High and selective cytotoxicity of ex vivo expanded allogeneic human natural killer cells from peripheral blood against bladder cancer: implications for natural killer cell instillation after transurethral resection of bladder tumor

Sep 21, 2026

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Non-muscle-invasive bladder cancer (NMIBC) treated by transurethral resection of bladder tumor (TURBT) followed by intravesical chemotherapy or Bacillus Calmette–Guérin (BCG) shows high recurrence and treatment-related side effects, creating demand for alternative instillation options. Expanded allogeneic human natural killer cells from peripheral blood are evaluated for bladder cancer killing. In vitro and patient-derived organoid and xenograft models demonstrate potent, high-selectivity cytotoxicity against bladder cancer cells with limited toxicity to normal urothelial cells, alongside cytokine and chemokine-mediated T-cell recruitment, supporting translational intravesical NK therapy after TURBT for NMIBC.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages19

Reading Time48 min

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Abstract

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Wang et al. Journal of Experimental & Clinical Cancer Research (2024) 43:24 [https://doi.org/10.1186/s13046-024-02955-7](https://doi.org/10.1186/s13046-024-02955-7) Journal of Experimental & Clinical Cancer Research RESEARCH Open Access High and selective cytotoxicity of ex vivo expanded allogeneic human natural killer cells from peripheral blood against bladder cancer: implications for natural killer cell instillation after transurethral resection of bladder tumor Fangming Wang1, Gang Zhang1, Tianli Xu2, Jianlin Ma2, Jing Wang2, Shuai Liu2, Yuzhe Tang1, Song Jin1, Jianxing Li 1* and Nianzeng Xing3,4* Abstract Background Non-muscle-invasive bladder cancer (NMIBC) is treated with transurethral resection of bladder tumor (TURBT) followed by intravesical instillation of chemotherapy or Bacillus Calmette–Guérin therapy. However, these treatments have a high recurrence rate and side effects, emphasizing the need for alternative instillations. Previously, we revealed that expanded allogeneic human natural killer (NK) cells from peripheral blood are a promising cellular therapy for prostate cancer. However, whether NK cells exhibit a similar killing effect in bladder cancer (BCa) remains unknown. Methods Expansion, activation, and cryopreservation of allogeneic human NK cells obtained from peripheral blood were performed as we previously described. In vitro cytotoxicity was evaluated using the cell counting kit-8 . The levels of perforin, granzyme B, interferon-γ, tumor necrosis factor-α, and chemokines (C-C-motif ligand [CCL]1, CCL2, CCL20, CCL3L1, and CCL4; C-X-C-motif ligand [CXCL]1, CXCL16, CXCL2, CXCL3, and CXCL8; and X-motif ligand 1 and 2) were determined using enzyme-linked immunosorbent assay. The expression of CD107a, major histocompatibility complex class I (MHC-I), MHC-I polypeptide-related sequences A and B (MICA/B), cytomegalovirus UL16-binding protein-2/5/6 (ULBP-2/5/6), B7-H6, CD56, CD69, CD25, killer cell Ig-like receptors (KIR)2DL1, KIRD3DL1, NKG2D, NKp30, NKp46, and CD16 of NK cells or BCa and normal urothelial cells were detected using flow cytometry. Cytotoxicity was evaluated using lactate dehydrogenase assay in patient-derived organoid models. BCa growth was monitored in vivo using calipers in male NOD-scid IL2rg−/− mice subcutaneously injected with 5637 and NK cells. Differential gene *Correspondence: Jianxing Li[ljxa01048@btch.edu.cn](ljxa01048@btch.edu.cn)[ ](ljxa01048@btch.edu.cn)Nianzeng Xing [xingnianzeng@126.com](xingnianzeng@126.com) Full list of author information is available at the end of the article © The Author(s) 2024. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit [http://creativecommons.org/licenses/by/4.0/](http://creativecommons.org/licenses/by/4.0/. The)[. The](http://creativecommons.org/licenses/by/4.0/. The)[ ](http://creativecommons.org/licenses/by/4.0/. The)Creative Commons Public Domain Dedication waiver ([http://creativecommons.org/publicdomain/zero/1.0/](http://creativecommons.org/publicdomain/zero/1.0/)) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Wang et al. Journal of Experimental & Clinical Cancer Research (2024) 43:24 Page 2 of 19 expressions were inve

FAQs

Why is an alternative to intravesical chemotherapy or BCG needed for NMIBC?

TURBT followed by intravesical chemotherapy or Bacillus Calmette–Guérin therapy has high recurrence rates and side effects, motivating exploration of other instillation approaches.

How were the allogeneic NK cells prepared and assessed for cytotoxicity?

Allogeneic NK cells from peripheral blood were expanded, activated, and cryopreserved. Cytotoxicity and immune activation markers were measured using assays such as CCK-8, LDH, ELISA, and flow cytometry, including patient-derived organoid testing.

What evidence supports that expanded NK cells are effective and selective against bladder cancer?

The study reports higher in vitro cytotoxicity toward bladder cancer lines, tumor effects in patient-derived organoids and xenograft mouse models, and limited toxicity toward normal urothelial cells, along with chemokine secretion that supports T-cell chemotaxis.

Sep 21, 2026

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