RESUMEN
The deficiency of CITRIN, the liver mitochondrial aspartate-glutamate carrier (AGC), is the cause of four human clinical phenotypes, neonatal intrahepatic cholestasis caused by CITRIN deficiency (NICCD), silent period, failure to thrive and dyslipidemia caused by CITRIN deficiency (FTTDCD), and citrullinemia type II (CTLN2). Clinical symptoms can be traced back to disruption of the malate-aspartate shuttle due to the lack of citrin. A potential therapy for this condition is the expression of aralar, the AGC present in brain, to replace citrin. To explore this possibility we have first verified that the NADH/NAD+ ratio increases in hepatocytes from citrin(-/-) mice, and then found that exogenous aralar expression reversed the increase in NADH/NAD+ observed in these cells. Liver mitochondria from citrin (-/-) mice expressing liver specific transgenic aralar had a small (~ 4-6 nmoles x mg prot-1 x min-1) but consistent increase in malate aspartate shuttle (MAS) activity over that of citrin(-/-) mice. These results support the functional replacement between AGCs in the liver. To explore the significance of AGC replacement in human therapy we studied the relative levels of citrin and aralar in mouse and human liver through absolute quantification proteomics. We report that mouse liver has relatively high aralar levels (citrin/aralar molar ratio of 7.8), whereas human liver is virtually devoid of aralar (CITRIN/ARALAR ratio of 397). This large difference in endogenous aralar levels partly explains the high residual MAS activity in liver of citrin(-/-) mice and why they fail to recapitulate the human disease, but supports the benefit of increasing aralar expression to improve the redox balance capacity of human liver, as an effective therapy for CITRIN deficiency.
RESUMEN
Background: The COVID-19 vaccine candidate CVnCoV comprises sequence-optimized mRNA encoding SARS-CoV-2 S-protein encapsulated in lipid nanoparticles. In this phase 2a study, we assessed reactogenicity and immunogenicity of two or three doses in younger and older adults. Methods: Younger (18-60 years) and older (>60 years) adults were enrolled in two sites in Panama and Peru to receive either 6 or 12 µg doses of CVnCoV or licensed control vaccines 28 days apart; subsets received a 12 µg booster dose on Day 57 or Day 180. Solicited adverse events (AE) were reported for 7 days and unsolicited AEs for 4 weeks after each vaccination, and serious AEs (SAE) throughout the study. Humoral immunogenicity was measured as neutralizing and receptor binding domain (RBD) IgG antibodies and cellular immunogenicity was assessed as CD4+/CD8 + T cell responses. Results: A total of 668 participants were vaccinated (332 aged 18-60 years and 336 aged > 60 years) including 75 who received homologous booster doses. Vaccination was well tolerated with no vaccine-related SAEs. Solicited and unsolicited AEs were mainly mild to moderate and resolved spontaneously. Both age groups demonstrated robust immune responses as neutralizing antibodies or RBD-binding IgG, after two doses, with lower titers in the older age group than the younger adults. Neither group achieved levels observed in human convalescent sera (HCS), but did equal or surpass HCS levels following homologous booster doses. Following CVnCoV vaccination, robust SARS-CoV-2 S-protein-specific CD4 + T-cell responses were observed in both age groups with CD8 + T-cell responses in some individuals, consistent with observations in convalescing COVID-19 patients after natural infection. Conclusions: We confirmed that two 12 µg doses of CVnCoV had an acceptable safety profile, and induced robust immune responses. Marked humoral immune responses to homologous boosters suggest two doses had induced immune memory.
RESUMEN
Gene therapy is becoming an increasingly valuable tool to treat many genetic diseases with no or limited treatment options. This is the case for hundreds of monogenic metabolic disorders of hepatic origin, for which liver transplantation remains the only cure. Furthermore, the liver contains 10-15% of the body's total blood volume, making it ideal for use as a factory to secrete proteins into the circulation. In recent decades, an expanding toolbox has become available for liver-directed gene delivery. Although viral vectors have long been the preferred approach to target hepatocytes, an increasing number of non-viral vectors are emerging as highly efficient vehicles for the delivery of genetic material. Herein, we review advances in gene delivery vectors targeting the liver and more specifically hepatocytes, covering strategies based on gene addition and gene editing, as well as the exciting results obtained with the use of RNA as a therapeutic molecule. Moreover, we will briefly summarise some of the limitations of current liver-directed gene therapy approaches and potential ways of overcoming them.