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Genetically engineered and enucleated human mesenchymal stromal cells for the targeted delivery of therapeutics to diseased tissue

Nature Biomedical Engineering volume 6, pages 882–897 (2022)Cite this article

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Abstract

Targeting the delivery of therapeutics specifically to diseased tissue enhances their efficacy and decreases their side effects. Here we show that mesenchymal stromal cells with their nuclei removed by density-gradient centrifugation following the genetic modification of the cells for their display of chemoattractant receptors and endothelial-cell-binding molecules are effective vehicles for the targeted delivery of therapeutics. The enucleated cells neither proliferate nor permanently engraft in the host, yet retain the organelles for energy and protein production, undergo integrin-regulated adhesion to inflamed endothelial cells, and actively home to chemokine gradients established by diseased tissues. In mouse models of acute inflammation and of pancreatitis, systemically administered enucleated cells expressing two types of chemokine receptor and an endothelial adhesion molecule enhanced the delivery of an anti-inflammatory cytokine to diseased tissue (with respect to unmodified stromal cells and to exosomes derived from bone-marrow-derived stromal cells), attenuating inflammation and ameliorating disease pathology. Enucleated cells retain most of the cells’ functionality, yet acquire the cargo-carrying characteristics of cell-free delivery systems, and hence represent a versatile delivery vehicle and therapeutic system.

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Fig. 1: Enucleated cells retain important cellular functions.
Fig. 2: Cargocytes have chemotaxis activities and migrate better through confined spaces.
Fig. 3: Cargocytes interact with adhesion molecules.
Fig. 4: Bioengineered cargocytes actively and specifically home to the inflamed ear.
Fig. 5: Bioengineered cargocytes efficiently deliver bioactive IL-10 to inflamed ears and ameliorate local inflammation.
Fig. 6: Bioengineered cargocytes ameliorate Caerulein-induced acute pancreatitis.

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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 author on reasonable request. Source data are provided with this paper.

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Acknowledgements

This work was supported by the National Institutes of Health grants (R01 CA097022, CA184594 and CA182495 (R.L.K.) and R01 HL082792, R01 GM137605 and U54 CA210184 (J.L.)), the National Science Foundation (CAREER Award CBET-1254846 to J.L.), the Hartwell Foundation (J.D.B.) and the Center for Drug Discovery and Innovation at UC San Diego (R.L.K.). C.N.A. was supported by NIH 5T32 OD17863-4. Imaging and analysis were in part performed at the UCSD School of Medicine Microscopy Core (NS047101) and UCSD Moores Cancer Center Tissue Technology Shared Resource Core (CCSG Grant P30CA23100). Human primary bone marrow MSCs were obtained from Texas A&M Health Science Center College of Medicine Institute for Regenerative Medicine (NIH P40OD011050). This work was performed in part at the Cornell NanoScale Science & Technology Facility (CNF), a member of the National Nanotechnology Coordinated Infrastructure (NNCI), which is supported by the National Science Foundation (Grant NNCI-2025233). We thank Dr W. Zhao and Dr A. Ségaliny of UC Irvine, and D. Thomas, Dr L. de Siqueira Neto, Dr M. Ruchhoeft and Dr L. Zhao of UCSD for thoughtful discussion.

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  1. These authors contributed equally: Huawei Wang, Christina N. Alarcón.

Authors and Affiliations

  1. Department of Pathology, University of California, San Diego, La Jolla, CA, USA

    Huawei Wang, Christina N. Alarcón, Bei Liu, Felicia Watson, Stephen Searles, Calvin K. Lee, Willie Pi, Dale Allen, Jack D. Bui & Richard L. Klemke

  2. Moores Cancer Center, University of California, San Diego, La Jolla, CA, USA

    Huawei Wang, Christina N. Alarcón, Bei Liu, Felicia Watson, Willie Pi, Dale Allen & Richard L. Klemke

  3. Biomedical Sciences Program, University of California, San Diego, La Jolla, CA, USA

    Christina N. Alarcón & Stephen Searles

  4. Nancy E. and Peter C. Meinig School of Biomedical Engineering and Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, NY, USA

    Jeremy Keys & Jan Lammerding

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Contributions

H.W. and R.L.K. designed and supervised the project. H.W., C.N.A., B.L., F.W., S.S., C.K.L., J.K., W.P. and D.A. performed the experiments. H.W., C.N.A., S.S. and J.K. analysed the data. C.N.A. read the blinded histology. H.W., C.N.A. and R.L.K. wrote the paper. J.D.B. and J.L. revised the paper.

Corresponding author

Correspondence to Richard L. Klemke.

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Competing interests

Cargocyte is the trademark of Cytonus Therapeutics. R.L.K. is the co-founder and equity holder of Cytonus Therapeutics. H.W. and R.L.K. are co-inventors of a related patent (WO 2019/032628 A1, US 10,927,349 B2) and H.W., W.P. and R.L.K. are co-inventors of another related patent (PCT/US21/016,919). Both patents were filed by UCSD.

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Peer review information Nature Biomedical Engineering thanks Zhen Gu, Di Yu and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Video

Cargocyteslifeact-RFP treated as in Fig. 2d and allowed to move through constrictions along an FBS gradient in a microfluidic device.

Video

MSClifeact-RFP treated as in Fig. 2d and allowed to move through constrictions along an FBS gradient in a microfluidic device.

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Wang, H., Alarcón, C.N., Liu, B. et al. Genetically engineered and enucleated human mesenchymal stromal cells for the targeted delivery of therapeutics to diseased tissue. Nat. Biomed. Eng 6, 882–897 (2022). https://doi.org/10.1038/s41551-021-00815-9

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