Role of HCN ion channels in neuropathic pain: a combined animal and human study

Pain is commonly classified into three distinct varieties, each with different underlying causes. Acute pain, such as caused by a sudden injury or burn, is due to direct excitation of nociceptive (pain-sensitive) endings by the painful stimulus. Inflammatory pain follows an injury, and is caused by an action on nociceptive nerve endings of inflammatory mediators such as prostaglandin E2 and bradykinin released from stressed or injured tissues. Neuropathic pain is caused by direct injury of the sensory nerves themselves. Common conditions which cause neuropathic pain include diabetic neuropathy, the after-effects of a Herpes Zoster outbreak, or shingles, and some forms of cancer chemotherapy. Neuropathic pain is also thought to be a contributor to the pain felt in other common conditions, such as lower back pain and end-stage cancer pain. Neuropathic pain is poorly treated by currently available pharmaceuticals, with even the first-line treatments giving relief in only one third of patients.

Recent experiments in the applicant lab have shown that neuropathic pain is initiated by an ion channel, known as HCN2, which is present in nociceptors. Activation of HCN2 causes an inward current to flow into nociceptors, producing a low-level barrage of nerve impulses in these pain-sensitive nerve fibres. The evidence that this activity is the causative agent in neuropathic pain comes from two types of experiment: when HCN2 is deleted genetically from nociceptors neuropathic pain is not initiated; and when it is blocked pharmacologically neuropathic pain is reversed. The aim of the present study is to take these observations further in both animal and human studies.

In animal studies we will examine the effect of genetic deletion of HCN2 after neuropathic pain has been established. This experiment is important because there is some evidence that neuropathic pain has an initial inflammatory phase followed by a long-term phase which many scientists attribute to events in the spinal cord. Does deletion of HCN2 at long times reverse neuropathic pain? If it does this will imply that the causative event at long as well as short times is activity initiated by HCN2 in peripheral nociceptors.

The effect of pharmacological block of HCN2 has been to date investigated only with one blocker, ZD7288. Two others are currently in clinical use or in late-stage trials and therefore could be used to alleviate human neuropathic pain. Are they effective in animals? These blockers are known to slow the heart, and in fact this is their clinical use - but is there a therapeutic window in which pain is relieved without cardiac effects? Finally, is pain in animal models of common human causes of neuropathic pain, such as diabetic neuropathy, alleviated by deletion or pharmacological block of HCN2?

In complementary human studies we will investigate the effect of a clinically approved HCN2 blocker, ivabradine, on neuropathic pain in both healthy volunteers and in patients. In initial studies we will induce a shortlived pain state, thought to share features with neuropathic pain, by application of capsaicin, which produces a localised burning sensation. Is mechanical hypersensitivity alleviated by ivabradine? If these studies are successful we will investigate the effect of ivabradine on patients in the Addenbrooke's Hospital Pain Clinic. Is some relief obtained with ivabradine? If only some patients see relief, can we explain the differences in terms of the characteristics of their pain?

Successful completion of both arms of this project will bring closer the possibility of alleviating neuropathic pain by pharmacological block of HCN2.

Recent experiments in the applicant lab have identified the ion channel HCN2 as an important initiator of neuropathic pain, which opens up the possibility of developing HCN2-selective blockers as novel analgesics with efficacy in neuropathic pain. Some aspects of this work need to be extended in animal models before this idea can be regarded as proven, as detailed below. We also propose to use a clinically approved non-selective HCN blocker, ivabradine, currently employed to treat stable angina, to investigate whether HCN block can in principle relieve neuropathic pain.

The animal studies will address the following:
1) Our previous studies have shown that nociceptor-specific deletion of HCN2 prevents the onset of neuropathic pain. We use an inducible Cre-lox system to test whether deletion reverses established neuropathic pain.
2) In complementary experiments we will investigate pharmacological block of neuropathic pain using established non-selective HCN blockers. Are they able to reverse even long-established neuropathic pain?
3) Is there a therapeutic window which will allow suppression of neuropathic pain without interference with the cardiac rhythm through block of the cardiac isoform, HCN4?
4) Are genetic deletion or pharmacological block of HCN2 effective in animal models of common human neuropathic pain conditions e.g. diabetic neuropathy?

Human studies will address the following:
5) Does ivabradine reverse mechanical allodynia in a model of neuropathic pain implemented in healthy human volunteers by application of a capsaicin patch?
6) Does ivabradine reverse neuropathic pain in patients? We do not expect a full effect because the dose will be limited by cardiac side effects caused by HCN4 block, but any effect will provide valuable proof of principle for the development of future HCN2-selective blockers.

Who will benefit from this research?

The research will investigate the potential utility of HCN blockers, and in particular blockers of the HCN2 ion channel, in the treatment of neuropathic pain. The outcome is likely to have considerable practical importance in the study of pain and in the development of novel analgesics. Work leading up to this application has identified HCN2 as a novel target for the control of inflammatory and neuropathic pain. The present proposal will further amplify and reinforce this work. In separate applications we are applying for funding to take this idea forward in collaboration with a company with facilities for high-throughput ion channel screening, with the objective of developing novel analgesics. If this work is successful the beneficiaries will be the many people who suffer from chronic inflammatory and neuropathic pain and whose pain is inadequately treated by currently available analgesics.

The general public has tremendous curiosity about pain, as is shown by the number of invitations I receive to give talks about my work and the general area of pain on radio and TV programmes, and in person to popular audiences. We are able to satisfy the natural curiosity of the general public, and they are therefore also beneficiaries of this work.

How will they benefit from this research?

The interest of the pharmaceutical industry in this area is shown by a current grant awarded to my lab from Organon, Inc (now Merck Sharp & Dohme) and the applicant has in the past had support from Merck Sharp & Dohme and from Glaxo SmithKline. An understanding of the involvement of ion channels such as HCN2 in pain is of particular interest to pharmaceutical companies.

Timescales for developing innovations arising from this research may not be long. An analogy can be drawn with the heat-sensitive ion channel TRPV1, which was published in 1997. Its importance in inflammatory pain was recognised as soon as genetically deleted mice were shown (in 2000) to exhibit an absence of inflammatory hyperalgesia. Pharmaceutical companies immediately began the development of antagonists and 10 years later a large number are in clinical trials, with several candidates well advanced into phase 2 trials. A similar rapid timescale of development has been seen with TRPA1, which was cloned in 2004 and for which several antagonists have already entered clinical trials with a view to treating bronchospasm in asthmatics. The analgesic effects of deletion of HCN2 are much more striking than either of these two examples, as in addition to a reduction of inflammatory pain, we have shown that deletion of this ion channel abolishes the much more intractable neuropathic pain. We can envisage a similarly rapid timescale for the development of analgesics based on antagonists of HCN2.