PH engineered NGF (nerve growth factor) antibody and medical application thereof

An engineering and antibody technology, applied in the direction of antibody, genetic engineering, application, etc., can solve problems such as low efficacy

Active Publication Date: 2018-06-19
FUNDAMENTA THERAPEUTICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, the effector functions of mouse antibodi

Method used

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  • PH engineered NGF (nerve growth factor) antibody and medical application thereof
  • PH engineered NGF (nerve growth factor) antibody and medical application thereof
  • PH engineered NGF (nerve growth factor) antibody and medical application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Example 1: Preparation of Rat Monoclonal Antibody

[0082] Generation of murine monoclonal antibodies against human NGF: Balb / C mice were immunized with purified recombinant human NGF extracellular domain His fusion protein (huNGF-His) (SEQ ID NO:1). Human NGF-His antigen: synthesized and expressed using P01138 in the Uniprot database as a template.

[0083] huNGF-His, recombinant human NGF protein (SEQ ID NO: 1):

[0084] MSMLFYTLITAFLIGIQAEPHSESNVPAGHTIPQAHWTKLQHSLDTALRRARSSAPAAAIAARVAGQTRNITVD PRLFKKRRLRSPRVLFSTQPPREAADTQDLDFEVGGAAPFNRTHRSKRSSSHPIFHRGEFSVCDSVSVWVGDKTTAT DIKGKEVMVLGEVNINNSVFKQYFFETKCRDPNPVDSGCRGIDSKHWNSYCTTTHTFVKALTMDGKQAAWRFIRIDT ACVCVLSRKAVRRA HHHHHH.

[0085] For the immunization using human NGF extracellular domain His fusion protein, the purified antigens used were high dose (50ug) and low dose (10ug) respectively. The mice were immunized three times with complete Freund's adjuvant on days 0, 14, and 35, and the immune response was moni...

Embodiment 2

[0086] Example 2: Molecular Cloning of Monoclonal Antibody Hybridoma Cells

[0087] The variable regions of the heavy and light chains of the rat monoclonal antibody N11 were cloned and amplified by PCR with high-fidelity Phusion enzyme. At least 28 different clones were selected for each amplification and sequenced. The common sequences obtained are as follows:

[0088] N11-VH (SEQ ID NO: 2)

[0089] QVQLKESGPGLVQPSQTLSLTCTVSGFSLTNNNVNWVRQATGRGLEWMGGVWAGGATDYNSALKSRLTITRDTSKSQVFLKMHSLQSEDTATYYCARDGGYSSSTLYAMDAWGQGTSVTVSS

[0090] N11-VL (SEQ ID NO: 3)

[0091] DIQMTQSPASLSASLGETVTIECRASEDIYNALAWYQQKPGKSPQLLIYNTDTLHTGVPSRFSGSGSGTQYSLKINSLQSEDVASYFCQHYFGYPRTFGGGTKLELK

[0092] It contains the following CDR sequences:

[0093]

[0094] 0

[0095] Recombinant N11 chimeric antibody full-length heavy chain sequence (SEQ ID NO: 10)

[0096] PDWTWRVFCLLAVAPGAHS QVQLKESGPGLVQPSQTLSLTCTVSGFSLTNNNVNWVRQATGRGLEWMGGVWAG GATDYNSALKSRLTITRDTSKSQVFLKMHSLQSEDTATYYCARDGGYSSSTLYAMDA...

Embodiment 4

[0099] Example 4: Mouse anti-humanization

[0100] Through conventional CDR grafting and back-mutation of adjacent amino acids in the framework region, a humanized G11-H1L1 antibody was obtained. The protein sequence of its heavy and light chains is as follows.

[0101] Heavy chain variable region sequence of recombinant G11-H1L1 humanized antibody (SEQ ID NO: 12)

[0102] QVQLQESGPGLVKPSETLSLTCTVSGFSLTNNNWSWIRQPPGKGLEWIGYVWAGGATNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCARDGGYSSSTLYAMDAWGQGTLVTVSS

[0103] Light chain variable region sequence (SEQ ID NO: 13) of recombinant G11-H1L1 humanized antibody

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PUM

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Abstract

The invention provides an NGF (nerve growth factor) antibody as well as an antigen binding fragment and a medical application thereof and also provides a chimeric antibody, a humanized antibody and apH engineered antibody which contain antibody CDR areas, medical composition containing a human NGF antibody and an antigen binding fragment thereof as well as an application of the antibody in preparation of drugs for treating and/or preventing pains including postoperation pain, rheumatic arthritis pain and osteoarthritis pain.

Description

technical field [0001] The present invention relates to an NGF antibody, an antigen-binding fragment, a chimeric antibody comprising the CDR region of the NGF antibody, a humanized antibody and a pH engineered antibody, a pharmaceutical composition comprising the NGF antibody and its antigen-binding fragment, and the like Use as pain treatment and prophylaxis. Background technique [0002] Nerve growth factor (NGF) was the first identified neurotrophin, and its role in the development and survival of the peripheral and central nerves has been fully characterized. NGF has been shown to be a critical survival and maintenance factor in the development of peripheral sympathetic and embryonic sensory and basal forebrain cholinergic neurons. Smeyne et al., Nature 368: 246249 (1994) and Crowley et al., Cell 76: 1001 1011 (1994). NGF upregulates the expression of neuropeptides in sensory neurons (Lindsay and Harmer, Nature 337:362364 (1989)), and via two distinct membrane-bound re...

Claims

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Application Information

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IPC IPC(8): C07K16/22C12N15/13C12N15/85A61K39/395A61P19/02A61P29/00A61P19/08A61P25/00A61P3/06
CPCA61K2039/505C07K16/22C07K2317/24C07K2317/56
Inventor 李俊
Owner FUNDAMENTA THERAPEUTICS INC
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