recombineering-based-manufacturing-engineered-viral-vectors-for

Recombineering-based Manufacturing of Engineered Viral Vectors for Research and Therapy

The increasing demand for engineered viral vectors in both basic and translational research has underscored the need for flexible, rapid, and scalable ...

CEINGE Biotecnologie Avanzate Franco Salvatore S.C.aR.L.

University of Naples Federico II

Sensitive detection of altered proteins expression in plasma membranes is of fundamental importance, for both diagnostic and prognostic purposes. Surface-Enhanced Raman Scattering (SERS) has proven to be a quite sensitive approach to detect proteins, even in very diluted samples. However, proteins detection in complex environment, such as the cellular membrane, is still a challenge. Herein, we demonstrate a SERS-based platform to reveal the overexpression of target proteins in cell membranes. As a proof of concept, we implemented ectopic expression of carbonic anhydrase IX (CA IX) and epidermal growth factor receptor (EGFR) in the plasma membrane of the SKOV3 tumor cell line. Our outcomes demonstrate that SERS signals from cells put in contact with a hyperuniform SERS substrate allow highlighting subtle differences in the biochemical composition of cell membranes, normally hidden in spontaneous Raman confocal microscopy. This opens new opportunities for a label-free membrane analysis and bio-sensing in a broader sense.

Revealing membrane alteration in cellsoverexpressing CA IX and EGFR by Surface-Enhanced Raman Scattering.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Publisher Correction: Revealing membrane alteration in cells overexpressing CA IX and EGFR by Surface-Enhanced Raman Scattering.

Different innate immune pathways converge to Stimulator of interferon genes (STING) and trigger type I interferon responses after recognition of abnormal nucleic acids in the cells. This non-redundant function renders STING a major player in immunosurveillance, and an emerging target for cancer and infectious diseases therapeutics. Beyond somatic mutations that often occur in cancer, the human gene encoding STING protein, TMEM173 (STING1), holds great genetic heterogeneity; R232, HAQ (R71H-G230A-R293Q) and H232 are the most common alleles. Although some of these alleles are likely to be hypomorphic, their function is still debated, due to the available functional assessments, which have been performed in biased biological systems. Here, by using genetic background-matched models, we report on the functional evaluation of R232, HAQ and H232 variants on STING function, and on how these genotypes affect the susceptibility to clinically relevant viruses, thus supporting a potential contributing cause to differences in inter-individual responses to infections. Our findings also demonstrate a novel toll-like receptor-independent role of STING in modulating monocytic cell function and differentiation into macrophages. We further supported the interplay of STING1 variants and human biology by demonstrating how monocytes bearing the H232 allele were impaired in M1/M2 differentiation, interferon response and antigen presentation. Finally, we assessed the response to PD-1 inhibitor in a small cohort of melanoma patients stratified according to STING genotype. Given the contribution of the STING protein in sensing DNA viruses, bacterial pathogens and misplaced cancer DNA, these data may support the development of novel therapeutic options for infectious diseases and cancer.

The common H232 STING allele shows impaired activities in DNA sensing, susceptibility to viral infection, and in monocyte cell function, while the HAQ variant possesses wild-type properties.

Carbonic anhydrases constitute a family of metalloenzymes vital for maintaining acid-base balance and regulating pH in physio-pathological processes. These findings suggest carbonic anhydrases as potential therapeutic targets for treating pH-associated disorders, including cerebral ischemia, to mitigate hypoxia- and reoxygenation-induced neuronal damage. A focus on carbonic anhydrase IX showed that ischemic stress altered subcellular distributions of this enzyme in rodent neuronal populations. Given the enzyme's canonical membrane localization, we implemented pharmacological inhibition using a membrane-impermeant sulfonamide inhibitor in neuronal models of brain ischemia. The treatments exerted neuroprotective effects on neurons from  knockout mice. Moreover, administration of the sulfonamide inhibitor to rats subjected to transient middle cerebral artery occlusion decreased infarct volumes and improved neurological deficits. Our results support the involvement of carbonic anhydrase IX in postischemic damage and pave the way for possible pharmacological interventions with selective inhibitors in the management of brain ischemia.

Pharmacological inhibition of carbonic anhydrases with a positively charged pyridinium sulfonamide phenocopies the neuroprotective effects of  genetic ablation in a murine setting of oxygen/glucose deprivation followed by re-oxygenation and is associated with improved neuronal function in ischemic rats.

Article	Year
Revealing membrane alteration in cellsoverexpressing CA IX and EGFR by Surface-Enhanced Raman Scattering. Scientific reports\| PubMed ID: 30755643	2019
Publisher Correction: Revealing membrane alteration in cells overexpressing CA IX and EGFR by Surface-Enhanced Raman Scattering. Scientific reports\| PubMed ID: 31209245	2019
The common H232 STING allele shows impaired activities in DNA sensing, susceptibility to viral infection, and in monocyte cell function, while the HAQ variant possesses wild-type properties. Scientific reports\| PubMed ID: 37945588	2023
Pharmacological inhibition of carbonic anhydrases with a positively charged pyridinium sulfonamide phenocopies the neuroprotective effects of genetic ablation in a murine setting of oxygen/... Heliyon\| PubMed ID: 40028587	2025

Nicola Zambrano

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