Cisplatin Mouse Models: Treatment, Toxicity and Translatability

Abstract

Cisplatin is one of the most widely used chemotherapeutic drugs in the treatment of a wide range of pediatric and adult malignances. However, it has various side effects which limit its use. Cisplatin mouse models are widely used in studies investigating cisplatin therapeutic and toxic effects. However, despite numerous promising results, no significant improvement in treatment outcome has been achieved in humans. There are many drawbacks in the currently used cisplatin protocols in mice. In the paper, the most characterized cisplatin protocols are summarized together with weaknesses that need to be improved in future studies, including hydration and supportive care. As demonstrated, mice respond to cisplatin treatment in similar ways to humans. The paper thus aims to illustrate the complexity of cisplatin side effects (nephrotoxicity, gastrointestinal toxicity, neurotoxicity, ototoxicity and myelotoxicity) and the interconnectedness and interdependence of pathomechanisms among tissues and organs in a dose- and time-dependent manner. The paper offers knowledge that can help design future studies more efficiently and interpret study outcomes more critically. If we want to understand molecular mechanisms and find therapeutic agents that would have a potential benefit in clinics, we need to change our approach and start to treat animals as patients and not as tools.

Keywords: cisplatin; gut; kidney; mouse; mouse model; nerve system; toxicity; treatment; tumors.

Figure 1

Schematic presentation of clinical signs and kidney function in mice after a nephrotoxic dose of cisplatin. First, two days after a single high dose of cisplatin (10–13 mg/kg; ip), minimal structural changes in the proximal tubules (P3) can be detected (i.e., mitochondria alterations, focal loss of the microvillus brush border, pycnotic nuclei, increased cytoplasmic vesicles) [63,64]. More obvious changes such as loss of the brush border or necrotic cells sloughing into the tubular lumen are usually seen 3–4 days after injection and changes are located in all parts of the proximal tubules (P1–3)[63,64,65]. Depending on the dose, increased BUN/Cr are usually observed 3–7 days after cisplatin injection [66,67,68,69], and if nephrotoxicity is reversible, BUN/Cr return to the baseline levels within 14 days [70]. In such cases, the first signs of structural regeneration can be observed 7 days after cisplatin injection [64,71]. A single high dose of cisplatin (B6D2F1: 8 mg/kg, 10 mg/kg, 12 mg/kg, 14 mg/kg; ip) induces dose-dependent weight loss (11–26%), reticulocytopenia with the lowest levels of body weight and reticulocytes observed 6 days after cisplatin injection. Necrosis in kidney tubular cells can be seen up to 10–22 days post-treatment [72]. When a lethal dose is used, death may occur within 10 days [73] and the time course of AKI development or mortality can occur slightly faster, but still 1–2 days after cisplatin injection. Cisplatin (F1 CBAxC57BL, 12 mg/kg, ip) induces lymphocytopenia, thrombocytopenia and anemia. Cisplatin exhibits cytotoxicity to spleen (CFU-S), granulocyte–macrophage (CFU-C) colony-forming units and mononuclear cells (MNC) in bone marrow and white blood cells (WBC) (adapted and modified from Nowrousian et al. [74]) Legend: P1–3 denotes kidney proximal tubules parts 1–3, DT—distal tubules; BB—brush border; BM—basal membrane; BUN- blood urea nitrogen; Cr—serum creatinine; AST—aspartate aminotransferase; WBC—white blood cells; Hb—hemoglobin; GFR—glomerular filtration rate; CFU—colony-forming unit; MNC—mononuclear cells; ER—endoplasmic reticulum.

Figure 2

Cisplatin causes acute and chronic effects in the gastrointestinal tract. A single injection of cisplatin causes pica, a rodent-specific behavior of nausea, which reflects as a progressive reduction in food intake, increase in non-nutritive material intake (for instance bedding) and decreased gastric motility [107]. As a result the stomach is full of bedding and markedly enlarged/distended (white arrow) [114]. Reduction in food (68%) and water intake (45%) and an increase in stomach content (threefold) is evident from day 2 on (C57BL/6; 6 mg/kg ip) [114]. First morphological changes in the small intestinal mucosa (i.e., apoptosis, necrosis, decreased number of goblet cells, shortened villi and inflammatory cell infiltration) can be seen 1 day after a single cisplatin injection (B6D2F1: 8 mg/kg, 10 mg/kg, 12 mg/kg, 14 mg/kg; ip; d1,3,6,10,14) followed by reduced mucosal digestive function (depletion in maltase, sucrose, disaccharidase activity and reduced absorption) [108,115]. Depression in crypt cell production is already evident 2h after cisplatin and is maximal between 12 and 24 h post-treatment (CBA: 10 mg/kg, ip). Cisplatin causes lesions also in the colon mucosa, however, they appear later and are less severe [72]. The severity of gastrointestinal damage and mucosal dysfunction is dose-dependent and can persist up to 10 days after a single sub-lethal dose of cisplatin (B6D2F1: 8 mg/kg, 10 mg/kg, 12 mg/kg, 14 mg/kg; ip; d1,3,6,10,14) [72]. Mucosal recovery is slow, first signs of recovery can be observed 7 days post-treatment [72]. Repeated cisplatin administration (C57BL/6; 4 mg/kg/week for 4 weeks, ip; ↓20% BW) besides gut lesions (↑IL-1β and IL-10) also causes delayed pica, [55] and alterations in the ENS seen as loss of neurons in the myenteric ganglia of mouse gastric fundus (total and nNOS⁺) [56] and colon (neurons (total, ChAT⁺, nNOS⁺) and gial cells (SOX-10⁺, GFAP⁺, S100β⁺) [55]. Circulation and the nervous system are the main pathways for communication between the gut, the kidney and the brain in health or disease (the brain–gut–kidney axis). Legend: BW—body weight; ChAT—choline acetyltransferase; ENS—enteric nerve system; GFAP—glial fibrillary acidic protein; NET—neutrophil extracellular traps; nNOS—neuronal nitric oxide synthase; ip—intraperitonealy.

