Bioconjugale
Chemistry SEPTEMBER/OCTOBER 1992 Volume 3, Number 5 0 Copyright 1992 by the American Chemical Society
REVIEWS Conjugates of Anticancer Agents and Polymers: Advantages of Macromolecular Therapeutics in Vivo Hiroshi Maeda,'p+ Len W. Seymour,tli and Yoichi Miyamotot Department of Microbiology, Kumamoto University School of Medicine, 2-2-1 Honjo, Kumamoto 860, Japan, and Cancer Research Campaign's Polymer-Controlled Drug Delivery Group, Department of Biological Sciences, Keele University, Keele, Staffordshire ST5 5BG, U.K. Received June 11, 1992 1. INTRODUCTION
There is a pressing need for the development of more effective and yet less toxic drugs for the treatment of diseases such as cancer and AIDS. The use of sophisticated macromolecular drugs offers many new therapeutic strategies (I+), although so far investigation and development of such materials has been surprisingly limited. Synthetic macromolecular therapeutic agents have never previously been used in clinical practice, although naturally-occurring macromolecules have been used routinely. Immunoglobulins, growth hormones, insulin, interferons, plasma albumin, fibrinogen, plasminogen activator, heparins, chondroitin sulfate, etc., are all widely used, and their basic therapeutic principle is for the supplementation of deficient patients. Hence molecular size alone should not be a factor prohibiting the development of synthetic macromolecular drugs. One reason for the slow development is probably a lack of perception of the potential advantages and therapeutic benefits, although another may be connected with the perceived complexity and diverse chemical structures of the materials involved. Interest in synthetic polymer-drug conjugates can be traced back before the 19509, but the understanding of pharmacology, polymer chemistry, subcellular/cellular biology, and purification technology was then so poorly developed that the precise requirements for a macromolecular drug could be neither identified nor met at that
* Author to whom correspondence should be addressed. +
Kumamoto University School of Medicine.
* Keele University.
5 Current address: Department of Clinical Oncology, Queen Elizabeth Hospital, University of Birmingham, Edgbaston, Birmingham B15 2TH, U.K.
1043-1802/92/2903-0351$03.00/0
time. One exception may be polyvinylpyrrolidone iodine complex (Isodine) (61,which was developed as a topical antiseptic agent and even now remains one of the best antiseptics available. In recent years great advances have been made in pharmacology,purification technology,and allied sciences, and the development of sophisticated macromolecular drugs is now a real possibility. As the clinical benefits of this new approach become gradually established, the field is likely to achieve an increasing momentum of growth. Macromolecular anticancer drugs are usually either conjugates of proteins with polymers (e.g. SMANCS') or immunoglobulins (e.g. antibody-ricin conjugates), or conjugates of low molecular weight drugs with synthetic polymers (e.g. HPMA) or proteins (e.g. IgG or albumin). The common aspects of the polymer conjugates are that the polymers function as carriers or stabilizers, frequently resulting in decreased drug toxicity, altered biodistribution, and mostly increased therapeutic efficacy. Firstly we shall review mechanistic principles of tumor targeting of polymer-drug conjugates in relation to the pathophysiology of tumor tissue, followed by brief comments on representative polymer-conjugated anticancer drugs. ~~
Abbreviations used: SMA, styrene-maleic acidlanhydride copolymer;SMANCS,poly(styrene-co-maleicacid n-butyl ester)conjugated neocarzinostatin; HPMA, N-(2-hydroxypropyl)methacrylamide copolymer; t l p , plasma half-life in vivo; NCS, neocarzinostatin; EPR effect, enhanced permeability and retention effect; HMKG, high molecular weight kininogen; Hyp3BK, [hydro~yprolyl~l bradykinin; PEG, polyethyleneglycol; SOD, superoxide dismutase; DIVEMA-NCS, divinylether-maleic acid copolymer- or pyran copolymer-conjugated NCS; LDm, dose giving 50% lethality; A7-NCS, an anti-human colorectal carci0 1992 American Chemical Society
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Table I. Plasma Clearance Time of Various Proteins, Polymer-Conjugated or Modified Proteins, and Synthetic Polymers protein or polymer type of polymer or modification daltons X 10-3 tlp tll10 test animal refs none 12 neocarzinostatin (NCS) 1.8min 15min mouse 8.70 19 min SMA"-conjugatedNCS 16 5h mouse 8,70 SMANCS 13.7 30 min 5 min 109 none mouse ribonuclease 18 min 109 cross-linked 5h mouse 27 ribonuclease dimer 80 min rabbit dextran 127 47,48 dextran-SBTI 5 min 34 min DTPACPICr mouse 29 8 ovomucoid 5 min 30 min rat 32 8 Cuz+,Zn2+superoxide dismutase (SOD) none SMA conjugate >5h 40 >10 h rat 9 SOD-SMA" divinyl ether-maleic acid conjugate 43 30 min >10 h mouse 71 SOD-DIVEMAd 3h polyvinyl alcohol (low mol wt) 67 >10 h SOD-PVAL~ mouse UD 7.8 h >15 h polyvinyl alcohol (high mol wt) mouse 118 UD SOD-PVAH~ 25 min 66 min succinyl gelatin mouse 92 UD SOD-suc-gelating 15 min 74 min rat 50 49 none bilirubin oxidase 5h 70 48 h PEG PEGh-bilirubin oxidase rat 49 3-4 days' 68 none serum albumin mouse 8 2h Evans blue dye serum albumin mouse 30 h 8, 11 25 min 4h formaldehyde-modified serum albumin formaldehyde/125I rat 110 65 X (2-8) 1.5-3.4 h rat none 42 L-asparaginase PEGz-linked 56 h 11days mouse >500 L-asparaginase-PEG 42 iodination/1251 72 h 148 HPMA mouse UD iodination/1251 25 h HPMA mouse >72 556 UD DTPA/Wr 60 h immunoglobulin G 150 rat 8 iodination/1251 140 h az-macroglobulin 180 X 4 22 days mouse 111 iodination/1251 2.5 min 180 X 2 20 min mouse az-macroglobulin-plasmin complex 111 0 SMA, styrene-maleicacid/anhydridecopolymer butylester, Mr 1600. It confersalbumin binding capacity. SBTI, Kunitz type. DIVEMA, divinyl ether-maleic acid (pyran) copolymer, Mr 5600. PVAL,polyvinyl alcohol, Mr 4500. e PVAH,polyvinyl alcohol, Mr 11200. Suc-gelatin, succinylatedgelatin,Mr 10 000.8 PEG, polyethylene glycol,Mr 6000. DTPA, diethylenetriaminepentaacetic acid. Human albumin in humans: 19 days. j HPMA, N-(2-hydroxypropyl)methacrylamidecopolymer. UD, unpublished data. ~~
