Biology subjects

Garcia-Escudero, R.

Publications and source records attributed to Garcia-Escudero, R..

11 recordsLinked to original sources

bioRxiv2026

Functional proteomics identifies targetable cancer-associated fibroblast programs in head and neck cancer

Head and neck squamous cell carcinoma (HNSCC) remains clinically challenging, with limited molecularly targeted options and a strong dependence on the tumor microenvironment. Cancer-associated fibroblasts (CAFs) are major stromal regulators that shape tumor progression, extracellular matrix remodeling, invasion, and therapeutic response. However, how CAF heterogeneity and plasticity translate into distinct tumor-promoting functions and targetable vulnerabilities remains insufficiently defined. Here, we integrated patient-matched primary CAFs and normal fibroblasts with 3D functional assays, tumor-stroma co-culture models, quantitative extracellular matrix analysis, whole-proteome profiling, and pharmacological perturbation. Primary fibroblast populations displayed marked interpatient heterogeneity and context-dependent plasticity in invasion, contractility, and responsiveness to tumor-derived signals, whereas enhanced fibronectin-rich matrix deposition and disorganization emerged as a conserved CAF-associated feature. Both normal fibroblasts and CAFs promoted HNSCC cell invasion in a population-dependent manner, whereas CAFs consistently induced less compact and more dispersed tumor nest architectures. Integrative functional analyses identified distinct CAF phenotypes characterized by either invasive and matrix-remodeling activity or high responsiveness to tumor-derived cues. Proteomic profiling revealed recurrent enrichment of adhesion, cytoskeletal, and extracellular matrix programs and guided the selection of pharmacological inhibitors aimed at modulating specific CAF-mediated pro-tumoral functions. Pharmacological targeting selectively altered these functions: CHI3L1 inhibition disrupted fibronectin matrix deposition, broad phosphodiesterase inhibition increased matrix alignment, and FZD7 inhibition consistently blocked tumor-induced CAF invasion across all tested populations. These findings define functionally distinct and pharmacologically targetable CAF programs in HNSCC and support stromal-directed interventions as a rational component of future combination treatment strategies.

cancer biology↗

bioRxiv2024

The G-Protein Couple Receptor Kinase 2 (GRK2) Orchestrates Hair Follicle Homeostasis

Tightly regulated cell-cell and cell-niche intercommunications via intertwined signaling networks are involved in maintaining normal hair follicle (HF) homeostasis, cycling and cell fate determination. However, knowledge of specific mechanisms by which hair loss takes place under pathological situations is needed. Using a keratinocyte-specific knockout mouse model, we uncover that the G-protein-coupled receptor kinase 2 (GRK2) signaling node plays a key role in HF homeostasis. Epidermal GRK2 ablation causes alterations during anagen induction, giving rise to abnormal cyst-like structures. HF-linked cysts display aberrant growth and differentiation patterns as well as lineage infidelity, displaying features of abortive HFs unable to fully acquire canonical hallmarks. Cysts triggered by GRK2 deletion displace the dermal papilla away from the bulge and promote irreversible changes in HF stem cell architecture, leading to bulge destruction and hair loss. Our data provide unforeseen roles of GRK2 in epidermal physiology and uncover mechanisms linking dystrophic follicular cysts formation with hair loss, with potential connections to pathogenic processes operating in immune-mediated alopecias.

cell biology↗

bioRxiv2023

Non-viral CRISPR/Cas9 Mutagenesis for Streamlined Generation of Mouse Lung Cancer Models

