NCS-1 is a Regulator of Calcium Signaling in Health and Disease

Abstract

Neuronal Calcium Sensor-1 (NCS-1) is a highly conserved calcium binding protein which contributes to the maintenance of intracellular calcium homeostasis and regulation of calcium-dependent signaling pathways. It is involved in a variety of physiological cell functions, including exocytosis, regulation of calcium permeable channels, neuroplasticity and response to neuronal damage. Over the past 30 years, continuing investigation of cellular functions of NCS-1 and associated disease states have highlighted its function in the pathophysiology of several disorders and as a therapeutic target. Among the diseases that were found to be associated with NCS-1 are neurological disorders such as bipolar disease and non-neurological conditions such as breast cancer. Furthermore, alteration of NCS-1 expression is associated with substance abuse disorders and severe side effects of chemotherapeutic agents. The objective of this article is to summarize the current body of evidence describing NCS-1 and its interactions on a molecular and cellular scale, as well as describing macroscopic implications in physiology and medicine. Particular attention is paid to the role of NCS-1 in development and prevention of chemotherapy induced peripheral neuropathy (CIPN).

1. Introduction

Calcium (Ca²⁺) is a ubiquitous second messenger molecule that regulates multiple mammalian cell functions, ranging from Ca²⁺ waves during fertilization at the beginning of the life cycle, modulation of transients and oscillations during cell cycle progression and cell division, and apoptotic cell death vis-à-vis high intracellular Ca²⁺ [1, 2]. In order to achieve this versatile role as a signaling molecule, a cell specific expression of Ca²⁺ binding proteins, channels and transporters as well as the involvement of multiple storage compartments within the cell must collaborate and coordinate [3–5]. The intracellular resting calcium concentration is maintained through active transport into storage compartments including the endoplasmic reticulum (ER) and the mitochondria as well as transport out of the cell across the plasma membrane. Furthermore, Ca²⁺ binding proteins enable distinct temporal and spatial signaling patterns through binding of free cytosolic Ca²⁺. This allows interaction with numerous target proteins [6, 7].

This review will give a brief outline of the functional role of Neuronal Calcium Sensor-1 (NCS-1, frequenin in non-mammals) in neuronal and extra-neuronal tissues. Primarily, we will discuss the role of NCS-1 in the development of chemotherapy-induced peripheral neuropathy (CIPN) and implications of the existing body of evidence with regard to possible treatments.

2. NCS-1 Properties and Molecular Functions

2.1 General Properties

NCS-1 is a member of the Neuronal Calcium Sensor (NCS) family of Ca²⁺ binding proteins [ 8]. In the human genome NCS proteins are encoded by 14 genes, however, amino acid sequences are highly conserved and can also be found in single-celled microorganisms [9, 10]. Essentially all members of the NCS family contain 2-3 functional Ca²⁺ binding domains, as well as a N-terminal myristoylation site [11].

NCS-1 was first discovered in the Drosophila nervous system [12] and has since been shown to be expressed in organisms from yeast to humans and in most neuronal cell types [13–15], as well as endocrine, heart, smooth muscle, and gastrointestinal tract tissue [16]. In humans the highest expression levels of NCS-1 were found in the cerebral cortex [17], however, expression in other regions, the hippocampus [18–20] and dorsal root ganglion cells [21], influence important functions. NCS-1 contains four helix-loop-helix structural domains called EF hand motifs [22]. This motif represents a Ca²⁺-binding site and is one of the most common structural motifs in animal genomes [23]. In human NCS-1 only three of the four EF hands bind Ca²⁺ because the first EF hand lacks Ca²⁺-coordinating amino acids (Asp or Glu) [22]. This pseudo-EF hand stabilizes the three functional EF hands and cleavage within this domain by the Ca²⁺ dependent protease calpain destabilizes the protein [24]. The 2x2 structural arrangement of the four EF-hands divides NCS-1 into a myristoylated N-terminal pair (EF1, EF2) and a C-terminal pair (EF3, EF4) which are connected by a hinge loop. Ca²⁺ binding enables a large positional shift in the C-terminal pair, exposing a wide hydrophobic crevice that can interact with target proteins [22, 25]. The pairing of EF hand motifs is ideally suited for high-affinity Ca²⁺ binding so that subtle Ca²⁺ signals can be detected [23, 26]. This makes NCS-1 a Ca²⁺ sensor protein rather than merely a Ca²⁺ binding protein [27]. Ca²⁺ binding as well as N-terminal myristoylation was originally described as a mechanism to enable on-off switch responses [23], however, this function has lately been challenged [28–30]. A better way to describe NCS-1 is as a facilitator that allows rapid transduction of Ca²⁺ signals. A second Frq-encoding gene (frq2) [31, 32] with distinct functions was found in Drosophila [31, 33]. Similarly, a second variant of NCS-1 was identified in humans from RNA screens where the N-terminal 22 amino acids are lacking which results in altered Ca²⁺ binding. This variant does not appear to be expressed and no functional role was found [34].

