The Influence of Carbohydrate Structure on the Clearance of Recombinant Tissue-Type Plasminogen Activator

A Hotchkiss; C J Refino; C K Leonard; J V O'Connor; C Crowley; J McCabe; K Tate; G Nakamura; D Powers; A Levinson; M Mohler; M W Spellman

doi:10.1055/s-0038-1647041

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1988; 60(02): 255-261
DOI: 10.1055/s-0038-1647041

Original Article

Schattauer GmbH Stuttgart

The Influence of Carbohydrate Structure on the Clearance of Recombinant Tissue-Type Plasminogen Activator

A Hotchkiss

¹The Department of Pharmacological Sciences, Genentech, Inc., South San Francisco, CA, USA

,

C J Refino

¹The Department of Pharmacological Sciences, Genentech, Inc., South San Francisco, CA, USA

,

C K Leonard

²The Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, CA, USA

,

J V O'Connor

²The Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, CA, USA

,

C Crowley

³The Department of Cell Genetics, Genentech, Inc., South San Francisco, CA, USA

,

J McCabe

¹The Department of Pharmacological Sciences, Genentech, Inc., South San Francisco, CA, USA

,

K Tate

⁴The Department of Molecular Biology, Genentech, Inc., South San Francisco, CA, USA

,

G Nakamura

²The Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, CA, USA

,

D Powers

⁴The Department of Molecular Biology, Genentech, Inc., South San Francisco, CA, USA

,

A Levinson

⁴The Department of Molecular Biology, Genentech, Inc., South San Francisco, CA, USA

,

M Mohler

¹The Department of Pharmacological Sciences, Genentech, Inc., South San Francisco, CA, USA

,

M W Spellman

⁴The Department of Molecular Biology, Genentech, Inc., South San Francisco, CA, USA

› Author Affiliations

Abstract

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Summary

Modification of the carbohydrate structures of recombinant tissue-type plasminogen activator (rt-PA) can increase or decrease its rate of clearance in rabbits. When rt-PA was treated with sodium periodate to oxidize carbohydrate residues, the rate of clearance was decreased from 9.6 ± 1.9 ml min⁻¹ kg⁻¹ to 3.5 ± 0.6 ml min⁻¹ kg⁻¹ (mean ± SD, n = 5). A similar change in the clearance of rt-PA was introduced by the use of endo-β-N-acetyl- glucosaminidase H (Endo-H), which selectively removes high mannose asparagine-linked oligosaccharides; the clearance of Endo-H-treated rt-PA was 5.0 ± 0.5 ml min⁻¹ kg⁻¹. A mutant of rt-PA was produced with an amino acid substitution at position 117 (Asn replaced with Gin) to remove a potential glycosylation site that normally contains a high mannose structure. The clearance of this material was also decreased, similar to the periodate and Endo-H-treated rt-PA. Conversely, when rt-PA was produced in the CHO 15B cell line, which can produce only high mannose oligosaccharide structures on glycoproteins, the clearance was increased by a factor of 1.8. These results demonstrate that the removal of rt-PA from the blood depends significantly upon the nature of its oligosaccharide structures.

Keywords

Tissue-type plasminogen activator - Plasminogen activator - Pharmacokinetics - CHO 15 B cells - High mannose oligosaccharides

