Calcium Signalling in Breast Cancer Associated Bone Pain

Abstract

Calcium (Ca²⁺) is involved as a signalling mediator in a broad variety of physiological processes. Some of the fastest responses in human body like neuronal action potential firing, to the slowest gene transcriptional regulation processes are controlled by pathways involving calcium signalling. Under pathological conditions these mechanisms are also involved in tumoral cells reprogramming, resulting in the altered expression of genes associated with cell proliferation, metastatisation and homing to the secondary metastatic site. On the other hand, calcium exerts a central function in nociception, from cues sensing in distal neurons, to signal modulation and interpretation in the central nervous system leading, in pathological conditions, to hyperalgesia, allodynia and pain chronicization. It is well known the relationship between cancer and pain when tumoral metastatic cells settle in the bones, especially in late breast cancer stage, where they alter the bone micro-environment leading to bone lesions and resulting in pain refractory to the conventional analgesic therapies. The purpose of this review is to address the Ca²⁺ signalling mechanisms involved in cancer cell metastatisation as well as the function of the same signalling tools in pain regulation and transmission. Finally, the possible interactions between these two cells types cohabiting the same Ca²⁺ rich environment will be further explored attempting to highlight new possible therapeutical targets.

Keywords: allodynia; bone; calcium; cancer; hyperalgesia; metastasis; nociception; pain; signalling.

Figure 1

Schematic overview of the main Ca²⁺ signalling mechanisms. Calcium release from the intracellular calcium stores is mainly mediated by inositol-1,4,5-triphosphate receptors (IP₃Rs) and ryanodine receptors (RYRs), sensitive to signals present both inside and outside the endoplasmic/sarcoplasmic reticulum (ER/SR). Inositol-1,4,5-triphosphate (IP₃) is produced by different isoforms of phospholipase C (PLC) and binding to IP₃Rs can trigger calcium release. The CD38 ADP ribosyl cyclase has both synthase (S) and hydrolase (H) functions producing cyclic ADP ribose (cADPr) and nicotinic acid adenine dinucleotide phosphate (NAADP) and has been proposed to act as a cellular metabolism sensor since ATP and NADH can inhibit the hydrolase enzymatic function. The exact way cADPr and NAADP influence calcium release is still unclear but they seem to indirectly act on both RYRs and IP₃Rs. cADPr might enhance sarco(endo)plasmic reticulum Ca²⁺-ATPase (SERCA) pump activity, increasing the level of Ca²⁺ in the ER/SR, sensitising the RYRs. NAADP triggers calcium release from a channel located on lysosomes-related organelles, locally increasing the intracellular calcium concentration which can in turn result in direct stimulation of RYRs and IP₃Rs or in an indirect stimulation through an increase of Ca²⁺ concentration in ER/SR similarly to what happens with cADPr. PI, phosphatidylinositol-4,5-bisphosphate. Adapted from [5].

Figure 2

Calcium sources and sinks. Calcium can enter neurons from the extracellular space through many membrane channels as NMDARs and AMPARs, P2XRs, ASICs, TRP channels and VDCCs while PMCA pumps and NCX Ca²⁺ exchanger extrude calcium to the extracellular environment to maintain the basal membrane potential and regulating the intracellular calcium concentration. The activation of menbrane Gq-coupled GPCRs and TKRs can trigger calcium release from the ER via IP₃Rs and ryanodine receptors (RYRs) where again SERCA and NCX are responsible for calcium re-uptake. Mitochondrial NCXL and MCU contribute to cytoplasmic calcium signalling by releasing and removing it respectively, while all calcium binding proteins (CaBP) act as calcium buffers amongst all stimuli, dynamically releasing and removing calcium ions depending on the concentration. Through nuclear pores cytosolic calcium can enter the nuclear space where it can regulate the activity of transcription factors acting therefore on genes expression. Adapted from [14].

Figure 3

Vicious cycle of osteolytic metastases. Tumoural cells secrete soluble factors as interleukines (IL), tumour-necrosis factor alpha (TNF-$\alpha$) and parathyroid-hormone related peptide (PTHrP), promoting osteoblasts mediated receptor activator of nuclear factor kappa-B ligand (RANKL) secretion. RANKL stimulates pre-osteoclasts differentiation into fully mature osteoclasts increasing therefore the bone matrix degradation and the release of the soluble factors trapped in it, like transforming-growth factor beta (TGF-$\beta$), bone morphogenic protein (BMP), fibroblasts growth factor (FGF), insuline-like growth factor (IGF) and platelet-derived growth factor (PDGF). Finally, these growth factors stimulate tumoural growth resulting therefore in a feed forward vicious cycle.

Figure 4

Mechanisms of extracellular acidification: Tumoural cells secrete H⁺ ions and import HCO₃⁻ to counter the intracellular acidification resulting from the increased glycolysis (Warburg Effect). This actively regulated process involves many plasma membrane proteins like HCO₃⁻ transporters, co-transporters and exchangers, proton pupms and exchangers, and monocarboxylate transporters (MCT). At the same time osteoclasts actively secrete protons to degrade the minaralized bone matrix acidifying the bone extracellular environment. Bicarbonate transporters (BTs), sodium-bicarbonate co-transporters (NBCs). anions exchangers (AEs), sodium-hydrogen exchangers (NHEs), vacuolar proton-translocating ATPase (V-ATPase), carbonic anhydrase 2 (CA2), glucose transporter 1 (GLUT1). Adapted from [134].