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31.
Sutherland MS Sanderson RJ Gordon KA Andreyka J Cerveny CG Yu C Lewis TS Meyer DL Zabinski RF Doronina SO Senter PD Law CL Wahl AF 《The Journal of biological chemistry》2006,281(15):10540-10547
The chimeric anti-CD30 monoclonal antibody cAC10, linked to the antimitotic agents monomethyl auristatin E (MMAE) or F (MMAF), produces potent and highly CD30-selective anti-tumor activity in vitro and in vivo. These drugs are appended via a valine-citrulline (vc) dipeptide linkage designed for high stability in serum and conditional cleavage and putative release of fully active drugs by lysosomal cathepsins. To characterize the biochemical processes leading to effective drug delivery, we examined the intracellular trafficking, internalization, and metabolism of the parent antibody and two antibody-drug conjugates, cAC10vc-MMAE and cAC10vc-MMAF, following CD30 surface antigen interaction with target cells. Both cAC10 and its conjugates bound to target cells and internalized in a similar manner. Subcellular fractionation and immunofluorescence studies demonstrated that the antibody and antibody-drug conjugates entering target cells migrated to the lysosomes. Trafficking of both species was blocked by inhibitors of clathrin-mediated endocytosis, suggesting that drug conjugation does not alter the fate of antibody-antigen complexes. Incubation of cAC10vc-MMAE or cAC10vc-MMAF with purified cathepsin B or with enriched lysosomal fractions prepared by subcellular fractionation resulted in the release of active, free drug. Cysteine protease inhibitors, but not aspartic or serine protease inhibitors, blocked antibody-drug conjugate metabolism and the ensuing cytotoxicity of target cells and yielded enhanced intracellular levels of the intact conjugates. These findings suggest that in addition to trafficking to the lysosomes, cathepsin B and perhaps other lysosomal cysteine proteases are requisite for drug release and provide a mechanistic basis for developing antibody-drug conjugates cleavable by intracellular proteases for the targeted delivery of anti-cancer therapeutics. 相似文献
32.
Doronina SO Mendelsohn BA Bovee TD Cerveny CG Alley SC Meyer DL Oflazoglu E Toki BE Sanderson RJ Zabinski RF Wahl AF Senter PD 《Bioconjugate chemistry》2006,17(1):114-124
We have previously shown that antibody-drug conjugates (ADCs) consisting of cAC10 (anti-CD30) linked to the antimitotic agent monomethylauristatin E (MMAE) lead to potent in vitro and in vivo activities against antigen positive tumor models. MMAF is a new antimitotic auristatin derivative with a charged C-terminal phenylalanine residue that attenuates its cytotoxic activity compared to its uncharged counterpart, MMAE, most likely due to impaired intracellular access. In vitro cytotoxicity studies indicated that mAb-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl-MMAF (mAb-L1-MMAF) conjugates were >2200-fold more potent than free MMAF on a large panel of CD30 positive hematologic cell lines. As with cAC10-L1-MMAE, the corresponding MMAF ADC induced cures and regressions of established xenograft tumors at well tolerated doses. To further optimize the ADC, several new linkers were generated in which various components within the L1 linker were either altered or deleted. One of the most promising linkers contained a noncleavable maleimidocaproyl (L4) spacer between the drug and the mAb. cAC10-L4-MMAF was approximately as potent in vitro as cAC10-L1-MMAF against a large panel of cell lines and was equally potent in vivo. Importantly, cAC10-L4-MMAF was tolerated at >3 times the MTD of cAC10-L1-MMAF. LCMS studies indicated that drug released from cAC10-L4-MMAF was the cysteine-L4-MMAF adduct, which likely arises from mAb degradation within the lysosomes of target cells. This new linker technology appears to be ideally suited for drugs that are both relatively cell-impermeable and tolerant of substitution with amino acids. Thus, alterations of the linker have pronounced impacts on toxicity and lead to new ADCs with greatly improved therapeutic indices. 相似文献
33.
Phil Senter 《Palaeontology》2003,46(3):613-621
The course of the nasolacrimal duct, interdental plate morphology, and most details of tooth and denticle morphology have not previously been described in non–archosauriform reptilkes. Here I describe these details in the Triassic archosauriform Euparkeria capensis. The nasolacrimal canal opens orbitally via a pair of foramina between the lacrimal and prefrontal. The canal arches over the antorbital fenestra, as in archosaurs. The term ‘interdental unit’ is introduced for the unit composed of an interdental septum and its accompanying interdental plate. There is no demarcation between interdental plate and septum in E. capensis. The interdental units are heavily pitted on exposed surfaces. Like teeth, they are implanted in the dental groove and are separate from the surrounding bone and from each other. They are well positioned to serve as spacers between teeth, and to resist sagittal forces on teeth during prey capture. The teeth of E. capensis are labiolingually compressed, except for the nearly conical premaxillary teeth and mesialmost dentary tooth. Lateral teeth are serrated on mesial and distal keels. The denticles are low, rounded, and separated by grooves, and are slightly larger on the distal keel. Tooth morphology suggests carnivorous habits for Euparkeria. 相似文献
34.