Functional analysis in mouse models is necessary to establish the involvement of a set of genetic variations in tumor development. Many lung cancer models have been developed using genetic techniques to create gain- or loss-of-function alleles in genes involved in tumorigenesis; however, because of their labor- and time-intensive nature, these models are not suitable for quick and flexible hypothesis testing. Here we introduce a lung mutagenesis platform that utilizes CRISPR/Cas9 RNPs delivered via cationic polymers. This approach allows for the simultaneous inactivation of multiple genes. We validate the effectiveness of this system by targeting a group of tumor suppressor genes, specifically Rb1, Rbl1, Pten, and Trp53, which were chosen for their potential to cause lung tumors, namely Small Cell Lung Carcinoma (SCLC). This polymer-based delivery platform enables the modeling of lung tumorigenesis independently of the genetic background, thus simplifying and expediting the process without the need for modifying the mouse germline or creating custom viral vectors. SignificanceThe development of models to rapidly introduce gene mutations into lung tissue to study their impact on tumor growth is critical for advancing the functional genomics of lung cancer. While previous methods using viral vectors and genetic manipulation in mice have been time-consuming and expensive, here we describe a new technique using cationic polymers as non-viral carriers for CRISPR/Cas9 delivery to induce cancer driving mutations that streamlines this process. This approach mimics natural mutations in lung cancer and accelerates the generation of accurate tumor models. Our study demonstrates the effectiveness of this method in generating small cell lung cancer (SCLC) by modifying four tumor suppressor genes in different mouse genetic backgrounds. This innovative strategy holds promise for faster and more cost-effective cancer modeling. Graphical AbstracSmall Cell Lung Cancer (SCLC) tumors are rapidly generated in any mouse genetic background by using cationic polymers to simultaneously deliver Cas9 and gRNAs targeting the Rb1, Rbl1, Pten and Trp53 tumor-suppressor genes to the adult airway respiratory system in vivo. Addition of the frt guide in the RC::FLTG mice provides a tdTomato gene editing reporter. This study shows the feasibility of rapidly generating lung cancer mouse models via somatic genome engineering through delivery of all CRISPR components in the form of nanoparticles. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=94 SRC="FIGDIR/small/572771v1_ufig1.gif" ALT="Figure 1"> View larger version (13K): org.highwire.dtl.DTLVardef@d12eeeorg.highwire.dtl.DTLVardef@160472corg.highwire.dtl.DTLVardef@1e0c371org.highwire.dtl.DTLVardef@13f33c1_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

Fanconi anemia-isogenic head and neck cancer cell line pairs - a basic and translational science resource

Fanconi anemia (FA) is a heritable malformation, bone marrow failure and cancer predisposition syndrome that confers an exceptionally high risk of developing carcinomas arising in squamous mucosal epithelia lining the mouth, proximal esophagus, vulva and anus. The origin of these cancers is not understood, and no effective way has been identified to prevent or delay their appearance. FA-associated carcinomas are also therapeutically challenging, as they may be multi-focal and stage-advanced at diagnosis making surgical control challenging. Moreover, individuals with FA have systemic DNA damage hypersensitivity and thus an elevated risk of toxicity when treated with standard-of-care therapies such as DNA cross-linking drugs and ionizing radiation. We developed the Fanconi Anemia Cancer Cell Line Resource (FA-CCLR) in order to foster new research on the origins, treatment, and prevention of FA-associated cancers. The FA-CCLR consists of FANC-isogenic head and neck squamous cell carcinoma (HNSCC) cell line pairs from cancers arising in individuals with FA, or newly engineered from sporadic HNSCC cell lines. Molecular, cellular, and biochemical analyses were used to demonstrate the causal dependence of key FA-associated phenotypes on FANC genotype, expression and pathway activity. These FANC-isogenic cell line pairs are available to academic and non-profit investigators, with ordering information available at the Fanconi Anemia Research Materials Resource and Repository at Oregon Health & Sciences University, Portland OR. SignificanceWe have generated new isogenic cancer cell line models to investigate the origins, treatment and prevention of Fanconi anemia-associated squamous carcinomas that target the oral mucosa, proximal esophagus, and anogenital region.

cancer biology↗

bioRxiv2022

Fanca and p53 genes cooperate to suppress oral SCC in transgenic mice

Fanconi anemia (FA) patients frequently develop oral squamous cell carcinoma (OSCC). This cancer in FA patients is diagnosed within the first 3-4 decades of life, very often preceded by lesions that suffer a malignant transformation. In addition, they respond poorly to current treatments due to toxicity or multiple recurrences. Translational research of new chemopreventive agents and therapeutic strategies has been unsuccessful partly due to scarcity of disease models or failure to fully reproduce the disease. Here we report that Fanca gene knockout mice (Fanca-/-) frequently display pre-malignant lesions in the oral cavity. Moreover, when these animals were crossed with animals having conditional deletion of Trp53 gene in oral mucosa (K14cre;Trp53F2-10/F2-10), they spontaneously developed OSCC with a high penetrance and a median latency of less than ten months. Tumors were well differentiated and expressed markers of squamous differentiation, such as keratins K5 and K10. In conclusion, Fanca and Trp53 genes cooperate to suppress oral cancer in mice, and Fanca-/-;K14cre;Trp53F2-10/F2-10 mice constitute the first animal model of spontaneous OSCC in FA.

