ABSTRACT Natural Killer cells are immune leukocytes required for responses against tumor cells and virus-infected cells. In the last decade, natural killer cells have emerged as promising tools in cancer therapy, and clinical studies on patients treated with natural killer cells have revealed increased rates of disease-free survival. In this article, we review results from the major clinical trials that have used natural killer cells for cancer treatment, including their global distribution. We also discuss the major mechanisms of natural killer cell activation and expansion and focus on the advantages and disadvantages of each mechanism for clinical applications. Although natural killer cells can be isolated from several sources, primary natural killer cells are most commonly used in clinical trials. However, the frequency of natural killer cells available in peripheral and cord blood is low, necessitating development of methods for expansion of natural killer cells for clinical use. The development of a platform for the expansion of large-scale good manufacturing practice-compliant natural killer cells has limitations as several methods for natural killer cell activation and expansion yield conflicting results. Only techniques using feeder cells can produce large numbers of cells, allowing the “off-the-shelf” use of natural killer cells. However, advances in cell culture have supported the development of feeder-free platforms for natural killer cell expansion, which is fundamental for improving the safety of this type of cell therapy. virusinfected virus infected decade therapy diseasefree disease free survival article treatment distribution applications sources However low largescale scale practicecompliant practice compliant offtheshelf off shelf “off-the-shelf feederfree

ABSTRACT Objective Phase 1 clinical trial to determine feasibility, safety, and efficacy of a new advanced cell therapy product for treatment of knee articular cartilage injuries. Methods Three participants with knee focal chondral lesions were included, with no signs of osteoarthritis. Chondrocytes were obtained through knee arthroscopy, cultured in collagen membrane for 3 weeks at the laboratory, subjected to tests to release the cell therapy product, and implanted. All patients underwent a specific 3-month rehabilitation protocol, followed by assessments using functional and imaging scales. The main outcome was the incidence of severe adverse events. Results Three participants were included and completed the 2-year follow-up. There was one severe adverse event, venous thrombosis of distal leg veins, which was no associated with therapy, was treated and left no sequelae. The clinical and radiological scales showed improvement in the three cases. Conclusion The preliminary results, obtained with the described methodology, allow concluding that this product of advanced cell therapy is safe and feasible. ReBEC platform registration number: RBR-6fgy76

ABSTRACT To describe a case of autologous chondrocyte implantation after cell culture contamination by Mycoplasma pneumoniae and the measures taken to successfully complete cell therapy in a patient with focal chondral lesion. A 45-year-old male patient, complaining of chronic pain on the knee and no history of trauma. He had a chondral lesion in the trochlear region of the femur and clinical tests compatible with pain in the anterior compartment of the knee. Conservative treatment failed to alleviate symptoms. Surgical treatment was indicated, but due to the size of the lesion, membrane-assisted autologous chondrocyte implantation was the technique of choice. Cartilage biopsies were collected from the intercondylar region of the distal femur. After isolation, chondrocytes were expanded ex vivo in a trained laboratory, for three weeks, and seeded onto a commercially available collagen membrane prior to implantation in the knee. Two days before surgery, a cell culture sample tested positive for Mycoplasma pneumoniae. The source of contamination was found to be autologous blood serum, extracted from the patient´s peripheral vein, and used to supplement the cell culture medium. After treating the patient with antibiotics, all procedures were repeated and the new final cell product, free from contaminants, was successfully implanted. We discuss the strategies available to deal with this situation, and describe the results of this particular case, which led to modifications in the autologous chondrocyte implant protocol.