Amyloidogenesis: What Do We Know So Far?

The study of protein aggregation, and amyloidosis in particular, has gained considerable interest in recent times. Several neurodegenerative diseases, such as Alzheimer's (AD) and Parkinson's (PD) show a characteristic buildup of proteinaceous aggregates in several organs, especially the brain. Despite the enormous upsurge in research articles in this arena, it would not be incorrect to say that we still lack a crystal-clear idea surrounding these notorious aggregates. In this review, we attempt to present a holistic picture on protein aggregation and amyloids in particular. Using a chronological order of discoveries, we present the case of amyloids right from the onset of their discovery, various biophysical techniques, including analysis of the structure, the mechanisms and kinetics of the formation of amyloids. We have discussed important questions on whether aggregation and amyloidosis are restricted to a subset of specific proteins or more broadly influenced by the biophysiochemical and cellular environment. The therapeutic strategies and the significant failure rate of drugs in clinical trials pertaining to these neurodegenerative diseases have been also discussed at length. At a time when the COVID-19 pandemic has hit the globe hard, the review also discusses the plausibility of the far-reaching consequences posed by the virus, such as triggering early onset of amyloidosis. Finally, the application(s) of amyloids as useful biomaterials has also been discussed briefly in this review.

AA-amyloidosis in cats (Felis catus) housed in shelters.

Systemic AA-amyloidosis is a protein-misfolding disease characterized by fibril deposition of serum amyloid-A protein (SAA) in several organs in humans and many animal species. Fibril deposits originate from abnormally high serum levels of SAA during chronic inflammation. A high prevalence of AA-amyloidosis has been reported in captive cheetahs and a horizontal transmission has been proposed. In domestic cats, AA-amyloidosis has been mainly described in predisposed breeds but only rarely reported in domestic short-hair cats. Aims of the study were to determine AA-amyloidosis prevalence in dead shelter cats. Liver, kidney, spleen and bile were collected at death in cats from 3 shelters. AA-amyloidosis was scored. Shedding of amyloid fibrils was investigated with western blot in bile and scored. Descriptive statistics were calculated. In the three shelters investigated, prevalence of AA-amyloidosis was 57.1% (16/28 cats), 73.0% (19/26) and 52.0% (13/25), respectively. In 72.9% of cats (35 in total) three organs were affected concurrently. Histopathology and immunofluorescence of post-mortem extracted deposits identified SAA as the major protein source. The duration of stay in the shelters was positively associated with a histological score of AA-amyloidosis (B = 0.026, CI95% = 0.007-0.046; p = 0.010). AA-amyloidosis was very frequent in shelter cats. Presence of SAA fragments in bile secretions raises the possibility of fecal-oral transmission of the disease. In conclusion, AA-amyloidosis was very frequent in shelter cats and those staying longer had more deposits. The cat may represent a natural model of AA-amyloidosis.

Is amyloid involved in acute neuroinflammation? A CSF analysis in encephalitis.

INTRODUCTION: Several investigations have argued for a strong relationship between neuroinflammation and amyloid metabolism but it is still unclear whether inflammation exerts a pro-amyloidogenic effect, amplifies the neurotoxic effect of amyloid, or is protective. METHODS: Forty-two patients with acute encephalitis (ENC) and 18 controls underwent an extended cerebrospinal fluid (CSF) panel of inflammatory, amyloid (Aß40, 42, and 38, sAPP-α, sAPP-ß), glial, and neuronal biomarkers. Linear and non-linear correlations between CSF biomarkers were evaluated studying conditional independence relationships. RESULTS: CSF levels of inflammatory cytokines and neuronal/glial markers were higher in ENC compared to controls, whereas the levels of amyloid-related markers did not differ. Inflammatory markers were not associated with amyloid markers but exhibited a correlation with glial and neuronal markers in conditional independence analysis. DISCUSSION: By an extensive CSF biomarkers analysis, this study showed that an acute neuroinflammation state, which is associated with glial activation and neuronal damage, does not influence amyloid homeostasis.