Background
The Yellow Cat Member of the Cedar Mountain Formation (Early Cretaceous, Barremian?) of Utah has yielded a rich dinosaur fauna, including the basal therizinosauroid theropod Falcarius utahensis at its base. Recent excavation uncovered a new possible therizinosauroid taxon from a higher stratigraphic level in the Cedar Mountain Formation than F. utahensis.Methodology/Principal Findings
Here we describe a fragmentary skeleton of the new theropod and perform a phylogenetic analysis to determine its phylogenetic position. The skeleton includes fragments of vertebrae, a scapula, forelimb and hindlimb bones, and an ischium. It also includes several well-preserved manual unguals. Manual and pedal morphology show that the specimen is distinct from other theropods from the Cedar Mountain Formation and from previously described therizinosauroids. It is here named as the holotype of a new genus and species, Martharaptor greenriverensis. Phylogenetic analysis places M. greenriverensis within Therizinosauroidea as the sister taxon to Alxasaurus + Therizinosauridae, although support for this placement is weak.Conclusions/Significance
The new specimen adds to the known dinosaurian fauna of the Yellow Cat Member of the Cedar Mountain Formation. If the phylogenetic placement is correct, it also adds to the known diversity of Therizinosauroidea. 相似文献35.
Activation of prodrugs by antibody-enzyme conjugates: a new approach to cancer therapy 总被引:1,自引:0,他引:1
P D Senter 《FASEB journal》1990,4(2):188-193
A new strategy for the delivery of cytotoxic agents to solid tumors is described in which monoclonal antibodies are used as carriers for enzymes to tumor cell surfaces. The enzymes are chosen for their abilities to convert relatively noncytotoxic drug precursors (pro-drugs) into active anticancer drugs. The drugs thus formed can then penetrate into nearby tumor cells, resulting in cell death. A number of prodrugs have been developed that can be transformed into active anti-cancer drugs by enzymes of both mammalian and non-mammalian origin. The enzymes have been shown to localize into tumors when linked to monoclonal antibodies that bind to tumor-associated antigens. In vivo studies indicate that MAb-enzyme/prodrug combinations can result in antitumor activities significantly greater than those of the prodrugs or drugs given alone. This is most likely due to the generation of large amounts of active drug at the tumor site. 相似文献
36.
Identification and activities of human carboxylesterases for the activation of CPT-11, a clinically approved anticancer drug. 总被引:1,自引:0,他引:1
CPT-11 is a clinically approved anticancer drug used for the treatment of advanced colorectal cancer. Upon administration, the carbamate side chain of the drug is hydrolyzed, resulting in the release of SN-38, an agent that has approximately 1000-fold increased cytotoxic activity. Since only a very small percentage of the injected dose of CPT-11 is converted to SN-38, there is a significant opportunity to improve its therapeutic efficacy and to diminish its systemic toxicity by selectively activating the drug within tumor sites. We envisioned that a mAb-human enzyme conjugate for CPT-11 activation would be of interest, particularly since the conjugate would likely be minimally immunogenic, and the prodrug is clinically approved. Toward this end, it was necessary to identify the most active human enzyme that could convert CPT-11 to SN-38. We isolated enzymes from human liver microsomes based on their abilities to effect the conversion and identified human carboxylesterase 2 (hCE-2) as having the greatest specific activity. hCE-2 was 26-fold more active than human carboxylesterase 1 and was 65% as active as rabbit liver carboxylesterase, the most active CPT-11 hydrolyzing enzyme known. The anti-p97 mAb 96.5 was linked to hCE-2, forming a conjugate that could bind to antigen-positive cancer cells and convert CPT-11 to SN-38. Cytotoxicity assays established that the conjugate led to the generation of active drug, but the kinetics of prodrug activation (48 pmol x min(-1) x mg(-1) was insufficient for immunologically specific prodrug activation. These results confirm the importance of hCE-2 for CPT-11 activation and underscore the importance of enzyme kinetics for selective prodrug activation. 相似文献
37.
Cephalosporin prodrugs of paclitaxel for immunologically specific activation by L-49-sFv-beta-lactamase fusion protein 总被引:3,自引:0,他引:3
Vrudhula VM Kerr DE Siemers NO Dubowchik GM Senter PD 《Bioorganic & medicinal chemistry letters》2003,13(3):539-542
Paclitaxel conjugates of 7-phenylacetamidocephalosporanic acid were prepared as prodrugs for site specific activation by targeted beta-lactamase. Immunologically specific activation of the prodrug 5 containing 3,3-dimethyl-4-amino-butyric acid as linker in combination with the fusion protein L-49-sFv-beta-lactamase was demonstrated in vitro on 3677 melanoma cells. 相似文献
38.
Reductively activated disulfide prodrugs of paclitaxel 总被引:1,自引:0,他引:1
Vrudhula VM MacMaster JF Li Z Kerr DE Senter PD 《Bioorganic & medicinal chemistry letters》2002,12(24):571-3594
A series of unsymmetrical polar disulfide prodrugs 2–5 of paclitaxel were designed and synthesized as reductively activated prodrugs. These compounds behaved as prodrugs in vitro on L2987 lung carcinoma cells. In vivo evaluation in mice demonstrated that the mutual prodrug 5 with captopril exhibited significant regressions and cures. 相似文献
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40.
Epithelial invasion and cell lysis by virulent strains of Streptococcus suis is enhanced by the presence of suilysin 总被引:18,自引:0,他引:18
Pamela M Nortona Charlotte Rolpha Philip N Warda Robert W Bentleya James A Leigha 《FEMS immunology and medical microbiology》1999,26(1):25-35
Streptococcus suis is an important pathogen of pigs causing arthritis, pneumonia and meningitis and is an occupational disease of farmers and those in the meat industry. As with other streptococci, both virulent and avirulent strains of S. suis are frequently carried asymptomatically in the tonsillar crypts and nasal cavities. Little is known about the process by which virulent strains cross the mucosal epithelia to generate systemic disease and whether this process requires expression of specific bacterial virulence factors. Although putative virulence factors have been postulated, no specific role in the disease process has yet been demonstrated for these factors. This study is the first demonstration that virulent strains of S. suis both invade and lyse HEp-2 cells, a continuous laryngeal epithelial cell line, and that at least one bacterial virulence factor, suilysin, is involved in this process. 相似文献