Enhancement of anti-sarcoma immunity by NK cells engineered with mRNA for expression of a EphA2-targeted CAR

IF 7.9 1区医学 Q1 MEDICINE, RESEARCH & EXPERIMENTAL

Clinical and Translational Medicine Pub Date : 2025-01-06 DOI:10.1002/ctm2.70140

Pui Yeng Lam, Natacha Omer, Josh K. M. Wong, Cui Tu, Louisa Alim, Gustavo R. Rossi, Maria Victorova, Hannah Tompkins, Cheng-Yu Lin, Ahmed M. Mehdi, Amos Choo, Melissa R. Elliott, Elaina Coleborn, Jane Sun, Timothy Mercer, Orazio Vittorio, Lachlan J. Dobson, Alexander D. McLellan, Andrew Brooks, Zewen Kelvin Tuong, Seth W. Cheetham, Wayne Nicholls, Fernando Souza-Fonseca-Guimaraes

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These cancers share common challenges, including high rates of metastasis, recurrence or treatment resistance, leading to a 5-year survival rate of approximately 20% for patients with advanced disease stages. Despite the critical need, therapeutic advancements have been limited over the past three decades. The advent of chimeric antigen receptor (CAR)-based immunotherapies offers a promising avenue for novel treatments. However, CAR-T cells have faced significant challenges and limited success in treating solid tumours due to issues such as poor tumour infiltration, immunosuppressive tumour microenvironments and off-target effects. 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Transient messenger RNA (mRNA) transfection of NK cells is a safe approach in genetic engineering, with further chemical modifications to mRNA enhancing stability of temporal EphA2-CAR expression in NK cells, thereby promoting prolonged protein expression. Additionally, in vivo EphA2-CAR-NK cells showed promising anti-cancer activity in rhabdomyosarcoma and osteosarcoma mouse models.</p>\n </section>\n \n <section>\n \n <h3> Conclusions</h3>\n \n <p>The study provides a foundational basis for the clinical evaluation of EphA2-targeted CAR-NK cell therapy across a spectrum of paediatric sarcomas. The enhanced anti-tumour effects observed in vitro/vivo suggests potential for improved therapeutic outcomes in hard-to-cure paediatric sarcomas.</p>\n </section>\n \n <section>\n \n <h3> Key points</h3>\n \n <div>\n <ul>\n \n <li>Addressing unmet clinical needs in paediatric Sarcomas. Paediatric sarcomas, including rhabdomyosarcoma, Ewing sarcoma, and osteosarcoma, exhibit poor survival rates in advanced disease stages. The lack of significant therapeutic progress over the past three decades necessitates innovative treatment approaches.</li>\n \n <li>Advancing immunotherapy with CAR-NK cells. Natural killer (NK) cells modified with chimeric antigen receptors (CARs) represent a promising strategy to overcome the limitations of CAR-T cells, particularly in solid tumours. CAR-NK cells are associated with enhanced tumour targeting, reduced off-target effects, and improved safety profiles.</li>\n \n <li>EphA2 as a therapeutic target. EphA2, a receptor overexpressed in multiple paediatric sarcomas, is identified as a viable target for CAR-based immunotherapy due to its critical role in tumour progression and angiogenesis.</li>\n \n <li>Innovations in mRNA-based engineering. 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引用次数: 0

Abstract

Background

Paediatric sarcomas, including rhabdomyosarcoma, Ewing sarcoma and osteosarcoma, represent a group of malignancies that significantly contribute to cancer-related morbidity and mortality in children and young adults. These cancers share common challenges, including high rates of metastasis, recurrence or treatment resistance, leading to a 5-year survival rate of approximately 20% for patients with advanced disease stages. Despite the critical need, therapeutic advancements have been limited over the past three decades. The advent of chimeric antigen receptor (CAR)-based immunotherapies offers a promising avenue for novel treatments. However, CAR-T cells have faced significant challenges and limited success in treating solid tumours due to issues such as poor tumour infiltration, immunosuppressive tumour microenvironments and off-target effects. In contrast, the adaptation of CAR technology for natural killer (NK) cells has demonstrated potential in both haematological and solid tumours, suggesting a new therapeutic strategy for paediatric sarcomas.

