Kinetics of Internalization and Cytotoxicity of Transferrin‐Neocarzinostatin Conjugate in Human Leukemia Cell Line, K562
ABSTRACT Human serum transferrin was conjugated with an anticancer‐active polypeptide, neocarzinostatin, by using N‐succinimidy1‐3‐(2‐pyridyldithio)propionate. The conjugate consisted of 1.8 mol of neocarzinostatin per 1 mol of transferrin on average and retained cytotoxic activity against human tum...
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| Veröffentlicht in: | Cancer science 1990-01, Vol.81 (1), p.91-99 |
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| creator | Kohgo, Yutaka Kondo, Hitoshi Kato, Junji Sasaki, Katsunori Tsushima, Nobuyasu Nishisato, Takuji Hirayama, Michiaki Fujikawa, Koshi Shintani, Naoaki Mogi, Yoshihiro Niitsu, Yoshiro |
| description | ABSTRACT
Human serum transferrin was conjugated with an anticancer‐active polypeptide, neocarzinostatin, by using N‐succinimidy1‐3‐(2‐pyridyldithio)propionate. The conjugate consisted of 1.8 mol of neocarzinostatin per 1 mol of transferrin on average and retained cytotoxic activity against human tumor cells. This conjugate was capable of binding to the transferrin receptor of human myelogenous leukemia K562 cells and was internalized by endocytosis. The LD50 values of the conjugate and neocarzinostatin alone in the presence of excess native bovine transferrin were 0.20 μ/ml and 1.80 μ/ml, respectively, suggesting that the effect of the conjugate was greater than that of neocarzinostatin alone. A pulse‐chase experiment using 125I‐labeled conjugate revealed that 25% of the internalized conjugate was degraded in lysosomes and the rest was recycled back to the cell surface without degradation. About 75% of this conjugate recycled back to the cell surface in 18.3 min (3.4 min for receptor binding and 14.9 min for recycling to the cell surface through the acidosomes), while the rest was delivered from the cell surface to the lysosome in 19.6 min. This phenomenon was confirmed by chasing the radioactivity in subcellular fractions separated by Percoll density gradient centrifugation. Therefore, it was concluded that this conjugate is internalized specifically by transferrin receptors and is at least partly transferred to and accumulated in lysosomal compartments, resulting in the inhibition of cellular DNA synthesis. |
| doi_str_mv | 10.1111/j.1349-7006.1990.tb02512.x |
| format | Article |
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Human serum transferrin was conjugated with an anticancer‐active polypeptide, neocarzinostatin, by using N‐succinimidy1‐3‐(2‐pyridyldithio)propionate. The conjugate consisted of 1.8 mol of neocarzinostatin per 1 mol of transferrin on average and retained cytotoxic activity against human tumor cells. This conjugate was capable of binding to the transferrin receptor of human myelogenous leukemia K562 cells and was internalized by endocytosis. The LD50 values of the conjugate and neocarzinostatin alone in the presence of excess native bovine transferrin were 0.20 μ/ml and 1.80 μ/ml, respectively, suggesting that the effect of the conjugate was greater than that of neocarzinostatin alone. A pulse‐chase experiment using 125I‐labeled conjugate revealed that 25% of the internalized conjugate was degraded in lysosomes and the rest was recycled back to the cell surface without degradation. About 75% of this conjugate recycled back to the cell surface in 18.3 min (3.4 min for receptor binding and 14.9 min for recycling to the cell surface through the acidosomes), while the rest was delivered from the cell surface to the lysosome in 19.6 min. This phenomenon was confirmed by chasing the radioactivity in subcellular fractions separated by Percoll density gradient centrifugation. Therefore, it was concluded that this conjugate is internalized specifically by transferrin receptors and is at least partly transferred to and accumulated in lysosomal compartments, resulting in the inhibition of cellular DNA synthesis.