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With 5-10 newly diagnosed patients per 100,000 people every year, glioblastoma is the most common malignant primary brain tumor. Despite extensive research activity in the last decades, clinical effectiveness of the currently available therapy standard of surgery, radiochemotherapy and tumor-treating fields is still limited and mean survival rates in unselected collectives are only about one year. Accordingly, there is an urgent need to explore new therapeutic options. The current standard of care includes surgery followed by radiation therapy in combination with the alkylating chemotherapeutic agent Temozolomide. Even with successful initial therapy, tumor recurrence is still inevitable. Currently, there are no defined recommendations for clinical management of the disease in the event of tumor recurrence. Only 20-30% of patients qualify for a second surgical resection, while other options include retreatment with Temozolomide, CCNU (Lomustine) or Regorafenib and enrollment in a clinical trial.
The development of immunotherapies for glioblastoma, in particular, has been the focus of intense preclinical and clinical efforts. However, low numbers of mutations and a highly immunosuppressive tumor microenvironment result in glioblastoma being considered an immunologically “cold” tumor. Strategies successfully established in mutagen-induced tumors with antibodies directed against the PD-1, PD-L1 or CTLA-A4 immune checkpoints have therefore failed in glioblastoma.
Cellular immunotherapies based on chimeric antigen receptor (CAR)-technology have emerged as an alternative powerful option to tackle immunologically “cold” tumors. Several CAR-T cell products targeting glioma antigens have been developed and some evidence of clinical activity has been demonstrated. Natural killer (NK) cells as carriers of CAR constructs have several advantages over T cells, including a much lower risk of neurotoxicity and better interaction with immune cells in the microenvironment. Based on the human NK cell line NK-92, a clinical-grade product, suitable as an off-the-shelf therapeutic, has been developed. The NK-92/5.28.z clone (CAR-NK) expresses a CAR based on the HER2-specific antibody FRP5 in addition to signal-enhancing CD28 and CD3ζ domains. Similar to several other tumor entities, overexpression of the growth factor receptor HER2 is often found in glioblastoma patients. Because of its substantial role in the regulation of cell proliferation, survival, differentiation, angiogenesis and invasion, this receptor is classified as an oncogene. HER2 overexpression plays a major role in the malignant transformation of cells and its oncogenic potential has been studied in detail in breast cancer. However, HER2 expression was also found in up to 80% of glioblastomas, which correlates with an impaired probability of survival. Under physiological conditions, HER2 is not expressed in the adult central nervous system, making it a promising target antigen for glioblastoma immunotherapy.
In previous projects, it has already been shown that these CAR-NK cells exhibit a high and specific lytic activity towards HER2+ glioblastoma cells. While repetitive intratumoral injections of CAR-NK cells already significantly extended symptom-free survival in murine orthotopic xenograft models, CAR-NK cell therapy in immunocompetent mice promotes an endogenous anti-tumor immune response which improves tumor control and provides persisting anti-tumor immunity after therapy of early-stage tumors. However, in more advanced tumor models, efficacy is limited and induction of the checkpoint-molecule PD-L1 in response to CAR-NK-cell therapy was identified as a key mechanism of therapy resistance.
Immunotherapy employing the intravenous administration of checkpoint inhibitors has already revolutionized the treatment of various malignant diseases such as melanoma or lung cancer. In particular, the approach of cancer immunotherapy has focused on the systemic administration of antibodies directed against immune checkpoints such as PD-1, PD-L1 and CTLA-4. In glioblastoma, both tumor cells and microglia, the brain-resident macrophages, express PD-L1, which hinders the activation of CD8+ and CD4+ T cells. Therefore, immunotherapy directed against the PD-1/PD-L1 axis represents a promising approach for the treatment of glioblastoma. One problem, however, is the severe toxicity caused by the systemic effects of checkpoint inhibitors, since the immune response is stimulated not only in tumor tissue but also in healthy organs. Serious side effects such as colitis, hepatitis, pancreatitis or hypophysitis, including numerous deaths, have been reported.
This study aimed to improve the efficacy of CAR-NK cell therapy by combining it with adeno-associated virus (AAV)-mediated transfer of anti-PD-1 antibodies as a strategy to enable local combination therapy to control intracranial tumors.
AAVs carrying a payload coding for an anti-PD-1 immunoadhesin (aPD-1) retargeted to HER2-expressing cells by fusion of so-called Designed Ankyrin Repeat Proteins (DARPins) with a viral capsid protein were employed for this to focus checkpoint inhibitor therapy to the tumor area, resulting in high intratumoral and low systemic drug concentrations. ...