With complements: C3 inhibition in the clinic.

C3 is a key complement protein, located at the nexus of all complement activation pathways. Extracellular, tissue, cell-derived, and intracellular C3 plays critical roles in the immune response that is dysregulated in many diseases, making it an attractive therapeutic target. However, challenges such as very high concentration in blood, increased acute expression, and the elevated risk of infections have historically posed significant challenges in the development of C3-targeted therapeutics. This is further complicated because C3 activation fragments and their receptors trigger a complex network of downstream effects; therefore, a clear understanding of these is needed to provide context for a better understanding of the mechanism of action (MoA) of C3 inhibitors, such as pegcetacoplan. Because of C3's differential upstream position to C5 in the complement cascade, there are mechanistic differences between pegcetacoplan and eculizumab that determine their efficacy in patients with paroxysmal nocturnal hemoglobinuria. In this review, we compare the MoA of pegcetacoplan and eculizumab in paroxysmal nocturnal hemoglobinuria and discuss the complement-mediated disease that might be amenable to C3 inhibition. We further discuss the current state and outlook for C3-targeted therapeutics and provide our perspective on which diseases might be the next success stories in the C3 therapeutics journey.

Infection by the Helminth Parasite Fasciola hepatica Requires Rapid Regulation of Metabolic, Virulence, and Invasive Factors to Adjust to Its Mammalian Host.

The parasite Fasciola hepatica infects a broad range of mammals with impunity. Following ingestion of parasites (metacercariae) by the host, newly excysted juveniles (NEJ) emerge from their cysts, rapidly penetrate the duodenal wall and migrate to the liver. Successful infection takes just a few hours and involves negotiating hurdles presented by host macromolecules, tissues and micro-environments, as well as the immune system. Here, transcriptome and proteome analysis of ex vivo F. hepatica metacercariae and NEJ reveal the rapidity and multitude of metabolic and developmental alterations that take place in order for the parasite to establish infection. We found that metacercariae despite being encased in a cyst are metabolically active, and primed for infection. Following excystment, NEJ expend vital energy stores and rapidly adjust their metabolic pathways to cope with their new and increasingly anaerobic environment. Temperature increases induce neoblast proliferation and the remarkable up-regulation of genes associated with growth and development. Cysteine proteases synthesized by gastrodermal cells are secreted to facilitate invasion and tissue degradation, and tegumental transporters, such as aquaporins, are varied to deal with osmotic/salinity changes. Major proteins of the total NEJ secretome include proteases, protease inhibitors and anti-oxidants, and an array of immunomodulators that likely disarm host innate immune effector cells. Thus, the challenges of infection by F. hepatica parasites are met by rapid metabolic and physiological adjustments that expedite tissue invasion and immune evasion; these changes facilitate parasite growth, development and maturation. Our molecular analysis of the critical processes involved in host invasion has identified key targets for future drug and vaccine strategies directed at preventing parasite infection.

The Zoonotic Helminth Parasite Fasciola hepatica: Virulence-Associated Cathepsin B and Cathepsin L Cysteine Peptidases Secreted by Infective Newly Excysted Juveniles (NEJ).

Fasciolosis caused by Fasciola hepatica is a major global disease of livestock and an important neglected helminthiasis of humans. Infection arises when encysted metacercariae are ingested by the mammalian host. Within the intestine, the parasite excysts as a newly excysted juvenile (NEJ) that penetrates the intestinal wall and migrates to the liver. NEJ excystment and tissue penetration are facilitated by the secretion of cysteine peptidases, namely, cathepsin B1 (FhCB1), cathepsin B2 (FhCB2), cathepsin B3 (FhCB3) and cathepsin L3 (FhCL3). While our knowledge of these peptidases is growing, we have yet to understand why multiple enzymes are required for parasite invasion. Here, we produced functional recombinant forms of these four peptidases and compared their physio-biochemical characteristics. Our studies show great variation of their pH optima for activity, substrate specificity and inhibitory profile. Carboxy-dipeptidase activity was exhibited exclusively by FhCB1. Our studies suggest that, combined, these peptidases create a powerful hydrolytic cocktail capable of digesting the various host tissues, cells and macromolecules. Although we found several inhibitors of these enzymes, they did not show potent inhibition of metacercarial excystment or NEJ viability in vitro. However, this does not exclude these peptidases as targets for future drug or vaccine development.

Quantitative fluorescein angiographic analysis of choroidal neovascular membranes: validation and correlation with visual function.

PURPOSE: To compare computerized analysis with traditional grading methods in the analysis of fluorescein angiograms from patients with choroidal neovascularization (CNV) due to age-related macular degeneration (AMD) and to examine the clinical relevance of parameters generated by computerized analysis by testing their relationships with clinical measures of vision. METHODS: Custom quantitative fluorescein analysis (QFA) software was used to analyze 62 angiograms from patients with CNV for whom distance visual acuity (DVA) data were available. On applying QFA, we obtained three mathematical parameters for each lesion component: pixel area (PA), integrated intensity (II), and positive fluorescence (PF). Quotients (Q) were derived for the latter two parameters by correcting against background (b) or optic nerve (o) fluorescence (IIQ(b), IIQ degrees , PFQ(b), and PFQ degrees ). The new metrics were compared with traditional grading parameters of classic CNV and lesion area. The relationships of both sets of angiographic data with measures of vision were explored by regression analyses. RESULTS: Weighted kappa between QFA and traditional grading for lesion subtype assessment was high (kappa = 0.7). Regression analyses with PA, IIQ(b), IIQ degrees , PFQ(b), and PFQ degrees for each lesion descriptor (leakage, classic CNV, occult CNV, total lesion) as independent variables and DVA as the dependent variable showed that in every case PFQ(b) exhibited the most significant relationship with vision (adjusted r(2) = 0.26). Parameter estimates showed that for a change of 30 units on the PFQ(b) for classic CNV, a loss of 20 letters of DVA may be expected. No parameters from traditional grading methods showed statistically significant relationships with DVA. CONCLUSIONS: The markers of dynamic change in area and intensity of fluorescence exhibited stronger relationships with visual function than did area measurements alone.