Degree of immunoglobulin kappa light chain glycosylation of anti-spike SARS CoV-2 antibodies correlates with COVID-19 severity. (preprint)

Glycosylation of antibodies and the effects this has on inflammatory responses has concentrated predominately on the study of glycosylation moieties found in the Fc region of heavy chains. Light chain glycosylation and their ratios are relatively understudied. Nevertheless, variable glycosylation and ratio of {kappa} and {lambda} light chains have been associated with worse prognosis in myeloma and in tissue deposition amyloidosis. The {kappa} {lambda} light chains, of antibodies binding to SARS-CoV2 nucleocapsid and spike protein were analysed, using MALDI-ToF MS, in respect to their intensity, ratios, glycosylation patterns and any pattern changes correlating with COVID-19 severity. The molecular masses and signal intensity of {kappa} and {lambda} glycosylated and non-glycosylated light chains were measured for immunoglobulins isolated from plasma of seropositive and seronegative health care workers (HCW), and convalescent patients who had suffered from acute respiratory distress syndrome (ARDS). Overall, there was no significant changes in {kappa} to {lambda} ratio of total IgG (via protein G capture) antibodies between the groups. A non-statistically significant trend towards {lambda} light chains was found in antibodies against SARS CoV-2 Nucleocapsid and Spike proteins. However, detailed analysis of the molecular forms found a significant increase and bias towards un-glycosylated light chains and in particular un-glycosylated {kappa} light chains, in antibodies against SAR-CoV-2 spike protein, from convalescent COVID-ARDS patients. Here we have demonstrated a bias towards un-glycosylated {kappa} chains in anti-spike antibodies in those who suffered from ARDS as a result of SARS-CoV2 infection 3 months after recovery. How this relates to the immunopathology of COVID-19 requires further study.

Herpesviral encephalitis associated with bortezomib use in a patient with multiple myeloma and associated light-chain amyloidosis.

INTRODUCTION: Bortezomib is proteasome inhibitor used in multiple myeloma treatment. The reactivation of herpes simplex virus (HSV) and varicella-zoster virus (VZV) during bortezomib-based therapy is a well-known adverse event. Antiviral prophylaxis is mandatory. Nevertheless, reports of herpesviral encephalitis are scarce. CASE REPORT: A 57-year-old multiple myeloma patient who during CyBorD protocol (Bortezomib, cyclophosphamide, and dexamethasone), after a transient suspension of antiviral prophylaxis presented progressive headaches unresponsive to conventional analgesics, asthenia, fever, episodic visual hallucinations, and vesicular lesions in the right supraorbital and frontal region. Herpetic encephalitis was diagnosed after detecting herpes zoster in cerebrospinal fluid. MANAGEMENT & OUTCOME: The patient was treated with acyclovir 500mg every 6 hours for 21 days, and subsequent valacyclovir prophylaxis achieving an excellent clinical evolution. Anti-myeloma treatment was changed to lenalidomide and dexamethasone achieving a durable complete response. Herpesviral encephalitis is a rare but severe complication associated with the use of Bortezomib, especially when patients did not receive acyclovir prophylaxis. However, a rapid detection based on the clinical suspicion, and the prompt start of treatment, may lead to overcome this adverse event.

Adequate Antibody Response to COVID-19 Vaccine in Patients with Monoclonal Gammopathies and Light Chain Amyloidosis.

OBJECTIVE: Determine the COVID-19 seroconversion rate for patients with multiple myeloma receiving a COVID-19 vaccine. MATERIALS AND METHODS: After 45 patients received their second COVID-19 vaccine dose, their serum IgG antibodies were measured: 22 with monoclonal gammopathy (MG) of unknown significance, 3 with smoldering myeloma, 2 with light chain amyloidosis, and 18 with MG (9 in remission, 6 out of remission, and 3 with free light-chain gammopathy alone). A second serum specimen was retained for 16 patients with MG. Their antibody levels were compared to those of 78 uninfected healthy vaccinated control patients. RESULTS: Three patients with MG had low antibody levels on blood collected 98, 100, and 113 days after the initial vaccine dose (2 with MG of unknown significance and 1 with hypogammaglobulemia). The other 40 patients with MG (seroconversion rate 93%) and both patients with amyloidosis produced antibodies. Relative to days after vaccination, patients with MG had lower antibody levels than control patients. CONCLUSION: After receiving a COVID-19 vaccine, most patients with MG produce anti-SARS-CoV-2 antibodies comparable to levels in uninfected vaccinated healthy control patients.

The year in cardiovascular medicine 2021: imaging.

This article reviews the most relevant literature published in 2021 on the role of cardiovascular imaging in cardiovascular medicine. Coronavirus disease 2019 (COVID-19) continued to impact the healthcare landscape, resulting in reduced access to hospital-based cardiovascular care including reduced routine diagnostic cardiovascular testing. However, imaging has also facilitated the understanding of the presence and extent of myocardial damage caused by the coronavirus infection. What has dominated the imaging literature beyond the pandemic are novel data on valvular heart disease, the increasing use of artificial intelligence (AI) applied to imaging, and the use of advanced imaging modalities in both ischaemic heart disease and cardiac amyloidosis.

Amyloid processing in COVID-19-associated neurological syndromes.

