Zinc-Assisted Microscale Granules Made of the SARS-CoV-2 Spike Protein Trigger Neutralizing, Antivirus Antibody Responses

Protein secretory granules; SARS-CoV-2; AntibodiesGránulos secretores de proteínas; SARS-CoV-2; AnticuerposGrànuls secretors de proteïnes; SARS-CoV-2; AnticossosThe development of new and more efficient vaccination approaches is a constant need, due to the pressure of historical and emerging infectious diseases and the limited efficacy and universality of the current vaccination technologies. Peptides and recombinant proteins have been explored for decades as subunit vaccines for bacterial and viral infections, presented either as soluble protein species or as virus-like assemblies. Recently, synthetic secretory protein-only microscale granules have been developed as dynamic depots for sustained protein release. They are based on the reversible coordination between ionic Zn and histidine residues, which promotes a fast formation of granular particles in vitro out of soluble protein and a slow release of such building block protein in vivo through the physiological chelation of the metal. Such an endocrine-like platform represents a new drug delivery system fully validated in oncology by which soluble and functional protein drugs are progressively released from the granules and made available for antitumor activities upon subcutaneous administration. By exploring such an approach for immunization here, microparticles made of a recombinant form of the receptor binding domain (RBD) of SARS-CoV-2 were tested as an antigen delivery system for induction of antibody responses against the virus upon administration of the material in the absence of added adjuvants. Also, the comparison between protein materials produced in bacterial, mammalian, or insect cell factories has demonstrated a moderate impact of protein glycosylation on the final immunological performance of the system. Therefore, we propose the consideration of synthetic protein secretory granules as a new sustainable immunization platform based on fully manageable, self-organized, and self-formulated immunogens, aimed at reducing the dosage, costs, and complexity of vaccination regimens.The authors appreciate the financial support received from AGAUR (2020PANDE00003 and 2021SGR00092 to A.V.), from CIBER-Consorcio Centro de Investigación Biomédica en Red- (CB06/01/0014), Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación, through intramural projects (NANOSARS to E.P. and NANOREMOTE to E.V.). We also appreciate the support from AEI for the development of multimeric recombinant drugs (PID2019-105416RB-I00/AEI/10.13039/501100011033 and PDC2022-133858-I00 to E.V., PID2019-107298RB-C22 to N.F.-M., PID2020-116174RB-I00 to A.V. and PID2022-1368450 OB-10/AEI/10.13039/501100011033 to A.V. and E.V.). M.T.P.F. received a FAPESP fellowship (No. 2021/08528-0) and L.C.S.F. received a FAPESP research grant (No. 2020/05204-7). A.V. received an ICREA ACADEMIA award

Zinc-Assisted Microscale Granules Made of the SARS-CoV-2 Spike Protein Trigger Neutralizing, Antivirus Antibody Responses

Altres ajuts: acords transformatius de la UABThe development of new and more efficient vaccination approaches is a constant need, due to the pressure of historical and emerging infectious diseases and the limited efficacy and universality of the current vaccination technologies. Peptides and recombinant proteins have been explored for decades as subunit vaccines for bacterial and viral infections, presented either as soluble protein species or as virus-like assemblies. Recently, synthetic secretory protein-only microscale granules have been developed as dynamic depots for sustained protein release. They are based on the reversible coordination between ionic Zn and histidine residues, which promotes a fast formation of granular particles in vitro out of soluble protein and a slow release of such building block protein in vivo through the physiological chelation of the metal. Such an endocrine-like platform represents a new drug delivery system fully validated in oncology by which soluble and functional protein drugs are progressively released from the granules and made available for antitumor activities upon subcutaneous administration. By exploring such an approach for immunization here, microparticles made of a recombinant form of the receptor binding domain (RBD) of SARS-CoV-2 were tested as an antigen delivery system for induction of antibody responses against the virus upon administration of the material in the absence of added adjuvants. Also, the comparison between protein materials produced in bacterial, mammalian, or insect cell factories has demonstrated a moderate impact of protein glycosylation on the final immunological performance of the system. Therefore, we propose the consideration of synthetic protein secretory granules as a new sustainable immunization platform based on fully manageable, self-organized, and self-formulated immunogens, aimed at reducing the dosage, costs, and complexity of vaccination regimens

Recombinant Proteins for Assembling as Nano- and Micro-Scale Materials for Drug Delivery : A Host Comparative Overview

