The accumulation of platinum compounds and their metabolites in the dorsal root ganglion (DRG) after their systemic administration and formation of platinum-DNA adducts are considered key steps in neurotoxicity development (Figure 1)[2,11]. The presence of an abundant fenestrated capillary network and the absence of blood-brain barrier in DRG allow platinum drugs to preferentially accumulate in DRG with easy access to sensory neurons[2,11,12].

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Figure 1 Proposed mechanisms of platinum-induced neurotoxicity. Dorsal root ganglion (DRG) is the main target of platinum drugs that preferentially accumulate in DRG neurons. Membrane transporters, copper transporter-1 (CTR1) and organic cation transporter-2 (OCT2), can facilitate the cellular uptake of platinum drugs. Platinum-DNA adducts inhibit replication and transcription, which results in caspase activation and subsequent cell death. Neuronal mitochondrial damage leads to cellular ATP depletion and increased reactive oxygen species (ROS) production. The voltage-gated sodium (Na⁺), potassium (K⁺) and calcium (Ca²⁺) channels dysfunction, and the enhanced expression and responsiveness of transient receptor potential channels (TRPA1, transient receptor potential ankyrin-1; TRPM8, transient receptor potential melastatin 8; TRPV1, transient receptor potential vanilloid 1) play an important role in the development of platinum-induced neurotoxicity.

Recently, it was demonstrated that the uptake of platinum drugs into DRG neurons may be facilitated by two different types of neuronal membrane transporters: Copper transporter-1 (CTR1) and organic cation transporter-2 (OCT2)[13-15]. The overexpression of these transporters in neurons, therefore, can contribute to the development or aggravation of neurotoxicity. For example, a 16- to 35-fold increase in the cellular oxaliplatin uptake was observed in neurons overexpressing mouse OCT2 or human OCT2, and this process resulted in significantly increased DNA platination and neurotoxicity[15].

Once the platinum drugs reach the neuronal cell nucleus, they attack the nuclear DNA to form adducts. They usually form same types of adducts on the same DNA sites, including 1,2-intrastrand d (GpG) (between adjacent guanine bases on the same DNA strand) and 1,2-intrastrand d (ApG) (between adenine and adjacent guanine bases on the same DNA strand) crosslinks. A correlation between adduct levels and the degree of neurotoxicity has been reported[16]. The platinum-DNA adduct levels produced by cisplatin were found to be approximately three times higher than those generated by equimolar oxaliplatin doses. Concordantly, in vitro cisplatin caused significantly more neuronal cell death than oxaliplatin[16]. DNA repair ability of DRG neurons for adducts (primarily performed by the nucleoid excision repair) is an important factor determining neurotoxicity severity[17]. Chronic cisplatin administration resulted in an accelerated accumulation of unrepaired platinum-DNA adducts in DRG neurons of DNA repair-deficient mice, which induced early neurophysiological alterations and led to an increase in neuronal cell death[17].

Inhibition of the global transcriptional activity of DRG neurons is one of the major consequences of DNA adduct formation[18]. DRG neurons need a high level of active transcription to sustain their large size, high metabolism, and long axons. Therefore, platinum-induced DNA damage leads to neuronal atrophy and disruption of their distant axonal connections[18].

Several preclinical studies have reported that platinum-induced DNA damage also induces apoptosis and neuron loss in DRG both in vivo and in vitro[19-22]. Cisplatin has been shown to initiate several apoptotic events in neuronal cells, including p53 activation, Bax translocation, mitochondrial cytochrome c release, and activation of caspase-3 and caspase-9. Gill and Windebank demonstrated that following exposure to cisplatin, DRG neurons attempt to re-enter the cell cycle from G0 phase, and this event can be a prelude to triggering neuronal cell death[22].

Mitochondrial dysfunction in DRG neurons was first described as a potential mechanism for platinum drugs neurotoxicity by Podratz et al[23]. They demonstrated that cisplatin also directly binds to mitochondrial DNA with similar binding affinity as nuclear DNA. Cisplatin-mitochondrial DNA adducts inhibit mitochondrial DNA transcription and replication, and cause morphological changes in the mitochondria. This can lead to disruption of the electron transport chain, loss of adenosine triphosphate (ATP) generation, energy failure, and overproduction of reactive oxygen species. All these events cause the opening of mitochondrial permeability transition pores, mitochondrial membrane depolarization, intracellular calcium accumulation, and expression of apoptotic proteins.