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References

References
1 Hoylaerts M, Rijken DC, Lijnen HR, Collen D. Kinetics of the activation of plasminogen by human tissue plasminogen activator. Role of fibrin.. J Biol Chem 1982; 257: 2912-2919
2 Rijken DC, Hoylaerts M, Collen D. Fibrinolytic properties of one- chain and two-chain human extrinsic (tissue-type) plasminogen activator.. J Biol Chem 1982; 257: 2920-2925
3 Rijken DC, Wijngaards G, Zaal-de JongM, Welbergen J. Purification and partial characterization of plasminogen activator from human uterine tissue.. Biochim Biophys Acta 1979; 580: 140-153
4 Binder BR, Spragg J, Austen KF. Purification and characterization of human vascular plasminogen activator derived from blood vessel perfusates.. J Biol Chem 1979; 254: 1998-2003
5 Rijken DC, Collen D. Purification and characterization of the plasminogen activator secreted by human melanoma cells in culture.. J Biol Chem 1981; 256: 7035-7041
6 Wallen P, Pohl G, Bergsdorf N, Ranby M, Ny T, Jornvall H. Purification and characterization of a melanoma cell plasminogen activator.. Eur J Biochem 1983; 132: 681-686
7 Pennica D, Holmes WE, Kohr WJ, Harkins RN, Vehar GA, Ward CA, Bennett WF, Yelverton E, Seeburg PH, Heyneker HL, Goeddel DV, Collen D. Cloning and expression of human tissue-type plasminogen activator cDNA in E. coli. . Nature (London) 1983; 301: 214-221
8 Williams DO, Borer J, Braunwald E, Chesebro JH, Cohen LS, Dalen J, Dodge HT, Francis CK, Knatterud G, Ludbrook P, Markis JE, Mueller H, Desvigne-Nickens P, Passamani ER, Powers ER, Rao AK, Roberts R, Ross A, Ryan TJ, Sobel BE, Winniford M, Zaret B. Intravenous recombinant tissue-type plasminogen activator in patients with acute myocardial infarction: a report from the NHLBI thrombolysis in myocardial infarction trial. Circulation 1986; 73: 338-346
9 Graor RA, Risius B, Lucas FV, Young JR, Ruschhaupt WF, Beven EG, Grossbard EB. Thrombolysis with recombinant human tissue- type plasminogen activator in patients with peripheral artery and bypass graft. Circulation 1986; 74 (suppl 1) 115-120
10 Collen D, Bounameaux H, De CockF, Lijnen HR, Verstraete M. Analysis of coagulation and fibrinolysis during intravenous infusion of recombinant human tissue-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986; 73: 511-517
11 The thrombolysis in myocardial infarction (TIMI) trial. Phase I findings. TIMI Study Group. New Engl J Med 1985; 312: 932-936
12 Risius B, Graor RA, Geisinger MA, Zelch MG, Lucas FV, Young JR, Grossbard EB. Recombinant human tissue-type plasminogen activator for thrombolysis in peripheral arteries and bypass grafts.. Radiology 1986; 160: 183-188
13 Goldhaber SZ, Vaughan DE, Markis JE, Selwyn AP, Meyerovitz MF, Loscalzo J, Kim DS, Kessler CM, Dawley DL. et al Acute pulmonary embolism treated with tissue plasminogen activator.. Lancet 1986; 2: 886-899
14 Korninger C, Stassen JM, Collen D. Turnover of human extrinsic (tissue-type) plasminogen activator in rabbits.. Thromb Haemostas 1981; 46: 658-661
15 Nilsson S, Einarsson M, Ekvarn S, Haggroth L, Mattsson CH. Turnover of tissue plasminogen activator in normal and hepatectomized rabbits.. Thromb Res 1985; 39: 511-521
16 Bounameaux H, Stassen JM, Seghers C, Collen D. Influence of fibrin and liver blood flow on the turnover and the systemic fibrinogenolytic effects of recombinant human tissue-type plasminogen activator in rabbits.. Blood 1986; 67: 1493-1497
17 Baughman RA. Pharmacokinetics of tissue plasminogen activator.. In: Tissue Plasminogen Activator in Thrombolytic Therapy. Sobel B, Grossbard E, Collen D. (eds) Marcel Dekker; New York : 1987. pp 41-53
