Identification of 6-Anilino Imidazo[4,5-]pyridin-2-ones as Selective DNA-Dependent Protein Kinase Inhibitors and Their Application as Radiosensitizers.

The dominant role of non-homologous end-joining in the repair of radiation-induced double-strand breaks identifies DNA-dependent protein kinase (DNA-PK) as an excellent target for the development of radiosensitizers. We report the discovery of a new class of imidazo[4,5-]pyridine-2-one DNA-PK inhibitors. Structure-activity studies culminated in the identification of as a nM DNA-PK inhibitor with excellent selectivity for DNA-PK compared to related phosphoinositide 3-kinase (PI3K) and PI3K-like kinase (PIKK) families and the broader kinome, and displayed DNA-PK-dependent radiosensitization of HAP1 cells.

Design, Synthesis and Anticancer Evaluation of Nitroimidazole Radiosensitisers.

The role of hypoxic tumour cells in resistance to radiotherapy, and in suppression of immune response, continues to endorse tumour hypoxia as a bona fide, yet largely untapped, drug target. Radiotherapy innovations such as stereotactic body radiotherapy herald new opportunities for classical oxygen-mimetic radiosensitisers. Only nimorazole is used clinically as a radiosensitiser, and there is a dearth of new radiosensitisers in development.

Spin Trapping Hydroxyl and Aryl Radicals of One-Electron Reduced Anticancer Benzotriazine 1,4-Dioxides.

Hypoxia in tumors results in resistance to both chemotherapy and radiotherapy treatments but affords an environment in which hypoxia-activated prodrugs (HAP) are activated upon bioreduction to release targeted cytotoxins. The benzotriazine 1,4-di--oxide (BTO) HAP, tirapazamine (TPZ, ), has undergone extensive clinical evaluation in combination with radiotherapy to assist in the killing of hypoxic tumor cells. Although compound did not gain approval for clinical use, it has spurred on the development of other BTOs, such as the 3-alkyl analogue, SN30000, .

Radiosensitisation of SCCVII tumours and normal tissues in mice by the DNA-dependent protein kinase inhibitor AZD7648.

Background And Purpose: Inhibitors of DNA-dependent protein kinase (DNA-PK) are effective radiation sensitisers in preclinical tumours, but little is known about risks of normal tissue radiosensitisation. Here, we evaluate radiosensitisation of head and neck squamous cell carcinoma (HNSCC) cells by DNA-PK inhibitor AZD7648 under oxia and anoxia in vitro, and tumour (SCCVII), oral mucosa and small intestine in mice.

Materials And Methods: Radiosensitisation of human (UT-SCC-54C) and murine (SCCVII) HNSCC cells by AZD7648 under oxia and anoxia was evaluated by clonogenic assay.

Spatially-resolved pharmacokinetic/pharmacodynamic modelling of bystander effects of a nitrochloromethylbenzindoline hypoxia-activated prodrug.

Purpose: Hypoxia-activated prodrugs (HAPs) have the potential for eliminating chemo- and radiation-resistant hypoxic tumour cells, but their activity is often compromised by limited penetration into hypoxic zones. Nitrochloromethylbenzindoline (nitroCBI) HAPs are reduced in hypoxic cells to highly cytotoxic DNA minor groove alkylating aminoCBI metabolites. In this study, we investigate whether a lead nitroCBI, SN30548, generates a significant bystander effect through the diffusion of its aminoCBI metabolite and whether this compensates for any diffusion limitations of the prodrug in tumour tissue.

Schedule-dependent potentiation of chemotherapy drugs by the hypoxia-activated prodrug SN30000.

Hypoxia-activated prodrugs (HAPs) are hypothesized to improve the therapeutic index of chemotherapy drugs that are ineffective against tumor cells in hypoxic microenvironments. SN30000 (CEN-209) is a benzotriazine di--oxide HAP that potentiates radiotherapy in preclinical models, but its combination with chemotherapy has not been explored. Here we apply multiple models (monolayers, multicellular spheroids and tumor xenografts) to identify promising SN30000/chemotherapy combinations (with chemotherapy drugs before, during or after SN30000 exposure).

Functional CRISPR and shRNA Screens Identify Involvement of Mitochondrial Electron Transport in the Activation of Evofosfamide.