Cisplatin may also impair mitochondrial transport dynamics in neurons[24]. Proper mitochondrial transport in neurons is critical to cellular homeostasis. A new study in Drosophila has shown that cisplatin can significantly reduce mitochondrial movement frequency in axons[24]. This is probably caused by both ATP depletion and cellular calcium accumulation.

Some studies have demonstrated that cisplatin can alter the expression of mitochondrial fusion and fission proteins in peripheral nerves[25]. These proteins regulate mitochondrial shape, size, and number. Bobylev et al[25] detected a significant decrease in the mitochondrial fusion protein mitofusin 2 expression levels in DRG and tibial nerves of cisplatin-treated mice, resulting in mitochondrial swelling and vacuolization.

Oxaliplatin exhibits a tetrodotoxin-like inhibitory effect on the neuron voltage-gated sodium (Na⁺) channels[26-30]. It remarkably slows their inactivation and reduces the peak Na⁺ current, leading to an increase in the duration of the relative refractory period of sensory neurons. Oxaliplatin may also affect the Na⁺ channels indirectly via the chelation of extracellular calcium ions by its metabolite oxalate (diaminocyclohexane-platinum-C2O4)[26]. Because of Na⁺ channel dysfunction, sensory neurons become hyperexcitable and eventually generate spontaneous ectopic discharges.

Oxaliplatin can display isoform-specific effects on voltage-gated Na⁺ channels leading to the development of unique neuropathy symptoms such as cold-aggravated peripheral pain[31,32]. It has been suggested that oxaliplatin-induced Nav1.6 dysfunction may play a role in cold allodynia development[33,34]. Cooling in the presence of oxaliplatin increased Nav1.6-mediated persistent and resurgent Na⁺ currents in large-diameter DRG neurons and resulted in the generation of action potential burst firing[31].

Peripheral nerve axonal excitability studies performed before and immediately after oxaliplatin administration have confirmed the above mentioned in vitro findings and revealed acute abnormalities in sensory nerve function related to Na⁺ channel dysfunction, including decreased refractoriness and increased superexcitability[35]. Interestingly, it was shown that these excitability abnormalities can be detected in the initial oxaliplatin treatment cycles and may serve as a predictive tool to identify patients who are more likely to develop moderate or severe neurotoxicity.

Kagiava et al[33] suggested that altered voltage-gated potassium channel activity may be involved in oxaliplatin-induced neurotoxicity development. In their study, the effects of oxaliplatin on the compound action potential of rat sciatic nerve were observed to be similar to those with the potassium channel blockers 4-aminopyridine and tetraethylammonium. Oxaliplatin was found to cause broadening of action potentials and repetitive firing, suggesting its antagonistic effect on neuronal fast and slow potassium channels. This finding is indirectly supported by Sittl et al[34]. They showed that enhancement of axonal potassium conductance by flupirtine may reduce oxaliplatin-induced peripheral nerve hyperexcitability.

Conversely, voltage-gated potassium channels are unlikely to be the primary target for oxaliplatin because patch-clamp studies failed to show any effect of oxaliplatin on Shaker-type potassium channels[36]. Kagiava et al[37] found some evidence indicating that potassium channel dysfunction during oxaliplatin treatment can occur due to malfunction of the gap junction (GJ) channels and hemichannels in myelinated fibers. According to their findings, oxaliplatin causes prolonged opening of GJ channels and hemichannels, leading to excessive potassium accumulation in the periaxonal space and its osmotic swelling. This event is likely to have a disturbing effect on the voltage-gated potassium channel function.

Cisplatin does not appear to have a prominent effect on the neuronal sodium or potassium channel function. Initial studies using whole cell patch-clamp electrophysiological technique reported that cisplatin decreases the calcium channel currents, particularly in small-diameter neurons of rat DRG[38]. However, a new study revealed an increase in calcium influx through N-type calcium channels in rat DRG neurons after exposure to cisplatin[39]. This was mainly caused by the upregulation of the N-type calcium channels. Increased intracellular calcium levels led to caspase-3 activation and apoptosis induction.

