Abstract
Oxaliplatin is an effective anti-cancer drug for the treatment of colorectal cancer and other solid tumours. However, the compound is associated with dose-limiting effects. Uniquely for oxaliplatin, patients report acute cold-evoked paraesthesias and cold allodynia during or immediately after infusion of the drug. In addition, everyday tasks such as buttoning shirts, fastening zips and touching cool surfaces is associated with discomfort and pain. A chronic neuropathy develops over repeated cycles of treatment with a stocking-and-glove distribution. The aim of my project was to identify novel neuronal and ionic mechanisms in acute oxaliplatin-induced hypersensitivities and paraesthesias. The six resulting chapters focus on in vivo, in vitro and ex vivo mechanistic studies of oxaliplatin-induced neuropathy.We examined the effects of oxaliplatin in vivo and established a translational mouse model. From this, we were able to isolate DRG neurons for mechanistic in vitro studies using [Ca2+]i imaging. We observed a modestly increased cold sensitivity in isolated neurons from treated mice, and in naïve DRG neurons treated with oxaliplatin. Oxaliplatin also directly activated a very small proportion of neurons. However, these effects appeared unlikely to accurately reflect the proportion of neurons responsible for the symptoms observed in mice in vivo or those reported by patients in the clinic. Therefore, we used the skin-saphenous nerve preparation to record the activity of single afferent nerve fibres stimulated at the peripheral nerve terminal.
Acute and direct oxaliplatin treatment sensitised all A fibre classes to cold ramps and notably, resulted in cold-evoked action potential bursts. In addition, novel cold-evoked responses were seen in normally cold-insensitive C fibres. Oxaliplatin also changed the adaptation properties of all A fibres, such that impulses and action potential bursts were displayed outside mechanical application periods. Interestingly, the acute effects of oxaliplatin at the nerve terminal level was different to recordings made in fibres from oxaliplatin-treated mice. Here, we showed increased spontaneous activity and C fibres being most affected by cold ramps, rather than A fibres. It is likely that differences in the duration and effective concentration of oxaliplatin exposure may explain at least part of these differences.
In order to identify novel ion channel targets involved in oxaliplatin evoked hyperexcitability, we used a multielectrode array in a 48-well assay format to study cultured cortical neurons. Oxaliplatin produced a concentration-dependent increase in network bursts in these cultures. In addition, we identified a novel voltage-gated potassium channel (Kv2.1) that is required for oxaliplatin-induced excitability. We demonstrate that inhibition of Kv2.1 fully reversed oxaliplatin-induced cold hypersensitivity in vivo and in single fibre recordings in naïve and treated preparations. Oxaliplatin also directly produced a hyperpolarising shift of the Kv2.1 channel activation kinetics by ~9 mV, in patch-clamp experiments of Kcnb1 transfected HEK293 cells. Finally, we report that oxaliplatin produced reversible and irreversible oxidation of cysteine residues in the model protein, human serum albumin. We hypothesise that such covalent modification are likely to be central for the side effects produced by oxaliplatin, and may also be responsible for the positive modulation of Kv2.1 described here. However, further experiments are required to confirm this. We have comprehensively assessed the effect of oxaliplatin acute and chronic treatment on peripheral sensory neurons and identified possible mechanisms that underlie acute neuropathy.
| Date of Award | 1 Nov 2021 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | David Andersson (Supervisor) & Stuart Bevan (Supervisor) |