Cancers

Research

23 pages, 5213 KiB

Open AccessArticle

Optimizing the Procedure to Manufacture Clinical-Grade NK Cells for Adoptive Immunotherapy

by Adrián Fernández, Alfonso Navarro-Zapata, Adela Escudero, Nerea Matamala, Beatriz Ruz-Caracuel, Isabel Mirones, Alicia Pernas, Marta Cobo, Gema Casado, Diego Lanzarot, Carlos Rodríguez-Antolín, María Vela, Cristina Ferreras, Carmen Mestre, Aurora Viejo, Alejandra Leivas, Joaquín Martínez, Lucía Fernández and Antonio Pérez-Martínez

Cancers 2021, 13(3), 577; https://doi.org/10.3390/cancers13030577 - 2 Feb 2021

Cited by 15 | Viewed by 8610

Abstract

Natural killer (NK) cells represent promising tools for cancer immunotherapy. We report the optimization of an NK cell activation–expansion process and its validation on clinical-scale. Methods: RPMI-1640, stem cell growth medium (SCGM), NK MACS and TexMACS were used as culture mediums. Activated and [...] Read more.

Natural killer (NK) cells represent promising tools for cancer immunotherapy. We report the optimization of an NK cell activation–expansion process and its validation on clinical-scale. Methods: RPMI-1640, stem cell growth medium (SCGM), NK MACS and TexMACS were used as culture mediums. Activated and expanded NK cells (NKAE) were obtained by coculturing total peripheral blood mononuclear cells (PBMC) or CD45RA⁺ cells with irradiated K562mbIL15-41BBL or K562mbIL21-41BBL. Fold increase, NK cell purity, activation status, cytotoxicity and transcriptome profile were analyzed. Clinical-grade NKAE cells were manufactured in CliniMACS Prodigy. Results: NK MACS and TexMACs achieved the highest NK cell purity and lowest T cell contamination. Obtaining NKAE cells from CD45RA⁺ cells was feasible although PBMC yielded higher total cell numbers and NK cell purity than CD45RA⁺ cells. The highest fold expansion and NK purity were achieved by using PBMC and K562mbIL21-41BBL cells. However, no differences in activation and cytotoxicity were found when using either NK cell source or activating cell line. Transcriptome profile showed to be different between basal NK cells and NKAE cells expanded with K562mbIL21-41BBL or K562mbIL15-41BBL. Clinical-grade manufactured NKAE cells complied with the specifications from the Spanish Regulatory Agency. Conclusions: GMP-grade NK cells for clinical use can be obtained by using different starting cells and aAPC. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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19 pages, 2728 KiB

Open AccessArticle

TIM-3 Expression Is Downregulated on Human NK Cells in Response to Cancer Targets in Synergy with Activation

by Tram N. Dao, Sagar Utturkar, Nadia Atallah Lanman and Sandro Matosevic

Cancers 2020, 12(9), 2417; https://doi.org/10.3390/cancers12092417 - 26 Aug 2020

Cited by 18 | Viewed by 3845

Abstract

Among natural killer (NK) cell receptors, the T-cell immunoglobulin and mucin-containing domain (TIM-3) has been associated with both inhibitory and activating functions, depending on context and activation pathway. Ex vivo and in vitro, expression of TIM-3 is inducible and depends on activation stimulus. [...] Read more.

Among natural killer (NK) cell receptors, the T-cell immunoglobulin and mucin-containing domain (TIM-3) has been associated with both inhibitory and activating functions, depending on context and activation pathway. Ex vivo and in vitro, expression of TIM-3 is inducible and depends on activation stimulus. Here, we report that TIM-3 expression can be downregulated on NK cells under specific conditions. When NK cells are exposed to cancer targets, they synergize with stimulation conditions to induce a substantial decrease in TIM-3 expression on their surface. We found that such downregulation occurs following prior NK activation. Downregulated TIM-3 expression correlated to lower cytotoxicity and lower interferon gamma (IFN-γ) expression, fueling the notion that TIM-3 might function as a benchmark for human NK cell dysfunction. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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18 pages, 1756 KiB

Open AccessArticle

Genetic and Molecular Basis of Heterogeneous NK Cell Responses against Acute Leukemia

by Dhon Roméo Makanga, Francesca Da Rin de Lorenzo, Gaëlle David, Catherine Willem, Léa Dubreuil, Nolwenn Legrand, Thierry Guillaume, Pierre Peterlin, Amandine Lebourgeois, Marie Christine Béné, Alice Garnier, Patrice Chevallier, Ketevan Gendzekhadze, Anne Cesbron, Katia Gagne, Béatrice Clemenceau and Christelle Retière

Cancers 2020, 12(7), 1927; https://doi.org/10.3390/cancers12071927 - 16 Jul 2020

Cited by 16 | Viewed by 2825

Abstract

Natural killer (NK) cells are key cytotoxic effectors against malignant cells. Polygenic and polymorphic Killer cell Immunoglobulin-like Receptor (KIR) and HLA genes participate in the structural and functional formation of the NK cell repertoire. In this study, we extensively investigated the anti-leukemic potential [...] Read more.

