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Adeno-associated viral (AAV) vectors are a leading candidate for the delivery of CRISPR-Cas9 for therapeutic genome editing in vivo. However, AAV-based delivery involves persistent expression of the Cas9 nuclease, a bacterial protein. Recent studies indicate a high prevalence of neutralizing antibodies and T cells specific to the commonly used Cas9 orthologs from Streptococcus pyogenes (SpCas9) and Staphylococcus aureus (SaCas9) in humans. We tested in a mouse model whether pre-existing immunity to SaCas9 would pose a barrier to liver genome editing with AAV packaging CRISPR-Cas9. Although efficient genome editing occurred in mouse liver with pre-existing SaCas9 immunity, this was accompanied by an increased proportion of CD8+ T cells in the liver. This cytotoxic T cell response was characterized by hepatocyte apoptosis, loss of recombinant AAV genomes, and complete elimination of genome-edited cells, and was followed by compensatory liver regeneration. Our results raise important efficacy and safety concerns for CRISPR-Cas9-based in vivo genome editing in the liver., Graphical Abstract, Li and colleagues demonstrated that pre-existing immunity to Cas9 poses a significant barrier to liver-directed genome editing with AAV-CRISPR. They found that AAV expression of Cas9 elicited a robust CD8+ T cell response, resulting in elimination of edited hepatocytes. This raises important safety and efficacy concerns for in vivo editing with CRISPR-Cas9.

Background Atherosclerosis is a slowly progressing, chronic multifactorial disease characterized by the accumulation of lipids, inflammatory cells, and fibrous tissue that drives to the formation of asymmetric focal thickenings in the tunica intima of large and mid-sized arteries. Despite the high therapeutic potential of ApoA-1 proteins, the purification and delivery into the disordered organisms of these drugs is still limited by low efficiency in these processes. Methods and results We report here a novel production and delivery system of anti-atherogenic APOA-1Milano muteins (APOA-1M) by means of genetically modified rice plants. APOA-1M, delivered as protein extracts from transgenic rice seeds, significantly reduced macrophage activation and foam cell formation in vitro in oxLDL-loaded THP-1 model. The APOA-1M delivery method and therapeutic efficacy was tested in healthy mice and in Apoe−/− mice fed with high cholesterol diet (Western Diet, WD). APOA-1M rice milk significantly reduced atherosclerotic plaque size and lipids composition in aortic sinus and aortic arch of WD-fed Apoe−/− mice as compared to wild type rice milk-treated, WD-fed Apoe−/− mice. APOA-1M rice milk also significantly reduced macrophage number in liver of WD-fed Apoe−/− mice as compared to WT rice milk treated mice. Translational impact The delivery of therapeutic APOA-1M full length proteins via oral administration of rice seeds protein extracts (the ‘rice milk’) to the disordered organism, without any need of purification, might overcome the main APOA1-based therapies' limitations and improve the use of this molecules as therapeutic agents for cardiovascular patients.

In the framework of the UA1 upgrading program a hybrid amplifier suitable for amplification of high capacitance detector charges has been developed. The adopted scheme, although very classical [V. Radeka, IEEE Trans. Nucl. Sci. NS-21 (1974) 51], will be discussed and performance presented.

The problem of handling large numbers of analog signals in fast elementary particle detectors has been approached using parallel input to an optical linear array, through a LED matrix. This method results in a much smaller cost per channel than conventional instrumentation. A prototype system is described and the results of some tests are reported. The availability of charge coupled devices (CCD) promises further interesting developments for this technique that will also be discussed.