Is γ-secretase a beneficial inactivating enzyme of the toxic APP C-terminal fragment C99?

Abstract

Genetic, biochemical, and anatomical grounds led to the proposal of the amyloid cascade hypothesis centered on the accumulation of amyloid beta peptides (Aβ) to explain Alzheimer's disease (AD) etiology. In this context, a bulk of efforts have aimed at developing therapeutic strategies seeking to reduce Aβ levels, either by blocking its production (γ- and β-secretase inhibitors) or by neutralizing it once formed (Aβ-directed immunotherapies). However, so far the vast majority of, if not all, clinical trials based on these strategies have failed, since they have not been able to restore cognitive function in AD patients, and even in many cases, they have worsened the clinical picture. We here propose that AD could be more complex than a simple Aβ-linked pathology and discuss the possibility that a way to reconcile undoubted genetic evidences linking processing of APP to AD and a consistent failure of Aβ-based clinical trials could be to envision the pathological contribution of the direct precursor of Aβ, the β-secretase-derived C-terminal fragment of APP, βCTF, also referred to as C99. In this review, we summarize scientific evidences pointing to C99 as an early contributor to AD and postulate that γ-secretase should be considered as not only an Aβ-generating protease, but also a beneficial C99-inactivating enzyme. In that sense, we discuss the limitations of molecules targeting γ-secretase and propose alternative strategies seeking to reduce C99 levels by other means and notably by enhancing its lysosomal degradation.

Keywords: APP C-terminal fragments; C99; clinical trials; toxicity; β-secretase; γ-secretase.

Figure 1

APP metabolism.A, represents the α/γ-secretase pathway in which APP is first cleaved by the α-secretase producing the soluble fragment sAPPα and the membrane-embedded C-terminal fragment, C83 that is then cleaved by γ-secretase to release the cytosolic AICD fragment and p3 peptide, respectively. B, represents the β or β′/γ-secretase pathway in which APP is first cleaved by the β-secretase at β or β′ site to liberate the soluble fragments sAPPβ or sAPPβ’ and the membrane-embedded C99 or C89 fragments, respectively. Then γ-secretase cleavage releases the cytosolic AICD fragment and Aβ peptides (Aβ1-40 or Aβ1-42) or Aβ11–40, respectively. Note that in physiological conditions, β-secretase mainly cleaves at the β’ site, but some FAD mutations shift the cleavage to the β-site to produce more C99. C, represents the ƞ/γ-secretase pathway in which ƞ-secretase cleavage releases the soluble fragment sAPPƞ and the membrane-embedded ƞCTF, which is processed by α- or β-secretase, thus generating Aƞα or Aƞβ peptides, respectively.

Figure 2

γ-secretase.A, displays a schematic representation of presenilins (PS1 or PS2) composed of nine transmembrane domains and harboring the catalytic core of the complex with two aspartyl residues in the transmembrane domains TM-6 and TM-7. During maturation, presenilin undergoes endoproteolysis, and the resulting N-terminal fragment (PS-NTF) and C-terminal fragment (PS-CTF) remain associated. B, represents the multimeric γ-secretase complex composed of four membrane proteins: presenilin (green), nicastrin (blue) composed of a single transmembrane and a large glycosylated N-terminal ectodomain, Aph-1 (purple) composed of seven transmembrane domains, and Pen-2 (pink) composed of two transmembrane domains. The substrate C99 (yellow) interacts with γ-secretase to form an enzyme substrate complex. C, represents the successive proteolysis of C99 transmembrane domain by γ-secretase. First, an endoproteolytic cleavage occurs close to the membrane–cytosol interface at two possible ε sites yielding long Aβ peptides Aβ48 or Aβ49, respectively. Then a carboxypeptidase activity of γ-secretase trims Aβ48 and Aβ49 each 3 or 4 amino acid at successive ζ-, γ, and γ′-sites leading to two Aβ product lines: Aβ49→Aβ46→Aβ43→Aβ40 and Aβ48→Aβ45→Aβ42→ Aβ38, respectively.

Figure 3

Schematic models for C99 accumulation. A, displays a schematic model of C99 fate in the absence or presence of γ-secretase cleavage. In physiological conditions, γ-secretase cleavage of C99 (blue/red sticks) leads to the generation of Aβ and AICD, whereas when γ-secretase activity is impaired (inhibitors or PS mutations), C99 accumulates and aggregates. B, displays a schematic view of C99 generation and accumulation in conditions of blocked γ-secretase. APP (long blue/red bar) maturates and traffics through the Golgi to the plasma membrane (blue arrows), where a small part of it is processed by α-secretase (non-amyloidogenic pathway). APP that escapes α-secretase-mediated cleavage is endocytosed into early endosomes and some of it is recycled to the plasma membrane, either directly or through the Golgi network (open black arrows). Within early endosomes, some APP is cleaved by β-secretase generating C99 (short blue/red bar), which then can undergo γ-secretase cleavage, thereby releasing the Aβ peptide and AICD. Some C99 escapes from γ-secretase cleavage and is incorporated in ILVs (intra luminal vesicles), which are either released as exosomes after the fusion of multivesicular bodies (MVBs) with the plasma membrane (1) or degraded after their fusion of MVBs with lysosomes or with autophagosomes, which then fuse with lysosomes (2). The presence of APP and PS mutations or γ-secretase inhibitors and/or lowered lysosomal degradation lead to lysosomal, autolysosomal, and exosomal C99 accumulation. C99 has also been found to accumulate in MAMs and mitochondria.

Figure 4

C99 toxicity. C99 accumulation can be a consequence of either the presence of FAD mutations or a lowered degradation (possibly linked to risk factors). C99 firstly accumulates in endosomes and can itself be a direct cause of a dysfunction of the endosomal–lysosomal–autophagic (EAL) degradation pathway, thus leading to a vicious pathological cycle, in which more C99 accumulates, aggregates, and becomes neurotoxic by causing synaptic dysfunction, inflammation, mitochondrial dysfunction, and exosomal spread. Therapeutic strategies aimed at reducing C99 levels could be through either BACE1 inhibitors and/or BACE1 aptamers, which should be expected to reduce its production or autophagic/lysosomal activators or other drugs interfering with C99 degradation, which should increase its degradation.