Biology subjects

Guillen, G.

Publications and source records attributed to Guillen, G..

6 recordsLinked to original sources

bioRxiv2025

Immunological evaluation in African Green Monkeys of two nasal vaccine candidates to induce a broad immunity against coronaviruses

The COVID-19 pandemic has had a profound impact on the world. While the disease is currently under control, the emergence of new coronavirus variants with pandemic potential underscores the need for more universal coronavirus vaccines. This study evaluates the immunogenicity in non-human primates of two broad-spectrum nasal vaccine candidates called PanCoV. The vaccine preparations are based on highly conserved regions of SARS-CoV-2, specifically the nucleocapsid (N) and the S2 subunit from Spike protein, which include several relevant T-cell epitopes and have the potential to form virus-like particles. PanCoV1 candidate contains the N protein, while the PanCoV2 comprises a chimeric protein containing the C-terminal domain of N protein fused to a fragment of S2 subunit. Both vaccine candidates also include the receptor-binding domain (RBD) from Spike protein and the ODN-39M as CpG adjuvant. The results demonstrate that both nasal vaccine candidates can boost the anti-RBD immune response and induce anti-N immunity, this systemic response shows cross-reactivity with SARS-CoV, MERS-CoV, and H-CoV antigens. In addition, a neutralizing response against SARS-CoV-2 that persisted for almost three months after the last dose was induced. A mucosal-specific and cross-reactive IgA response was also observed, mainly in monkeys vaccinated with PanCoV2. Furthermore, the specific IFN{gamma}-secreting response observed in the vaccinated groups peripheral blood mononuclear cells highlights the ability of these candidates to stimulate cellular immunity with a Th1 bias. PanCoV vaccines emerge as promising candidates for use as a booster alternative, with the potential to amplify and broaden the scope of the immune response against coronaviruses.

immunology↗

bioRxiv2025

A nasal vaccine candidate, based on S2 and N proteins from SARS-CoV-2, generates a broad antibody response systemic and at lower respiratory tract.

Since the beginning of the COVID-19 pandemic various groups around the world have intensively worked in the development of vaccine candidates against SARS-CoV-2. Several vaccines have been approved in the past years, the majority are based on the Spike or RBD proteins and employs parenteral administration routes. Considering the recent history of Coronavirus zoonotic events, causing serious human health problems, the generation of vaccines with a broad scope of protection and the potential to cut / reduce the transmission remains in the spotlight. The current global pandemic preparedness initiatives have promoted also the preclinical evaluation of a new group of Coronavirus vaccines. In the present work a nasal vaccine candidate based on two highly conserved Sarbecovirus proteins, S2 and nucleocapsid (N), is evaluated in two different mice strains. The vaccine preparation, containing a CpG ODN as adjuvant, was able to generate high antibody titers against both antigens, in sera and bronchoalveolar lavages. This antibody response results cross-reactive to S2 from SARS-CoV-1 and MERS-CoV, and to N from SARS-CoV-1. However, a very low neutralizing capacity was found in the sera of the immunized mice when a pseudoviral system assay was used. On the other hand, the vaccine preparation induces, at systemic compartment, IFN {gamma} secretion, and a marked IgG2a response, specific against both proteins; a profile consistent with the development of a Th1 pattern. Although further evaluations should be done, including protection assays, the demonstrated cross-reactivity level and mucosal response constitutes promising features of this vaccine candidate.

immunology↗

bioRxiv2024

Combined inhibition of MDM2 and PARP lead to a synergistic anti-tumoral response in p53 wild-type rhabdomyosarcoma models

Targeting MDM2-p53 interaction to enhance p53 activity represents a promising antitumoral strategy for p53 wild-type pediatric tumors, including soft-tissue sarcomas. However, results from early-phase clinical trials in hematological and solid cancers have shown limited efficacy of MDM2 inhibitors, underscoring the necessity to explore more effective combinations with other agents. In this study, we provide results from a 22-drug combination screening demonstrating the therapeutic potential of combining siremadlin (MDM2 inhibitor) and olaparib (PARP inhibitor) for treating p53 wild-type rhabdomyosarcoma (p53 WT RMS). Cell survival, cell death, and apoptosis analysis revealed synergistic effects of combining siremadlin plus olaparib in vitro. The combination of both drugs led to a significant increase in p53 activity, as evidenced by p53 accumulation and K382 acetylation, along with an increased expression of bona fide p53 targets. Furthermore, treatment with both drugs resulted in significant reduction of tumor growth and increased mice overall survival when compared to single treatments and control in cell line-derived orthotopic xenograft models. Overall, our study demonstrates, for the first time, the synergistic effect of combining siremadlin plus olaparib in inhibiting p53 WT RMS tumor growth in vitro and in vivo. Our findings support the potential to study the combination of both drugs in clinical trials and warrants further investigation in other tumors with similar molecular features.