2.2 Molecular Functions

NCS-1 regulates many different cellular functions including exocytosis [11], neurite outgrowth [35], neuroprotection [36], axonal regeneration [37], nuclear Ca²⁺ regulation [38], and is implicated in development and disease. NCS-1 binds to a number of proteins (Table 1, adapted from [7]) but the functional implications of many of the interactions are still to be determined. Depending on whether NCS-1 is in a Ca²⁺ bound or unbound state, it can physically interact with a distinct set of proteins [6]. Once bound, NCS-1 activates downstream signaling pathways [7].

Table 1.

List of Proteins that bind NCS-1 (Table adapted from [7])

NCS-1 Interacting Protein	Abbreviation	Effect on target	Functional consequence	References
Ca2+-bound state
ADP-ribosylation factor 1	ARF1	Competes for PI4KIIIβ activation	Regulation of TGN to plasma membrane traffic	[48, 49]
α1 subunit of voltage-gated Ca2+ channel 2.1	CaV2.1	Activates Ca2+-dependent facilitation of channel	Increases facilitation of neurotransmitter release	[50, 51]
Dopamine D2 Receptor	D2R	Inhibits internalisation of receptor	Promotes spatial memory formation	[18, 52, 53]
Interleukin-1 receptor accessory protein-like 1	IL1RAPL	?	Regulates N-type channels, secretion and neurite elongation	[54, 55]
Phosphatidylinositol 4-kinase IIIß	PI4KIIIß	Activates the enzyme	Regulation of TGN to plasma membrane traffic	[48, 56]
apo/unbound state
Adenosine A2 receptor	ADORA2A	?	Increases receptor signaling	[57]
G-Protein coupled receptor Kinase 2	GRK2	Inhibits kinase activity	Inhibits receptor internalisation	[52]
Inositol 1,4,5-trisphosphate receptor	InsP3R	Enhances receptor activity	Increases calcium signaling in neurons and heart	[46, 58, 59]
Resistance to inhibitors of cholinesterase-8A/ Synembryn-A	Ric8A	?	Increases synapse number and synaptic release probability	[33]
Transient Receptor Protein 5	TRPC5	Activates channel	Retards neurite growth	[60]

TGN: trans-Golgi network

The ER resident inositol 1,4,5-trisphosphate receptor (InsP₃R) binds NCS-1 and has been identified as a key player in many Ca²⁺ dependent cellular events [39]. Loss of InsP₃R regulation is associated with pathological conditions ranging from epileptic seizures and ataxia [40] to bile duct obstruction [41]. An important aspect of InsP₃R signaling pathways is that regulation of the channel activity is modulated by Ca²⁺ itself [42]. In addition to NCS-1, a number of other Ca²⁺ binding proteins interact with the InsP₃R as activators, modulators, and inhibitors [43–45].