18 Verstraete M, Bounameaux H, De CockF, Van De WerfF, Collen D. Pharmacokinetics and systemic fibrinogenolytic effects of recombinant human tissue-type plasminogen activator (rt-PA) in humans.. J Pharm Exp Ther 1985; 235: 506-512
19 Ashwell G, Hartford J. Carbohydrate-specific receptors in the liver.. Annu Rev Biochem 1982; 51: 531-554
20 Spellman MW, Ferguson CK, O’Connor JV. Characterization of N-linked oligosaccharides from recombinant tissue-type plasminogen activator. XHIth Int Carbohydrate Symposium, Abstr B122. 1986
21 Pohl G, Kenne L, Nilsson B, Einarsson M. Isolation and characterization of three different carbohydrate chains from melanoma tissue plasminogen activator.. Eur J Biochem 1987; 170: 69-75
22 Fuchs HE, Berger H, Pizzo SV. Hepatic clearance of tissue-type plasminogen activator in rats.. Blood 1985; 65: 539-544
23 Emeis JJ, van den HoogenCM, Jense D. Hepatic clearance of tissue-type plasminogen activator in rats.. Thromb Haemostas 1985; 54: 661-664
24 Einarsson M, Smedsrod B, Pertoft H. Endocytosis of tissue plasminogen activator (t-PA) by rat liver cells.. Thromb Haemostas 1985; 54: 270 (Abstr).
25 Bakhit C, Lewis D, Billings R, Malfroy B. Cellular catabolism of recombinant tissue-type plasminogen activator. Identification and characterization of a novel high affinity uptake system on rat hepatocytes.. J Biol Chem 1987; 262: 8716-8720
26 Li E, Kornfeld S. Structure of the altered oligosaccharide present in glycoproteins from a clone of Chinese hamster ovary cells deficient in N-acetylglucosaminyltransferase activity.. J Biol Chem 1978; 253: 6426-6431
27 Graham FL, Van Der EBA. A new technique for the assay of infectivity of human adenovirus 5 DNA.. Virology 1973; 52: 456-467
28 Albersheim P, Nevins DJ, English PD, Karr A. A method for the analysis of sugars in plant cell-wall polysaccharides by gas-liquid chromatography. Carbohydrate Res 1967; 5: 340-345
29 Nesser JR, Schweizer JF. A quantitative determination by capillary gas-liquid chromatography of neutral and amino sugars (as o-methyloxime acetates), and a study on hydrolytic conditions for glycoproteins and polysaccharides in order to increase sugar recoveries.. Anal Biochem 1984; 142: 58-67
30 Mohler MA, Refino CJ, Chen SA, Chen AB, Hotchkiss AJ. D-Phe-Pro-Arg-chloromethylketone: its potential use in inhibiting the formation of in vitro artifacts in blood collected during tissue-type plasminogen activator thrombolytic therapy.. Thromb Haemostas 1986; 56: 160-164
31 Benninger A, Baughman B. FEATHER, an interactive curve stripping program. Pharm Res 1986; 3: 120s
32 Gibaldi M, Perrier D. Pharmacokinetics.. Marcel Dekker Inc.; New York : 1982. 2nd ed, pp 433-444
33 Tolleshaug H, Berg T, Blomhoff R. Uptake of mannose-terminated glycoproteins in isolated rat liver cells. Evidence for receptor-mediated endocytosis in hepatocytes.. Biochem J 1984; 223: 151-160
34 Collen D, Tytgat G, Verstraete M. Semi-automatic method for routine evaluation of fibrinolytic components.. J Clin Pathol 1968; 21: 705-707
35 Collen D, Stassen JM, Marafino BJ, Builder S, DeCock F, Ogez J, Tajiri D, Pennica D, Bennett WF, Salwa J. et al. Biological properties of human tissue-type plasminogen activator obtained by expression of recombinant DNA in mammalian cells.. J Pharm Exp Ther 1984; 231: 146-152
36 Mohler MA, Tate K, Bringman TS, Fuller G, Keyt B, Vehar G, Hotchkiss AJ. Circulatory metabolism of recombinant tissue-type plasminogen activator in monkeys and rabbits. Fibrinolysis 1988; 2: 17-23