Altres ajuts: CIBER - Consorcio Centro de Investigación Biomédica en Red CB06/01/0014; María Zambrano postdoctoral researcher contract (677904)By following simple protein engineering steps, recombinant proteins with promising applications in the field of drug delivery can be assembled in the form of functional materials of increasing complexity, either as nanoparticles or nanoparticle-leaking secretory microparticles. Among the suitable strategies for protein assembly, the use of histidine-rich tags in combination with coordinating divalent cations allows the construction of both categories of material out of pure polypeptide samples. Such molecular crosslinking results in chemically homogeneous protein particles with a defined composition, a fact that offers soft regulatory routes towards clinical applications for nanostructured protein-only drugs or for protein-based drug vehicles. Successes in the fabrication and final performance of these materials are expected, irrespective of the protein source. However, this fact has not yet been fully explored and confirmed. By taking the antigenic RBD domain of the SARS-CoV-2 spike glycoprotein as a model building block, we investigated the production of nanoparticles and secretory microparticles out of the versions of recombinant RBD produced by bacteria (Escherichia coli), insect cells (Sf9), and two different mammalian cell lines (namely HEK 293F and Expi293F). Although both functional nanoparticles and secretory microparticles were effectively generated in all cases, the technological and biological idiosyncrasy of each type of cell factory impacted the biophysical properties of the products. Therefore, the selection of a protein biofabrication platform is not irrelevant but instead is a significant factor in the upstream pipeline of protein assembly into supramolecular, complex, and functional materials

Recombinant Proteins for Assembling as Nano- and Micro-Scale Materials for Drug Delivery: A Host Comparative Overview

By following simple protein engineering steps, recombinant proteins with promising applications in the field of drug delivery can be assembled in the form of functional materials of increasing complexity, either as nanoparticles or nanoparticle-leaking secretory microparticles. Among the suitable strategies for protein assembly, the use of histidine-rich tags in combination with coordinating divalent cations allows the construction of both categories of material out of pure polypeptide samples. Such molecular crosslinking results in chemically homogeneous protein particles with a defined composition, a fact that offers soft regulatory routes towards clinical applications for nanostructured protein-only drugs or for protein-based drug vehicles. Successes in the fabrication and final performance of these materials are expected, irrespective of the protein source. However, this fact has not yet been fully explored and confirmed. By taking the antigenic RBD domain of the SARS-CoV-2 spike glycoprotein as a model building block, we investigated the production of nanoparticles and secretory microparticles out of the versions of recombinant RBD produced by bacteria (Escherichia coli), insect cells (Sf9), and two different mammalian cell lines (namely HEK 293F and Expi293F). Although both functional nanoparticles and secretory microparticles were effectively generated in all cases, the technological and biological idiosyncrasy of each type of cell factory impacted the biophysical properties of the products. Therefore, the selection of a protein biofabrication platform is not irrelevant but instead is a significant factor in the upstream pipeline of protein assembly into supramolecular, complex, and functional materials

Impact of late presentation of HIV infection on short-, mid- and long-term mortality and causes of death in a multicenter national cohort : 2004-2013

To analyze the impact of late presentation (LP) on overall mortality and causes of death and describe LP trends and risk factors (2004-2013). Cox models and logistic regression were used to analyze data from a nation-wide cohort in Spain. LP is defined as being diagnosed when CD4 < 350 cells/ml or AIDS. Of 7165 new HIV diagnoses, 46.9% (CI:45.7-48.0) were LP, 240 patients died.First-year mortality was the highest (aHR = 10.3[CI:5.5-19.3]); between 1 and 4 years post-diagnosis, aHR = 1.9(1.2-3.0); an

Prediction of long-term outcomes of HIV-infected patients developing non-AIDS events using a multistate approach

Outcomes of people living with HIV (PLWH) developing non-AIDS events (NAEs) remain poorly defined. We aimed to classify NAEs according to severity, and to describe clinical outcomes and prognostic factors after NAE occurrence using data from CoRIS, a large Spanish HIV cohort from 2004 to 2013. Prospective multicenter cohort study. Using a multistate approach we estimated 3 transition probabilities: from alive and NAE-free to alive and NAE-experienced ("NAE development"); from alive and NAE-experienced to death ("Death after NAE"); and from alive and NAE-free to death ("Death without NAE"). We analyzed the effect of different covariates, including demographic, immunologic and virologic data, on death or NAE development, based on estimates of hazard ratios (HR). We focused on the transition "Death after NAE". 8,789 PLWH were followed-up until death, cohort censoring or loss to follow-up. 792 first incident NAEs occurred in 9.01% PLWH (incidence rate 28.76; 95% confidence interval [CI], 26.80-30.84, per 1000 patient-years). 112 (14.14%) NAE-experienced PLWH and 240 (2.73%) NAE-free PLWH died. Adjusted HR for the transition "Death after NAE" was 12.1 (95%CI, 4.90-29.89). There was a graded increase in the adjusted HRs for mortality according to NAE severity category: HR (95%CI), 4.02 (2.45-6.57) for intermediate-severity; and 9.85 (5.45-17.81) for serious NAEs compared to low-severity NAEs. Male sex (HR 2.04; 95% CI, 1.11-3.84), ag