells, palivizumab-producing CHO cells, infliximab-producing CHO cells, basiliximab-producing CHO cells, tocilizumab-producing CHO cells, gemtuzumab ozogamicin-producing CHO cells, bevacizumab-producing CHO cells, ibritumomab tiuxetan-producing CHO cells, adalimumab-producing CHO cells, cetuximab-producing CHO cells, ranibizumab-producing CHO cells, omalizumab-producing CHO cells, eculizumab-producing CHO cells, panitumumab-producing CHO cells, ustekinumab-producing CHO cells, golimumab-producing CHO cells, canakinumab-producing CHO cells, denosumab-producing CHO cells, mogamulizumab-producing CHO cells, certolizumab pegol-producing CHO cells, ofatumumab-producing CHO cells, pertuzumab-producing CHO cells, trastuzumab emtansine-producing CHO cells, brentuximab vedotin-producing CHO cells, natalizumab-producing CHO cells, nivolumab-producing CHO cells, alemtuzumab-producing CHO cells, secukinumab-producing CHO cells, ramucirumab-producing CHO cells, ipilimumab-producing CHO cells, evolocumab-producing CHO cells, mepolizumab-producing CHO cells, alirocumab-producing CHO cells, ixekizumab-producing CHO cells, brodalumab-producing CHO cells, idarucizumab-producing CHO cells, elotuzumab-producing CHO cells, pembrolizumab-producing CHO cells, sarilumab-producing CHO cells, bezlotoxumab-producing CHO cells, belimumab-producing CHO cells, daratumumab-producing CHO cells, avelumab-producing CHO cells, dupilumab-producing CHO cells, atezolizumab-producing CHO cells, benralizumab-producing CHO cells, inotuzumab ozogamicin-producing CHO cells, emicizumab-producing CHO cells, guselkumab-producing CHO cells, durvalumab-producing CHO cells, obinutuzumab-producing CHO cells and vedolizumab-producing CHO cells.

### Claim 16 (depends on 1)

The alginate gel fiber according to claim 1, wherein the base material comprised in the core layer is a collagen solution or a collagen gel.

### Claim 17 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the antibody-producing cells 6 are cells selected from the group consisting of the muromonab-CD3-producing CHO cells, trastuzumab-producing CHO cells, rituximab-producing CHO cells, palivizumab-producing CHO cells, infliximab-producing CHO cells, basiliximab-producing CHO cells, tocilizumab-producing CHO cells, gemtuzumab ozogamicin-producing CHO cells, bevacizumab-producing CHO cells, ibritumomab tiuxetan-producing CHO cells, adalimumab-producing CHO cells, cetuximab-producing CHO cells, ranibizumab-producing CHO cells, omalizumab-producing CHO cells, eculizumab-producing CHO cells, panitumumab-producing CHO cells, ustekinumab-producing CHO cells, golimumab-producing CHO cells, canakinumab-producing CHO cells, denosumab-producing CHO cells, mogamulizumab-producing CHO cells, certolizumab pegol-producing CHO cells, ofatumumab-producing CHO cells, pertuzumab-producing CHO cells, trastuzumab emtansine-producing CHO cells, brentuximab vedotin-producing CHO cells, natalizumab-producing CHO cells, nivolumab-producing CHO cells, alemtuzumab-producing CHO cells, secukinumab-producing CHO cells, ramucirumab-producing CHO cells, ipilimumab-producing CHO cells, evolocumab-producing CHO cells, mepolizumab-producing CHO cells, alirocumab-producing CHO cells, ixekizumab-producing CHO cells, brodalumab-producing CHO cells, idarucizumab-producing CHO cells, elotuzumab-producing CHO cells, pembrolizumab-producing CHO cells, sarilumab-producing CHO cells, bezlotoxumab-producing CHO cells, belimumab-producing CHO cells, daratumumab-producing CHO cells, avelumab-producing CHO cells, dupilumab-producing CHO cells, atezolizumab-producing CHO cells, benralizumab-producing CHO cells, inotuzumab ozogamicin-producing CHO cells, emicizumab-producing CHO cells, guselkumab-producing CHO cells, durvalumab-producing CHO cells, obinutuzumab-producing CHO cells and vedolizumab-producing CHO cells.

### Claim 18 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the base material comprised in the core layer is a collagen solution or a collagen gel.

### Claim 19 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the divalent metal ion is calcium ions.

### Claim 20 (depends on 8)

The method for manufacturing an alginate gel fiber according to claim 8, wherein the solution comprising the divalent metal ion is a calcium chloride aqueous solution.

### Claim 21

An alginate gel fiber comprising:
a core layer; and
a shell layer disposed on the core layer,
wherein the core layer comprises antibody-producing cells and a base material, the shell layer comprises an alginate gel, and the shell layer has a greater mechanical strength than the core layer.