N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4
Purpose Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipe...
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| Veröffentlicht in: | Pharmaceutical research 2022-10, Vol.39 (10), p.2541-2554 |
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| creator | Masui, Sho Yonezawa, Atsushi Yokoyama, Kotoko Iwamoto, Noriko Shimada, Takashi Onishi, Akira Onizawa, Hideo Fujii, Takayuki Murakami, Kosaku Murata, Koichi Tanaka, Masao Nakagawa, Shunsaku Hira, Daiki Itohara, Kotaro Imai, Satoshi Nakagawa, Takayuki Hayakari, Makoto Matsuda, Shuichi Morinobu, Akio Terada, Tomohiro Matsubara, Kazuo |
| description | Purpose
Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipeptidyl peptidase-4 (DPP-4). The purpose of this study was to investigate the biotransformation of ETN in humans and mice and evaluate its effects on functional properties
.
Methods
An analytical method using liquid chromatography-mass spectrometry (LC–MS/MS) was established. The N-terminal heterogeneity of ETN was assessed in the serum of patients with rheumatoid arthritis or mice receiving ETN. The
in vitro
N-terminal truncation was explored using recombinant DPP-4. The binding affinity to TNF-α or TNF-β was investigated using an in-house enzyme-linked immunosorbent assay.
Results
In the formulations, about 90% of ETN had an intact N-terminus, while the N-terminal truncated form was most abundant in the serum of the patients with rheumatoid arthritis and mice. Recombinant human DPP-4 cleaved two amino acids from the N-terminus of ETN
in vitro
. Sitagliptin, a DPP-4 inhibitor, inhibited N-terminal truncation both
in vivo
and
in vitro
. However, N-terminal truncation did not affect the binding ability to TNF-α or TNF-β and the pharmacokinetics of ETN. ETN biosimilars exhibited similar characteristics to the reference product
in vivo
and
in vitro
.
Conclusions
ETN undergoes N-terminal truncation in the body, and DPP-4 cleaves exogenous ETN via N-terminal proteolysis. The application of an MS-based assay will detect novel biotransformation of therapeutic proteins. |
| doi_str_mv | 10.1007/s11095-022-03371-2 |
| format | Article |
| fullrecord | <record><control><sourceid>gale_proqu</sourceid><recordid>TN_cdi_proquest_miscellaneous_2704867498</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><galeid>A722215626</galeid><sourcerecordid>A722215626</sourcerecordid><originalsourceid>FETCH-LOGICAL-c436t-8e387ce571c13699f85e0359d85f7ef0c2f3fe1a8408a0f78a53353ca10ba6893</originalsourceid><addsrcrecordid>eNp9kU1LXDEUhoO04NT6B1xdcOMmevJ1kyzFz4JUFxXchZg5kcidm2lyZ3H_vRlHkJZSskhOeJ7DCy8hRwxOGYA-q4yBVRQ4pyCEZpTvkQVTWlAL8ukLWYDmkhot2T75VusrABhm5YLc_6QTllUaN7XLsbua_Igl4HrqUu0eSp4wD_OUgh-GeTsHrBWX3fPcXaZ1w9JyHrqH94evSOV38jX6oeLhx31AHq-vfl3c0rv7mx8X53c0SNFP1KAwOqDSLDDRWxuNQhDKLo2KGiMEHkVE5o0E4yFq45UQSgTP4Nn3xooDcrLbuy759wbr5FapBhyGlj9vquMapOm1tKahx3-hr3lTxpauUVyCtlayT-rFD-jSGPNUfNgudeeac85Uz_tGnf6DameJqxTyiDG1_z8EvhNCybUWjG5d0sqX2TFw2-rcrjrXqnPv1TneJLGTaoPHFyyfif9jvQFeyZlI</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>2724079941</pqid></control><display><type>article</type><title>N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4</title><source>SpringerLink Journals - AutoHoldings</source><creator>Masui, Sho ; Yonezawa, Atsushi ; Yokoyama, Kotoko ; Iwamoto, Noriko ; Shimada, Takashi ; Onishi, Akira ; Onizawa, Hideo ; Fujii, Takayuki ; Murakami, Kosaku ; Murata, Koichi ; Tanaka, Masao ; Nakagawa, Shunsaku ; Hira, Daiki ; Itohara, Kotaro ; Imai, Satoshi ; Nakagawa, Takayuki ; Hayakari, Makoto ; Matsuda, Shuichi ; Morinobu, Akio ; Terada, Tomohiro ; Matsubara, Kazuo</creator><creatorcontrib>Masui, Sho ; Yonezawa, Atsushi ; Yokoyama, Kotoko ; Iwamoto, Noriko ; Shimada, Takashi ; Onishi, Akira ; Onizawa, Hideo ; Fujii, Takayuki ; Murakami, Kosaku ; Murata, Koichi ; Tanaka, Masao ; Nakagawa, Shunsaku ; Hira, Daiki ; Itohara, Kotaro ; Imai, Satoshi ; Nakagawa, Takayuki ; Hayakari, Makoto ; Matsuda, Shuichi ; Morinobu, Akio ; Terada, Tomohiro ; Matsubara, Kazuo</creatorcontrib><description>Purpose
Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipeptidyl peptidase-4 (DPP-4). The purpose of this study was to investigate the biotransformation of ETN in humans and mice and evaluate its effects on functional properties
.