</description><identifier>ISSN: 0910-5050</identifier><identifier>ISSN: 1347-9032</identifier><identifier>EISSN: 1349-7006</identifier><identifier>EISSN: 1876-4673</identifier><identifier>DOI: 10.1111/j.1349-7006.1990.tb02512.x</identifier><identifier>PMID: 2139019</identifier><identifier>CODEN: GANNA2</identifier><language>eng</language><publisher>Oxford, UK: Blackwell Publishing Ltd</publisher><subject>Antibiotics, Antineoplastic - metabolism ; Antineoplastic agents ; Azides - pharmacology ; Biological and medical sciences ; Cell surface ; Centrifugation ; Centrifugation, Density Gradient ; Conjugate ; Cytotoxicity ; DNA biosynthesis ; Endocytosis ; Humans ; Internalization ; Iodine Radioisotopes ; Kinetics ; Leukemia ; Leukemia, Erythroblastic, Acute - drug therapy ; Leukemia, Erythroblastic, Acute - metabolism ; Lysosomes ; Lysosomes - metabolism ; Medical sciences ; Myeloid leukemia ; Neocarzinostatin ; Pharmacology. Drug treatments ; Propionic acid ; Protein turnover ; Radioactivity ; Receptors, Transferrin - metabolism ; Sodium Azide ; Targetting ; Transferrin ; Transferrin - metabolism ; Transferrin - therapeutic use ; Transferrin receptor ; Transferrin receptors ; Tumor cells ; Tumor Cells, Cultured ; Zinostatin - metabolism ; Zinostatin - therapeutic use</subject><ispartof>Cancer science, 1990-01, Vol.81 (1), p.91-99</ispartof><rights>1993 INIST-CNRS</rights><rights>Copyright John Wiley & Sons, Inc. Jan 1990</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c5981-bcc9d3b5e4b5766fcfd28c46eb521416731fe437f906f040905ed7c7146cb22a3</citedby><cites>FETCH-LOGICAL-c5981-bcc9d3b5e4b5766fcfd28c46eb521416731fe437f906f040905ed7c7146cb22a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5917957/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5917957/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,315,729,782,786,887,1419,4028,27932,27933,27934,45583,45584,53800,53802</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=4615578$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2139019$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kohgo, Yutaka</creatorcontrib><creatorcontrib>Kondo, Hitoshi</creatorcontrib><creatorcontrib>Kato, Junji</creatorcontrib><creatorcontrib>Sasaki, Katsunori</creatorcontrib><creatorcontrib>Tsushima, Nobuyasu</creatorcontrib><creatorcontrib>Nishisato, Takuji</creatorcontrib><creatorcontrib>Hirayama, Michiaki</creatorcontrib><creatorcontrib>Fujikawa, Koshi</creatorcontrib><creatorcontrib>Shintani, Naoaki</creatorcontrib><creatorcontrib>Mogi, Yoshihiro</creatorcontrib><creatorcontrib>Niitsu, Yoshiro</creatorcontrib><title>Kinetics of Internalization and Cytotoxicity of Transferrin‐Neocarzinostatin Conjugate in Human Leukemia Cell Line, K562</title><title>Cancer science</title><addtitle>Jpn J Cancer Res</addtitle><description>ABSTRACT
