Perez-Sanchez J, Middleton SJ, Pattison LA, Hilton H, Ali Awadelkareem M , et al.
Science translational medicine •
Hyperexcitability in sensory neurons is known to underlie many of the maladaptive changes associated with persistent pain. Chemogenetics has shown promise as a means to suppress such excitability, yet chemogenetic approaches suitable for human applications are needed. PSAM-GlyR is a modular system based on the human α7 nicotinic acetylcholine and glycine receptors, which responds to inert chemical ligands and the clinically approved drug varenicline. Here, we demonstrated the efficacy of this channel in silencing both mouse and human sensory neurons by the activation of large shunting conductances after agonist administration. Virally mediated expression of PSAM-GlyR in mouse sensory neurons produced behavioral hyposensitivity upon agonist administration, which was recovered upon agonist washout. Stable expression of the channel led to similar reversible suppression of pain-related behavior even after 10 months of viral delivery. Mechanical and spontaneous pain readouts were also ameliorated by PSAM-GlyR activation in acute and joint pain inflammation mouse models. Furthermore, suppression of mechanical hypersensitivity generated by a spared nerve injury model of neuropathic pain was also observed upon activation of the channel. Effective silencing of behavioral hypersensitivity was reproduced in a human model of hyperexcitability and clinical pain: PSAM-GlyR activation decreased the excitability of human-induced pluripotent stem cell-derived sensory neurons and spontaneous activity due to a gain-of-function Na1.7 mutation causing inherited erythromelalgia. Our results demonstrate the contribution of sensory neuron hyperexcitability to neuropathic pain and the translational potential of an effective, stable, and reversible humanized chemogenetic system for the treatment of pain.
Myeloproliferative neoplasms are a heterogeneous group of disorders resulting from the abnormal proliferation of one or more terminal myeloid cells-established complications include thrombosis and haemorrhagic events; however, there is limited evidence to suggest an association with arterial hypertension. Herein, we report two independent cases of severe hypertension in JAK2 mutation-positive myeloproliferative neoplasms. . Case 1: a 39-year-old male was referred to our specialist hypertension unit with high blood pressure (BP) (200/120 mmHg), erythromelalgia, and headaches. We recorded elevated serum creatinine levels (146 M) and panmyelosis. Bone marrow biopsy confirmed JAK2-mutation-positive polycythaemia vera. Renal imaging revealed renal artery stenosis. Aspirin, long-acting nifedipine, interferon-alpha 2A, and renal artery angioplasty were employed in management. BP reached below target levels to an average of 119/88 mmHg. Renal parameters normalised gradually alongside BP. Case 2: a 45-year-old male presented with high BP (208/131 mmHg), acrocyanosis, (vasculitic) skin rashes, and nonhealing ulcers. Fundoscopy showed optic disc blurring in the left eye and full blood count revealed thrombocytosis. Bone marrow biopsy confirmed JAK2-mutation-positive essential thrombocytosis. No renal artery stenosis was found. Cardiac output was measured at 5 L/min using an inert gas rebreathing method, providing an estimated peripheral vascular resistance of 1840 dynes/s/cm. BP was well-controlled (reaching 130/70 mmHg) with CCBs. These presentations highlight the utility of full blood count analysis in patients with severe hypertension. Hyperviscosity and constitutive JAK-STAT activation are amongst the proposed pathophysiology linking myeloproliferative neoplasms and hypertension. Further experimental and clinical research is necessary to identify and understand possible interactions between BP and myeloproliferative neoplasms.
Emery EC, Habib AM, Cox JJ, Nicholas AK, Gribble FM , et al.