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2. PLASMA HALF-LIFE OF MACROMOLECULAR DRUGS
The profile of plasma concentration of drugs is generally of great pharmacological and therapeutic significance. To attain a high concentration in any peripheral target tissue or organ following systemic administration, a high plasma concentration (generally measured as the area under the clearance curve) is essential (1-3). Consequently it is important that the material demonstrates good blood and tissue compatibility. For this purpose a neutral or slightly negative electric charge appears to be optimal since polycationic polymers are rapidly captured by the first pass effect and also during circulation (7). The reason for this is that the endothelial surfaces of the blood vessels are covered with negatively charged components such as chondroitin sulfate, heparan sulfate, and glycocalyx. The bioadhesiveness of polycationic drugs may make them more suitable for topical or local application than for intravenous injection. Molecular size is another important parameter. Small proteins less than 40 000 Da are cleared rapidly into the urine, with a plasma half-life in vivo (tip) of usually less than 5 min in mice (e.g. superoxide dismutase, 30 kDa) (1-4,8, 9) (Table I). On the other hand, low molecular weight drugs which bind to albumin or other large plasma proteins exhibit relatively high plasma concentrations for prolonged periods. Conjugation of the small anticancer protein neocarzinostatin (NCS, 12 kDa) with two copolymers of styrene-maleic acid/anhydride (SMA, 1.6 kDa), a conjugate designated SMANCS, has a molecular mass noma monoclonal antibody A7-conjugated NCS; TIC, a ratio of the value obtained in the treatment group to that in the control group; DOX, doxorubicin; OXD-DOX, oxidized dextran-DOX conjugate;DIVEMA-DOX, divinyl ether-maleic acid copolymerDOX conjugate; MMC-D, mitomycin C-dextran conjugate; MMC-DCat,cationic MMC-D; MMC-D,, anionic MMC-D.
of only 15.5 kDa. However, conjugation results in a prolongation of plasma tl/z in mice from 1.9 min for NCS to 20 min for the conjugate (8, 91,as a result of binding of SMANCS to albumin (10). Systematic studies using plasma proteins or synthetic polymers with various molecular weights have shown clearly that noncationic materials larger in size than the renal threshold frequently display extended plasma circulation times (11, 12, and unpublished data, Seymour et al.). 3. INCREASED PERMEABILITY OF TUMOR VASCULATURE
In inflammatory conditions the permeability of the blood vessels is greatly increased by factors acting on endothelial cells and opening the tight intercellular junctions. These factors include agents such as bradykinin, histamine, prostaglandins, and tumor necrosis factor. This has also been shown to be the case with certain microbial infections, where the bradykinin-generating cascade is activated and hence edema is formed (13). In tumor tissue there are at least two substances known to be involved in modulation of vascular permeability. Vascular permeability factor (VPF), initially described by Dvorak et al. (14,15),is a protein of about 38 kDa with sequence homology to platelet derived growth factor (PDGF) (16,17).VPF is produced by a range of cancer cells and also by pituitary follicular cells (18, 19). This factor possesses endothelial growth (angiogenic) activity and mitogenic activity in vivo, and is thought to be involved in a range of physiological processes such as inflammation and wound healing (20,211. Bradykinin (or kinin) is generated from high molecular weight kininogen (HMKG) by limited proteolysis of kallikrein (22-24). Kallikrein is generated from prekallikrein, a proenzyme, by the action of activated Hageman factor (also called clotting factor XII). The cascade of the proteolytic reactions to generate bradykinin is shown in
Bioconlugafe Chem., Vol. 3, No. 5, 1992 353
Review
trypsin inhibitor
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Figure 1. Kinin-generatingand -degrading cascade and points of inhibitionby various inhibitors: CPNI,carboxypeptidase N inhibitor; ACEI, angiotensin converting enzyme inhibitor (from ref 23 with permission). Bradykinin
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Table 11. Accumulation of ‘%-Labeled Iodinated Fatty Acida
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tumor 1252.58 130.94 muscle 17.02 skin liver (adjacent) 566.25 6.89 mes lymph node liver (remote) 28.95) 4.44 cer lymph nodes sm intestine 1.06 2.02 thymus lung 2.66 2.57 serum kidney 1.61 plasma cells stomach 10.97 1.72 bone marrow heart 2.65 lg intestine 0.35 1.06 urine (exc) spleen 2.39 3.28 urine (vesical) 1.31 bile bladder 0.28 0.46 brain
