Scientists at Yale University have developed an anticancer platform that harnesses CRISPR technology to activate multiple cancer-related genes, which signals the body’s immune system to recognize, attack, and kill the cancer cells. Initial tests showed that the CRISPR activation (CRISPRa) platform, termed multiplexed activation of endogenous genes as an immunotherapy (MAEGI), effectively reduced or eliminated a range of cancer types in mice, including tumors that were located far away from the primary, treated tumor site.
“This is an entirely new form of immunotherapy,” said Sidi Chen, PhD, assistant professor of genetics and senior author of the team’s study, which is reported in Nature Immunology, in a paper titled, “Multiplexed activation of endogenous genes by CRISPRa elicits potent antitumor immunity.”
Immunotherapies that direct the body’s immune system against tumors have revolutionized cancer therapy. Major types of immunotherapy include checkpoint blockade, adoptive cell transfer, human recombinant cytokines, and cancer vaccines, the authors explained. Immunotherapies recognize the mutant or abnormal peptides that are generated by expression of the cancer cells’ mutated genes. This flags the cells as “nonself,” or foreign, and triggers an immune attack. However, this basic strategy isn’t infallible, and cancers can escape immune cell scrutiny. “Recognition of tumor-associated antigens (TAAs) formed by mutations and dysregulated gene expression programs is an essential step for cancer immunotherapy,” the authors noted. However, “ … these mutant products might not be expressed at levels sufficient to elicit an effective T cell–mediated response, and cancer cells often downregulate antigen presentation to escape immune recognition … the spontaneous immune recognition of tumor antigens is often insufficient to elicit effective immune responses, because the abnormal products may not be adequately presented.”
To address cancer’s immunotherapy escape routes, Chen’s lab developed a technology that combines adeno-associated viral (AAV) gene therapy and CRISPR gene-editing technology. But instead of finding and editing target DNA sites and inserting new genes, the MAEGI approach can search for and highlight cancer-related genes expressed by particular types of cancer cells, and then act like a homing beacon to pinpoint their location and amplify the signals, marking them for immune attack and destruction. The approach effectively turns a cold tumor (lacking immune cells) into a hot tumor (with immune cells), recruiting effector T cells to the tumor and remodeling the tumor microenvironment.
“We reasoned that augmenting the expression and thus the presentation of endogenous antigens in tumors could amplify the ‘nonself’ identity of cancer cells, thereby flagging them for immune destruction,” the team commented. This strategy is the molecular equivalent of dressing tumor cells in highly visible orange jumpsuits, which allows the immune system to quickly find and eradicate them, Chen suggested. “And once those cells are identified, the immune system immediately recognizes them if they show up in the future.”
For their studies reported in Nature Immunology, the Yale University team harnessed CRISPR activation technology, which pairs catalytically inactivated Cas9 (dCas9) with single guide RNAs (sgRNAs), to regulate the expression and presentation of potentially thousands of cancer-specific genes that express immunogenic antigens. They developed a number of iterations of the platform, including genome-scale system, AAV-g-MAEGI, which demonstrated antitumor effects in mice bearing melanoma or pancreatic tumors. “Thus, AAV-mediated delivery of genome-scale MAEGI elicits host immune responses against established tumors across multiple aggressive cancer types,” they wrote.
The researchers also generated an AAV-mediated precision version of the MAEGI system, AAV-p-MAEGI, which incorporated an sgRNA library targeting more than a thousand gene mutations specifically in E0771 breast cancer cells. Tests showed that 44% of E0771 TNBC-bearing animals demonstrated a near complete response (nCR), or complete response (CR) following treatment using AAV-p-MAEGI. Encouragingly, all nine of the AAV-p-MAEGI-treated animals that had exhibited a CR rejected a subsequent rechallenge with E0771 three months after their initial tumor transplantation, and the animals all retained long-term complete remission for more than 180 days, “indicating that AA-p-MAEGI had induced potent and durable antitumor responses,” the authors reported.
The team then tested the system in mice carrying both “local” and “distant” breast tumors. After administration of AAV-p-MAEGI only to the local tumors, antitumor effects were observed both against these local tumors and against the distant tumors that hadn’t directly received AAV-p-MAEGI. “A 67% CR rate was observed at distant tumors after AAV-p-MAEGI treatment,” the authors wrote. Their collective cell and in vivo studies showed that MAEGI effectively increases target gene expression and antigen presentation, increases T cell effector function, recruits CD4+ and CD8+ T cells to the tumor microenvironment, and promotes both local and systemic antitumor T cell responses, the authors commented.
The new system could feasibly be effective against many cancer types, including those currently resistant to immunotherapy. “… we demonstrated that direct activation of endogenous mutant genes through CRISPRa amplifies the ‘nonself’ signals of tumor cells, inducing potent antitumor adaptive immunity,” the investigators concluded. “As a potential viral gene therapy in oncology, administration of AAV-p-MAEGI could be performed to primary tumors, metastatic sites, or residual disease as a surgery adjuvant … Importantly, as AAV-MAEGI activates endogenous genes, the acquisition of additional mutations by the tumor, which may have driven relapse by nullifying existing antitumor immunological memory, can still be targeted by MAEGI.” A next stage of studies will need to optimize the system for simpler manufacturing and prepare for clinical trials in cancer patients.