NCS-1 modulates the InsP₃R by increasing InsP₃-gated channel activity without altering the amplitude of the single-channel currents [46]. This modulation is Ca²⁺ dependent, so that high levels of Ca²⁺ increase the channel open probability more than low Ca²⁺ [46]. NCS-1 alone, without InsP_3, is not sufficient to activate the InsP₃R, showing that NCS-1 acts as a modulator, rather than as a direct agonist. The specificity of the functional effects was emphasized by the lack of effect of NCS-1 on the activity of the ryanodine receptor (RyR). An EF-hand mutant of NCS-1 (E120Q) [47] with reduced Ca²⁺-binding ability, was still able to increase the open probability of the InsP₃R. However, the additional amplification that was seen with increased Ca²⁺ was no longer observed [46]. Measurements of Ca²⁺ transients in cells over-expressing NCS-1 confirmed the effects of increased NCS-1 observed with the isolated InsP₃R incorporated into planar lipid bilayers [46].

3. NCS-1 in disorders and diseases

The range of molecular interactions listed in Table 1 makes NCS-1 a likely contributor in pathophysiology (Table 2). Post mortem studies found increased levels of NCS-1 in the prefrontal cortex of bipolar and schizophrenic patients [61]. Furthermore, NCS-1 has been implicated in sleep-wake disturbance symptoms in bipolar disease [62] and this interaction could contribute the effectiveness of lithium in treating bipolar disorder patients [63]. Mutations in Interleukin-1 Receptor Accessory Protein-Like 1 (IL1RAPL1), one of NCS-1’s target proteins, are associated with a wide range of neurological impairments [64]. The same study found a missense mutation of NCS-1 (R102Q) in an autistic patient [64]. This mutation was further characterized and it appears to disrupt target protein interaction [65, 66]. The interaction of NCS-1 with the Dopamine D2 Receptor [67] supports the implication of NCS-1 in the susceptibility to cocaine addiction [68] and the effectiveness of nicotine replacement therapy [69]. In Parkinson’s disease (PD), increased levels of NCS-1 mRNA were measured in substantia nigra neurons [70]. In a rat model, a significant decrease of NCS-1 levels in the prefrontal cortex was proposed to contribute to cognitive impairment during sepsis [71]. These studies convey preliminary insights in the complex roles for NCS-1 and they validate in vitro pathways.

Table 2.

Diseases associated with NCS-1

Field of study	Disorder/Sensory System	Organism	NCS-1’s Role/Observation	References
Neuro-Psychiatry	Bipolar disorder & Schizophrenia	human, rat	>50% higher levels of NCS-1 in the PFC compared to normal controls Chronic treatment with typical or atypical antipsychotics does not change NCS-1 expression in five brain regions: prefrontal cortex, hippocampus, striatum, cortex and cerebellum	[61, 74–80]
			Modulation of gamma band oscillations in the PPN in a concentration-dependent manner
			Lithium reduces effects of over expressed NCS -1 on PPN neurons
	Autistic spectrum disorder (ASD)	human	Rare missense mutation was identified in one autistic patient during study	[64, 65]
	X linked mental retardation	drosophil a	Potential drug target at the NCS-1/Ric8a interface	[30, 54 ]
	Addiction	human	Genetic polymorphisms are associated with cocaine addiction in African-Americans but not European Americans	[68, 69]
			Variations of the NCS-1 gene influence the efficacy of nicotine replacement therapy
	ADHD	rat	Methylphenidate induces changes in expression levels in rat hippocampus, prefrontal cortex and cerebellum	[81]
	Depression/Motivation	mouse	Deficiency appeared to result in anxiety- and depressive-like behaviors as well as in decreased willingness to work for food	[82, 83]
	Insomnia	rat	Dysregulation may lead to increased activity in PPN neurons. Potential mediated target for modulation of hyperarousal	[84]
	Parkinson’s disease (PD)	human, mouse	Post-mortem PD brains show increased NCS-1 mRNA suggesting increased NCS-1/D2- autoreceptor signalling in PD	[70, 85]
			Potential target in modulating the neuron activity and vulnerability to degeneration in PD
	CNS trauma	rat	Potential intracellular target for therapeutic intervention following injury to the central nervous system; NCS-1-induced neurite sprouting	[37]
	Neuro-degeneration (ND)	in vitro	Misfolding at physiological Ca2+ levels suggests potential link between Ca2+ dysregulation, protein misfolding and ND	[86]
	Memory	mouse	Deficiency shows impaired spatial learning and memory function as well as reduced exploration	[18–20, 87]
			Overexpression selectively in the adult murine dentate gyrus promotes a specific form of exploratory behavior
			Regulation of genes that are related to intrinsically motivated exploration, something that could be considered akin to curiosity.
			Up regulation of expression in the hippocampus through swimming training promotes memory
Oncology	CIPN	mouse	Prevention target through lithium pretreatment	[24, 46, 88 - 95]
	Taxol-induced cardiac arrhythmia	rat, mouse	Increased expression in cardiomyocytes after treatment with Paclitaxel leads to an acceleration of Ca2+ oscillations	[58]
	Breast cancer	human	Outcome predictor	[72]
Cardiology	Cardiac hypertrophy and stress	mouse	Increased expression in early stages of cardiac hypertrophy and potential mediator of hormone-induced progression of hypertrophy in adult hearts. Mediator of stress tolerance in cardiomyocytes; Upregulation in hearts after ischemia-reperfusion	[59, 73]
Infectious Diseases	Sepsis	rat	Low expression in prefrontal cortex may be associated with the pathophysiology of cognitive impairment during sepsis	[71]
Gastro-enterology	Colitis	rat, ENS	Selective loss of NCS-1 expression after DNBS-induced colitis	[96]
Urology	Erectile dysfunction (ED)	rat, penile tissue	Potential target in the treatment of ED, up-regulation after administration of tadalafil	[97]
Developmenta l	Olfactory system	mouse	Expression in olfactory epithelium during development, down-regulation of axonal expression after synapse formation	[27]
Biology
	Eye	chick/rat	Neuronal process outgrowth and synaptogenesis in the retina	[98, 99]
	Inner ear	zebrafish	Signaling pathway for semicircular canal formation, Knockdown of NCS-1a mRNA blocked formation of semicircular canals.	[100, 101]
	Heart	mouse	High expression in the heart during fetal period and decline after birth; regulator of excitation-contraction coupling in fetal and neonatal hearts through enhancement of Ca(2+) signals	[59, 102, 103]