Evofosfamide (TH-302) is a hypoxia-activated DNA-crosslinking prodrug currently in clinical development for cancer therapy. Oxygen-sensitive activation of evofosfamide depends on one-electron reduction, yet the reductases that catalyze this process in tumors are unknown. We used RNA sequencing, whole-genome CRISPR knockout, and reductase-focused short hairpin RNA screens to interrogate modifiers of evofosfamide activation in cancer cell lines.

An Intratumor Pharmacokinetic/Pharmacodynamic Model for the Hypoxia-Activated Prodrug Evofosfamide (TH-302): Monotherapy Activity is Not Dependent on a Bystander Effect.

Tumor hypoxia contributes to resistance to anticancer therapies. Hypoxia-activated prodrugs (HAPs) selectively target hypoxic cells and their activity can extend to well-oxygenated areas of tumors via diffusion of active metabolites. This type of bystander effect has been suggested to be responsible for the single agent activity of the clinical-stage HAP evofosfamide (TH-302) but direct evidence is lacking.

Bystander Effects of Hypoxia-Activated Prodrugs: Agent-Based Modeling Using Three Dimensional Cell Cultures.

Intra-tumor heterogeneity represents a major barrier to anti-cancer therapies. One strategy to minimize this limitation relies on bystander effects via diffusion of cytotoxins from targeted cells. Hypoxia-activated prodrugs (HAPs) have the potential to exploit hypoxia in this way, but robust methods for measuring bystander effects are lacking.

Evofosfamide for the treatment of human papillomavirus-negative head and neck squamous cell carcinoma.

Evofosfamide (TH-302) is a clinical-stage hypoxia-activated prodrug of a DNA-crosslinking nitrogen mustard that has potential utility for human papillomavirus (HPV) negative head and neck squamous cell carcinoma (HNSCC), in which tumor hypoxia limits treatment outcome. We report the preclinical efficacy, target engagement, preliminary predictive biomarkers and initial clinical activity of evofosfamide for HPV-negative HNSCC. Evofosfamide was assessed in 22 genomically characterized cell lines and 7 cell line-derived xenograft (CDX), patient-derived xenograft (PDX), orthotopic, and syngeneic tumor models.

Next-Generation Hypoxic Cell Radiosensitizers: Nitroimidazole Alkylsulfonamides.

Innovations in the field of radiotherapy such as stereotactic body radiotherapy, along with the advent of radio-immuno-oncology, herald new opportunities for classical oxygen-mimetic radiosensitizers. The role of hypoxic tumor cells in resistance to radiotherapy and in suppression of immune response continues to endorse tumor hypoxia as a bona fide, yet largely untapped, drug target. Only nimorazole is used clinically as a radiosensitizer, and there is a dearth of new radiosensitizers in development.

Radical Chemistry and Cytotoxicity of Bioreductive 3-Substituted Quinoxaline Di-N-Oxides.

The radical chemistry and cytotoxicity of a series of quinoxaline di-N-oxide (QDO) compounds has been investigated to explore the mechanism of action of this class of bioreductive drugs. A series of water-soluble 3-trifluoromethyl (4-10), 3-phenyl (11-19), and 3-methyl (20-21) substituted QDO compounds were designed to span a range of electron affinities consistent with bioreduction. The stoichiometry of loss of QDOs by steady-state radiolysis of anaerobic aqueous formate buffer indicated that one-electron reduction of QDOs generates radicals able to initiate chain reactions by oxidation of formate.

Efficient Protocol for the Identification of Hypoxic Cell Radiosensitisers.

An evolution in radiotherapy practice is leading to greater use of stereotactic body radiotherapy (SBRT), raising the prospect of increased hypoxic cell radioresistance. New clinical interest in nitroimidazole radiosensitisers, combined with appropriate biomarkers, signals a revival for radiosensitisers in the context of SBRT. Our interest in modifiers of radiation therapy led us to revisit this area and we have identified a new class of nitroimidazole radiosensitiser.

Novel nitroimidazole alkylsulfonamides as hypoxic cell radiosensitisers.

A novel class of nitroimidazole alkylsulfonamides have been prepared and evaluated as hypoxia-selective cytotoxins and radiosensitisers. The sulfonamide side chain markedly influences the physicochemical properties of the analogues: lowering aqueous solubility and raising the electron affinity of the nitroimidazole group. The addition of hydroxyl or basic amine groups increased aqueous solubility, with charged amine groups contributing to increased electron affinity.