Methods

This study developed and validated a novel CAR-NK cell therapy targeting the ephrin type-A receptor-2 (EphA2) antigen, which is highly expressed in various paediatric sarcomas.

Results

CAR expression was successfully detected on the surface of NK cells post-electroporation, indicating successful transfection. Significantly, EphA2-specific CAR-NK cells demonstrated enhanced cytotoxic activity against several paediatric sarcoma cell lines in vitro, including those of rhabdomyosarcoma, Ewing sarcoma and osteosarcoma, compared to unmodified NK cells. Transient messenger RNA (mRNA) transfection of NK cells is a safe approach in genetic engineering, with further chemical modifications to mRNA enhancing stability of temporal EphA2-CAR expression in NK cells, thereby promoting prolonged protein expression. Additionally, in vivo EphA2-CAR-NK cells showed promising anti-cancer activity in rhabdomyosarcoma and osteosarcoma mouse models.

Conclusions

The study provides a foundational basis for the clinical evaluation of EphA2-targeted CAR-NK cell therapy across a spectrum of paediatric sarcomas. The enhanced anti-tumour effects observed in vitro/vivo suggests potential for improved therapeutic outcomes in hard-to-cure paediatric sarcomas.

Key points

Addressing unmet clinical needs in paediatric Sarcomas. Paediatric sarcomas, including rhabdomyosarcoma, Ewing sarcoma, and osteosarcoma, exhibit poor survival rates in advanced disease stages. The lack of significant therapeutic progress over the past three decades necessitates innovative treatment approaches.
Advancing immunotherapy with CAR-NK cells. Natural killer (NK) cells modified with chimeric antigen receptors (CARs) represent a promising strategy to overcome the limitations of CAR-T cells, particularly in solid tumours. CAR-NK cells are associated with enhanced tumour targeting, reduced off-target effects, and improved safety profiles.
EphA2 as a therapeutic target. EphA2, a receptor overexpressed in multiple paediatric sarcomas, is identified as a viable target for CAR-based immunotherapy due to its critical role in tumour progression and angiogenesis.
Innovations in mRNA-based engineering. This study demonstrates the feasibility of transient mRNA transfection to engineer NK cells for CAR expression, offering a non-integrative and safer alternative to viral transduction. Enhancements in mRNA stability through chemical modifications, can further optimise protein expression.
Preclinical efficacy of EphA2-CAR NK cells. EphA2-specific CAR-NK cells exhibit superior cytotoxicity against sarcoma cell lines in vitro and demonstrate significant anti-tumour activity in in vivo mouse models of rhabdomyosarcoma and osteosarcoma.
Clinical translation potential. The findings establish a strong preclinical rationale for the clinical evaluation of EphA2-targeted CAR-NK therapy as a novel immunotherapeutic option for paediatric sarcomas.
Future research directions: Combining EphA2-CAR NK cells with immune checkpoint inhibitors or other immunomodulatory agents could further enhance therapeutic outcomes and durability. Advanced preclinical models mimicking human tumour microenvironments are needed to refine and optimise this therapeutic approach.

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用mRNA表达epha2靶向CAR的NK细胞增强抗肉瘤免疫

