title: Natural killer cell-based adoptive immunotherapy eradicates and drives differentiation of chemoresistant bladder cancer stem-like cells.
abstract: High-grade non-muscle invasive bladder cancer (NMIBC) has a high risk of recurrence and progression to muscle-invasive forms, which seems to be largely related to the presence of tumorigenic stem-like cell populations that are refractory to conventional therapies. Here, we evaluated the therapeutic potential of Natural Killer (NK) cell-based adoptive immunotherapy against chemoresistant bladder cancer stem-like cells (CSCs) in a pre-clinical relevant model, using NK cells from healthy donors and NMIBC patients.
  
  
  
  Cytokine-activated NK cells from healthy donors and from high-grade NMIBC patients were phenotypically characterized and assayed in vitro against stem-like and bulk differentiated bladder cancer cells. Stem-like cells were isolated from two bladder cancer cell lines using the sphere-forming assay. The in vivo therapeutic efficacy was evaluated in mice bearing a CSC-induced orthotopic bladder cancer. Animals were treated by intravesical instillation of interleukin-activated NK cells. Tumor response was evaluated longitudinally by non-invasive bioluminescence imaging.
  
  
  
  NK cells from healthy donors upon activation with IL-2 and IL-15 kills indiscriminately both stem-like and differentiated tumor cells via stress ligand recognition. In addition to cell killing, NK cells shifted CSCs towards a more differentiated phenotype, rendering them more susceptible to cisplatin, highlighting the benefits of a possible combined therapy. On the contrary, NK cells from NMIBC patients displayed a low density on NK cytotoxicity receptors, adhesion molecules and a more immature phenotype, losing their ability to kill and drive differentiation of CSCs. The local administration, via the transurethral route, of activated NK cells from healthy donors provides an efficient tumor infiltration and a subsequent robust tumoricidal activity against bladder cancer with high selective cytolytic activity against CSCs, leading to a dramatic reduction in tumor burden from 80 % to complete remission.
  
  
  
  Although pre-clinical, our results strongly suggest that an immunotherapeutic strategy using allogeneic activated NK cells from healthy donors is effective and should be exploited as a complementary therapeutic strategy in high-risk NMIBC patients to prevent tumor recurrence and progression.
authors: Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences (IBILI), Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|CNC.IBILI, University of Coimbra, Coimbra, Portugal.
authors: Margarida
authors: M
authors: Ferreira-Teixeira
authors: Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences (IBILI), Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|CNC.IBILI, University of Coimbra, Coimbra, Portugal.
authors: Daniela
authors: D
authors: Paiva-Oliveira
authors: Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences (IBILI), Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|Urology and Renal Transplantation Department, Coimbra University Hospital Centre (CHUC), Coimbra, Portugal.
authors: Belmiro
authors: B
authors: Parada
authors: Institute of Immunology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
authors: Vera
authors: V
authors: Alves
authors: Service of Anatomical Pathology, Coimbra University Hospital Centre (CHUC), Coimbra, Portugal.|Institute of Anatomical and Molecular Pathology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
authors: Vitor
authors: V
authors: Sousa
authors: Viral Immunobiology, Institute of Experimental Immunology, University of Zürich, Zürich, Switzerland.
authors: Obinna
authors: O
authors: Chijioke
authors: Viral Immunobiology, Institute of Experimental Immunology, University of Zürich, Zürich, Switzerland.
authors: Christian
authors: C
authors: Münz
authors: Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences (IBILI), Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|CNC.IBILI, University of Coimbra, Coimbra, Portugal.
authors: Flávio
authors: F
authors: Reis
authors: Institute of Immunology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|Center of Investigation in Environment, Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, Coimbra, Portugal.|Immunology and Oncology Laboratory, Center for Neurosciences and Cell Biology (CNC), University of Coimbra, Coimbra, Portugal.
authors: Paulo
authors: P
authors: Rodrigues-Santos
authors: Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences (IBILI), Faculty of Medicine, University of Coimbra, Coimbra, Portugal. cgomes@fmed.uc.pt.|CNC.IBILI, University of Coimbra, Coimbra, Portugal. cgomes@fmed.uc.pt.|Center of Investigation in Environment, Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, Coimbra, Portugal. cgomes@fmed.uc.pt.
authors: Célia
authors: C
authors: Gomes
mesh_terms: D000368:Aged; D000818:Animals; D002454:Cell Differentiation / Q000276:immunology; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000494:pharmacology; D005260:Female; D006801:Humans; D016130:Immunophenotyping; D016219:Immunotherapy, Adoptive / Q000379:methods*; D019409:Interleukin-15 / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D020014:K562 Cells; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology* / Q000637:transplantation*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D009364:Neoplasm Recurrence, Local / Q000473:pathology; D014411:Neoplastic Stem Cells / Q000276:immunology* / Q000473:pathology; D001749:Urinary Bladder Neoplasms / Q000276:immunology / Q000473:pathology / Q000628:therapy*
chemical_list: C569274:IL15 protein, human; D019409:Interleukin-15; D007376:Interleukin-2; D002945:Cisplatin
keywords: Bladder cancer; Cancer stem cells; Immunotherapy; Natural killer cells