Research

11 pages, 3473 KiB

Open AccessArticle

O-Glycan-Dependent Interaction between MUC1 Glycopeptide and MY.1E12 Antibody by NMR, Molecular Dynamics and Docking Simulations

by Ryoka Kokubu, Shiho Ohno, Hirohide Kuratani, Yuka Takahashi, Noriyoshi Manabe, Hiroki Shimizu, Yasunori Chiba, Kaori Denda-Nagai, Makoto Tsuiji, Tatsuro Irimura and Yoshiki Yamaguchi

Int. J. Mol. Sci. 2022, 23(14), 7855; https://doi.org/10.3390/ijms23147855 - 16 Jul 2022

Cited by 3 | Viewed by 1802

Abstract

Anti-mucin1 (MUC1) antibodies have been widely used for breast cancer diagnosis and treatment. This is based on the fact that MUC1 undergoes aberrant glycosylation upon cancer progression, and anti-MUC1 antibodies differentiate changes in glycan structure. MY.1E12 is a promising anti-MUC1 antibody with a [...] Read more.

Anti-mucin1 (MUC1) antibodies have been widely used for breast cancer diagnosis and treatment. This is based on the fact that MUC1 undergoes aberrant glycosylation upon cancer progression, and anti-MUC1 antibodies differentiate changes in glycan structure. MY.1E12 is a promising anti-MUC1 antibody with a distinct specificity toward MUC1 modified with an immature O-glycan (NeuAcα(2-3)Galβ(1-3)GalNAc) on a specific Thr. However, the structural basis for the interaction between MY.1E12 and MUC1 remains unclear. The aim of this study is to elucidate the mode of interaction between MY.1E12 and MUC1 O-glycopeptide by NMR, molecular dynamics (MD) and docking simulations. NMR titration using MUC1 O-glycopeptides suggests that the epitope is located within the O-linked glycan and near the O-glycosylation site. MD simulations of MUC1 glycopeptide showed that the O-glycosylation significantly limits the flexibility of the peptide backbone and side chain of the O-glycosylated Thr. Docking simulations using modeled MY.1E12 Fv and MUC1 O-glycopeptide, suggest that V_H mainly contributes to the recognition of the MUC1 peptide portion while V_L mainly binds to the O-glycan part. The V_H/V_L-shared recognition mode of this antibody may be used as a template for the rational design and development of anti-glycopeptide antibodies. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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18 pages, 11509 KiB

Open AccessArticle

MG-Pe: A Novel Galectin-3 Ligand with Antimelanoma Properties and Adjuvant Effects to Dacarbazine

by Stellee M. P. Biscaia, Cassiano Pires, Francislaine A. R. Lívero, Daniel L. Bellan, Israel Bini, Silvina O. Bustos, Renata O. Vasconcelos, Alexandra Acco, Marcello Iacomini, Elaine R. Carbonero, Martin K. Amstalden, Fábio R. Kubata, Richard D. Cummings, Marcelo Dias-Baruffi, Fernanda F. Simas, Carolina C. Oliveira, Rilton A. Freitas, Célia Regina Cavichiolo Franco, Roger Chammas and Edvaldo S. Trindade

Int. J. Mol. Sci. 2022, 23(14), 7635; https://doi.org/10.3390/ijms23147635 - 11 Jul 2022

Cited by 2 | Viewed by 1869

Abstract

Melanoma is a highly metastatic and rapidly progressing cancer, a leading cause of mortality among skin cancers. The melanoma microenvironment, formed from the activity of malignant cells on the extracellular matrix and the recruitment of immune cells, plays an active role in the [...] Read more.

Melanoma is a highly metastatic and rapidly progressing cancer, a leading cause of mortality among skin cancers. The melanoma microenvironment, formed from the activity of malignant cells on the extracellular matrix and the recruitment of immune cells, plays an active role in the development of drug resistance and tumor recurrence, which are clinical challenges in cancer treatment. These tumoral metabolic processes are affected by proteins, including Galectin-3 (Gal-3), which is extensively involved in cancer development. Previously, we characterized a partially methylated mannogalactan (MG-Pe) with antimelanoma activities. In vivo models of melanoma were used to observe MG-Pe effects in survival, spontaneous, and experimental metastases and in tissue oxidative stress. Analytical assays for the molecular interaction of MG-Pe and Gal-3 were performed using a quartz crystal microbalance, atomic force microscopy, and contact angle tensiometer. MG-Pe exhibits an additive effect when administered together with the chemotherapeutic agent dacarbazine, leading to increased survival of treated mice, metastases reduction, and the modulation of oxidative stress. MG-Pe binds to galectin-3. Furthermore, MG-Pe antitumor effects were substantially reduced in Gal-3/KO mice. Our results showed that the novel Gal-3 ligand, MG-Pe, has both antitumor and antimetastatic effects, alone or in combination with chemotherapy. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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15 pages, 3692 KiB

