The Spectrum of C9orf72-mediated Neurodegeneration and Amyotrophic Lateral Sclerosis

Abstract

The discovery that a hexanucleotide repeat expansion in C9orf72 is the most numerous genetic variant of both amyotrophic lateral sclerosis and frontotemporal dementia has opened a rapidly growing field, which may provide long hoped for advances in the understanding and treatment of these devastating diseases. In this review we describe the various phenotypes, clinical and pathological, associated with expansion of C9orf72, which go beyond amyotrophic lateral sclerosis and frontotemporal dementia to include neurodegeneration more broadly. Next we take a step back and summarize the current understanding of the C9orf72 expansion and its protein products at a molecular level. Three mechanisms are prominent: toxicity mediated directly by RNA transcribed from the repeat; toxicity mediated by dipeptide repeat proteins translated from the repeat sequence; and haploinsufficiency resulting from reduced transcription of the C9orf72 exonic sequence. A series of exciting advances have recently described how dipeptide repeat proteins might interfere with the normal role of the nucleolus in maturation of RNA binding proteins and in production of ribosomes. Importantly, these mechanisms are unlikely to be mutually exclusive. We draw attention to the fact that clinical and pathological similarities to other genetic variants without a repeat expansion must not be overlooked in ascribing a pathogenic mechanism to C9orf72-disease. Finally, with a view to impact on patient care, we discuss current practice with respect to genetic screening in patients with and without a family history of disease, and the most promising developments towards therapy that have been reported to date.

Fig. 1

Characteristic pathology of C9orf72 amyotrophic lateral sclerosis (ALS). Motor neurons of the spinal cord display typical TAR DNA binding protein-43 (TDP-43) pathology, including cytoplasmic TDP-43 positive skeins and compact inclusions [(A), anterior horn, stained with 3,3' diaminobenzidine (DAB) for pTDP-43, scale bar 100 μm] and Bunina bodies [(B), anterior horn, hematoxylin and eosin stain, scale bar 100 μm]. However, in addition, patients with C9orf72 ALS display p62-positive cytoplasmic inclusions in extramotor areas [(C), hippocampus dentate gyrus, DAB stained for p62, scale bar 100 μm], which also stain for dipeptide repeat protein [(D), cerebellar granule neurons, stained for poly-GA and 4',6'-diamidino-2-phenylindole (DAPI), scale bar 3 μm]. Numerous tissues also show nuclear RNA foci [(E), cerebellar granule neurons, stained for (GGGGCC)₃ and DAPI, foci are indicated by arrowheads, scale bar 3 μm]

Fig. 2

Proposed mechanisms of toxicity mediated by protein sequestration to RNA foci. Various proteins have been shown to be sequestered to RNA foci transcribed either in the sense or antisense direction from the GGGGCC repeat sequence. Proposed mechanisms include sequestration of proteins important to mRNA splicing with consequent disruption of RNA splicing; sequestration of adenosine deaminase, RNA-specific, B2 (ADARB2) (nonfunctional) leading to increased susceptibility to excitotoxicity; and sequestration of nucleolin producing nucleolar stress

Fig. 3

Proposed mechanisms of toxicity mediated via translation of dipeptide repeat protein (DPR). Both sense and antisense, or GGGGCC and GGCCCC repeat RNA species are observed to be translated into DPRs. The absence of traditional ATG start sites has led to the proposal that this occurs via a repeat-associated initial or repeat-associated non-ATG (RAN) translation. The various DPR sequences are proposed to be toxic via a number of mechanisms, including disruption of normal processing of RNA-binding proteins associated with the nucleolus leading to nucleolar stress and/or disruption of normal RNA splicing; or disruption of normal protein processing pathways leading to formation of protein inclusions and toxic protein stress. Implicated DPR species are shown for each mechanism

Fig. 4

Proposed therapeutic approaches for C9orf72 disease. At this point, 2 therapeutic approaches have been proposed and partially demonstrated for C9orf72 disease. First, an antisense oligonucleotide approach whereby the introduced molecule forms a double-stranded complement with the transcribed repeat sequence and thus targets it for degradation by RNAse H. This has been reported to reverse the formation of RNA foci and rescue certain molecular phenotypes. Second, a small molecule approach targeting the secondary structure of GGGGCC repeat RNA, which is thought to be in equilibrium between a hairpin and a G-quadruplex conformation, has been shown to disrupt both formation of RNA foci and translation of dipeptide repeat proteins (DPR)