Biology subjects

Sanchez-Guijo, F.

Publications and source records attributed to Sanchez-Guijo, F..

6 recordsLinked to original sources

bioRxiv2025

Targeting SOS1 synergistically enhances efficacy of BCR/ABL tyrosine kinase inhibitors and overcomes resistance in chronic myeloid leukemia

ABSTRACTDisease persistence and therapeutic resistance remain a significant challenge in chronic myelogenous leukemia (CML). Here, we evaluated the therapeutic impact of SOS1 inhibition by its specific pharmacological inhibitor BI-3406 as single-agent or in combination with BCR/ABL tyrosine kinase inhibitors (TKI) like imatinib in preclinical models of CML including p210BCR/ABL mice, human CML cell lines, and patient-derived bone marrow cells. In p210BCR/ABL mice, treatment with BI-3406 or imatinib was well-tolerated in vivo after single or combined use of the drugs. Treatment with imatinib alone significantly improved survival and corrected various hematological parameters of disease burden, while the combination with BI-3406 therapy yielded even more pronounced benefits, including a substantial increase in median survival, marked reductions in peripheral white blood cell and neutrophil counts, and a notable decrease in leukemia stem cells within the bone marrow. Additionally, the combination led to further spleen size reduction and restoration of normal splenic architecture. Human CML cell lines and primary cells from CML patients subjected to combined treatment with BI-3406 and imatinib or later-generation TKI drugs showed significantly reduced proliferation and enhanced apoptosis as compared to single-agent-treated cultures, revealing a strong synergistic therapeutic behavior of the BI-3406 +TKI combinations. Remarkably, the combined treatments including BI-3406 significantly restored imatinib sensitivity in CML patient cells harboring imatinib-resistant mutations. Cellular signaling and transcriptomics profiling suggested coordinated attenuation of RAS and RAC downstream signals as a mechanistic basis for the observed therapeutic responses. Our findings highlight the synergistic therapeutic behavior of BI-3406 and underscore the benefit of SOS1 pharmacological targeting as a novel strategy enhancing efficacy and overcoming resistance to TKIs in CML.

cancer biology↗

bioRxiv2025

Single-Cell Multiomics Reveals Regulatory Mechanisms of CAR T Cell Persistence and Dysfunction in Multiple Myeloma

Understanding the mechanisms that drive chimeric antigen receptor (CAR) T cell function and persistence in multiple myeloma (MM) remains a critical challenge for improving therapeutic outcomes. In this study, we applied single-cell multiomics and gene regulatory network (GRN) analysis to characterize the transcriptional dynamics and clonal evolution of BCMA-targeted CAR T cells in longitudinally collected bone marrow (BM) and peripheral blood (PB) samples from MM patients. Our results revealed that CAR T cells infiltrating BM exhibited a more activated and exhausted phenotype compared to their PB counterparts, with key transcriptional regulators driving these changes. Dysregulation in the effector-to-memory transition led to an increased presence of terminally differentiated CAR T cells, correlating with poor persistence. Additionally, we identified a hyperexpanded CAR T clone in the BM of a patient with partial response, marked by elevated IL10 expression. Functional analyses demonstrated that stimulation of endogenous TCR enhanced IL10 production, potentially contributing to impaired CAR T cell proliferation and persistence. These findings uncover critical regulatory mechanisms influencing CAR T cell dynamics, offering new insights into improving CAR T cell persistence and therapeutic efficacy in MM and highlights potential molecular targets for optimizing CAR T cell therapy in patients with MM.

genomics↗

bioRxiv2021

Transcriptional regulation of HSCs in Aging and MDS reveals DDIT3 as a Potential Driver of Transformation

Myelodysplastic syndromes (MDS) are hematopoietic stem cell (HSC) malignancies characterized by ineffective hematopoiesis, with increased incidence in elderly individuals. In this work, we analyzed the transcriptome of human HSCs purified from young and elderly healthy donors, as well as MDS patients, identifying transcriptional alterations following eight different patterns of expression. While aging-associated lesions seemed to predispose HSCs to myeloid transformation, disease-specific alterations may trigger MDS development. Among MDS-specific lesions, we detected the upregulation of the transcription factor DDIT3. Overexpression of DDIT3 in human healthy HSCs induced an MDS-like transcriptional state, and a delay in erythropoiesis. Such effect was associated with downregulation of transcription factors required for normal erythropoiesis, and with a failure in the activation of their transcriptional programs. Moreover, DDIT3 knockdown in CD34+ cells from MDS patients with anemia was able to restore erythropoiesis. These results identify DDIT3 as a driver of dyserythropoiesis, and a potential therapeutic target to restore the inefficient erythropoiesis characterizing MDS patients. STATEMENT OF SIGNIFICANCEThis study defines how human aging and MDS development are characterized by transcriptional alterations in HSCs that follow different patterns, some of which may contribute to myeloid transformation. Among them, we demonstrate how MDS-specific upregulation of DDIT3 in HSCs induces dyserythropoiesis, while its knockdown in HSPCs from MDS patients restores proper erythroid differentiation.