Methods
An analytical method using liquid chromatography-mass spectrometry (LC–MS/MS) was established. The N-terminal heterogeneity of ETN was assessed in the serum of patients with rheumatoid arthritis or mice receiving ETN. The
in vitro
N-terminal truncation was explored using recombinant DPP-4. The binding affinity to TNF-α or TNF-β was investigated using an in-house enzyme-linked immunosorbent assay.
Results
In the formulations, about 90% of ETN had an intact N-terminus, while the N-terminal truncated form was most abundant in the serum of the patients with rheumatoid arthritis and mice. Recombinant human DPP-4 cleaved two amino acids from the N-terminus of ETN
in vitro
. Sitagliptin, a DPP-4 inhibitor, inhibited N-terminal truncation both
in vivo
and
in vitro
. However, N-terminal truncation did not affect the binding ability to TNF-α or TNF-β and the pharmacokinetics of ETN. ETN biosimilars exhibited similar characteristics to the reference product
in vivo
and
in vitro
.
Conclusions
ETN undergoes N-terminal truncation in the body, and DPP-4 cleaves exogenous ETN via N-terminal proteolysis. The application of an MS-based assay will detect novel biotransformation of therapeutic proteins.</description><identifier>ISSN: 0724-8741</identifier><identifier>EISSN: 1573-904X</identifier><identifier>DOI: 10.1007/s11095-022-03371-2</identifier><language>eng</language><publisher>New York: Springer US</publisher><subject>Arthritis ; Biochemistry ; Biological products ; Biomedical and Life Sciences ; Biomedical Engineering and Bioengineering ; Biomedicine ; Biotransformation ; Enzyme-linked immunosorbent assay ; Enzymes ; Etanercept ; Fc receptors ; Fusion protein ; Immunoglobulin G ; Liquid chromatography ; Mass spectrometry ; Mass spectroscopy ; Medical colleges ; Medical Law ; Medical research ; Medicine, Experimental ; N-Terminus ; Original Research Article ; Peptidase ; Pharmacokinetics ; Pharmacology/Toxicology ; Pharmacy ; Proteolysis ; Rheumatoid arthritis ; Rheumatoid factor ; Sitagliptin ; Tumor necrosis factor ; Tumor necrosis factor-TNF ; Tumor necrosis factor-α</subject><ispartof>Pharmaceutical research, 2022-10, Vol.39 (10), p.2541-2554</ispartof><rights>The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature 2022. Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.</rights><rights>COPYRIGHT 2022 Springer</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><cites>FETCH-LOGICAL-c436t-8e387ce571c13699f85e0359d85f7ef0c2f3fe1a8408a0f78a53353ca10ba6893</cites><orcidid>0000-0002-8057-6768</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s11095-022-03371-2$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s11095-022-03371-2$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,780,784,27924,27925,41488,42557,51319</link.rule.ids></links><search><creatorcontrib>Masui, Sho</creatorcontrib><creatorcontrib>Yonezawa, Atsushi</creatorcontrib><creatorcontrib>Yokoyama, Kotoko</creatorcontrib><creatorcontrib>Iwamoto, Noriko</creatorcontrib><creatorcontrib>Shimada, Takashi</creatorcontrib><creatorcontrib>Onishi, Akira</creatorcontrib><creatorcontrib>Onizawa, Hideo</creatorcontrib><creatorcontrib>Fujii, Takayuki</creatorcontrib><creatorcontrib>Murakami, Kosaku</creatorcontrib><creatorcontrib>Murata, Koichi</creatorcontrib><creatorcontrib>Tanaka, Masao</creatorcontrib><creatorcontrib>Nakagawa, Shunsaku</creatorcontrib><creatorcontrib>Hira, Daiki</creatorcontrib><creatorcontrib>Itohara, Kotaro</creatorcontrib><creatorcontrib>Imai, Satoshi</creatorcontrib><creatorcontrib>Nakagawa, Takayuki</creatorcontrib><creatorcontrib>Hayakari, Makoto</creatorcontrib><creatorcontrib>Matsuda, Shuichi</creatorcontrib><creatorcontrib>Morinobu, Akio</creatorcontrib><creatorcontrib>Terada, Tomohiro</creatorcontrib><creatorcontrib>Matsubara, Kazuo</creatorcontrib><title>N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4</title><title>Pharmaceutical research</title><addtitle>Pharm Res</addtitle><description>Purpose
Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipeptidyl peptidase-4 (DPP-4). The purpose of this study was to investigate the biotransformation of ETN in humans and mice and evaluate its effects on functional properties
.