Open AccessArticle

Proscillaridin A Sensitizes Human Colon Cancer Cells to TRAIL-Induced Cell Death

by Manami Semba, Shinji Takamatsu, Sachiko Komazawa-Sakon, Eiji Miyoshi, Chiharu Nishiyama, Hiroyasu Nakano and Kenta Moriwaki

Int. J. Mol. Sci. 2022, 23(13), 6973; https://doi.org/10.3390/ijms23136973 - 23 Jun 2022

Cited by 1 | Viewed by 1698

Abstract

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytotoxic cytokine that induces cancer cell death by binding to TRAIL receptors. Because of its selective cytotoxicity toward cancer cells, TRAIL therapeutics, such as recombinant TRAIL and agonistic antibodies targeting TRAIL receptors, have garnered attention [...] Read more.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytotoxic cytokine that induces cancer cell death by binding to TRAIL receptors. Because of its selective cytotoxicity toward cancer cells, TRAIL therapeutics, such as recombinant TRAIL and agonistic antibodies targeting TRAIL receptors, have garnered attention as promising cancer treatment agents. However, many cancer cells acquire resistance to TRAIL-induced cell death. To overcome this issue, we searched for agents to sensitize cancer cells to TRAIL-induced cell death by screening a small-molecule chemical library consisting of diverse compounds. We identified a cardiac glycoside, proscillaridin A, as the most effective TRAIL sensitizer in colon cancer cells. Proscillaridin A synergistically enhanced TRAIL-induced cell death in TRAIL-sensitive and -resistant colon cancer cells. Additionally, proscillaridin A enhanced cell death in cells treated with TRAIL and TRAIL sensitizer, the second mitochondria-derived activator of caspase mimetic. Proscillaridin A upregulated TRAIL receptor expression, while downregulating the levels of the anti-cell death molecules, cellular FADD-like IL-1β converting enzyme-like inhibitor protein and Mcl1, in a cell type-dependent manner. Furthermore, proscillaridin A enhanced TRAIL-induced cell death partly via O-glycosylation. Taken together, our findings suggest that proscillaridin A is a promising agent that enhances the anti-cancer efficacy of TRAIL therapeutics. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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16 pages, 2520 KiB

Open AccessArticle

Unique Glycoform-Dependent Monoclonal Antibodies for Mouse Mucin 21

by Jun Nishida, Shigeyuki Shichino, Tatsuya Tsukui, Mayumi Hoshino, Tomoko Okada, Kyoko Okada, Yuri Yi, Seiko Toraya-Brown, Miho Mochizuki, Ryouta Koizumi, Katrin Ishii-Schrade, Kaori Denda-Nagai and Tatsuro Irimura

Int. J. Mol. Sci. 2022, 23(12), 6718; https://doi.org/10.3390/ijms23126718 - 16 Jun 2022

Cited by 2 | Viewed by 1798

Abstract

Mucin 21(Muc21)/epiglycanin is expressed on apical surfaces of squamous epithelia and has potentially protective roles, which are thought to be associated with its unique glycoforms, whereas its aberrant glycosylation is implicated in the malignant behaviors of some carcinomas. Despite the importance of glycoforms, [...] Read more.