cancer biology↗

bioRxiv2021

Uncovering perturbations in human hematopoiesis associated with healthy aging and myeloid malignancies at single cell resolution

Early hematopoiesis is a continuous process in which hematopoietic stem and progenitor cells (HSPCs) gradually differentiate toward specific lineages. Aging and myeloid malignant transformation are characterized by changes in the composition and regulation of HSPCs. In this study, we used single cell RNA sequencing (scRNAseq) to characterize an enriched population of human hematopoietic stem and progenitor cells (HSPC) obtained from young and elderly healthy individuals. Based on their transcriptional profile, we identified changes in the proportions of progenitor compartments during aging, and differences in their functionality, as evidenced by gene set enrichment analysis. Trajectory inference revealed that altered gene expression dynamics accompanied cell differentiation, which could explain age-associated changes in hematopoiesis. Next, we focused on key regulators of transcription by constructing gene regulatory networks and detected regulons that were specifically active in elderly individuals. Using previous findings in healthy cells as a reference, we analyzed scRNA-seq data obtained from patients with myelodysplastic syndrome (MDS) and detected specific alterations of the expression dynamics of genes involved in erythroid differentiation in all patients with MDS such as TRIB2. In addition, the comparison between transcriptional programs and GRN regulating normal HSPCs and MDS HSPCs allowed identification of regulons that were specifically active in MDS cases such as SMAD1, HOXA6, POU2F2 and RUNX1 suggesting a role of these TF in the pathogenesis of the disease. In summary, we demonstrate that the combination of single cell technologies with computational analysis tools enable the study of a variety of cellular mechanisms involved in early hematopoiesis and can be used to dissect perturbed differentiation trajectories associated with aging and malignant transformation. Furthermore, the identification of abnormal regulatory mechanisms associated with myeloid malignancies could be exploited for personalized therapeutic approaches in individual patients.

genomics↗

bioRxiv2020

CRISPR/Cas9 technology abolishes the BCR/ABL1 oncogene effect in chronic myeloid leukemia and restores normal hematopoiesis

Chronic myeloid leukemia (CML) is a hematopoietic stem cell disease produced by a unique oncogenic event involving the constitutively active tyrosine kinase (TK) BCR/ABL1. TK activity explains most features of CML, such as tumor development and maintenance. TK-inhibitory (TKI) drugs have changed its prognosis and natural history. Unfortunately, the ABL1 gene persists unaffected by TKIs, leukemic stem cells (LSCs) remains, resistant mutations arise and adverse effects may occur during treatment. To address this problem, we have designed a potential therapeutic alternative with CRISPR/Cas9 genome editing nucleases that target LSCs. The strategy was successfully developed in murine and human cell lines and finally was evaluated in primary LSCs isolated from CML transgenic mice and from CML patients. Mouse CML-LSCs edited were orthotopic transplanted in immunodeficient NSG niches where restored the normal hematopoiesis. Importantly, patient-derived xenografts with CD34+-LSCs edited, repopulated and restored the normal hematopoiesis in immunodeficient NSG niches. We show, for the first time, how CRISPR technology efficiently interrupts the BCR/ABL1 oncogene in murine and human LSCs to provide a significant therapeutic benefit. We propose human CML as a potential candidate for CRISPR therapy, providing proof-of-principle for genome editing in CML patients, and open new avenues for the application of this technique in other fusion genes. Key pointsCRISPR system destroys BCR/ABL oncogene and induces a therapeutic benefit in a CML mouse model and CML patient derived xenografts.

cancer biology↗

bioRxiv2019

Splice donor site sgRNAs enhance CRISPR/Cas9-mediated knockout efficiency

CRISPR/Cas9 enables the generation of knockout cell lines and null zygotes by inducing site-specific double-stranded breaks. In most cases the DSB is repaired by non-homologous end joining, resulting in small nucleotide insertions or deletions that can be used to construct knockout alleles. However, these mutations do not produce the desired null result in all cases, but instead generate a similar, functionally active protein. This effect could limit the therapeutic efficiency of gene therapy strategies based on abrogating oncogene expression, and therefore needs to be considered carefully. If there is an acceptable degree of efficiency of CRISPR/Cas9 delivery to cells, the key step for success lies in the effectiveness of a specific sgRNA at knocking out the oncogene, when only one sgRNA can be used. This study shows that the null effect could be increased with an sgRNA targeting the splice donor site (SDS) of the chosen exon. Following this strategy, the generation of null alleles would be facilitated in two independent ways: the probability of producing a frameshift mutation and the probability of interrupting the canonical mechanism of pre-mRNA splicing. In these contexts, we propose to improve the loss-of-function yield driving the CRISPR system at the SDS of critical exons.

molecular biology↗