Methods
An analytical method using liquid chromatography-mass spectrometry (LC–MS/MS) was established. The N-terminal heterogeneity of ETN was assessed in the serum of patients with rheumatoid arthritis or mice receiving ETN. The
in vitro
N-terminal truncation was explored using recombinant DPP-4. The binding affinity to TNF-α or TNF-β was investigated using an in-house enzyme-linked immunosorbent assay.
Results
In the formulations, about 90% of ETN had an intact N-terminus, while the N-terminal truncated form was most abundant in the serum of the patients with rheumatoid arthritis and mice. Recombinant human DPP-4 cleaved two amino acids from the N-terminus of ETN
in vitro
. Sitagliptin, a DPP-4 inhibitor, inhibited N-terminal truncation both
in vivo
and
in vitro
. However, N-terminal truncation did not affect the binding ability to TNF-α or TNF-β and the pharmacokinetics of ETN. ETN biosimilars exhibited similar characteristics to the reference product
in vivo
and
in vitro
.
Conclusions
ETN undergoes N-terminal truncation in the body, and DPP-4 cleaves exogenous ETN via N-terminal proteolysis. The application of an MS-based assay will detect novel biotransformation of therapeutic proteins.</description><subject>Arthritis</subject><subject>Biochemistry</subject><subject>Biological products</subject><subject>Biomedical and Life Sciences</subject><subject>Biomedical Engineering and Bioengineering</subject><subject>Biomedicine</subject><subject>Biotransformation</subject><subject>Enzyme-linked immunosorbent assay</subject><subject>Enzymes</subject><subject>Etanercept</subject><subject>Fc receptors</subject><subject>Fusion protein</subject><subject>Immunoglobulin G</subject><subject>Liquid chromatography</subject><subject>Mass spectrometry</subject><subject>Mass spectroscopy</subject><subject>Medical colleges</subject><subject>Medical Law</subject><subject>Medical research</subject><subject>Medicine, Experimental</subject><subject>N-Terminus</subject><subject>Original Research Article</subject><subject>Peptidase</subject><subject>Pharmacokinetics</subject><subject>Pharmacology/Toxicology</subject><subject>Pharmacy</subject><subject>Proteolysis</subject><subject>Rheumatoid arthritis</subject><subject>Rheumatoid factor</subject><subject>Sitagliptin</subject><subject>Tumor necrosis factor</subject><subject>Tumor necrosis factor-TNF</subject><subject>Tumor necrosis factor-α</subject><issn>0724-8741</issn><issn>1573-904X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2022</creationdate><recordtype>article</recordtype><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><recordid>eNp9kU1LXDEUhoO04NT6B1xdcOMmevJ1kyzFz4JUFxXchZg5kcidm2lyZ3H_vRlHkJZSskhOeJ7DCy8hRwxOGYA-q4yBVRQ4pyCEZpTvkQVTWlAL8ukLWYDmkhot2T75VusrABhm5YLc_6QTllUaN7XLsbua_Igl4HrqUu0eSp4wD_OUgh-GeTsHrBWX3fPcXaZ1w9JyHrqH94evSOV38jX6oeLhx31AHq-vfl3c0rv7mx8X53c0SNFP1KAwOqDSLDDRWxuNQhDKLo2KGiMEHkVE5o0E4yFq45UQSgTP4Nn3xooDcrLbuy759wbr5FapBhyGlj9vquMapOm1tKahx3-hr3lTxpauUVyCtlayT-rFD-jSGPNUfNgudeeac85Uz_tGnf6DameJqxTyiDG1_z8EvhNCybUWjG5d0sqX2TFw2-rcrjrXqnPv1TneJLGTaoPHFyyfif9jvQFeyZlI</recordid><startdate>20221001</startdate><enddate>20221001</enddate><creator>Masui, Sho</creator><creator>Yonezawa, Atsushi</creator><creator>Yokoyama, Kotoko</creator><creator>Iwamoto, Noriko</creator><creator>Shimada, Takashi</creator><creator>Onishi, Akira</creator><creator>Onizawa, Hideo</creator><creator>Fujii, Takayuki</creator><creator>Murakami, Kosaku</creator><creator>Murata, Koichi</creator><creator>Tanaka, Masao</creator><creator>Nakagawa, Shunsaku</creator><creator>Hira, Daiki</creator><creator>Itohara, Kotaro</creator><creator>Imai, Satoshi</creator><creator>Nakagawa, Takayuki</creator><creator>Hayakari, Makoto</creator><creator>Matsuda, Shuichi</creator><creator>Morinobu, Akio</creator><creator>Terada, Tomohiro</creator><creator>Matsubara, Kazuo</creator><general>Springer US</general><general>Springer</general><general>Springer Nature B.V</general><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>7RV</scope><scope>7TK</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>BENPR</scope><scope>CCPQU</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>K9.