On March 3, 2022 Autolus Therapeutics plc (Nasdaq: AUTL), a clinical-stage biopharmaceutical company developing next-generation programmed T cell therapies, reported the publication of an article in BioTechniques describing a novel technology that provides for very low levels of expression of one gene module, while maintaining high levels of expression of other gene modules expressed from the same promotor1 (Press release, Autolus, MAR 3, 2022, View Source [SID1234609482]).
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This technical paper describes a method of achieving very low levels of transgene expression in multi-cistronic mammalian expression systems. This is achieved via the insertion of a stop codon and translational readthrough motif (TRM) between the transgenes of an mRNA encoding a multi-cistronic cassette. The TRM helps to suppress the stop codon, facilitating continued translation of the downstream transgene at reduced levels compared with the upstream transgene. This system addresses a fundamental challenge in cell therapy when highly potent receptors, cytokines or toxins are expressed, which, at normal levels of expression, would be unsafe for patients.
"Throughout the history of gene-therapy the primary focus was on engineering mammalian expression cassettes driving high levels of transgene expression," said James Sillibourne, director of synthetic genomics at Autolus. "However, with a very potent or toxic transgene, you need a very low level of expression. Up until now, an easy way of achieving this was not available. Building on mechanisms known from bacteria and viruses, we have developed a reliable way of tightly controlling low levels of transgene expression."
IL-12 is a potent anti-tumor cytokine. However, the majority of clinical studies involving treatment of patients with IL-12 have been associated with severe systemic side effects and significant toxicities for patients.
"Our approach to solid tumors combines multiple gene modules in CAR T cells to drive the desired set of properties we believe are essential to maximize anti-tumor activity without increasing toxicity. Selectively adjusting expression levels became an important technology to establish therapeutic windows," added Martin Pule, chief scientific officer and founder of Autolus. "In this paper we successfully applied this technology for highly restricted IL-12 release which increases CAR T anti-tumor activity in an immunocompetent mouse model without inducing systemic toxicity."
1. Sillibourne JE, Agliardi G, Righi M et al. A compact and simple method of achieving differential transgene expression by exploiting translational readthrough. BioTechniques doi: 10.2144/btn-2021-0079 (2022) (Epub ahead of print). The full publication in BioTechniques can be viewed here.