Phthalocyanine Tetrasulfonates Affect the Amyloid Formation and Cytotoxicity of α-Synuclein

Abstract

Alpha-synuclein is a pathological component of Parkinson's disease by constituting the filamentous component of Lewy bodies. Phthalocyanine (Pc) effects on the amyloidosis of alpha-synuclein have been examined. The copper complex of phthalocyanine tetrasulfonate (PcTS-Cu(2+)) caused the self-oligomerization of alpha-synuclein while Pc-Cu(2+) did not affect the protein, indicating that introduction of the sulfonate groups was critical for the selective protein interaction. The PcTS-Cu(2+) interaction with alpha-synuclein has occurred predominantly at the N-terminal region of the protein with a K(d) of 0.83 microM apart from the hydrophobic NAC (non-Abeta component of Alzheimer's disease amyloid) segment. Phthalocyanine tetrasulfonate (PcTS) lacking the intercalated copper ion also showed a considerable affinity toward alpha-synuclein with a K(d) of 3.12 microM, and its binding site, on the other hand, was located at the acidic C-terminus. These mutually exclusive interactions between PcTS and PcTS-Cu(2+) toward alpha-synuclein resulted in distinctive features on the kinetics of protein aggregation, morphologies of the final aggregates, and their in vitro cytotoxicities. The PcTS actually suppressed the fibrous amyloid formation of alpha-synuclein, but it produced the chopped-wood-looking protein aggregates. The aggregates showed rather low toxicity (9.5%) on human neuroblastoma cells (SH-SY5Y). In fact, the PcTS was shown to effectively rescue the cell death of alpha-synuclein overexpressing cells caused by the lactacystin treatment as a proteasome inhibitor. The anti-aggregative and anti-amyloidogenic properties of PcTS were also demonstrated with alcohol dehydrogenase, glutathione S-transferase, and amyloid beta/A4 protein under their aggregative conditions. The PcTS-Cu(2+), on the other hand, promoted the protein aggregation of alpha-synuclein, which gave rise to the fibrillar protein aggregates whose cytotoxicity became significant to 35.8%. Taken together, the data provided in this study indicate that PcTS/PcTS-Cu(2+) could be considered as possible candidates for the development of therapeutic or prophylactic strategies against the alpha-synuclein-related neurodegenerative disorders.

Toward the Discovery of Effective Polycyclic Inhibitors of α-Synuclein Amyloid Assembly

“…Several studies have focused recently on the role of small molecules that interact with AS and inhibit its aggregation-fibrillation in vitro and in vivo (19,20,23,(25)(26)(27)(28)(29). Polyphenols, flavonoids, porphyrins, and phthalocyanines are all polyaromatic scaffolds included in this group of compounds.…”

“…One of the most studied polyaromatic aggregation inhibitors is the cyclic tetrapyrrole phthalocyanine tetrasulfonate (PcTS). This compound has been shown to exhibit anti-scrapie activity in vitro and in vivo (17,22), disassembling of tau filaments (24), and inhibition of AS filament assembly, leading to the formation of nontoxic AS aggregates (29). We recently elucidated key aspects related to the structural and molecular basis behind the inhibitory interaction of this compound with AS (28).…”

Dequalinium-induced Protofibril Formation of α-Synuclein

“…This abnormal behavior could be due to possible cooperative interaction of the compound to the protein and subsequent nonspecific protein aggregation, which might suppress the ladder formation on the gel at the higher concentrations of C14-DQ. The phenomenon of ligand-induced self-oligomerization of ␣-synuclein has been demonstrated to be a highly selective process between the protein and various small chemicals, such as eosin, erythrosine B, Coomassie Brilliant Blue (G and R), and the coppercontaining phthalocyanine tetrasulfonate in addition to A␤25-35 and the divalent copper ion (26,36,37,45,51,53). The dequalinium interaction of ␣-synuclein, therefore, could be also considered as another highly selective process of molecular recognition.…”

“…In particular, we have been interested in screening for ␣-synuclein interactive small chemicals that could be used to control the amyloidosis eventually. Recently, phthalocyanine tetrasulfonate, previously proposed as an antiscrapie agent (35), was shown to interact with ␣-synuclein via selective binding to the acidic C terminus, and it prevented the cytotoxicity on SH-SY5Y cells caused by the overexpression of ␣-synuclein in the presence of a proteasomal inhibitor, lactacystin (36). The copper complex of phthalocyanine tetrasulfonate-Cu 2ϩ , on the other hand, influenced the protein to be self-oligomerized by interacting with the N-terminal region and stimulated the amyloid formation (36).…”

“…Recently, phthalocyanine tetrasulfonate, previously proposed as an antiscrapie agent (35), was shown to interact with ␣-synuclein via selective binding to the acidic C terminus, and it prevented the cytotoxicity on SH-SY5Y cells caused by the overexpression of ␣-synuclein in the presence of a proteasomal inhibitor, lactacystin (36). The copper complex of phthalocyanine tetrasulfonate-Cu 2ϩ , on the other hand, influenced the protein to be self-oligomerized by interacting with the N-terminal region and stimulated the amyloid formation (36). Coomassie Brilliant Blue-G and -R augmented the protein aggregation of ␣-synuclein and gave rise to two different shapes of the aggregates, such as worm-like and filamentous structures, respectively (26).…”