ADHD: attention deficit hyperactivity disorder, CIPN: Chemotherapy induced peripheral neuropathy, CNS: Central nervous system, DNBS: dinitrobenzene sulfonic acid, ENS: Enteric Nervous System, PPN: pedunculopontine nucleus

In addition to the involvement of NCS-1 in neuro-psychiatric diseases, an up-regulation of NCS-1 mRNA was found in a variety of non-neurological diseases. For example, NCS-1 has been proposed to be a biomarker in aggressive breast cancer [72]. In the heart, NCS-1 mediates paclitaxel-induced cardiac arrhythmias through altered Ca²⁺ signaling via the InsP₃R [58]. It also contributes to stress tolerance in cardiomyocytes by activation of Ca²⁺ dependent survival pathways [73].

3.1 NCS-1, Paclitaxel, and Chemotherapy-Induced Peripheral Neuropathy

Drug development is driven by the aim to design a compound that has maximum efficacy with minimal side effects. Understanding the pharmacodynamics of the drug, as well as the pathophysiology of the associated side effects, can help to distinguish molecular targets responsible for wanted and unwanted effects. If the same pathway is responsible for both therapeutic and non-therapeutic effects, additional treatment regimens are needed. In some cases reduced timing or dosing has been utilized to alleviated the unwanted effects of treatment. In other cases the molecular cause of the side effect stems from a pathway that runs independently to the desired pathway. When a parallel pathway is responsible, the non-therapeutic target can be used to develop or identify compounds that attenuate the side effect without compromising the therapeutic effect (Fig. 2).

Fig. 2. A model of desired and unwanted drug effects.

After application, a drug can operate through several pathways that entail different molecular players and lead to distinct positive therapeutic effects (green) and undesired side effects (red). Because Pathway C (blue) mediates only side effects, it is an ideal target for pharmacological intervention. In contrast, intervention of ’Pathway B’ comes with the risk of compromising desired effects because this pathway leads to both wanted and undesired effects.

Paclitaxel, a commonly used chemotherapeutic agent [104], interacts with NCS-1 and upon binding initiates a pathway that is responsible for CIPN, a severe, dose limiting side effect. This pathway differs significantly from the established antineoplastic mechanism of action of this drug and can be targeted to prevent CIPN.

