Periodic Reporting for period 1 - BRAINHIB (Integrated drug discovery approach to generate brain-penetrant inhibitors of glioblastoma cell proliferation)
Reporting period: 2018-03-01 to 2020-02-29
Glioblastoma multiforme (GBM) is the most common and aggressive brain cancer. Without treatment the average survival following diagnosis is merely 3 months. Although clinically-approved kinase inhibitors present promising features to treat GBM, most of them do not cross the blood brain barrier (BBB), impeding their clinical use in GBM patients. Therefore, GBM is still an unmet medical need. We proposed an agile approach that combines ligand-based drug design of highly-focused compound libraries and first-in-class phenotypic assay for simultaneous screening of BBB potential toxicity and anticancer properties, in an iterative manner. This approach will allow a one-step selection of hit / lead compounds from in-house generated compound libraries that cross the BBB to accelerate the design of CNS-active anticancer drugs. Furthermore, the use of state-of-the-art phenotypic screening techniques facilitates the identification of the kinases and / or pathways that are involved in the observed phenotype, which could result in the discovery of novel GBM oncotargets and unknown modes of action. This highly innovative integrated approach has the potential to speed up preclinical drug discovery efforts in CNS diseases.
The BRAINHIB project focused on the development of novel brain penetrant kinase inhibitors for the treatment of glioblastoma multiforme. According to the DoA, the project was divided into three work packages (WP), each corresponding to one of the project objectives.
The first specific objective of the project was the design and synthesis and novel small molecule kinase inhibitors. To achieve this objective three different scaffolds were used as templates to design three different libraries upon modification of the key positions. Focus was given to the achievement of the physicochemical properties which allow a blood brain barrier penetration, such as pKa, PSA, ClogD, CLogP, HBD and molecular weight. A total of 99 novel compounds divided in 2 libraries, were synthesised.
The second specific objective was the phenotypic screening of compound libraries. To achieve this objective our approach was three fold: A first screening in two commercial glioma cell lines, a second screening in blood brain barrier simplified models and a third screening in patient-derived glioblastoma cells. Results of the first and second screening were used to inform further synthesis, while screening on patient-derived glioma spheroids was used as the validation of hits. Additionally, to newly synthesised compounds, two previously synthesised libraries were tested. A total of 199 final compounds were tested. The results of these screening campaigns informed further synthesis in an iterative manner. Potent hits with submicromolar potency against glioma cells were identified.
The third specific objective was the target deconvolution studies of most potent hits. For this specific objective we took advantage of clear relationships found on kinase profiling studies with a critical tumour driving pathway, to develop structure activity relationships. This research has been recently published as an open access article in Bioorganic and Medicinal Chemistry (Valero T, et al. Bioorg Med Chem 2020 1;28(1):115215). Preliminary results were and are being disseminated across scientific conferences, network groups and collaborators. Further original structures will be assessed for patentability before any kind of disclosure.
The first specific objective of the project was the design and synthesis and novel small molecule kinase inhibitors. To achieve this objective three different scaffolds were used as templates to design three different libraries upon modification of the key positions. Focus was given to the achievement of the physicochemical properties which allow a blood brain barrier penetration, such as pKa, PSA, ClogD, CLogP, HBD and molecular weight. A total of 99 novel compounds divided in 2 libraries, were synthesised.
The second specific objective was the phenotypic screening of compound libraries. To achieve this objective our approach was three fold: A first screening in two commercial glioma cell lines, a second screening in blood brain barrier simplified models and a third screening in patient-derived glioblastoma cells. Results of the first and second screening were used to inform further synthesis, while screening on patient-derived glioma spheroids was used as the validation of hits. Additionally, to newly synthesised compounds, two previously synthesised libraries were tested. A total of 199 final compounds were tested. The results of these screening campaigns informed further synthesis in an iterative manner. Potent hits with submicromolar potency against glioma cells were identified.
The third specific objective was the target deconvolution studies of most potent hits. For this specific objective we took advantage of clear relationships found on kinase profiling studies with a critical tumour driving pathway, to develop structure activity relationships. This research has been recently published as an open access article in Bioorganic and Medicinal Chemistry (Valero T, et al. Bioorg Med Chem 2020 1;28(1):115215). Preliminary results were and are being disseminated across scientific conferences, network groups and collaborators. Further original structures will be assessed for patentability before any kind of disclosure.
The progress beyond the state of the art includes the generation of novel phenotypic assays, a toolkit with the potential to improve the development of CNS-active drugs, and the development of patentable leads / drug candidates that can be applied in the treatment of GBM. This project has confirmed the PI3K/mTOR pathway as druggable axis for glioma treatment, has developed two well populated libraries of active compounds and has inspired new projects. As a potential added value of my research proposal, this pseudo target-agnostic strategy could find bioactive compounds among these libraries that inhibit kinases not previously associated with GBM oncogenesis. Therefore, the identification of such kinases may result in the discovery of novel GBM oncotargets, thereby increasing our understanding of GBM tumorigenicity and expanding the range of therapeutic opportunities available to fight this currently incurable disease.