Sensory neurons express various types of transient receptor potential (TRP) channels, including TRPA1, TRPM8, and TRVP1, which all play an important role in the generation and sensation of inflammatory and neuropathic pain[40-45].

Nassini et al[40] showed that oxaliplatin- and cisplatin-induced mechanical and cold hyperalgesia in rats are mediated by transient receptor potential ankyrin-1 (TRPA1), and TRPA1 activation is most likely caused by glutathione-sensitive molecules. Subsequently, Zhao et al[44] reported that oxaliplatin-induced cold hyperalgesia could be related to increased responsiveness of TRPA1. Pretreatment of the cultured DRG neurons with oxaliplatin resulted in an increase in the number of allyl-isothiocyanate (a TRPA1 agonist)-sensitive neurons.

The results of a recent study suggested that aluminum accumulation in DRG may augment oxaliplatin-induced neuropathic pain through activation of TRPA1 and stimulation of apoptotic cell death[46]. In this study, aluminum concentration of in DRG was greater in mice treated with aluminum chloride and oxaliplatin than in those treated with aluminum chloride alone.

Gauchan et al[43] revealed that oxaliplatin treatment increased the cold receptor transient receptor potential melastatin 8 (TRPM8) expression in rat DRG neurons, which resulted in enhanced sensitivity to cooling stimulation. Capsazepine, a blocker of both TRMP8 and TRPV1 channels, but not the selective TRV1 blocker 5’-Iodoresiniferatoxin, was able to inhibit oxaliplatin-induced cold allodynia. These findings suggested that TRPM8 plays a role in cold allodynia caused by oxaliplatin.

Ta et al[41] showed that mice DRG neurons treated with cisplatin or oxaliplatin displayed an increase in transient receptor potential vanilloid 1 (TRPV1), TRPA1, and TRMP8 mRNA expression. Trigeminal ganglion neurons from the cisplatin-treated animals showed increased TRPV1 and TRPA1 mRNA expression, and this was associated with enhanced heat and mechanical hypersensitivity. Conversely, oxaliplatin affected only TRPA1 expression, which induced cold and mechanic hypersensitivity.

Di Cesare Mannelli et al[47,48] first suggested a link between oxaliplatin-induced neuropathic pain and glial activation. In a rat model with oxaliplatin-induced peripheral neuropathy, they showed a transient activation of microglia and astrocytes in the spinal cord and supraspinal areas involved with pain modulation accompanied by a decrease in mechanical and thermal pain thresholds following intraperitoneal oxaliplatin administration[48]. Intrathecal co-administration of microglial inhibitor minocycline was able to prevent microglial activation, but had no effect on the response of astrocytes. The astrocytic activation could be inhibited by intrathecal injection of fluorocitrate, an astrocyte specific metabolic inhibitor. Fluorocitrate did not influence oxaliplatin-induced microglial activation. Both drugs increased pain tolerance, but fluorocitrate produced greater pain relief than minocycline. However, neither minocycline nor fluorocitrate prevented oxaliplatin-dependent morphological alterations in DRG neurons[48]. These findings provide some evidence for the participation of glial cells in oxaliplatin-induced neuropathy.

Oxaliplatin treatment was found to induce down regulation of alpha7 nicotinic acetylcholine receptor (nAChR) in the rat sciatic nerve, DRG, and spinal cord[49]. The administration of the selective alpha7 nAChR agonists (R)-ICH3 and PNU-282987 could prevent receptor down regulation and increase the pain threshold by oxaliplatin. These two agonists also could inhibit oxaliplatin-induced morphological changes in DRG and peripheral nerves, and upregulate glial cell density in the spinal cord, thalamus, and somatosensory area 1. CDP-choline, the other selective alpha7 nAChR agonist, was also found to be effective in reducing oxaliplatin-induced mechanical hyperalgesia when administered into the cerebral ventricles[50]. These findings suggested a neuroprotective role of alpha7 nAChR during oxaliplatin treatment.