Human serum transferrin was conjugated with an anticancer‐active polypeptide, neocarzinostatin, by using N‐succinimidy1‐3‐(2‐pyridyldithio)propionate. The conjugate consisted of 1.8 mol of neocarzinostatin per 1 mol of transferrin on average and retained cytotoxic activity against human tumor cells. This conjugate was capable of binding to the transferrin receptor of human myelogenous leukemia K562 cells and was internalized by endocytosis. The LD50 values of the conjugate and neocarzinostatin alone in the presence of excess native bovine transferrin were 0.20 μ/ml and 1.80 μ/ml, respectively, suggesting that the effect of the conjugate was greater than that of neocarzinostatin alone. A pulse‐chase experiment using 125I‐labeled conjugate revealed that 25% of the internalized conjugate was degraded in lysosomes and the rest was recycled back to the cell surface without degradation. About 75% of this conjugate recycled back to the cell surface in 18.3 min (3.4 min for receptor binding and 14.9 min for recycling to the cell surface through the acidosomes), while the rest was delivered from the cell surface to the lysosome in 19.6 min. This phenomenon was confirmed by chasing the radioactivity in subcellular fractions separated by Percoll density gradient centrifugation. Therefore, it was concluded that this conjugate is internalized specifically by transferrin receptors and is at least partly transferred to and accumulated in lysosomal compartments, resulting in the inhibition of cellular DNA synthesis.</description><subject>Antibiotics, Antineoplastic - metabolism</subject><subject>Antineoplastic agents</subject><subject>Azides - pharmacology</subject><subject>Biological and medical sciences</subject><subject>Cell surface</subject><subject>Centrifugation</subject><subject>Centrifugation, Density Gradient</subject><subject>Conjugate</subject><subject>Cytotoxicity</subject><subject>DNA biosynthesis</subject><subject>Endocytosis</subject><subject>Humans</subject><subject>Internalization</subject><subject>Iodine Radioisotopes</subject><subject>Kinetics</subject><subject>Leukemia</subject><subject>Leukemia, Erythroblastic, Acute - drug therapy</subject><subject>Leukemia, Erythroblastic, Acute - metabolism</subject><subject>Lysosomes</subject><subject>Lysosomes - metabolism</subject><subject>Medical sciences</subject><subject>Myeloid leukemia</subject><subject>Neocarzinostatin</subject><subject>Pharmacology. Drug treatments</subject><subject>Propionic acid</subject><subject>Protein turnover</subject><subject>Radioactivity</subject><subject>Receptors, Transferrin - metabolism</subject><subject>Sodium Azide</subject><subject>Targetting</subject><subject>Transferrin</subject><subject>Transferrin - metabolism</subject><subject>Transferrin - therapeutic use</subject><subject>Transferrin receptor</subject><subject>Transferrin receptors</subject><subject>Tumor cells</subject><subject>Tumor Cells, Cultured</subject><subject>Zinostatin - metabolism</subject><subject>Zinostatin - therapeutic use</subject><issn>0910-5050</issn><issn>1347-9032</issn><issn>1349-7006</issn><issn>1876-4673</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1990</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqVkc-O1CAcx4nRrOPqI5g0ajzZChTKYKLJplndzU70sp4JpXSltrALVGfm5CP4jD6JNNNM1JtwIOT7_f39APAMwQKl87ovUEl4ziCsCsQ5LGIDMUW42N4Dq6N0H6wgRzCnkMKH4FEIPYSIwQqfgBOMSg4RX4H9lbE6GhUy12WXNmpv5WD2MhpnM2nbrN5FF93WKBN3s-faSxs67b2xv378_Kidkn5vrAsxxdisdrafbmTUWfpcTKO02UZPX_VoZFbrYcg2qd6r7IpW-DF40Mkh6CfLewo-vz-_ri_yzacPl_XZJleUr1HeKMXbsqGaNJRVVae6Fq8VqXRDMSKoYiXqNClZx2HVQQI5pLpliiFSqQZjWZ6Cd4e8t1Mz6lZpG70cxK03o_Q74aQRfyvWfBE37pugHDFOWUrwckng3d2kQxSjCSoNI612UxCMM4QhXyfj83-MvZvmhQaBSVp-uoQm15uDS3kXgtfdsRUExcxX9GKGKGaIYuYrFr5im4Kf_jnMMXQBmvQXiy6DkkOXaCkTjjZSIUrZ3Onbg-27GfTuPxoQ9dk5R-Vvbs3Ecw</recordid><startdate>199001</startdate><enddate>199001</enddate><creator>Kohgo, Yutaka</creator><creator>Kondo, Hitoshi</creator><creator>Kato, Junji</creator><creator>Sasaki, Katsunori</creator><creator>Tsushima, Nobuyasu</creator><creator>Nishisato, Takuji</creator><creator>Hirayama, Michiaki</creator><creator>Fujikawa, Koshi</creator><creator>Shintani, Naoaki</creator><creator>Mogi, Yoshihiro</creator><creator>Niitsu, Yoshiro</creator><general>Blackwell