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Conjugation of glucosylated polymer chains to checkpoint blockade antibodies augments their efficacy and specificity for glioblastoma

Nature Biomedical Engineering volume 5, pages 1274–1287 (2021)Cite this article

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Abstract

Because of the blood–tumour barrier and cross-reactivity with healthy tissues, immune checkpoint blockade therapy against glioblastoma has inadequate efficacy and is associated with a high risk of immune-related adverse events. Here we show that anti-programmed death-ligand 1 antibodies conjugated with multiple poly(ethylene glycol) (PEG) chains functionalized to target glucose transporter 1 (which is overexpressed in brain capillaries) and detaching in the reductive tumour microenvironment augment the potency and safety of checkpoint blockade therapy against glioblastoma. In mice bearing orthotopic glioblastoma tumours, a single dose of glucosylated and multi-PEGylated antibodies reinvigorated antitumour immune responses, induced immunological memory that protected the animals against rechallenge with tumour cells, and suppressed autoimmune responses in the animals’ healthy tissues. Drug-delivery formulations leveraging multivalent ligand interactions and the properties of the tumour microenvironment to facilitate the crossing of blood–tumour barriers and increase drug specificity may enhance the efficacy and safety of other antibody-based therapies.

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Fig. 1: Characterization of reduction-activatable Gluc-S-aPD-L1.
Fig. 2: In vivo delivery efficacy of Gluc-S-aPD-L1.
Fig. 3: In vivo treatment efficacy of Gluc-S-aPD-L1.
Fig. 4: In vivo detection of reduction-induced PEG chain detachment.
Fig. 5: In vivo antitumour immune response.
Fig. 6: Gluc-S-aPD-L1 suppressed the incidence of irAEs in healthy tissues.

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Data availability

The main data supporting the results in this study are available within the paper and its Supplementary Information. The raw and analysed datasets generated during the study are too large to be publicly shared, but they are available for research purposes from the corresponding authors on reasonable request. Source data are provided with this paper.

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Acknowledgements

We thank Y. Tezuka and S. Fukushima for the technical assistance. This work was supported by the Center of Innovation Science and Technology-based Radical Innovation and Entrepreneurship Program (COI STREAM) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), JSPS KAKENHI Grant-in-Aid for Scientific Research (A) (JP18H04170) and Project for Cancer Research and Therapeutic Evolution (P-CREATE) (JP19cm0106202) from Japan Agency for Medical Research and Development (AMED).

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Authors and Affiliations

  1. Innovation Center of NanoMedicine, Kawasaki Institute of Industrial Promotion, Kawasaki, Japan

    Tao Yang, Yuki Mochida, Xueying Liu, Hang Zhou, Jinbing Xie, Hiroaki Kinoh & Kazunori Kataoka

  2. Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, Tokyo, Japan

    Yasutaka Anraku & Horacio Cabral

  3. Institute for Future Initiatives, The University of Tokyo, Tokyo, Japan

    Kazunori Kataoka

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  1. Tao Yang
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Contributions

T.Y. conducted the synthesis, characterization, cell studies, animal experiments and data analysis. X.L. helped in the construction of the orthotopic brain tumour model and animal experiments. Y.M. helped in the CD spectra test and antitumour immune response analysis. H.Z., J.X. and H.K. contributed to the FACS experiments and data analysis. Y.A. helped in the analysis and discussion on the heterogeneity of GLUT1 in the brain tumours. T.Y. and K.K. conceived the idea and designed the experiments. T.Y. and H.C. discussed the data and wrote the manuscript. K.K. commented on and revised the manuscript, and supervised the project.

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Correspondence to Horacio Cabral or Kazunori Kataoka.

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The authors declare no competing interests.

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Peer review information Nature Biomedical Engineering thanks Esther Chang, Dai Fukumura and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available.

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Extended data

Extended Data Fig. 1 Spatial analysis of native aPD-L1 and Gluc-S-aPD-L1 distribution in GL261 tumours.

a,b, Fluorescent images of the distribution of Alexa Fluor 647-labelled (a) native aPD-L1 and (b) Gluc0-S-aPD-L1 (cyan) in GL261 tumour sections stained with anti-CD31 antibody (red), anti-GLUT1 antibody (green) and DAPI (blue), white dash line indicated tumour boundary. Scale bar, 1,000 μm. c, Fluorescence intensity of native aPD-L1, Gluc0-S-aPD-L1 and Gluc25-S-aPD-L1 at tumour region at 24 h post-injection. d, Calculated fluorescence intensity ratio of tumour to normal brain in native aPD-L1, Gluc0-S-aPD-L1 and Gluc25-S-aPD-L1 groups at 24 h post-injection. Column heights and error bars represent means ± s.e.m. (n = 3).

Extended Data Fig. 2 Distribution and quantitative analysis of Gluc25-S-aPD-L1 in GL261 tumour regions with high and low GLUT1 levels.

a,b, Fluorescence images of GLUT1-rich areas (a) and GLUT1-low areas (b) in GL261 tumour sections stained with Alexa Fluor 647-labelled Gluc25-S-aPD-L1 (cyan), anti-CD31 antibody (red), anti-GLUT1 antibody (green) and DAPI (blue). 6 GLUT1-rich areas and 6 GLUT1-low areas from sections of 3 different tumours were selected. Scale bar, 100 μm. c, Blood vessel density and GLUT1 density in GLUT1-rich areas and GLUT1-low areas. d, Co-localization rate of blood vessel and GLUT1 in GLUT1-rich areas and GLUT1-low areas. e, Fluorescence intensity of Alexa Fluor 647-labelled Gluc25-S-aPD-L1 in GLUT1-rich areas and GLUT1-low areas. Data are means ± s.e.m. (n = 6). Statistical significance was calculated by student’s t-test.

Extended Data Fig. 3 In vivo antitumour efficacy of Gluc25-S-aPD-L1 in CT2A tumour models.

a, Timeline schedule for the treatment of CT2A tumour models. b, Bioluminescence images of CT2A tumour-bearing mice treated with saline, native aPD-L1, and Gluc25-S-aPD-L1 (aPD-L1: 1.5 mg per kg, n = 5). c, Region-of-interest analysis of bioluminescence intensities from whole brain. d, Survival curves for treated and control mice (n = 5). Statistical significance was calculated by log-rank test.

Extended Data Fig. 4 Quantitative analysis of chemoattractants production in GL261 tumour after various treatments.

a,b, Levels of chemoattractants such as Cxcl9 (a) and Cxcl10 (b) inside GL261 tumour of mice at 3 d post-injection of saline, native aPD-L1, Gluc0-S-aPD-L1 and Gluc25-S-aPD-L1 at the dose of 1.5 mg kg−1 aPD-L1 (n = 3 biologically independent samples). Data are presented as means ± s.e.m. Statistical significance was calculated by one-way analysis of variance (ANOVA) with Tukey’s post hoc test.

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Yang, T., Mochida, Y., Liu, X. et al. Conjugation of glucosylated polymer chains to checkpoint blockade antibodies augments their efficacy and specificity for glioblastoma. Nat Biomed Eng 5, 1274–1287 (2021). https://doi.org/10.1038/s41551-021-00803-z

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