Figure 3

Cisplatin can have long-term effects in the gastrointestinal tract (A). A case of penetrating ulcer (B, arrow and C) in a mouse that survived a single lethal dose of cisplatin (17 mg/kg). Three months after cisplatin recovery, body weight started to decrease, and the mouse was killed and autopsy performed. Inflammatory cells found in the kidney (D).

Figure 4

Cisplatin neurotoxicity. In mice, two cycles of cisplatin (2.3 mg/kg/daily for 5 days followed by 5 days recovery; 5d+5r/5d+5r; cumulative dose 23 mg/kg) resulted in reduced density of intraepidermal nerve fibers (IENF) (wk3, and wk5) [152,164] and epidermal Merkel cells [152] in the mouse plantar footpad. Merkel cells, mechanosensory cells actively involved in touch reception (tactile sensation), [165,166,167] are proposed to underlie sensory dysfunction in diabetic patients and animals [168]. In sensory nerves (sciatic, caudal, tibial) mild hypomyelination with few degenerating axons (reduced density of myelinated fibers without alterations in axon diameter) can be observed together with a slight decrease in the sensory nerve conduction velocity (SNCV; indication of demyelination) and the sensory nerve action potential (SNAP) [153]. In sensory neurons (trigeminal ganglia) cisplatin activated the transient receptor potential (TRP) channels (TRPA1, TRPV1) [151], a non-selective cation channels involved in chemical and thermal evoked pain sensation [169]. In the spinal cord (L4-L6) cisplatin activated microglia (Iba1), induced pro-inflammatory cytokines (IL-1β, IL-6, TNFα, iNOS, CD16, a marker of pro-inflammatory microglia (wk3) and increased protein levels of triggering receptor expressed on myeloid cells 2 (TREM2) and DNAX activating protein of 12 kDa (DAP12) (wk3) [152]. TREM2/DNAX is a receptor complex predominantly expressed on microglia in the central nervous system associated with neurodegenerative diseases and inflammatory response of microglia [152]. Cisplatin induced structural abnormalities in cerebral white matter (loss of neuronal dendritic spines and arborizations) [145,146] and reduced myelin density in the cingulated cortex [147]. It also [145] decreased cerebral neurogenesis (DCX⁺ cells) [146] but did not cause inflammation (IL1β, IL6, TNFα, GFAP, CD11b) [146] or microglia (Iba1⁻, GFAP⁻) activation [145]. However, decreased synaptic integrity (synaptophysin, vGlut2, vGAT) in the prefrontal cortex [148] and global functional neuronal connectivity in the mouse brain was found (fMRI) [147]. Cisplatin induced mitochondrial dysfunction and structural abnormalities in brain synaptosomes [147]. Mice treated with three cycles of cisplatin (protocol 2.3 mg/kg 5d + 5r/5d + 5r/5d + 5r; cumulative dose 34.5mg/kg) developed more severe impairment of mitochondrial transport and mitochondrial dysfunction in the hippocampus [149] (43% decrease in cytochrome C activity, ATP production, 96% increase in ROS, 29% decrease in mitochondrial membrane potential, impaired mitochondrial transport, reduced α-tubulin acetylation in the hippocampus, decrease in dendritic spine and synaptic density (vGlut1 and PSD95) [149]. Legend: DAP12—DNAX activating protein of 12 kDa; DRG—dorsal root ganglia; GFAP—glial fibrillary acidic protein; IENF—intraepidermal nerve fibers; IL—interleukine; Iba1—ionized calcium-binding adaptor molecule 1; iNOS—inducible nitric oxide synthase; L4-L6—lumbal vertebra; mtDNA—mitochondrial DNA; NER - nucleotide excision repair; Olig-2—oligodendrocyte lineage gene 2; ROS—reactive oxidative species; SNAP—sensory nerve action potential; SNCV—sensory nerve conduction velocity; TNFα—tumor necrosis factor alpha; TRP—transient receptor potential channels (TRPA1, TRPV1); TREM2—triggering receptor expressed on myeloid cells 2; vGlut2—vesicular glutamate transporter 2; vGAT—vesicular GABA transporter.

Figure 5

Schematic presentation of cisplatin toxicity in non-tumor cells in the body. Extent and intensity of oxidative stress, changes in signaling, metabolism, function, intensity of inflammation, activation of certain immune cell types, inflammatory and molecular crosstalk and response, type of cell death, etc., depend on cisplatin dose (single or cumulative) and severity of toxicity. ER—endoplasmic reticulum; mtDNA—mitochondrial DNA; ROS—reactive oxygen species.

Figure 6

Dose-dependent toxicity of cisplatin and factors affecting maximum tolerated dose (MTD), nephrotoxic and lethal dose.