cancer biology↗

bioRxiv2022

The Nucleocapsid Protein Of SARS-CoV-2, Combined With ODN-39M, Is A Potential Component For An Intranasal Bivalent Pancorona Vaccine

Despite the rapid development of vaccines and their reported efficacy for controlling the COVID-19 waves, two key challenges remain: the scope of the immunity against upcoming variants and zoonosis events, and the induction of mucosal immunity able to clear the virus in the upper respiratory tract for halting the transmission. The present study is aiming at assessing a potential component for a new generation of vaccines so as to overcome such limitations. The recombinant nucleocapsid (N) protein from SARS-CoV-2 Delta variant was combined with a phosphodiester backbone CpG ODN (ODN-39M), forming high molecular weight aggregates. The evaluation of its immunogenicity in Balb/C mice revealed that only administration by intranasal route induced a systemic cross-reactive Cell-Mediated-Immunity (CMI). In turn, this combination was able to induce anti-N IgA in lungs, which along with the specific IgG in sera and CMI in spleen, resulted cross-reactive against the nucleocapsid protein of SARS-CoV-1. Furthermore, the nasal administration of the N+ODN-39M preparation combined with the RBD Delta protein, as inductor of neutralizing Abs, enhanced the local and systemic immune response against RBD with a modulation toward a Th1 pattern. Taken together, these results make the N+ODN-39M preparation a suitable component for a future intranasal pancorona vaccine against Sarbecoviruses. Particularly, the bivalent vaccine formulation N+ODN-39M+RBD could be used as an effective nasal booster in previously vaccinated population.

immunology↗

bioRxiv2021

In-solution buffer-free digestion for the analysis of SARS-CoV-2 RBD proteins allows a full sequence coverage and detection of post-translational modifications in a single ESI-MS spectrum

Subunit vaccines based on the receptor-binding domain (RBD) of the spike protein of SARS-CoV-2, are among the most promising strategies to fight the COVID-19 pandemic. The detailed characterization of the protein primary structure by mass spectrometry (MS) is mandatory, as described in ICHQ6B guidelines. In this work, several recombinant RBD proteins produced in five expression systems were characterized using a non-conventional protocol known as in-solution buffer-free digestion (BFD). In a single ESI-MS spectrum, BFD allowed very high sequence coverage ([≥] 99 %) and the detection of highly hydrophilic regions, including very short and hydrophilic peptides (2-8 amino acids), the His6-tagged C-terminal peptide carrying several post-translational modifications at Cys538 such as cysteinylation, glutathionylation, cyanilation, among others. The analysis using the conventional digestion protocol allowed lower sequence coverage (80-90 %) and did not detect peptides carrying some of the above-mentioned post-translational modifications. The two C-terminal peptides of a dimer [RBD(319-541)-(His)6]2 linked by an intermolecular disulfide bond (Cys538-Cys538) with twelve histidine residues were only detected by BFD. This protocol allows the detection of the four disulfide bonds present in the native RBD and the low-abundance scrambling variants, free cysteine residues, O-glycoforms and incomplete processing of the N-terminal end, if present. Artifacts that might be generated by the in-solution BFD protocol were also characterized. BFD can be easily implemented and we foresee that it can be also helpful to the characterization of mutated RBD.

biochemistry↗

bioRxiv2020

Sequential combinations of chemotherapeutic agents with BH3 mimetics to treat rhabdomyosarcoma and avoid resistance.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in childhood and adolescence. Refractory/relapsed RMS patients present a bad prognosis that combined with the lack of specific biomarkers difficult the development of new therapies. We here utilize dynamic BH3 Profiling (DBP), a functional predictive biomarker that measures net changes in mitochondrial apoptotic signaling, to identify anti-apoptotic adaptations upon treatment. We use this information to guide the use of BH3 mimetics to specifically inhibit BCL-2 pro-survival proteins, defeat resistance and avoid relapse to therapy. Indeed, we found that BH3 mimetics that selectively target BCL-xL and MCL-1 synergistically enhance the effect of the clinically used chemotherapeutic agents vincristine and doxorubicin in RMS cells. We validated this strategy in vivo using a RMS patient-derived xenograft (PDX) model and observed a reduction on tumor growth with a tendency to its stabilization with the sequential combination of vincristine and the MCL-1 inhibitor S63845. Finally, we identified the molecular mechanism by which RMS cells acquire resistance to vincristine: through the anti-apoptotic protein MCL-1 for which we observed an enhanced binding between MCL-1 and BID after drug exposure, which is suppressed by the sequential addition of S63845. In conclusion, our findings validate the use of DBP as a functional assay to predict treatment effectiveness in RMS and provide a rationale for BH3 mimetic combination with chemotherapeutic agents to avoid tumor resistance, improve treatment efficiency and decrease undesired secondary effects.

cancer biology↗