Publishing Ltd</general><general>Japanese Cancer Association</general><general>John Wiley & Sons, Inc</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>8FE</scope><scope>8FH</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>LK8</scope><scope>M7P</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>199001</creationdate><title>Kinetics of Internalization and Cytotoxicity of Transferrin‐Neocarzinostatin Conjugate in Human Leukemia Cell Line, K562</title><author>Kohgo, Yutaka ; Kondo, Hitoshi ; Kato, Junji ; Sasaki, Katsunori ; Tsushima, Nobuyasu ; Nishisato, Takuji ; Hirayama, Michiaki ; Fujikawa, Koshi ; Shintani, Naoaki ; Mogi, Yoshihiro ; Niitsu, Yoshiro</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c5981-bcc9d3b5e4b5766fcfd28c46eb521416731fe437f906f040905ed7c7146cb22a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1990</creationdate><topic>Antibiotics, Antineoplastic - metabolism</topic><topic>Antineoplastic agents</topic><topic>Azides - pharmacology</topic><topic>Biological and medical sciences</topic><topic>Cell surface</topic><topic>Centrifugation</topic><topic>Centrifugation, Density Gradient</topic><topic>Conjugate</topic><topic>Cytotoxicity</topic><topic>DNA biosynthesis</topic><topic>Endocytosis</topic><topic>Humans</topic><topic>Internalization</topic><topic>Iodine Radioisotopes</topic><topic>Kinetics</topic><topic>Leukemia</topic><topic>Leukemia, Erythroblastic, Acute - drug therapy</topic><topic>Leukemia, Erythroblastic, Acute - metabolism</topic><topic>Lysosomes</topic><topic>Lysosomes - metabolism</topic><topic>Medical sciences</topic><topic>Myeloid leukemia</topic><topic>Neocarzinostatin</topic><topic>Pharmacology. Drug treatments</topic><topic>Propionic acid</topic><topic>Protein turnover</topic><topic>Radioactivity</topic><topic>Receptors, Transferrin - metabolism</topic><topic>Sodium Azide</topic><topic>Targetting</topic><topic>Transferrin</topic><topic>Transferrin - metabolism</topic><topic>Transferrin - therapeutic use</topic><topic>Transferrin receptor</topic><topic>Transferrin receptors</topic><topic>Tumor cells</topic><topic>Tumor Cells, Cultured</topic><topic>Zinostatin - metabolism</topic><topic>Zinostatin - therapeutic use</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kohgo, Yutaka</creatorcontrib><creatorcontrib>Kondo, Hitoshi</creatorcontrib><creatorcontrib>Kato, Junji</creatorcontrib><creatorcontrib>Sasaki, Katsunori</creatorcontrib><creatorcontrib>Tsushima, Nobuyasu</creatorcontrib><creatorcontrib>Nishisato, Takuji</creatorcontrib><creatorcontrib>Hirayama, Michiaki</creatorcontrib><creatorcontrib>Fujikawa, Koshi</creatorcontrib><creatorcontrib>Shintani, Naoaki</creatorcontrib><creatorcontrib>Mogi, Yoshihiro</creatorcontrib><creatorcontrib>Niitsu, Yoshiro</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>ProQuest Central Student</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Biological Science Collection</collection><collection>Biological Science Database</collection><collection>Access via ProQuest (Open Access)</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><collection>PubMed Central (Full Participant titles)</collection><jtitle>Cancer