背景：儿童肉瘤，包括横纹肌肉瘤、尤文氏肉瘤和骨肉瘤，是一组恶性肿瘤，对儿童和年轻人的癌症相关发病率和死亡率有重要影响。这些癌症面临着共同的挑战，包括高转移率、复发率或治疗耐药性，导致疾病晚期患者的5年生存率约为20%。尽管有迫切的需求，但在过去的三十年里，治疗方面的进展有限。基于嵌合抗原受体（CAR）的免疫疗法的出现为新型治疗提供了一条有希望的途径。然而，由于肿瘤浸润不良、免疫抑制肿瘤微环境和脱靶效应等问题，CAR-T细胞在治疗实体肿瘤方面面临着巨大的挑战和有限的成功。相比之下，CAR技术对自然杀伤（NK）细胞的适应性已经在血液病和实体肿瘤中显示出潜力，为儿科肉瘤提供了一种新的治疗策略。方法：本研究开发并验证了一种靶向ephrin a型受体-2 （EphA2）抗原的新型CAR-NK细胞疗法，该抗原在多种小儿肉瘤中高度表达。结果：电穿孔后NK细胞表面成功检测到CAR表达，表明转染成功。值得注意的是，与未修饰的NK细胞相比，epha2特异性CAR-NK细胞在体外对几种儿科肉瘤细胞系（包括横纹肌肉瘤、尤文氏肉瘤和骨肉瘤）显示出增强的细胞毒活性。转染NK细胞的瞬时信使RNA （mRNA）在基因工程中是一种安全的方法，对mRNA进行进一步的化学修饰可以增强NK细胞中EphA2-CAR暂时表达的稳定性，从而促进蛋白表达的延长。此外，EphA2-CAR-NK细胞在小鼠横纹肌肉瘤和骨肉瘤模型中显示出良好的抗癌活性。结论：本研究为epha2靶向CAR-NK细胞治疗儿科肉瘤的临床评估提供了基础。在体外/体内观察到的增强的抗肿瘤作用表明，在难以治愈的儿科肉瘤中，有可能改善治疗结果。重点：解决儿科肉瘤未满足的临床需求。小儿肉瘤，包括横纹肌肉瘤、尤文氏肉瘤和骨肉瘤，在疾病晚期表现出较低的生存率。在过去的三十年中缺乏显著的治疗进展需要创新的治疗方法。推进CAR-NK细胞免疫治疗。嵌合抗原受体修饰的自然杀伤细胞（NK）代表了克服CAR-T细胞局限性的一种有希望的策略，特别是在实体肿瘤中。CAR-NK细胞与增强肿瘤靶向性、减少脱靶效应和提高安全性有关。EphA2作为治疗靶点。EphA2是一种在多种小儿肉瘤中过表达的受体，由于其在肿瘤进展和血管生成中的关键作用，被确定为基于car的免疫治疗的可行靶点。基于mrna的工程创新。该研究证明了瞬时转染mRNA以工程NK细胞表达CAR的可行性，为病毒转导提供了一种非整合且更安全的替代方法。通过化学修饰增强mRNA的稳定性，可以进一步优化蛋白质表达。EphA2-CAR NK细胞的临床前疗效。epha2特异性CAR-NK细胞在体外对肉瘤细胞系表现出优越的细胞毒性，并在体内小鼠横纹肌肉瘤和骨肉瘤模型中表现出显著的抗肿瘤活性。临床翻译潜力。这些发现为临床评估epha2靶向CAR-NK疗法作为儿科肉瘤的一种新的免疫治疗选择奠定了强有力的临床前理论基础。未来研究方向：EphA2-CAR NK细胞联合免疫检查点抑制剂或其他免疫调节剂可进一步提高治疗效果和持久性。需要模拟人类肿瘤微环境的先进临床前模型来完善和优化这种治疗方法。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Clinical and Translational Medicine Multiple-

CiteScore

15.90

自引率

1.90%

发文量

450

审稿时长

4 weeks

期刊介绍： Clinical and Translational Medicine (CTM) is an international, peer-reviewed, open-access journal dedicated to accelerating the translation of preclinical research into clinical applications and fostering communication between basic and clinical scientists. It highlights the clinical potential and application of various fields including biotechnologies, biomaterials, bioengineering, biomarkers, molecular medicine, omics science, bioinformatics, immunology, molecular imaging, drug discovery, regulation, and health policy. With a focus on the bench-to-bedside approach, CTM prioritizes studies and clinical observations that generate hypotheses relevant to patients and diseases, guiding investigations in cellular and molecular medicine. The journal encourages submissions from clinicians, researchers, policymakers, and industry professionals.