Natural killer (NK) cells are key cytotoxic effectors against malignant cells. Polygenic and polymorphic Killer cell Immunoglobulin-like Receptor (KIR) and HLA genes participate in the structural and functional formation of the NK cell repertoire. In this study, we extensively investigated the anti-leukemic potential of NK cell subsets, taking into account these genetic parameters and cytomegalovirus (CMV) status. Hierarchical clustering analysis of NK cell subsets based on NKG2A, KIR, CD57 and NKG2C markers from 68 blood donors identified donor clusters characterized by a specific phenotypic NK cell repertoire linked to a particular immunogenetic KIR and HLA profile and CMV status. On the functional side, acute lymphoblastic leukemia (ALL) was better recognized by NK cells than acute myeloid leukemia (AML). However, a broad inter-individual disparity of NK cell responses exists against the same leukemic target, highlighting bad and good NK responders. The most effective NK cell subsets against different ALLs expressed NKG2A and represented the most frequent subset in the NK cell repertoire. In contrast, minority CD57⁺ or/and KIR⁺ NK cell subsets were more efficient against AML. Overall, our data may help to optimize the selection of hematopoietic stem cell donors on the basis of immunogenetic KIR/HLA for ALL patients and identify the best NK cell candidates in immunotherapy for AML. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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Review

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24 pages, 2152 KiB

Open AccessReview

Arming Immune Cells for Battle: A Brief Journey through the Advancements of T and NK Cell Immunotherapy

by Philipp Wendel, Lisa Marie Reindl, Tobias Bexte, Leander Künnemeyer, Vinzenz Särchen, Nawid Albinger, Andreas Mackensen, Eva Rettinger, Tobias Bopp and Evelyn Ullrich

Cancers 2021, 13(6), 1481; https://doi.org/10.3390/cancers13061481 - 23 Mar 2021

Cited by 22 | Viewed by 6820

Abstract

The promising development of adoptive immunotherapy over the last four decades has revealed numerous therapeutic approaches in which dedicated immune cells are modified and administered to eliminate malignant cells. Starting in the early 1980s, lymphokine activated killer (LAK) cells were the first ex [...] Read more.

The promising development of adoptive immunotherapy over the last four decades has revealed numerous therapeutic approaches in which dedicated immune cells are modified and administered to eliminate malignant cells. Starting in the early 1980s, lymphokine activated killer (LAK) cells were the first ex vivo generated NK cell-enriched products utilized for adoptive immunotherapy. Over the past decades, various immunotherapies have been developed, including cytokine-induced killer (CIK) cells, as a peripheral blood mononuclear cells (PBMCs)-based therapeutic product, the adoptive transfer of specific T and NK cell products, and the NK cell line NK-92. In addition to allogeneic NK cells, NK-92 cell products represent a possible “off-the-shelf” therapeutic concept. Recent approaches have successfully enhanced the specificity and cytotoxicity of T, NK, CIK or NK-92 cells towards tumor-specific or associated target antigens generated by genetic engineering of the immune cells, e.g., to express a chimeric antigen receptor (CAR). Here, we will look into the history and recent developments of T and NK cell-based immunotherapy. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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22 pages, 4027 KiB

Open AccessReview

Design and Implementation of NK Cell-Based Immunotherapy to Overcome the Solid Tumor Microenvironment

by Ishwar Navin, Michael T. Lam and Robin Parihar

Cancers 2020, 12(12), 3871; https://doi.org/10.3390/cancers12123871 - 21 Dec 2020

Cited by 20 | Viewed by 4379

Abstract

Natural killer (NK) cells are innate immune effectors capable of broad cytotoxicity via germline-encoded receptors and can have conferred cytotoxic potential via the addition of chimeric antigen receptors. Combined with their reduced risk of graft-versus-host disease (GvHD) and cytokine release syndrome (CRS), NK [...] Read more.

