Materials for Improving Immune Cell Transfection

Abstract

recognize and bind to tumor-associated antigens such as CD19 and epidermal growth factor receptor (EGFR) before releasing cytotoxic granules and effector cytokines to destroy cancer cells. [3] CAR-T therapy has shown remarkable success against CD19 expressing B cell hematological malignancies and, thus far, five products (Kymriah from Novartis; Yescarta and Tecartus from Kite Pharma/Gilead Sciences; and Breyanzi and Abecma from Bristol-Myers Squibb) have been approved by the US Food and Drug Administration (FDA) for clinical use. [4,5] Currently, there are at least 500 clinical trials registered on ClinicalTrials.gov database using engineered immune cells to treat various cancers including that of the brain, lungs, and skin.Despite the clinical success of CAR-T cells against B cell leukemia, CAR-T therapy still face tremendous challenges in manufacturing, safety, and affordability before it can become a viable clinical option. [6] One of the biggest technical difficulties is to transfect sensitive, primary T cells whereby biomolecules like oligonucleotides and proteins have to be delivered intracellularly and into the nuclei of cells. FDA-approved gold standard viruses and bulk electroporation suffer from low transfection efficiency while also perturbing the critical biological attributes of cells such as proliferation, metabolism, and gene expression. [7] This increases the time and costs for cell expansion, and without an efficient and cost-friendly transfection technology, the price (between USD 0.4-0.5 million per patient) for FDA-approved CAR treatments (Kymriah and Yescarta) will remain unaffordable and untimely. [8] The limitations in conventional transfection techniques have motivated the development of micro-and nanoplatforms such as microfluidics, nanoparticles, and high-aspect-ratio nanostructures to improve immune cell viability and throughput during transfection.Herein, we first provide an overview of CAR-T cell manufacturing, with emphasis on the science of transfection and limitations of traditional technology using viruses and bulk electroporation. There will also be discussion of other cell-based cancer immunotherapy using other types of promising immune cell types. Next, we describe emerging transfection platforms and companies that have been established to overcome gaps in CAR-T transfection. We then provide a list of assays constituting the polyfunctionalities of immune cells that we believe will help the field better assess the robustness and suitability of their transfection methods. Finally, we end off with existing challenges in CAR-T transfection and how overcoming these challenges can significantly enhance the clinical impact of CAR-T therapy.Chimeric antigen receptor T cell (CAR-T) therapy holds great promise for preventing and treating deadly diseases such as cancer. However, it remains challenging to transfect and engineer primary immune cells for clinical cell manufacturing. Conventional tools using viral vectors and bulk electroporation suffer from low efficiency while posing risks ...

High-throughput and high-content bioassay enables tuning of polyester nanoparticles for cellular uptake, endosomal escape, and systemic in vivo delivery of mRNA

“…These cell lines were chosen to validate our assay in a diverse range of cell types. In particular, B16-F10 cells were included as a model cancer cell line that is frequently used for mouse tumor studies ( 28 , 29 ); RAW macrophages were included to investigate gene delivery to immune cells, which many studies have demonstrated pose additional intracellular barriers to transfection ( 30 , 31 ); and 3T3 fibroblasts were included as a noncancerous, healthy control cell line. Our results showed highest mRNA transfection levels in B16-F10 cells, medium transfection in RAW 264.7 macrophages, and lowest transfection levels in NIH/3T3 fibroblasts (Fig.…”

Construction of the waaF Subunit and DNA Vaccine Against Escherichia coli in Cow Mastitis and Preliminary Study on Their Immunogenicity

“…There are many methods of transfection, and electroporation and liposome transfection are routinely used. Among them, electroporation has the highest transfection efficiency, which can reach more than 90% ( 33 ), but it can interfere with the key biological properties of cells, such as proliferation, metabolism, and gene expression ( 34 ). If the conditions of liposome transfection are optimized, the effect of liposome transfection is better than that of electroporation ( 35 ).…”

The start-ups taking nanoneedles into the clinic

“…Emerging researchers deserve active support to develop industry-relevant skills in the process of translating their findings, including the culture of entrepreneurship and investment, advanced manufacturing practice, and how to engage effectively with potential players. ❐ Roey Elnathan 1,2,3 ✉ , Andy Tay 4 , Nicolas H. Voelcker 2,3 and Ciro Chiappini using our company's CO 2 electrolysers as a model, we describe the challenges involved in incorporating nanomaterial catalysts into industrial-scale electrolysers and suggest ways to more efficiently realize the performance improvements of academic-scale novel nanomaterials at industrial scales.…”

“…Among these nanotechnologies, nanoneedles -vertical arrays of highaspect-ratio nanomaterials -have emerged as a simple, controllable and powerful tool to efficiently access cells with minimal perturbation 2 . Nanoneedles are rapidly emerging as competitive solutions for sensing 3 , and offer a path to transforming gene and cell therapies 4,5 . Realizing the transformative potential of nanoneedles is a work in progress.…”