3.2 Intracellular Ca²⁺ Signaling and Paclitaxel

Paclitaxel (Taxol) is a chemotherapeutic agent that is still widely used in ovarian, breast, and lung cancer, as well as Kaposi’s sarcoma [104]. The main mechanism of action of paclitaxel is the induction of mitotic arrest through stabilization and polymerization of microtubules [105]. Interaction with several proteins other than tubulin, including heat shock proteins and the antiapoptotic protein Bcl-2 [106, 107], has been reported but the functional consequences of these interactions are unclear [108]. In contrast, NCS-1 was found to be a binding partner of paclitaxel [90] and this binding was shown to initiate an undesired side effect of paclitaxel therapy, CIPN. CIPN is a severe, debilitating condition leading to numbness or pain and occurs in over 50 % of paclitaxel-treated patients [109]. Because there is no effective medication preventing or reversing CIPN, a targeted drug for this condition is urgently needed to improve quality of life of cancer survivors.

Application of paclitaxel to human neuroblastoma cells (SH-SY5Y) evoked an oscillatory cytosolic Ca²⁺ increase that was concentration dependent [90]. It is worth noting that many previous experiments were performed using high paclitaxel concentrations of 10–20 μM or higher, whereas recent experiments used paclitaxel concentrations of 1 μM which more closely represent the maximum plasma concentration of <1μM [110] observed in patients. These oscillations required NCS-1 expression in the cells and Ca²⁺ release from the ER [90]. Selective inhibition of the RyR resulted only in minor changes of the Ca²⁺ response to paclitaxel, supporting the hypothesis that the RyR is not a direct target of NCS-1 and paclitaxel [90, 111].

Previous work has attributed the paclitaxel side-effects primarily to opening of the mitochondrial permeability transition pore [112, 113]. With the opening of this pore intracellular Ca²⁺ would be increased and cell death pathways would be initiated. As paclitaxel binds directly to NCS-1, not the mitochondrial proteins, there would first be Ca²⁺ release from the NCS-1 regulated channel, the InsP₃R . The excess paclitaxel-induced Ca²⁺ release from the ER would lead to Ca²⁺ uptake and subsequent Ca²⁺ overload of the mitochondrial. The mitochondrial stress that develops as a consequence of overload would indeed lead to unwanted cellular responses and cell death. However, the initiating event is most likely release of Ca²⁺ from the ER.

3.3 Molecular Mechanism of CIPN Development in the Context of Paclitaxel Treatment

Experiments demonstrated an increased binding of NCS-1 to the InsP₃R if submicromolar concentrations of paclitaxel were present [90]. Furthermore, knockdown of NCS-1 abrogated Ca²⁺ responses to paclitaxel stimulation but not stimulation with ATP. This finding strongly suggests that paclitaxel induced Ca²⁺ oscillations depend on NCS-1 [114]. In addition to the acute treatment effects, chronic exposure of SH-SY5Y with submicromolar paclitaxel concentrations resulted in an attenuation of signaling without impairing cell viability, depletion of intracellular Ca²⁺ stores or degradation of InsP₃R [91] (Fig. 3). In these experiments, NCS-1 protein levels decreased after incubation with paclitaxel leading to a decrease in Ca²⁺ signaling [91], but no interaction between tubulin and NCS-1 could be demonstrated [91]. In contrast, the Ca²⁺ dependent protease calpain does show a time and dose dependent increase in activity in response to treatment with paclitaxel [89, 91]. It was shown that NCS-1 is a calpain target and inhibition of calpain activation also preserved Ca²⁺ signaling [91].

Fig. 3. Possible pathophysiological pathway of CIPN.