</scope><scope>KB0</scope><scope>M0S</scope><scope>M1P</scope><scope>NAPCQ</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0002-8057-6768</orcidid></search><sort><creationdate>20221001</creationdate><title>N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4</title><author>Masui, Sho ; Yonezawa, Atsushi ; Yokoyama, Kotoko ; Iwamoto, Noriko ; Shimada, Takashi ; Onishi, Akira ; Onizawa, Hideo ; Fujii, Takayuki ; Murakami, Kosaku ; Murata, Koichi ; Tanaka, Masao ; Nakagawa, Shunsaku ; Hira, Daiki ; Itohara, Kotaro ; Imai, Satoshi ; Nakagawa, Takayuki ; Hayakari, Makoto ; Matsuda, Shuichi ; Morinobu, Akio ; Terada, Tomohiro ; Matsubara, Kazuo</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c436t-8e387ce571c13699f85e0359d85f7ef0c2f3fe1a8408a0f78a53353ca10ba6893</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2022</creationdate><topic>Arthritis</topic><topic>Biochemistry</topic><topic>Biological products</topic><topic>Biomedical and Life Sciences</topic><topic>Biomedical Engineering and Bioengineering</topic><topic>Biomedicine</topic><topic>Biotransformation</topic><topic>Enzyme-linked immunosorbent assay</topic><topic>Enzymes</topic><topic>Etanercept</topic><topic>Fc receptors</topic><topic>Fusion protein</topic><topic>Immunoglobulin G</topic><topic>Liquid chromatography</topic><topic>Mass spectrometry</topic><topic>Mass spectroscopy</topic><topic>Medical colleges</topic><topic>Medical Law</topic><topic>Medical research</topic><topic>Medicine, Experimental</topic><topic>N-Terminus</topic><topic>Original Research Article</topic><topic>Peptidase</topic><topic>Pharmacokinetics</topic><topic>Pharmacology/Toxicology</topic><topic>Pharmacy</topic><topic>Proteolysis</topic><topic>Rheumatoid arthritis</topic><topic>Rheumatoid factor</topic><topic>Sitagliptin</topic><topic>Tumor necrosis factor</topic><topic>Tumor necrosis factor-TNF</topic><topic>Tumor necrosis factor-α</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Masui, Sho</creatorcontrib><creatorcontrib>Yonezawa, Atsushi</creatorcontrib><creatorcontrib>Yokoyama, Kotoko</creatorcontrib><creatorcontrib>Iwamoto, Noriko</creatorcontrib><creatorcontrib>Shimada, Takashi</creatorcontrib><creatorcontrib>Onishi, Akira</creatorcontrib><creatorcontrib>Onizawa, Hideo</creatorcontrib><creatorcontrib>Fujii, Takayuki</creatorcontrib><creatorcontrib>Murakami, Kosaku</creatorcontrib><creatorcontrib>Murata, Koichi</creatorcontrib><creatorcontrib>Tanaka, Masao</creatorcontrib><creatorcontrib>Nakagawa, Shunsaku</creatorcontrib><creatorcontrib>Hira, Daiki</creatorcontrib><creatorcontrib>Itohara, Kotaro</creatorcontrib><creatorcontrib>Imai, Satoshi</creatorcontrib><creatorcontrib>Nakagawa, Takayuki</creatorcontrib><creatorcontrib>Hayakari, Makoto</creatorcontrib><creatorcontrib>Matsuda, Shuichi</creatorcontrib><creatorcontrib>Morinobu, Akio</creatorcontrib><creatorcontrib>Terada, Tomohiro</creatorcontrib><creatorcontrib>Matsubara, Kazuo</creatorcontrib><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Nursing & Allied Health Database</collection><collection>Neurosciences Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central</collection><collection>ProQuest One Community College</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Nursing & Allied Health Database (Alumni Edition)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Nursing & Allied Health Premium</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>MEDLINE - Academic</collection><jtitle>Pharmaceutical