The Journal of neuroscience : the official journal of the Society for Neuroscience •
The importance of NaV1.7 (encoded by SCN9A) in the regulation of pain sensing is exemplified by the heterogeneity of clinical phenotypes associated with its mutation. Gain-of-function mutations are typically pain-causing and have been associated with inherited erythromelalgia (IEM) and paroxysmal extreme pain disorder (PEPD). IEM is usually caused by enhanced NaV1.7 channel activation, whereas mutations that alter steady-state fast inactivation often lead to PEPD. In contrast, nonfunctional mutations in SCN9A are known to underlie congenital insensitivity to pain (CIP). Although well documented, the correlation between SCN9A genotypes and clinical phenotypes is still unclear. Here we report three families with novel SCN9A mutations. In a multiaffected dominant family with IEM, we found the heterozygous change L245 V. Electrophysiological characterization showed that this mutation did not affect channel activation but instead resulted in incomplete fast inactivation and a small hyperpolarizing shift in steady-state slow inactivation, characteristics more commonly associated with PEPD. In two compound heterozygous CIP patients, we found mutations that still retained functionality of the channels, with two C-terminal mutations (W1775R and L1831X) exhibiting a depolarizing shift in channel activation. Two mutations (A1236E and L1831X) resulted in a hyperpolarizing shift in steady-state fast inactivation. To our knowledge, these are the first descriptions of mutations with some retained channel function causing CIP. This study emphasizes the complex genotype-phenotype correlations that exist for SCN9A and highlights the C-terminal cytoplasmic region of NaV1.7 as a critical region for channel function, potentially facilitating analgesic drug development studies.
Koenig J, Werdehausen R, Linley JE, Habib AM, Vernon J , et al.
PloS one •
The Nav1.7 voltage-gated sodium channel, encoded by SCN9A, is critical for human pain perception yet the transcriptional and post-transcriptional mechanisms that regulate this gene are still incompletely understood. Here, we describe a novel natural antisense transcript (NAT) for SCN9A that is conserved in humans and mice. The NAT has a similar tissue expression pattern to the sense gene and is alternatively spliced within dorsal root ganglia. The human and mouse NATs exist in cis with the sense gene in a tail-to-tail orientation and both share sequences that are complementary to the terminal exon of SCN9A/Scn9a. Overexpression analyses of the human NAT in human embryonic kidney (HEK293A) and human neuroblastoma (SH-SY5Y) cell lines show that it can function to downregulate Nav1.7 mRNA, protein levels and currents. The NAT may play an important role in regulating human pain thresholds and is a potential candidate gene for individuals with chronic pain disorders that map to the SCN9A locus, such as Inherited Primary Erythromelalgia, Paroxysmal Extreme Pain Disorder and Painful Small Fibre Neuropathy, but who do not contain mutations in the sense gene. Our results strongly suggest the SCN9A NAT as a prime candidate for new therapies based upon augmentation of existing antisense RNAs in the treatment of chronic pain conditions in man.
Cox JJ, Sheynin J, Shorer Z, Reimann F, Nicholas AK , et al.
Human mutation •
SCN9Aencodes the voltage-gated sodium channel Na(v)1.7, a protein highly expressed in pain-sensing neurons. Mutations in SCN9A cause three human pain disorders: bi-allelic loss of function mutations result in Channelopathy-associated Insensitivity to Pain (CIP), whereas activating mutations cause severe episodic pain in Paroxysmal Extreme Pain Disorder (PEPD) and Primary Erythermalgia (PE). To date, all mutations in SCN9A that cause a complete inability to experience pain are protein truncating and presumably lead to no protein being produced. Here, we describe the identification and functional characterization of two novel non-truncating mutations in families with CIP: a homozygously-inherited missense mutation found in a consanguineous Israeli Bedouin family (Na(v)1.7-R896Q) and a five amino acid in-frame deletion found in a sporadic compound heterozygote (Na(v)1.7-DeltaR1370-L1374). Both of these mutations map to the pore region of the Na(v)1.7 sodium channel. Using transient transfection of PC12 cells we found a significant reduction in membrane localization of the mutant protein compared to the wild type. Furthermore, voltage clamp experiments of mutant-transfected HEK293 cells show a complete loss of function of the sodium channel, consistent with the absence of pain phenotype. In summary, this study has identified critical amino acids needed for the normal subcellular localization and function of Na(v)1.7.
