Generation of Knock-out Primary and Expanded Human NK Cells Using Cas9 Ribonucleoproteins

Kararoudi, Meisam Naeimi; Dolatshad, Hamid; Trikha, Prashant; Hussain, Syed-Rehan A.; Elmas, Ezgi; Foltz, Jennifer A.; Moseman, Jena E.; Thakkar, Aarohi; Nakkula, Robin J.; Lamb, Margaret; Chakravarti, Nitin; McLaughlin, K. John; Lee, Dean A.

doi:10.3791/58237

Cited by 50 publications

(43 citation statements)

References 19 publications

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“…A major next wave of NK cell immunotherapy will very likely involve the genetic modification of NK cells to overcome specific inherent weakness or enable NK cells new function. Viral and non‐viral approaches to gene modification of NK cells may find application if the NK cells can be propagated to large numbers after small‐scale genetic engineering …”

Section: Lessons Learned and Continued Controversiesmentioning

confidence: 99%

Cellular therapy: Adoptive immunotherapy with expanded natural killer cells

Lee

2019

Immunological Reviews

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Adoptive immunotherapy with natural killer cells was pioneered 30 years ago in human clinical trials with the development of cytokine‐induced killer cells—unfractionated peripheral blood mononuclear cell (PBMC) populations activated overnight with IL‐2. Higher doses were subsequently made possible through the advent of steady‐state apheresis, allowing the collection of PBMC numbers equivalent to an entire adult blood volume, and increased purity made feasible through magnetic CD3‐depletion and/or CD56‐selection methods. Still, these approaches rarely achieved clinical dosing above a single infusion of 108 NK cells/kg, except with substantial donor‐recipient size mismatch (eg, parents donating cells to children). To address this shortcoming, leukemia cell lines with NK cell‐like function or ex vivo expansion approaches centered on the homeostatic cytokine IL‐15 were developed. Here, we describe the development of an ex vivo expansion system based on a feeder cell expressing membrane‐bound IL‐21 that enables log‐phase growth of primary NK cells for many weeks without inducing senescence, and describe the biology, correlative science, and translation to clinical trials for patients with leukemia, brain tumors, and solid tumors.

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Section: Lessons Learned and Continued Controversiesmentioning

confidence: 99%

Cellular therapy: Adoptive immunotherapy with expanded natural killer cells

Lee

2019

Immunological Reviews

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“…Immune NK cells play an important role in host immunity against cancer and viral infections. Despite the low efficiency of viral and non-viral delivery methods, several NK cells can be edited to enhance their persistence, cytotoxicity, and tumor targeting (117). Dendritic cells (DCs) play a critical role in T-cell response instructions, with triple knockout established as proof of concept (118).…”

Section: Advances In Genetic Engineering Of Immune Cellsmentioning

confidence: 99%

Using Gene Editing Approaches to Fine-Tune the Immune System

et al. 2020

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Genome editing technologies not only provide unprecedented opportunities to study basic cellular system functionality but also improve the outcomes of several clinical applications. In this review, we analyze various gene editing techniques used to finetune immune systems from a basic research and clinical perspective. We discuss recent advances in the development of programmable nucleases, such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat (CRISPR)-Cas-associated nucleases. We also discuss the use of programmable nucleases and their derivative reagents such as base editing tools to engineer immune cells via gene disruption, insertion, and rewriting of T cells and other immune components, such natural killers (NKs) and hematopoietic stem and progenitor cells (HSPCs). In addition, with regard to chimeric antigen receptors (CARs), we describe how different gene editing tools enable healthy donor cells to be used in CAR T therapy instead of autologous cells without risking graft-versus-host disease or rejection, leading to reduced adoptive cell therapy costs and instant treatment availability for patients. We pay particular attention to the delivery of therapeutic transgenes, such as CARs, to endogenous loci which prevents collateral damage and increases therapeutic effectiveness. Finally, we review creative innovations, including immune system repurposing, that facilitate safe and efficient genome surgery within the framework of clinical cancer immunotherapies.

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“…To overcome the suppressive effects of TGFβ on NK cells, Foltz et al have shown that TGFβ imprinting during NK cell activation and expansion decreases NK cell sensitivity to TGFβ suppression [117]. CRISPR modification of primary and expanded NK cells using electroporation of Cas9/RNP targeting TGFBR2 also has proven to be an effective mechanism to blunt NK cell inhibition upon exposure to TGFβ [118]. Using the CRISPR modification approach, there is evidence showing that gene-modified primary NK cells have enhanced antitumor effects, including SOCS3 and CISH knockout NK cells [119][120][121].…”

Section: Natural Killer Cell-based Therapiesmentioning

confidence: 99%

Immunotherapies for pediatric cancer: current landscape and future perspectives

Hutzen

Paudel

Kararoudi

et al. 2019

Cancer Metastasis Rev

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The advent of immunotherapy has revolutionized how we manage and treat cancer. While the majority of immunotherapy-related studies performed to date have focused on adult malignancies, a handful of these therapies have also recently found success within the pediatric space. In this review, we examine the immunotherapeutic agents that have achieved the approval of the US Food and Drug Administration for treating childhood cancers, highlighting their development, mechanisms of action, and the lessons learned from the seminal clinical trials that ultimately led to their approval. We also shine a spotlight on several emerging immunotherapeutic modalities that we believe are poised to have a positive impact on the treatment of pediatric malignancies in the near future.

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Generation of Knock-out Primary and Expanded Human NK Cells Using Cas9 Ribonucleoproteins

Cited by 50 publications

References 19 publications

Cellular therapy: Adoptive immunotherapy with expanded natural killer cells

Cellular therapy: Adoptive immunotherapy with expanded natural killer cells

Using Gene Editing Approaches to Fine-Tune the Immune System

Immunotherapies for pediatric cancer: current landscape and future perspectives

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