Mucin 21(Muc21)/epiglycanin is expressed on apical surfaces of squamous epithelia and has potentially protective roles, which are thought to be associated with its unique glycoforms, whereas its aberrant glycosylation is implicated in the malignant behaviors of some carcinomas. Despite the importance of glycoforms, we lack tools to detect specific glycoforms of mouse Muc21. In this study, we generated two monoclonal antibodies (mAbs) that recognize different glycoforms of Muc21. We used membrane lysates of Muc21-expressing TA3-Ha cells or Chinese hamster ovary (CHO)-K1 cells transfected with Muc21 as antigens. Specificity testing, utilizing Muc21 glycosylation variant cells, showed that mAb 1A4-1 recognized Muc21 carrying glycans terminated with galactose residues, whereas mAb 18A11 recognized Muc21 carrying sialylated glycans. mAb 1A4-1 stained a majority of mouse mammary carcinoma TA3-Ha cells in vitro and in engrafted tumors in mice, whereas mAb 18A11 recognized only a subpopulation of these. mAb 1A4-1 was useful in immunohistochemically detecting Muc21 in normal squamous epithelia. In conclusion, these mAbs recognize distinct Muc21 epitopes formed by combinations of peptide portions and O-glycans. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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11 pages, 3310 KiB

Open AccessArticle

Comprehensive Analysis of Oligo/Polysialylglycoconjugates in Cancer Cell Lines

by Masaya Hane, Ken Kitajima and Chihiro Sato

Int. J. Mol. Sci. 2022, 23(10), 5569; https://doi.org/10.3390/ijms23105569 - 16 May 2022

Cited by 2 | Viewed by 1611

Abstract

In cancer cells, cell-surface sialylation is altered, including a change in oligo/polysialic acid (oligo/polySia) structures. Since they are unique and rarely expressed in normal cells, oligo/polySia structures may serve as promising novel biomarkers and targets for therapies. For the diagnosis and treatment of [...] Read more.

In cancer cells, cell-surface sialylation is altered, including a change in oligo/polysialic acid (oligo/polySia) structures. Since they are unique and rarely expressed in normal cells, oligo/polySia structures may serve as promising novel biomarkers and targets for therapies. For the diagnosis and treatment of the disease, a precise understanding of the oligo/polySia structures in cancer cells is necessary. In this study, flow cytometric analysis and gene expression datasets were obtained from sixteen different cancer cell lines. These datasets demonstrated the ability to predict glycan structures and their sialylation status. Our results also revealed that sialylation patterns are unique to each cancer cell line. Thus, we can suggest promising combinations of antibody and cancer cell for glycan prediction. However, the precise prediction of minor glycans need to be further explored. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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15 pages, 3486 KiB

Open AccessArticle

Truncated O-Glycan-Bearing MUC16 Enhances Pancreatic Cancer Cells Aggressiveness via α4β1 Integrin Complexes and FAK Signaling

by Christabelle Rajesh, Satish Sagar, Ashok Kumar Rathinavel, Divya Thomas Chemparathy, Xianlu Laura Peng, Jen Jen Yeh, Michael A. Hollingsworth and Prakash Radhakrishnan

Int. J. Mol. Sci. 2022, 23(10), 5459; https://doi.org/10.3390/ijms23105459 - 13 May 2022

Cited by 8 | Viewed by 2436

Abstract

Elevated levels of Mucin-16 (MUC16) in conjunction with a high expression of truncated O-glycans is implicated in playing crucial roles in the malignancy of pancreatic ductal adenocarcinoma (PDAC). However, the mechanisms by which such aberrant glycoforms present on MUC16 itself promote an increased [...] Read more.

Elevated levels of Mucin-16 (MUC16) in conjunction with a high expression of truncated O-glycans is implicated in playing crucial roles in the malignancy of pancreatic ductal adenocarcinoma (PDAC). However, the mechanisms by which such aberrant glycoforms present on MUC16 itself promote an increased disease burden in PDAC are yet to be elucidated. This study demonstrates that the CRISPR/Cas9-mediated genetic deletion of MUC16 in PDAC cells decreases tumor cell migration. We found that MUC16 enhances tumor malignancy by activating the integrin-linked kinase and focal adhesion kinase (ILK/FAK)-signaling axis. These findings are especially noteworthy in truncated O-glycan (Tn and STn antigen)-expressing PDAC cells. Activation of these oncogenic-signaling pathways resulted in part from interactions between MUC16 and integrin complexes (α4β1), which showed a stronger association with aberrant glycoforms of MUC16. Using a monoclonal antibody to functionally hinder MUC16 significantly reduced the migratory cascades in our model. Together, these findings suggest that truncated O-glycan containing MUC16 exacerbates malignancy in PDAC by activating FAK signaling through specific interactions with α4 and β1 integrin complexes on cancer cell membranes. Targeting these aberrant glycoforms of MUC16 can aid in the development of a novel platform to study and treat metastatic pancreatic cancer. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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15 pages, 1649 KiB

