DNA adduct formation leading to kidney failure

Devon Barnes, Department of Pharmaceutical Sciences, Utrecht University

Manoe Janssen, Department of Pharmaceutical Sciences, Utrecht University

Rosalinde Masereeuw,  Department of Pharmaceutical Sciences, Utrecht University

Huan Yang, esqLABS GmbH

This Adverse Outcome Pathway (AOP) depicts a possible sequence of key events that associate how DNA adduct formation caused by platinum anticancer drugs can induce tubular necrosis, resulting in the occurrence of kidney failure as the adverse outcome (AO). Currently, cisplatin, carboplatin and oxaliplatin are the three most utilised Pt-based drugs used globally for the treatment of cancer. The cytotoxicity of these agents are primarily determined by their DNA adducts. Increased intracellular concentrations following uptake of platinum anticancer agents into the nephron, mostly by proximal convoluted tubules, evoking a cascade of negative cellular responses that can cause tubular necrosis, potentially resulting in kidney failure. The aquation of platinum anticancer agents following cellular uptake produces electrophilic intermediates that covalently bind to nucleophilic sites on DNA to form adducts that represent the molecular initiating event (MIE). The nephrotoxic response following the formation of DNA adducts leads to DNA damage (KE1) and mitochondrial dysfunction (KE2). These events promote the release of reaction oxygen species (ROS) (KE3) to induce oxidative stress (KE4), causing cell death (KE5) and inflammation (KE6). As these cells detach from the basement membrane, they are deposited in the tubular lumen. Tubular obstruction and inflammatory responses to proximal tubule insult can cause secondary toxicity and tubular necrosis (KE7), further amplifying kidney injury and a progressive decline of function, finally resulting in kidney failure (AO). This information is primarily based on mechanisms of actions previously described in cited literature sources and intended as a resource template for AOP development and data organization. The primary species of this AOP is humans, most often observed following chemotherapeutic intervention when treating a range of tumours and multiple malignancies; supported by data with potential applicability for other species.

DNA adduct formation leading to kidney failure

This Adverse Outcome Pathway (AOP) details the sequence of key events (KE) that connect DNA adduct formation to kidney failure. Platinum (Pt)-based antineoplastics that are routinely utilized as anticancer agents for the effective treatment of multiple cancer types, including breast, cervical, oesophageal, bladder, small cell lung and testicular cancer (1), can function as chemical stressors for the described pathway. Currently, cisplatin, carboplatin and oxaliplatin are the three most utilised Pt-based drugs used globally for the treatment of cancer, all possessing a similar chemical structure consisting of platinum, carrier and leaving groups, undergoing similar mechanisms of cytotoxicity. Renowned for being one of the most potent and effective chemotherapeutics available, preventing cancerous cells from multiplying by binding DNA strands, clinical application of Pt anticancer agents remains limited due to the moderate to life-threating severity of their adverse side effects, such as neurotoxicity, hepatoxicity, ototoxicity, cardiotoxicity and myelosuppression and dose-limiting nephrotoxicity (1, 2).

Nephrotoxicity associated with platinum antineoplastic chemotherapeutic intervention