Paclitaxel facilitates binding of NCS-1 to InsP₃R, thereby inducing Ca²⁺ oscillations from the ER. The increased cytosolic Ca²⁺ is further enhanced through activation of Ca²⁺ dependent TRPV4 channels. Ca²⁺ activates the Ca²⁺ dependent protease calpain, which cleaves and a number of other proteins. This creates a negative feedback loop and impairs signaling through the InsP₃R. This leads to an attenuation of Ca²⁺ oscillations. Impaired signaling and further degradation of proteins lead to axonal damage and cell death. Lithium and ibudilast both interact with NCS-1 and prevent enhanced InsP₃R activity and NCS-1 protein degradation. TRPV4 inhibitor HC-067047 reduces mechanical allodynia after paclitaxel treatment. BAPTA and other Ca2+ reducing agents have been shown to inhibit the pathway. Overexpression of calpastatin, an endogenous calpain inhibitor, as well as overexpression of NCS-1 mutations (I35H or I35A) were able to protect WT NCS-1 in human neuroblastoma cells.

The plasma membrane Ca²⁺ channel, transient receptor potential vanilloid 4 (TRPV4), has been implicated in the development of paclitaxel induced neuropathy in rats and knock-down of TRPV4 prevented the occurrence of CIPN [88]. Unlike most permeable Ca²⁺ channels, TRPV4 activity is potentiated by increasing intracellular Ca²⁺ concentrations [115]. Although a direct effect of paclitaxel on TRPV4 has not be shown, the response to increased intracellular Ca²⁺ suggests that TRPV4 channels could be opened when calcium is released from the ER and could augment the increased intracellular Ca²⁺ concentrations and further activate calpain. TRPA1 has also been implicated in the development of neuropathic pain [116].

In summary, these findings lead to a model that conflicts with previous models suggesting that paclitaxel induced CIPN is merely mediated through paclitaxel’s microtubule interfering effect [117–119] or a side effect of the vehicle use in paclitaxel treatment (Cremophor EL) [120]. It seems more likely that the unwanted effects of paclitaxel treatment are consequences of the modulation of an independent pathway that disrupts Ca²⁺ signaling (Fig. 3): In this Ca²⁺ model, paclitaxel facilitates binding of NCS-1 to InsP₃R, thereby inducing Ca²⁺ oscillations. The increased cytosolic Ca²⁺ may be further enhanced through activation of Ca²⁺ dependent TRPV4 channels (leading to influx of extracellular calcium). Ca²⁺ activates the Ca²⁺ dependent protease calpain, which in turn cleaves proteins, including NCS-1. This process creates a negative feedback loop, impairing signaling through the InsP₃R, leading to an attenuation and ultimately a cessation of Ca²⁺ oscillations. Impaired signaling and further degradation of proteins through activation of calpain and caspases will lead to irreversible axonal damage and cell death.

3.4 Prevention of NCS-1-mediated CIPN

3.4.1 Introduction

A deeper understanding of the molecular basis for CIPN allows serious investigations into clinical strategies to prevent this side effect of chemotherapy. Such improvements in treatment are needed because CIPN is a dose limiting side effect that reduces quality of life and can lead to interruption or discontinuation of therapy [121]. Only a few strategies to prevent CIPN have been investigated in the last decade, and the best supported treatments, supplementation of vitamin E [122, 123] and the administration of pifithrin-μ [124, 125], have limited success. The American Society of Clinical Oncology (ASCO) clinical practice guideline recommends no agents for the prevention of CIPN due to the lack of evidence [126]. Given this lack of effective treatments, prevention strategies are needed. The proposed NCS-1-dependent signaling pathway suggests several potential targets (Fig. 3).

Recovery of mild neuropathy can sometimes be achieved after treatment discontinuation [127]. Although these altered treatments are well established strategies to prevent CIPN, a potential downside is a reduction of antineoplastic effects. Co-administration of an inhibitor of the proposed pathway might be a viable alternative to prevent CIPN without reducing treatment efficacy.

3.4.2 Possible Interventions

Paclitaxel treatment leads to the activation of calpains [89, 91]. In mouse studies, inhibition of calpain resulted in protection against CIPN without compromising the antineoplastic effect [89]. However, inhibition of calpain in patients is limited due to the expression of this protease in most tissues [128, 129]. With the known cleavage site of NCS-1 [24], a drug based on the amino acid sequence could function as a competitive inhibitor and thus prevent cleavage of native NCS-1. This approach was shown to be successful in cell based assays [92, 130, 131]. These results highlight the importance of calpain in the development of CIPN and suggest that calpain inhibition, if specificity could be achieved, is a promising approach for prevention of CIPN. Similar to calpain, manipulation of Ca²⁺ is difficult because of the universality of this signaling pathways. In isolated cells, buffering of free cytosolic Ca²⁺ prevented degradation of NCS-1 after paclitaxel treatment [91]. However, the usefulness of this type of treatment in humans seems limited.