SARS-CoV-2 infection can damage the nervous system with multiple neurological manifestations described. However, there is limited understanding of the mechanisms underlying COVID-19 neurological injury. This is a cross-sectional exploratory prospective biomarker cohort study of 21 patients with COVID-19 neurological syndromes (Guillain-Barre Syndrome [GBS], encephalitis, encephalopathy, acute disseminated encephalomyelitis [ADEM], intracranial hypertension, and central pain syndrome) and 23 healthy COVID-19 negative controls. We measured cerebrospinal fluid (CSF) and serum biomarkers of amyloid processing, neuronal injury (neurofilament light), astrocyte activation (GFAp), and neuroinflammation (tissue necrosis factor [TNF] ɑ, interleukin [IL]-6, IL-1ß, IL-8). Patients with COVID-19 neurological syndromes had significantly reduced CSF soluble amyloid precursor protein (sAPP)-ɑ (p = 0.004) and sAPPß (p = 0.03) as well as amyloid ß (Aß) 40 (p = 5.2 × 10-8 ), Aß42 (p = 3.5 × 10-7 ), and Aß42/Aß40 ratio (p = 0.005) compared to controls. Patients with COVID-19 neurological syndromes showed significantly increased neurofilament light (NfL, p = 0.001) and this negatively correlated with sAPPɑ and sAPPß. Conversely, GFAp was significantly reduced in COVID-19 neurological syndromes (p = 0.0001) and this positively correlated with sAPPɑ and sAPPß. COVID-19 neurological patients also displayed significantly increased CSF proinflammatory cytokines and these negatively correlated with sAPPɑ and sAPPß. A sensitivity analysis of COVID-19-associated GBS revealed a non-significant trend toward greater impairment of amyloid processing in COVID-19 central than peripheral neurological syndromes. This pilot study raises the possibility that patients with COVID-19-associated neurological syndromes exhibit impaired amyloid processing. Altered amyloid processing was linked to neuronal injury and neuroinflammation but reduced astrocyte activation.

COVID-19 and Light Chain Amyloidosis, Adding Insult to Injury.

Light chain (AL) amyloidosis is a potentially fatal disease of monoclonal plasma cells that leads to accumulation of light chain amyloid fibrils, organ damage, and the manifestations of clinical disease. Meanwhile, coronavirus disease 2019 (COVID-19) is a disease caused by infection with the severe acute respiratory syndrome coronavirus 2 virus, with the potential to cause severe systemic illness and death. There is significant overlap in the demographics and comorbidities observed in AL amyloidosis and those associated with highest risk for severe morbidity and mortality due to COVID-19. This overlap creates unique challenges in caring for patients with AL amyloidosis, which are further compounded by the immunosuppressive nature of anti-plasma cell therapies, the need for frequent clinical assessments, and the exclusion of AL amyloidosis patients from initial COVID-19 vaccine trials. Herein, we highlight many of the relevant concerns related to COVID-19 and the treatment of AL amyloidosis, summarize a general approach for AL amyloidosis management amidst the ongoing COVID-19 pandemic, and discuss current guidance about COVID-19 vaccination of patients with AL amyloidosis.

Is amyloid fibrillation related to 3D domain swapping for the C-terminal domain of SARS-CoV main protease?

The C-terminal domain of SARS-CoV main protease (Mpro-C) can form 3D domain-swapped dimer by exchanging the α1-helices fully buried inside the protein hydrophobic core, under non-denaturing conditions. Here, we report that Mpro-C can also form amyloid fibrils under the 3D domain-swappable conditions in vitro, and the fibrils are not formed through runaway/propagated domain swapping. It is found that there are positive correlations between the rates of domain swapping dimerization and amyloid fibrillation at different temperatures, and for different mutants. However, some Mpro-C mutants incapable of 3D domain swapping can still form amyloid fibrils, indicating that 3D domain swapping is not essential for amyloid fibrillation. Furthermore, NMR H/D exchange data and molecular dynamics simulation results suggest that the protofibril core region tends to unpack at the early stage of 3D domain swapping, so that the amyloid fibrillation can proceed during the 3D domain swapping process. We propose that 3D domain swapping makes it possible for the unpacking of the amyloidogenic fragment of the protein and thus accelerates the amyloid fibrillation process kinetically, which explains the well-documented correlations between amyloid fibrillation and 3D domain swapping observed in many proteins.

Anaesthetic challenges in the spine surgery of a young Asian man with lumbar amyloidosis.