Within the kidney, the proximal tubular cells are most sensitive to the toxic effects of Pt-based drugs. They are responsible for the reabsorption of most nutrients and low molecular weight proteins, and approximately 70% of filtered solutes and water. Furthermore, they use active transport to clear the blood of toxic side products and drug compounds. Uptake of these compounds takes place at the basolateral side of the cells which is exposed to the blood. After entering the cells compounds are again secreted at the apical side of the cells into the kidney filtrate and leave the body with the urine. The toxicity of xenobiotics is directly related to their exposure to proximal tubule cells, their uptake through transport proteins like the SLC22A2/organic cation transporter 2 (OCT2), there reactivity in the cytoplasm and secretion through efflux transporters like multi-antimicrobial extrusion protein transporter-1 (MATE1) and multidrug resistance-associated protein 2 (MRP2) (3). This also implies that (genetic) variations in drug transporter activity can have implications in an individual’s susceptibility to (Pt)-based antineoplastics (4). Over time, several generations of Pt-based antineoplastics have been developed to make this group of compounds less nephrotoxic. Cisplatin was the first generation of Pt-based antineoplastics approved by the U.S. Food and Drug administration (FDA) in 1978. Classified as an alkylating agent, cisplatin was shown to inhibit cell division of bacteria and was developed as a treatment for mesotheliomas (5). However, issues arose following reports of cisplatin-induced nephrotoxicity, most notably the prevalence of acute kidney injury (AKI) and acute tubular necrosis (ATN) following treatment (6, 7). This increased susceptibility to severe nephrotoxicity during these early clinical trials led to the revision of cisplatin as an applicable chemotherapeutic agent. Therefore, much of the initial attempts of Pt drug development was to determine less-toxic analogues of cisplatin that retained similar levels of anticancer activity. The second-generation Pt drug, carboplatin, was developed to decrease the dose-limiting toxicity of cisplatin. Carboplatin’s mechanism of action is similar to that of cisplatin, however it was shown to have fewer and less severe side effects (8), benefiting from a reduced aquation rate due to its bidentate cyclobutane dicarboxylate ligand (9), and later approved by the FDA for use in 1989. Due to its decreased nephrotoxicity, carboplatin was identified as a more suitable option for aggressive, high-dose chemotherapy. However, carboplatin was also dose-limited following reports of cumulative anaemia and myelosuppression (10). Carboplatin is also a nephrotoxic drug, albeit much lower compared to cisplatin due to its increased stability (11). Nephrotoxicity had been reported in patients treated with intraperitoneal carboplatin, high-dose carboplatin, or in combination with other drugs (12), with AKI also shown to occur within days and often only partially reversible (13). The third-generation Pt drug oxaliplatin was developed to overcome both cisplatin and carboplatin toxicity. Oxaliplatin possesses a Pt complex with (1R,2R)-1,2-diaminocyclohexane (DACH) ligand and oxalate functioning as a leaving group. The bidentate oxalate considerably decreases the reactivity of oxaliplatin, limiting toxicity (14). However, although less nephrotoxic than both cisplatin and carboplatin, oxaliplatin can still induce proximal tubular cell damage (15). Furthermore, oxaliplatin has also been reported to cause several forms of nephrotoxicity, such as kidney tubular vacuolization (16) and acidosis (17-19). Despite the reduced incidence of nephrotoxicity, carboplatin and oxaliplatin, along with cisplatin, have all been reported to induce AKI and thrombotic microangiopathy (20).

References

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16.  Joybari AY, Sarbaz S, Azadeh P, Mirafsharieh SA, Rahbari A, Farasatinasab M, et al. Oxaliplatin-induced renal tubular vacuolization. Ann Pharmacother. 2014;48(6):796-800.

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20.  Gupta S, Portales-Castillo I, Daher A, Kitchlu A. Conventional Chemotherapy Nephrotoxicity. Adv Chronic Kidney Dis. 2021;28(5):402-14 e1.

This AOP is being developed as part of the ONTOX consortium. The aim of this consortium is to provide a generic strategy to create innovative new approach methodologies (NAMs) in order to predict systemic repeated dose toxicity effects that, upon combination with tailored exposure assessment, will enable human risk assessment. Part of this approach is the development of physiological maps, quantitative adverse outcome pathway networks and ontology frameworks. This project is funded by the European Union’s Horizon 2020 research and innovation programme under grant agreement No 963845. https://ontox-project.eu/project/

Within the ONTOX consortium in vitro test batteries are being developed to evaluate toxicity in the liver, (steatosis and cholestasis), kidneys (tubular necrosis and crystallopathy) and developing brain (neural tube closure and cognitive function defects). This AOP focusses on kidney tubular necrosis as a result of exposure to a DNA-adduct forming compound (Pt-based drugs).

The described AOP aims to deliver a mechanistic framework toward the advancement of in vitro assays that proficiently predict quantitative points of departure toward nephrotoxicity safety assessment. These assays could potentially form part of an integrated testing strategy to reduce the need for repeated dose toxicity studies. Producing quantitative data by measuring all KEs in a singular model following repeated, long-term exposure to a variety of concentrations of these chemical initiators could also expedite the advancement of computational predictive approaches. With prospective overlap between KEs, this AOP could provide further connections for expansive AOP networks toward modelling nephrotoxicity and kidney failure.