research</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Masui, Sho</au><au>Yonezawa, Atsushi</au><au>Yokoyama, Kotoko</au><au>Iwamoto, Noriko</au><au>Shimada, Takashi</au><au>Onishi, Akira</au><au>Onizawa, Hideo</au><au>Fujii, Takayuki</au><au>Murakami, Kosaku</au><au>Murata, Koichi</au><au>Tanaka, Masao</au><au>Nakagawa, Shunsaku</au><au>Hira, Daiki</au><au>Itohara, Kotaro</au><au>Imai, Satoshi</au><au>Nakagawa, Takayuki</au><au>Hayakari, Makoto</au><au>Matsuda, Shuichi</au><au>Morinobu, Akio</au><au>Terada, Tomohiro</au><au>Matsubara, Kazuo</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4</atitle><jtitle>Pharmaceutical research</jtitle><stitle>Pharm Res</stitle><date>2022-10-01</date><risdate>2022</risdate><volume>39</volume><issue>10</issue><spage>2541</spage><epage>2554</epage><pages>2541-2554</pages><issn>0724-8741</issn><eissn>1573-904X</eissn><abstract>Purpose
Biologics are structurally heterogeneous and can undergo biotransformation in the body. Etanercept (ETN) is a fusion protein composed of a soluble tumor necrosis factor (TNF) receptor and the Fc portion of human immunoglobulin G1. The N-terminus of ETN has a putative sequence cleaved by dipeptidyl peptidase-4 (DPP-4). The purpose of this study was to investigate the biotransformation of ETN in humans and mice and evaluate its effects on functional properties
.
Methods
An analytical method using liquid chromatography-mass spectrometry (LC–MS/MS) was established. The N-terminal heterogeneity of ETN was assessed in the serum of patients with rheumatoid arthritis or mice receiving ETN. The
in vitro
N-terminal truncation was explored using recombinant DPP-4. The binding affinity to TNF-α or TNF-β was investigated using an in-house enzyme-linked immunosorbent assay.
Results
In the formulations, about 90% of ETN had an intact N-terminus, while the N-terminal truncated form was most abundant in the serum of the patients with rheumatoid arthritis and mice. Recombinant human DPP-4 cleaved two amino acids from the N-terminus of ETN
in vitro
. Sitagliptin, a DPP-4 inhibitor, inhibited N-terminal truncation both
in vivo
and
in vitro
. However, N-terminal truncation did not affect the binding ability to TNF-α or TNF-β and the pharmacokinetics of ETN. ETN biosimilars exhibited similar characteristics to the reference product
in vivo
and
in vitro
.
Conclusions
ETN undergoes N-terminal truncation in the body, and DPP-4 cleaves exogenous ETN via N-terminal proteolysis. The application of an MS-based assay will detect novel biotransformation of therapeutic proteins.</abstract><cop>New York</cop><pub>Springer US</pub><doi>10.1007/s11095-022-03371-2</doi><tpages>14</tpages><orcidid>https://orcid.org/0000-0002-8057-6768</orcidid></addata></record> |
| fulltext | fulltext |
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| ispartof | Pharmaceutical research, 2022-10, Vol.39 (10), p.2541-2554 |
| issn | 0724-8741 1573-904X |
| language | eng |
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| source | SpringerLink Journals - AutoHoldings |
| subjects | Arthritis Biochemistry Biological products Biomedical and Life Sciences Biomedical Engineering and Bioengineering Biomedicine Biotransformation Enzyme-linked immunosorbent assay Enzymes Etanercept Fc receptors Fusion protein Immunoglobulin G Liquid chromatography Mass spectrometry Mass spectroscopy Medical colleges Medical Law Medical research Medicine, Experimental N-Terminus Original Research Article Peptidase Pharmacokinetics Pharmacology/Toxicology Pharmacy Proteolysis Rheumatoid arthritis Rheumatoid factor Sitagliptin Tumor necrosis factor Tumor necrosis factor-TNF Tumor necrosis factor-α |
| title | N-terminus of Etanercept is Proteolytically Processed by Dipeptidyl Peptidase-4 |
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