Primary amyloidosis is a rare systemic disorder often associated with multiple organ dysfunction. The most common form, light chain amyloidosis, has an estimated age-adjusted incidence of 5.1-12.8 cases per million person-years. Spine involvement is extremely uncommon. We present the case of a young Asian man with newly diagnosed amyloidosis involving the lumbar spine among multiple organs with a pathological vertebral fracture that required urgent spine surgery. We believe this is the first reported case to discuss the perianaesthetic challenges in the management of lumbar spine amyloidosis.

Presence of a SARS-CoV-2 Protein Enhances Amyloid Formation of Serum Amyloid A.

A marker for the severeness and disease progress of COVID-19 is overexpression of serum amyloid A (SAA) to levels that in other diseases are associated with a risk for SAA amyloidosis. To understand whether SAA amyloidosis could also be a long-term risk of SARS-CoV-2 infections, we have used long all-atom molecular dynamic simulations to study the effect of a SARS-CoV-2 protein segment on SAA amyloid formation. Sampling over 40 µs, we find that the presence of the nine-residue segment SK9, located at the C-terminus of the envelope protein, increases the propensity for SAA fibril formation by three mechanisms: it reduces the stability of the lipid-transporting hexamer shifting the equilibrium toward monomers, it increases the frequency of aggregation-prone configurations in the resulting chains, and it raises the stability of SAA fibrils. Our results therefore suggest that SAA amyloidosis and related pathologies may be a long-term risk of SARS-CoV-2 infections.

Transthyretin: From Structural Stability to Osteoarticular and Cardiovascular Diseases.

Transthyretin (TTR) is a tetrameric protein transporting hormones in the plasma and brain, which has many other activities that have not been fully acknowledged. TTR is a positive indicator of nutrition status and is negatively correlated with inflammation. TTR is a neuroprotective and oxidative-stress-suppressing factor. The TTR structure is destabilized by mutations, oxidative modifications, aging, proteolysis, and metal cations, including Ca2+. Destabilized TTR molecules form amyloid deposits, resulting in senile and familial amyloidopathies. This review links structural stability of TTR with the environmental factors, particularly oxidative stress and Ca2+, and the processes involved in the pathogenesis of TTR-related diseases. The roles of TTR in biomineralization, calcification, and osteoarticular and cardiovascular diseases are broadly discussed. The association of TTR-related diseases and vascular and ligament tissue calcification with TTR levels and TTR structure is presented. It is indicated that unaggregated TTR and TTR amyloid are bound by vicious cycles, and that TTR may have an as yet undetermined role(s) at the crossroads of calcification, blood coagulation, and immune response.

The effect of SARS-COV-2 Infections on Amyloid Formation of Serum Amyloid A (preprint)

A marker for the severeness and disease progress of COVID-19 is overexpression of serum amyloid A (SAA) to levels that in other diseases are associated with a risk for SAA amyloidosis. This secondary illness is characterized by formation and deposition of SAA amyloids in blood vessels, causing inflammation, thrombosis and sometimes organ failure, with symptoms resembling the multisystem inflammatory syndrome (MIS) observed in some COVID-19 survivors. Hence, in order to understand better the danger of SAA amyloidosis in the context of COVID-19 we have used molecular dynamic simulations to study the effect of a SARS-COV-2 protein segment on SAA amyloid formation. We find that presence of the nine-residue segment SK9, located on the Envelope protein, increases the propensity for SAA fibril formation by three mechanisms: it reduces the stability of the lipid-transporting hexamer shifting the equilibrium toward monomers, it increases the frequency of aggregation-prone configurations in the resulting chains, and it raises the stability of SAA fibrils. Our results therefore suggest that SAA amyloidosis -related pathologies are a long-term risk of SARS-COV-2 infections.

Stress Response Is the Main Trigger of Sporadic Amyloidoses.

Amyloidoses are a group of diseases associated with the formation of pathological protein fibrils with cross-ß structures. Approximately 5-10% of the cases of these diseases are determined by amyloidogenic mutations, as well as by transmission of infectious amyloids (prions) between organisms. The most common group of so-called sporadic amyloidoses is associated with abnormal aggregation of wild-type proteins. Some sporadic amyloidoses are known to be induced only against the background of certain pathologies, but in some cases the cause of amyloidosis is unclear. It is assumed that these diseases often occur by accident. Here we present facts and hypotheses about the association of sporadic amyloidoses with vascular pathologies, trauma, oxidative stress, cancer, metabolic diseases, chronic infections and COVID-19. Generalization of current data shows that all sporadic amyloidoses can be regarded as a secondary event occurring against the background of diseases provoking a cellular stress response. Various factors causing the stress response provoke protein overproduction, a local increase in the concentration or modifications, which contributes to amyloidogenesis. Progress in the treatment of vascular, metabolic and infectious diseases, as well as cancers, should lead to a significant reduction in the risk of sporadic amyloidoses.