science</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kohgo, Yutaka</au><au>Kondo, Hitoshi</au><au>Kato, Junji</au><au>Sasaki, Katsunori</au><au>Tsushima, Nobuyasu</au><au>Nishisato, Takuji</au><au>Hirayama, Michiaki</au><au>Fujikawa, Koshi</au><au>Shintani, Naoaki</au><au>Mogi, Yoshihiro</au><au>Niitsu, Yoshiro</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Kinetics of Internalization and Cytotoxicity of Transferrin‐Neocarzinostatin Conjugate in Human Leukemia Cell Line, K562</atitle><jtitle>Cancer science</jtitle><addtitle>Jpn J Cancer Res</addtitle><date>1990-01</date><risdate>1990</risdate><volume>81</volume><issue>1</issue><spage>91</spage><epage>99</epage><pages>91-99</pages><issn>0910-5050</issn><issn>1347-9032</issn><eissn>1349-7006</eissn><eissn>1876-4673</eissn><coden>GANNA2</coden><abstract>ABSTRACT
Human serum transferrin was conjugated with an anticancer‐active polypeptide, neocarzinostatin, by using N‐succinimidy1‐3‐(2‐pyridyldithio)propionate. The conjugate consisted of 1.8 mol of neocarzinostatin per 1 mol of transferrin on average and retained cytotoxic activity against human tumor cells. This conjugate was capable of binding to the transferrin receptor of human myelogenous leukemia K562 cells and was internalized by endocytosis. The LD50 values of the conjugate and neocarzinostatin alone in the presence of excess native bovine transferrin were 0.20 μ/ml and 1.80 μ/ml, respectively, suggesting that the effect of the conjugate was greater than that of neocarzinostatin alone. A pulse‐chase experiment using 125I‐labeled conjugate revealed that 25% of the internalized conjugate was degraded in lysosomes and the rest was recycled back to the cell surface without degradation. About 75% of this conjugate recycled back to the cell surface in 18.3 min (3.4 min for receptor binding and 14.9 min for recycling to the cell surface through the acidosomes), while the rest was delivered from the cell surface to the lysosome in 19.6 min. This phenomenon was confirmed by chasing the radioactivity in subcellular fractions separated by Percoll density gradient centrifugation. Therefore, it was concluded that this conjugate is internalized specifically by transferrin receptors and is at least partly transferred to and accumulated in lysosomal compartments, resulting in the inhibition of cellular DNA synthesis.</abstract><cop>Oxford, UK</cop><pub>Blackwell Publishing Ltd</pub><pmid>2139019</pmid><doi>10.1111/j.1349-7006.1990.tb02512.x</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | Cancer science, 1990-01, Vol.81 (1), p.91-99 |
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| source | MEDLINE; Access via Wiley Online Library; IngentaConnect Free/Open Access Journals; EZB-FREE-00999 freely available EZB journals; PubMed Central |
| subjects | Antibiotics, Antineoplastic - metabolism Antineoplastic agents Azides - pharmacology Biological and medical sciences Cell surface Centrifugation Centrifugation, Density Gradient Conjugate Cytotoxicity DNA biosynthesis Endocytosis Humans Internalization Iodine Radioisotopes Kinetics Leukemia Leukemia, Erythroblastic, Acute - drug therapy Leukemia, Erythroblastic, Acute - metabolism Lysosomes Lysosomes - metabolism Medical sciences Myeloid leukemia Neocarzinostatin Pharmacology. Drug treatments Propionic acid Protein turnover Radioactivity Receptors, Transferrin - metabolism Sodium Azide Targetting Transferrin Transferrin - metabolism Transferrin - therapeutic use Transferrin receptor Transferrin receptors Tumor cells Tumor Cells, Cultured Zinostatin - metabolism Zinostatin - therapeutic use |
| title | Kinetics of Internalization and Cytotoxicity of Transferrin‐Neocarzinostatin Conjugate in Human Leukemia Cell Line, K562 |
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