**3. Channel inhibitory molecules**

Findings about Nav1.7 gain-of-function mutations causing intense pain and lossof-function mutations of the same channel causing painlessness have put Nav1.7 in the therapeutic spotlight as an important new target for development of analgesics, for which a channel blocking antagonist is required.

Despite the peculiar and promising nature of the Nav1.7 channel, the use of ion channel blockers for the treatment of pain is not new. For years, blockers of sodium (Na+ ), calcium (Ca++) and potassium (K+ ) channels have been used, such as local anesthetics, antiarrhythmics or antiepileptics, as adjuvants in the treatment of neuropathic pain. The problem lies in the poor selectivity of these drugs, in the adverse cardiac, neurological, hematological, and digestive effects that limit their use, and in the inter-individual variability that, together with the particular susceptibility to develop chronic pain, limits their efficacy; therefore, the challenge facing Nav1.7

*Nav1.7 Sodium Channel: A Potential Analgesic Target DOI: http://dx.doi.org/10.5772/intechopen.105162*

**Figure 2.** *Nav1.7 inhibitory arylsulfonamide.*

inhibitory molecules is that they must be extremely selective, for which the three binding sites described above have been proposed. Some of the compounds studied so far and proposed as selective inhibitors of Nav1.7 are described below.

## **3.1 Small molecules as sodium channel blockers**

In recent years, small molecules have been developed that seek to selectively inhibit domain IV of the voltage sensor, arylsulfonamides are some of them. In 2012, it was shown that the arylsulfonamide named PF-04856264 and shown in **Figure 2** interacts with amino acid residues on an extracellular facing region of the homologous Domain 4 voltage sensor of Nav1.7, selectively inhibiting it (IC50, 28 nM) over Nav1.5; this is important due block of the Nav1.5 channel may lead to arrhythmia and thus limit the therapeutic potential of nonselective Nav1.7 inhibitors [11].

In 2016, Focken et al. reported a series of aryl sulfonamides that act as nanomolar potent, isoform-selective inhibitors of the human sodium channel hNav1.7. In this study, the optimization of these inhibitors is described. The main goal of this research was to improve potency against hNav1.7 while minimizing off-target safety concerns, so, they generated the compound of the **Figure 3**. This agent displayed significant effects in rodent models of acute and inflammatory pain and demonstrated that binding to the voltage sensor domain 4 site of Nav1.7 leads to an analgesic effect *in vivo* and corroborate the importance of hNav1.7 as a drug target for the treatment of pain [4].

Vixotrigine (**Figure 4**) is another Nav1.7 channel blocker under clinical investigation to treat peripheral neuropathic pain conditions, including trigeminal neuralgia, which is characterized by brief episodes of severe pain from one or more branches of the trigeminal nerve. Clinical trial results suggest that vixotrigine 150 mg three

**Figure 3.** *Sulfonamide that acts as an inhibitor of the human sodium channel hNaV1.7.*

**Figure 4.** *Vixotrigine is a voltage-dependent Nav blocker.*

times a day may be an effective and safe treatment for trigeminal neuralgia pain with only dizziness and headache as side effects, and even a phase III study suggests that a higher dose (250 mg three times a day) may provide additional benefit in those who do not respond adequately to the first dose [12, 13].

In 2017 Swain et al. described a series of acidic diaryl ether heterocyclic sulfonamides that are potent and sub-type selective Nav1.7 inhibitors. Theirs design strategies and results from pre-clinical PK and Clinical human microdose PK data are described leading to the discovery of the first sub-type selective Nav1.7 inhibitor clinical clinical candidate: 4-[2-(5-Amino-1H-pyrazol-4-yl)-4-chlorophenoxy]-5 chloro-2-fluoro-N-1,3-thiazol-4-ylbenzenesulfonamide (PF-05089771) which binds to a site in the voltage sensing domain. (**Figure 5**). This was the first potent and selective molecule which binds to the domain IV voltage sensor region of the sodium channel to progress into the clinic. The authors demonstrated that the development of selective agents such as PF-05089771 offers new potential therapeutic modelities for acute and chronic pain [14].

