Calcium-sensing receptor antagonist (calcilytic) NPS 2143 specifically blocks the increased secretion of endogenous Aβ42 prompted by exogenous fibrillary or soluble Aβ25–35 in human cortical astrocytes and neurons—Therapeutic relevance to Alzheimer's disease

Abstract

The "amyloid-β (Aβ) hypothesis" posits that accumulating Aβ peptides (Aβs) produced by neurons cause Alzheimer's disease (AD). However, the Aβs contribution by the more numerous astrocytes remains undetermined. Previously we showed that fibrillar (f)Aβ25-35, an Aβ42 proxy, evokes a surplus endogenous Aβ42 production/accumulation in cortical adult human astrocytes. Here, by using immunocytochemistry, immunoblotting, enzymatic assays, and highly sensitive sandwich ELISA kits, we investigated the effects of fAβ25-35 and soluble (s)Aβ25-35 on Aβ42 and Aβ40 accumulation/secretion by human cortical astrocytes and HCN-1A neurons and, since the calcium-sensing receptor (CaSR) binds Aβs, their modulation by NPS 2143, a CaSR allosteric antagonist (calcilytic). The fAβ25-35-exposed astrocytes and surviving neurons produced, accumulated, and secreted increased amounts of Aβ42, while Aβ40 also accrued but its secretion was unchanged. Accordingly, secreted Aβ42/Aβ40 ratio values rose for astrocytes and neurons. While slightly enhancing Aβ40 secretion by fAβ25-35-treated astrocytes, NPS 2143 specifically suppressed the fAβ25-35-elicited surges of endogenous Aβ42 secretion by astrocytes and neurons. Therefore, NPS 2143 addition always kept Aβ42/Aβ40 values to baseline or lower levels. Mechanistically, NPS 2143 decreased total CaSR protein complement, transiently raised proteasomal chymotrypsin activity, and blocked excess NO production without affecting the ongoing increases in BACE1/β-secretase and γ-secretase activity in fAβ25-35-treated astrocytes. Compared to fAβ25-35, sAβ25-35 also stimulated Aβ42 secretion by astrocytes and neurons and NPS 2143 specifically and wholly suppressed this effect. Therefore, since NPS 2143 thwarts any Aβ/CaSR-induced surplus secretion of endogenous Aβ42 and hence further vicious cycles of Aβ self-induction/secretion/spreading, calcilytics might effectively prevent/stop the progression to full-blown AD.

Amyloid β-Exposed Human Astrocytes Overproduce Phospho-Tau and Overrelease It within Exosomes, Effects Suppressed by Calcilytic NPS 2143—Further Implications for Alzheimer's Therapy

“…Wakabayashi et al (2006) reported the co-localization of Aβ and p-Tau in the subiculum and entorhinal cortex astrocytes of a patient with corticobasal degeneration. They interpreted this finding as follows: “ the phagocytosis of A β coincides with production of phospho-Tau in the same reactive astrocytes .” However, as we previously showed, untreated cortical untransformed adult human astrocytes produce basal amounts of Aβ 42 and, once challenged with exogenous fibrillar (f)Aβ 25−35 , an Aβ 42 proxy, make and release significantly greater amounts of endogenous Aβ 42 /Aβ 42 -os (Armato et al, 2013; Dal Prà et al, 2015; Chiarini et al, 2016). We also demonstrated exogenous fAβ 25−35 -os bind the astrocytes' and neurons' calcium-sensing receptors (CaSRs) (Dal Prà et al, 2014a,b) and activate their signaling pathways heightening the production and secretion of endogenous Aβ 42 /Aβ 42 -os.…”

“…We also demonstrated exogenous fAβ 25−35 -os bind the astrocytes' and neurons' calcium-sensing receptors (CaSRs) (Dal Prà et al, 2014a,b) and activate their signaling pathways heightening the production and secretion of endogenous Aβ 42 /Aβ 42 -os. In fact, a specific CaSR agonist (calcimimetic), NPS R-568 (Nemeth and Goodman, 2016), mimicked the enhancing effect of exogenous fAβ 25−35 -os on Aβ 42 /Aβ 42 -os secretion (Armato et al, 2013). Conversely, a highly selective CaSR antagonist (calcilytic), NPS 2143 (Nemeth and Goodman, 2016), fully quelled the fAβ 25−35 -os-induced surplus de novo production and secretion of Aβ 42 /Aβ 42 -os in both human neurons and astrocytes (Armato et al, 2013; Dal Prà et al, 2015; Chiarini et al, 2016).…”

“…In fact, a specific CaSR agonist (calcimimetic), NPS R-568 (Nemeth and Goodman, 2016), mimicked the enhancing effect of exogenous fAβ 25−35 -os on Aβ 42 /Aβ 42 -os secretion (Armato et al, 2013). Conversely, a highly selective CaSR antagonist (calcilytic), NPS 2143 (Nemeth and Goodman, 2016), fully quelled the fAβ 25−35 -os-induced surplus de novo production and secretion of Aβ 42 /Aβ 42 -os in both human neurons and astrocytes (Armato et al, 2013; Dal Prà et al, 2015; Chiarini et al, 2016). Interestingly, human MIC neuroblasts secrete Tau enclosed within exosomes which are found in human cerebrospinal fluid too (Saman et al, 2012).…”

“…Altogether, these data indicated the urgent need to reassess the relationship between Aβ peptides exposure and p-Tau production and release in adult human astrocytes and neurons. Therefore, we undertook a pilot study using as model cultured human astrocytes (Armato et al, 2013) whose preliminary results we herein report. Details on materials and methods we used are in Supplementary Materials.…”

Dantrolene, A Treatment for Alzheimer Disease?

“…43,57,74,108,109 Dantrolene, as an antagonist of RYRs on ER, mitigates AD pathology and may serve as a probe compound for future studies to treat Alzheimer's disease.…”