Targeted crystallization of mixed-charge nanoparticles in lysosomes induces selective death of cancer cells


Lysosomes have become an important target for anticancer therapeutics because lysosomal cell death bypasses the classical caspase-dependent apoptosis pathway, enabling the targeting of apoptosis- and drug-resistant cancers. However, only a few small molecules—mostly repurposed drugs—have been tested so far, and these typically exhibit low cancer selectivity, making them suitable only for combination therapies. Here, we show that mixed-charge nanoparticles covered with certain ratios of positively and negatively charged ligands can selectively target lysosomes in cancerous cells while exhibiting only marginal cytotoxicity towards normal cells. This selectivity results from distinct pH-dependent aggregation events, starting from the formation of small, endocytosis-prone clusters at cell surfaces and ending with the formation of large and well-ordered nanoparticle assemblies and crystals inside cancer lysosomes. These assemblies cannot be cleared by exocytosis and cause lysosome swelling, which gradually disrupts the integrity of lysosomal membranes, ultimately impairing lysosomal functions and triggering cell death.

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Fig. 1: Summary of how crystallization of mixed-charge NPs in cancer lysosomes leads to selective killing of cancer cells.
Fig. 2: Structure and pH-dependent aggregation of mixed-charge NPs.
Fig. 3: Engineering cancer-specific cytotoxicity by tuning the balance of surface charges on mixed-charge NPs.
Fig. 4: Cellular uptake and differential intracellular aggregation of mixed-charge NPs in cancerous versus normal cells.
Fig. 5: The impact of intracellular aggregation of mixed-charge NPs on lysosome organelles.
Fig. 6: Mixed-charge NPs accumulate in autophagic vesicles and are excluded from non-tumorigenic cells through exocytosis.

Data availability

The data used to generate results in the current study are available from the corresponding authors upon reasonable request.

Code availability

Code used to compute osmotic pressures and NP volume fractions in lysosomes described in Supplementary Note 3 is available in GitHub repository ( Code for spatial analysis of lysosome distributions (from Lysotracker images) along with the raw data are available from the GitHub repository (


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We thank Y.-K. Jeong for TEM imaging of [+/−] NPs in vitro. We also thank M.-S. Jeong at the Korea Basic Science Institute for the TEM analyses of [+/−] NPs in cells. This work was supported by the Institute of Basic Science, Republic of Korea (award no. IBS-R020-D1 to B.A.G.).

Author information




M.B., M.S., D.K. and K.K.-G. designed and performed experiments and analysed the data. Y.S. performed osmotic pressure calculations and wrote the codes for the analysis of microscopy images. S.K. and Y.-K.C. performed dark-field microscopy experiments and analysed data. S.L. characterized small-molecule ligands and prepared selected figures for publication at early stages of manuscript preparation. K.K.-G. and B.A.G. conceived and supervised research, designed experiments and wrote the paper. All authors read and corrected the manuscript.

Corresponding authors

Correspondence to Kristiana Kandere-Grzybowska or Bartosz A. Grzybowski.

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Competing interests

Invention disclosure (Patent No. 10-2020-0021515) describing this research has been made to the Institute for Basic Science, which sponsored this work.

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Supplementary information

Supplementary Information

Supplementary Figs. 1–30, Tables 1 and 2, Videos 1–10, Notes 1–5 and refs. 1–48.

Reporting Summary

Supplementary Video 1

Impact of mixed-charge nanoparticles on lysosomes in HT1080 fibrosarcoma cells.

Supplementary Video 2

Impact of mixed-charge nanoparticles on lysosomes in MDA-MB-231 adenocarcinoma cells.

Supplementary Video 3

Impact of mixed-charge nanoparticles on lysosomes in MCF7 breast carcinoma cells.

Supplementary Video 4

Impact of mixed-charge nanoparticles on lysosomes in non-cancerous mouse embryonic fibroblasts.

Supplementary Video 5

Impact of mixed-charge nanoparticles on lysosomes in non-cancerous epithelial MCF-10A cells.

Supplementary Video 6

Mixed-charge nanoparticle transport through endo-lysosomal system in non-cancerous MCF-10A cells: early events.

Supplementary Video 7

Mixed-charge nanoparticle transport through endo-lysosomal system in non-cancerous MCF-10A cells: late events.

Supplementary Video 8

Mixed-charge nanoparticle transport through endo-lysosomal system in MDA-MB-231 cancer cells: early events.

Supplementary Video 9

Mixed-charge nanoparticle transport through endo-lysosomal system in MDA-MB-231 cancer cells: late events.

Supplementary Video 10

Localization of mixed-charge nanoparticles to autolysosomes in non-cancerous MCF-10A cells.

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Borkowska, M., Siek, M., Kolygina, D. et al. Targeted crystallization of mixed-charge nanoparticles in lysosomes induces selective death of cancer cells. Nat. Nanotechnol. 15, 331–341 (2020).

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