Recently Wang et al. designed and synthesized a series of compounds with ethanoanthracene and aryl sulfonamide moieties. They detected the inhibitory activity of this compounds on sodium channels with electrophysiological techniques. They found that a compound of theirs potently inhibited Nav1.7 channels stably expressed in HEK293 cells (IC50 = 0.64 ± 0.30 nmol/L) and displayed a high Nav1.7/Nav1.5 selectivity [15].

## **3.2 Toxins as sodium channel blockers**

On the other hand, studies of different toxins have been carried out for therapeutic benefit, finding that they bind to DSV2, a different domain than small molecules.

**Figure 5.** *PF-05089771.*

## *Nav1.7 Sodium Channel: A Potential Analgesic Target DOI: http://dx.doi.org/10.5772/intechopen.105162*

For example, ProTx-II, a peptide from spider venom was the first inhibitor of Nav1.7 reported to have greater than 50-fold selectivity over the other Nav isoforms.

In 2010, Xiao et al. discovered that ProTx-II and huwentoxin-IV (HWTX-IV), cystine knot peptides from tarantula venoms, preferentially block hNav1.7. This group described that the mutation E818C increases ProTx-II's and HWTX-IV's IC50 for block of hNav1.7 currents by 4- and 400-fold, respectively. Also, they reported that ProTx-II, but not HWTX-IV, preferentially interacts with hNav 1.7 to impede fast inactivation by trapping the domain IV voltage-sensor in the resting configuration [16].

Knowing that Protoxin-II (ProTx2) from the Peruvian green velvet tarantula (**Figure 6**) is an inhibitor cystine-knot peptide and selective antagonist of the human Nav1.7 channel, in 2019, Xu et al. visualize ProTx2 in complex with voltage-sensor domain II (VSD2) from Nav1.7 using X-ray crystallography and cryoelectron microscopy. They described that membrane partitioning orients ProTx2 for unfettered access to VSD2, where ProTx2 interrogates distinct features of the Nav1.7 receptor site. ProTx2 positions two basic residues into the extracellular vestibule to antagonize S4 gating-charge movement through an electrostatic mechanism. ProTx2 has trapped activated and deactivated states of VSD2, revealing a remarkable ~10 A° translation of the S4 helix, providing a structural framework for activation gating in voltagegated ion channels. This work delivered key templates to design selective Nav channel antagonists. Here, the researchers verified that ProTx2 shows 30–100 times more selectivity on the Nav1.7 channel than on the other Nav isoforms. The authors also demonstrated that this toxin is the most potent inhibitor available, testing its structural bases in the quest to accelerate the design of new modulators [17].

Finally, in 2020, Pajouhesh et al. described the discovery and characterization of ST-2262, a Nav1.7 inhibitor that blocks the extracellular vestibule of the channel with an IC50 of 72 nM and greater than 200-fold selectivity over off-target sodium channel isoforms, Nav1.1–1.6 and Nav1.8. ST-2262 (**Figure 7**) was discovered through a rational design strategy aimed at identifying derivatives of natural bis-guanidinium toxins that preferentially inhibit hNav1.7 over other off-target hNav isoforms. The potency of ST-2262 against hNav1.7 stably expressed in HEK293 cells was assessed by manual patch clamp electrophysiology with a voltage-protocol that favors the resting state of the channel. Using a stimulation protocol with a holding potential of – 110 mV and a stimulus frequency of 0.33 Hz, the IC50 of ST-2262 against hNav1.7 was measured

**Figure 6.** *Peruvian Green Velvet Tarantula.*

**Figure 7.** *ST-2262 is a potent and selective inhibitor of hNav1.7 and it is a derivative of natural bis-guanidinium toxins.*

at 0.072 μM (95% confidence interval (CI) 0.064–0.082). The authors find that, in contrast to other Nav1.7 inhibitors that preferentially inhibit the inactivated state of the channel, ST-2262 is equipotent in a protocol that favors the resting state of the channel, a protocol that favors the inactivated state, and a high frequency protocol. In a nonhuman primate study, animals treated with ST-2262 exhibited reduced sensitivity to noxious heat. These findings establish the extracellular vestibule of the sodium channel as a viable receptor site for the design of selective ligands targeting Nav1.7 [18].