Natural killer (NK) cells are innate immune effectors capable of broad cytotoxicity via germline-encoded receptors and can have conferred cytotoxic potential via the addition of chimeric antigen receptors. Combined with their reduced risk of graft-versus-host disease (GvHD) and cytokine release syndrome (CRS), NK cells are an attractive therapeutic platform. While significant progress has been made in treating hematological malignancies, challenges remain in using NK cell-based therapy to combat solid tumors due to their immunosuppressive tumor microenvironments (TMEs). The development of novel strategies enabling NK cells to resist the deleterious effects of the TME is critical to their therapeutic success against solid tumors. In this review, we discuss strategies that apply various genetic and non-genetic engineering approaches to enhance receptor-mediated NK cell cytotoxicity, improve NK cell resistance to TME effects, and enhance persistence in the TME. The successful design and application of these strategies will ultimately lead to more efficacious NK cell therapies to treat patients with solid tumors. This review outlines the mechanisms by which TME components suppress the anti-tumor activity of endogenous and adoptively transferred NK cells while also describing various approaches whose implementation in NK cells may lead to a more robust therapeutic platform against solid tumors. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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29 pages, 891 KiB

Open AccessReview

Will Next-Generation Immunotherapy Overcome the Intrinsic Diversity and Low Immunogenicity of Sarcomas to Improve Clinical Benefit?

by Hui Yi Chew, Victor Chan, Fiona Simpson and Riccardo Dolcetti

Cancers 2020, 12(11), 3392; https://doi.org/10.3390/cancers12113392 - 16 Nov 2020

Cited by 4 | Viewed by 2910

Abstract

Sarcomas are a rare type of a heterogeneous group of tumours arising from mesenchymal cells that form connective tissues. Surgery is the most common treatment for these tumours, but additional neoadjuvant or adjuvant chemotherapy or radiation therapies may be necessary. Unfortunately, a significant [...] Read more.

Sarcomas are a rare type of a heterogeneous group of tumours arising from mesenchymal cells that form connective tissues. Surgery is the most common treatment for these tumours, but additional neoadjuvant or adjuvant chemotherapy or radiation therapies may be necessary. Unfortunately, a significant proportion of patients treated with conventional therapies will develop metastatic disease that is resistant to therapies. Currently, there is an urgent need to develop more effective and durable therapies for the treatment of sarcomas. In recent years immunotherapies have revolutionised the treatment of a variety of cancers by restoring patient anti-tumour immune responses or through the adoptive infusion of immune effectors able to kill and eliminate malignant cells. The clinicopathologic and genetic heterogeneity of sarcomas, together with the generally low burden of somatic mutations potentially generating neoantigens, are currently limited to broad application of immunotherapy for patients with sarcomas. Nevertheless, a better understanding of the microenvironmental factors hampering the efficacy of immunotherapy and the identification of new and suitable therapeutic targets may help to overcome current limitations. Moreover, the recent advances in the development of immunotherapies based on the direct exploitation or targeting of T cells and/or NK cells may offer new opportunities to improve the treatment of sarcomas, particularly those showing recurrence or resistance to standard of care treatments. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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21 pages, 1106 KiB

Open AccessReview

Natural Born Killers: NK Cells in Cancer Therapy

by S. Elizabeth Franks, Benjamin Wolfson and James W. Hodge

Cancers 2020, 12(8), 2131; https://doi.org/10.3390/cancers12082131 - 31 Jul 2020

Cited by 46 | Viewed by 9058

Abstract

Cellular therapy has emerged as an attractive option for the treatment of cancer, and adoptive transfer of chimeric antigen receptor (CAR) expressing T cells has gained FDA approval in hematologic malignancy. However, limited efficacy was observed using CAR-T therapy in solid tumors. Natural [...] Read more.

Cellular therapy has emerged as an attractive option for the treatment of cancer, and adoptive transfer of chimeric antigen receptor (CAR) expressing T cells has gained FDA approval in hematologic malignancy. However, limited efficacy was observed using CAR-T therapy in solid tumors. Natural killer (NK) cells are crucial for tumor surveillance and exhibit potent killing capacity of aberrant cells in an antigen-independent manner. Adoptive transfer of unmodified allogeneic or autologous NK cells has shown limited clinical benefit due to factors including low cell number, low cytotoxicity and failure to migrate to tumor sites. To address these problems, immortalized and autologous NK cells have been genetically engineered to express high affinity receptors (CD16), CARs directed against surface proteins (PD-L1, CD19, Her2, etc.) and endogenous cytokines (IL-2 and IL-15) that are crucial for NK cell survival and cytotoxicity, with positive outcomes reported by several groups both preclinically and clinically. With a multitude of NK cell-based therapies currently in clinic trials, it is likely they will play a crucial role in next-generation cell therapy-based treatment. In this review, we will highlight the recent advances and limitations of allogeneic, autologous and genetically enhanced NK cells used in adoptive cell therapy. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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Other