A selective antagonist of TRPV4 (HC-067047 [132]) has been used to confirm the role of TRPV4 in the development of CIPN in a mouse model [116]. This fits the proposed model (Fig 3) and thus TRPV4 antagonists are another class of potential drug to prevent CIPN. Additional inhibitors for TRPV4 are under investigation [133].

Prevention of NCS-1 degradation and protection of intracellular Ca²⁺ signaling was also observed when cells were pre-treated with either lithium or ibudilast [93] and binding of these drugs to NCS-1 did not interfere binding of NCS-1 to paclitaxel [93]. Addition of lithium or ibudilast did not alter tubulin assembly, implying no decrease of antineoplastic activity of paclitaxel in the presence of these potential CIPN preventers [93]. Lithium is a drug that is used to treat bipolar disorder [134–136]. Ibudilast, a phosphodiesterase inhibitor, is used to treat bronchial asthma [137]. Considering the successful administration in several cases of sensory-motor neuropathy [138–140]. Beneficial effects of ibudilast treatment in rat models of neuropathic pain were also reported [141, 142].

Based on these findings, the effects of lithium and ibudilast was tested in mice treated with paclitaxel [94]. A single prophylactic injection of lithium or ibudilast prior to paclitaxel treatment inhibited CIPN in mice without compromising the anti-neoplastic effects [94]. Myelin sheath destruction and macrophage infiltration could also be prevented with lithium pre-treatment [143]. Another pathway known to be affected by lithium, the serine/threonine kinase GSK-3-beta pathway [144, 145], did not appear to prevent CIPN nor did it alter the paclitaxel influence on the NCS-1/InsP₃R pathway [94].

4. Conclusion

NCS-1 regulates a variety of physiological pathways and it is increasingly recognized as a contributor to pathophysiology. Among NCS-1-associated diseases are neurological conditions including bipolar disease and non-neurological disorders such as cancer. These diseases related to altered NCS-1 function are logical consequences of changes in Ca²⁺ signaling and binding of NCS-1 to various cellular proteins.

For example, Ca²⁺-signaling pathways modulated by NCS-1 are important contributors to the clinical picture of paclitaxel induced peripheral neuropathy. The binding of paclitaxel to NCS-1 initiates a cascade that leads to decreased neuronal function and this alteration influences the progression to CIPN. This pathway appears to be parallel to the path used for the beneficial anti-neoplastic effects of chemotherapy. Because the development of CIPN by paclitaxel is independent of tumor shrinkage and because there is molecular understanding of the aspects of the NCS-1 pathway related to CIPN initiation, it has been possible to propose treatments that prevent this side effect of chemotherapy.

Fig. 1. Interaction of NCS-1 and InsP3R.

(A) Upon binding of InsP₃, the InsP₃R releases calcium from the ER. (B) NCS-1 alone is not sufficient to activate InsP₃R. (C) Binding of NCS-1 enhances InsP₃ mediated InsP₃R channel activity by increasing the open probability and mean open time. (D) Lithium offsets NCS-1 mediated enhancing effects on InsP₃R. (E) Paclitaxel binds the hydrophobic cleft of NCS-1 in calcium-bound state. Thereby it facilitates binding of NCS-1 to InsP₃R, thus inducing calcium oscillations. (F) Lithium (as well as ibudilast) inhibit paclitaxel mediated, NCS-1 dependent increases in InsP₃R channel activity, preventing calcium oscillations. Note that many NCS-1 molecules are membrane bound.

Highlights.

• –NCS-1 is a calcium binding protein interacting with many cellular components
• –Binding of NCS-1 to ion channels regulates calcium homeostasis
• –Alterations in NCS-1 signaling are associated with neurological diseases
• –Targeting NCS-1 is a viable therapeutic option in these disorders