Open AccessArticle

Development and Evaluation of a Robust Sandwich Immunoassay System Detecting Serum WFA-Reactive IgA1 for Diagnosis of IgA Nephropathy

by Yuta Uenoyama, Atsushi Matsuda, Kazune Ohashi, Koji Ueda, Misaki Yokoyama, Takuya Kyoutou, Kouji Kishi, Youichi Takahama, Masaaki Nagai, Takaaki Ohbayashi, Osamu Hotta and Hideki Matsuzaki

Int. J. Mol. Sci. 2022, 23(9), 5165; https://doi.org/10.3390/ijms23095165 - 05 May 2022

Cited by 2 | Viewed by 1655

Abstract

Aberrant glycosylation of IgA1 is involved in the development of IgA nephropathy (IgAN). There are many reports of IgAN markers focusing on the glycoform of IgA1. None have been clinically applied as a routine test. In this study, we established an automated sandwich [...] Read more.

Aberrant glycosylation of IgA1 is involved in the development of IgA nephropathy (IgAN). There are many reports of IgAN markers focusing on the glycoform of IgA1. None have been clinically applied as a routine test. In this study, we established an automated sandwich immunoassay system for detecting aberrant glycosylated IgA1, using Wisteria floribunda agglutinin (WFA) and anti-IgA1 monoclonal antibody. The diagnostic performance as an IgAN marker was evaluated. The usefulness of WFA for immunoassays was investigated by lectin microarray. A reliable standard for quantitative immunoassay measurements was designed by modifying a purified IgA1 substrate. A validation study using multiple serum specimens was performed using the established WFA-antibody sandwich automated immunoassay. Lectin microarray results showed that WFA specifically recognized N-glycans of agglutinated IgA1 in IgAN patients. The constructed IgA1 standard exhibited a wide dynamic range and high reactivity. In the validation study, serum WFA-reactive IgA1 (WFA+-IgA1) differed significantly between healthy control subjects and IgAN patients. The findings indicate that WFA is a suitable lectin that specifically targets abnormal agglutinated IgA1 in serum. We also describe an automated immunoassay system for detecting WFA+-IgA1, focusing on N-glycans. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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18 pages, 3535 KiB

Open AccessArticle

Therapeutic Effects of an Anti-sialyl Lewis X Antibody in a Murine Model of Allergic Asthma

by Wei Xiong, Wenxin Liu, Shogo Nishida, Daichi Komiyama, Wei Liu, Jotaro Hirakawa and Hiroto Kawashima

Int. J. Mol. Sci. 2021, 22(18), 9961; https://doi.org/10.3390/ijms22189961 - 15 Sep 2021

Cited by 1 | Viewed by 2397

Abstract

Asthma is an allergic disease that causes severe infiltration of leukocytes into the lungs. Leukocyte infiltration is mediated by the binding of sialyl Lewis X (sLe^x) glycans present on the leukocytes to E-and P-selectins present on the endothelial cells at the [...] Read more.

Asthma is an allergic disease that causes severe infiltration of leukocytes into the lungs. Leukocyte infiltration is mediated by the binding of sialyl Lewis X (sLe^x) glycans present on the leukocytes to E-and P-selectins present on the endothelial cells at the sites of inflammation. Here, we found that mouse eosinophils express sLe^x glycans, and their infiltration into the lungs and proliferation in the bone marrow were significantly suppressed by an anti-sLe^x monoclonal antibody (mAb) F2 in a murine model of ovalbumin-induced asthma. The percentage of eosinophils in the bronchoalveolar lavage fluid and bone marrow and serum IgE levels decreased significantly in the F2-administered mice. Levels of T helper type 2 (Th2) cytokines and chemokines, involved in IgE class switching and eosinophil proliferation and recruitment, were also decreased in the F2-administered mice. An ex vivo cell rolling assay revealed that sLe^x glycans mediate the rolling of mouse eosinophils on P-selectin-expressing cells. These results indicate that the mAb F2 exerts therapeutic effects in a murine model of allergen-induced asthma, suggesting that sLe^x carbohydrate antigen could serve as a novel therapeutic target for allergic asthma. Full article

(This article belongs to the Special Issue Glycobiology-Based Drug Development)

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