Jump to: Research, Review

16 pages, 2595 KiB

Open AccessBrief Report

Quantification of Immune Variables from Liquid Biopsy in Breast Cancer Patients Links Vδ2⁺ γδ T Cell Alterations with Lymph Node Invasion

by Stéphane Fattori, Laurent Gorvel, Samuel Granjeaud, Philippe Rochigneux, Marie-Sarah Rouvière, Amira Ben Amara, Nicolas Boucherit, Magali Paul, Marie Mélanie Dauplat, Jeanne Thomassin-Piana, Maria Paciencia-Gros, Morgan Avenin, Jihane Pakradouni, Julien Barrou, Emmanuelle Charafe-Jauffret, Gilles Houvenaeghel, Eric Lambaudie, François Bertucci, Anthony Goncalves, Carole Tarpin, Jacques A. Nunès, Raynier Devillier, Anne-Sophie Chretien and Daniel Olive Show full author list Hide full author list

Cancers 2021, 13(3), 441; https://doi.org/10.3390/cancers13030441 - 25 Jan 2021

Cited by 5 | Viewed by 3752

Abstract

The rationale for therapeutic targeting of Vδ2⁺ γδ T cells in breast cancer is strongly supported by in vitro and murine preclinical investigations, characterizing them as potent breast tumor cell killers and source of Th1-related cytokines, backing cytotoxic αβ T cells. Nonetheless, [...] Read more.

The rationale for therapeutic targeting of Vδ2⁺ γδ T cells in breast cancer is strongly supported by in vitro and murine preclinical investigations, characterizing them as potent breast tumor cell killers and source of Th1-related cytokines, backing cytotoxic αβ T cells. Nonetheless, insights regarding Vδ2⁺ γδ T cell phenotypic alterations in human breast cancers are still lacking. This paucity of information is partly due to the challenging scarcity of these cells in surgical specimens. αβ T cell phenotypic alterations occurring in the tumor bed are detectable in the periphery and correlate with adverse clinical outcomes. Thus, we sought to determine through an exploratory study whether Vδ2⁺ γδ T cells phenotypic changes can be detected within breast cancer patients’ peripheral blood, along with association with tumor progression. By using mass cytometry, we quantified 130 immune variables from untreated breast cancer patients’ peripheral blood. Supervised analyses and dimensionality reduction algorithms evidenced circulating Vδ2⁺ γδ T cell phenotypic alterations already established at diagnosis. Foremost, terminally differentiated Vδ2⁺ γδ T cells displaying phenotypes of exhausted senescent T cells associated with lymph node involvement. Thereby, our results support Vδ2⁺ γδ T cells implication in breast cancer pathogenesis and progression, besides shedding light on liquid biopsies to monitor surrogate markers of tumor-infiltrating Vδ2⁺ γδ T cell antitumor activity. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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11 pages, 3668 KiB

Open AccessBrief Report

PVR and ICAM-1 on Blast Crisis CML Stem and Progenitor Cells with TKI Resistance Confer Susceptibility to NK Cells

by Nayoung Kim, Mi-Yeon Kim, Young-Uk Cho, WenYong Chen, Kyoo-Hyung Lee and Hun Sik Kim

Cancers 2020, 12(7), 1923; https://doi.org/10.3390/cancers12071923 - 16 Jul 2020

Cited by 6 | Viewed by 2902

Abstract

The BCR-ABL1 fusion gene generating an oncogenic tyrosine kinase is a hallmark of chronic myeloid leukemia (CML), which can be successfully targeted by BCR-ABL1 tyrosine kinase inhibitors (TKIs). However, treatment-free remission has been achieved in a minority of patients due to evolving TKI [...] Read more.

The BCR-ABL1 fusion gene generating an oncogenic tyrosine kinase is a hallmark of chronic myeloid leukemia (CML), which can be successfully targeted by BCR-ABL1 tyrosine kinase inhibitors (TKIs). However, treatment-free remission has been achieved in a minority of patients due to evolving TKI resistance and intolerance. Primary or acquired resistance to the approved TKIs and progression to blast crisis (BC), thus, remain a major clinical challenge that requires alternative therapeutic strategies. Here, we first demonstrate that donor natural killer (NK) cells prepared using a protocol adopted in clinical trials can efficiently eliminate CML-BC blasts, with TKI resistance regardless of BCR-ABL1 mutations, and preferentially target CD34⁺CD38⁻ leukemic stem cells (LSC), a potential source of disease relapse. Mechanistically, the predominant expression of PVR, a ligand for the NK cell-activating DNAM-1 receptor, in concert with ICAM-1, a ligand for NK cell adhesion, confer this susceptibility to NK cells, despite the lack of ligands for NKG2D, a principal NK cell activating receptor, as an immune evasion mechanism. With these mechanistic insights, our findings provide a proof-of-concept that donor NK cell-based therapy is a viable strategy for overcoming TKI resistance in CML, particularly the advanced, multi-TKI-resistant CML with dismal outcome. Full article

(This article belongs to the Special Issue The Role of NK and T Cells in Cancer)

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