Creation and biochemical analysis of a broad-specific claudin binder

Takahashi, Azusa; Saito, Yumiko; Kondoh, Masuo; Matsushita, Kyohei; Krug, Susanne M.; Suzuki, Haruo; Tsujino, Hirofumi; Li, Xiangru; Aoyama, Hiroshi; Matsuhisa, Koji; Uno, Tadayuki; Fromm, Michael; Hamakubo, Takao; Yagi, Kiyohito

doi:10.1016/j.biomaterials.2012.01.017

Cited by 47 publications

(61 citation statements)

References 60 publications

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“…6. Flow cytometric analysis of Interaction between m19 and cL-Expressing L cells 44) CL-expressing L cells were treated with C-CPE or m19; labeled secondary antibodies were used to visualize bound C-CPE or m19 through flow cytometry. Solid histograms indicate the level of reactivity of c-cPE or m19; dotted histograms indicate background binding of the anti-His tag and FITc-labeled antibodies without c-cPE or m19.…”

Section: Developing CL Bindersmentioning

confidence: 99%

“…44) Because understanding the mode of action of m19 would yield information useful in the development of cL binders, we performed X-ray structural analysis to determine the structures and electrostatic surface maps of m19 and c-cPE. Whereas m19 and c-cPE were structurally similar, the electrostatic maps of their cL-binding regions were greatly different.…”

Section: Developing CL Bindersmentioning

confidence: 99%

“…The binding region of c-cPE is negatively charged, whereas that of m19 is positively charged. 44) analysis of a series of chimeras revealed that sensitivity to cPE was determined by the region from asn149 to Met160 in cLs (that is, the cPE sensitivity-related region; cPE-SR).…”

Section: Developing CL Bindersmentioning

confidence: 99%

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Recent Advances in Claudin-Targeting Technology

Nagase

Doyama

Yagi

et al. 2013

Biological & Pharmaceutical Bulletin

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Most malignant tumors are derived from epithelium, and pathologic microorganisms often invade the body through the mucosal epithelium. Thus epithelial tissues are potent targets for drug delivery. The tight junction (TJ) is the intercellular seal in epithelial cell sheets. Claudins (CLs) are a family of tetratransmembrane proteins with a molecular mass of approximately 23 kDa. CLs are key structural and sealing components of TJs. CLs are often overexpressed in malignant tumors. CL-4 is highly expressed in the epithelial cells covering mucosal immune tissues. Therefore CLs may be potent targets for drug delivery, cancer therapy, and mucosal vaccination. Herein, we overview a series of our studies using the C-terminal fragment of Clostridium perfringens enterotoxin to target and bind CLs; we also discuss the efficacy of CL-targeted drug delivery.

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Section: Developing CL Bindersmentioning

confidence: 99%

Section: Developing CL Bindersmentioning

confidence: 99%

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Recent Advances in Claudin-Targeting Technology

Nagase

Doyama

Yagi

et al. 2013

Biological & Pharmaceutical Bulletin

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“…We previously created m19, which can bind to CLDN-1/-2/-4/-5, by using the CLDN-3/-4-binding polypeptide C-CPE as a template (Takahashi et al, 2012). m19 tagged with histidine at the N terminus was prepared as described.…”

Section: Methodsmentioning

confidence: 99%

“…One, a C-CPE mutant polypeptide (m19), bound to several CLDNs, including CLDN-1, -2, -4, and -5, and reduced TJ integrity in the intestinal epithelium more than did C-CPE (Takahashi et al, 2012). We also created four anti-CLDN-1 monoclonal antibodies (mAbs; clones 2C1, 3A2, 5F2, 7A5) and have shown that they all bound CLDN-1, but not CLDN-2-9, and prevented CLDN-1-mediated hepatitis C virus infection in a human hepatocyte cell line (Fukasawa et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

Claudin-1 Binder Enhances Epidermal Permeability in a Human Keratinocyte Model

Nakajima

Nagase

Iida

et al. 2015

J Pharmacol Exp Ther

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Tight junctions (TJs) are complex biochemical structures that seal the intercellular space and prevent the free movement of solutes across epithelial cell sheets. Modulating the TJ seal is a promising option for increasing the transdermal absorption of drugs. Within TJs, the binding of the claudin (CLDN) family of tetratransmembrane proteins through cis-and trans-interactions is an integral part of seal formation. Because epidermal TJs contain CLDN-1 and CLDN-4, a binder for these CLDNs may be a useful modulator of the permeability of the epidermal barrier. Here, we investigated whether m19, which can bind to CLDN-1/-4 (also CLDN-2/-5), modulates the integrity of epidermal TJs and the permeability of cell sheets to solutes. Treatment of normal human epidermal keratinocytes (NHEKs) with the CLDN binder reduced the integrity of TJs. A CLDN-1-specific binder (a monoclonal antibody, clone 7A5) also weakened the TJ seal in NHEKs. Although m19 attenuated the TJ barrier in human intestinal epithelial cells (Caco-2), 7A5 did not. Treatment of NHEKs with 7A5 enhanced permeation of a paracellular permeation marker. These findings indicate that CLDN-1 is a potential target for modulating the permeability of the epidermis, and that our CLDN-1 binder is a promising candidate molecule for development as a dermal absorption enhancer.

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Targeting claudin‐overexpressing thyroid and lung cancer by modified Clostridium perfringens enterotoxin

et al. 2020

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Clostridium perfringens enterotoxin (CPE) can be used to eliminate carcinoma cells that overexpress on their cell surface CPE receptorsa subset of claudins (e.g., Cldn3 and Cldn4). However, CPE cannot target tumors expressing solely CPE-insensitive claudins (such as Cldn1 and Cldn5). To overcome this limitation, structure-guided modifications were used to generate CPE variants that can strongly bind to Cldn1, Cldn2 and/or Cldn5, while maintaining the ability to bind Cldn3 and Cldn4. This enabled (a) targeting of the most frequent endocrine malignancy, namely, Cldn1-overexpressing thyroid cancer, and (b) improved targeting of the most common cancer type worldwide, non-small-cell lung cancer (NSCLC), which is characterized by high expression of several claudins, including Cldn1 and Cldn5. Different CPE variants, including the novel mutant CPE-Mut3 (S231R/S313H), were applied on thyroid cancer (K1 cells) and NSCLC (PC-9 cells) models. In vitro, CPE-Mut3, but not CPEwt, showed Cldn1dependent binding and cytotoxicity toward K1 cells. For PC-9 cells, CPE-Mut3 improved claudin-dependent cytotoxic targeting, when compared to CPEwt. In vivo, intratumoral injection of CPE-Mut3 in xenograft models bearing K1 or PC-9 tumors induced necrosis and reduced the growth of both tumor types. Thus, directed modification of CPE enables eradication of tumor entities that cannot be targeted by CPEwt, for instance, Cldn1overexpressing thyroid cancer by using the novel CPE-Mut3.

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Creation and biochemical analysis of a broad-specific claudin binder

Cited by 47 publications

References 60 publications

Recent Advances in Claudin-Targeting Technology

Recent Advances in Claudin-Targeting Technology

Claudin-1 Binder Enhances Epidermal Permeability in a Human Keratinocyte Model

Targeting claudin‐overexpressing thyroid and lung cancer by modified Clostridium perfringens enterotoxin

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