cancer biology↗

bioRxiv2022

Ambra1 haploinsufficiency results in metabolic alterations and exacerbates age-associated retinal degeneration

Autophagy is a key process in the maintenance of cellular homeostasis. The age-dependent decline in retinal autophagy has been associated with photoreceptor degeneration. Retinal dysfunction can also result from damage to the retinal pigment epithelium (RPE), as the RPE-retina constitutes an important metabolic ecosystem that must be finely tuned to preserve visual function. While studies of mice lacking essential autophagy genes have revealed a predisposition to retinal degeneration, the consequences of a moderate reduction in autophagy, similar to that which occurs during physiological aging, remain unclear. Here, we describe a retinal phenotype consistent with accelerated aging in mice carrying a haploinsufficiency for Ambra1, a pro-autophagic gene. These mice show protein aggregation in the retina and RPE, metabolic underperformance, and premature vision loss. Moreover, Ambra1+/gt mice are more prone to retinal degeneration after RPE stress. These findings indicate that autophagy provides crucial support to RPE-retinal metabolism and protects the retina against stress and physiological aging.

cell biology↗

bioRxiv2020

Generating new FANCA-deficient HNSCC cell lines by genomic editing recapitulate the cellular phenotypes of Fanconi anemia

Fanconi anemia (FA) patients have an exacerbated risk of head and neck squamous cell carcinoma (HNSCC). Treatment is challenging as FA patients display enhanced toxicity to standard treatments, including radio/chemotherapy. Therefore better therapies as well as new disease models are urgently needed. We have used CRISPR/Cas9 editing tools in order to interrupt the human FANCA gene by the generation of insertions/deletions (indels) in exon 4 in two cancer cell lines from sporadic HNSCC having no mutation in FA-genes: CAL27 and CAL33 cells. Our approach allowed efficient editing, subsequent purification of single-cell clones, and Sanger sequencing validation at the edited locus. Clones having frameshift indels in homozygosis did not express FANCA protein and were selected for further analysis. When compared with parental CAL27 and CAL33, FANCA-mutant cell clones displayed a FA-phenotype as they i) are highly sensitive to DNA interstrand crosslink (ICL) agents such as mitomycin C (MMC) or cisplatin, ii) do not monoubiquitinate FANCD2 upon MMC treatment and therefore iii) do not form FANCD2 nuclear foci, and iv) they display increased chromosome fragility and G2 arrest after diepoxybutane (DEB) treatment. These FANCA-mutant clones display similar growth rates as their parental cells. Interestingly, mutant cells acquire phenotypes associated with more aggressive disease, such as increased migration in wound healing assays. Therefore, CAL27 and CAL33 cells with FANCA mutations are phenocopies of FA-HNSCC cells.

cancer biology↗

bioRxiv2020

Huntington's disease-specific mis-splicing captured by human-mouse intersect-RNA-seq unveils pathogenic effectors and reduced splicing factors

Deregulated alternative splicing has been implicated in a wide range of pathologies. Deep RNA-sequencing has revealed global mis-splicing signatures in multiple human diseases; however, for neurodegenerative diseases, these analyses are intrinsically hampered by neuronal loss and neuroinflammation in post-mortem brains. To infer splicing alterations relevant to Huntingtons disease (HD) pathogenesis, here we performed intersect-RNA-seq analyses of human post-mortem striatal tissue and of an early symptomatic mouse model in which neuronal loss and gliosis are not yet present. Together with a human/mouse parallel motif scan analysis, this approach allowed us to identify the shared mis-splicing signature triggered by the HD-causing mutation in both species and to infer upstream deregulated splicing factors. Moreover, we identified a plethora of downstream neurodegeneration-linked effector genes, whose aberrant splicing is associated with decreased protein levels in HD patients and mice. In summary, our intersect-RNA-seq approach unveiled the pathogenic contribution of mis-splicing to HD and could be readily applied to other neurodegenerative diseases for which bona fide animal models are available.

neuroscience↗

bioRxiv2019

The IDH-Tau-EGFR triad defines diffuse glioma pathology by controlling mesenchymal differentiation and neo-vascular fitness.

Classification of gliomas as wild-type or mutant IDH1/2 tumors has profound clinical implications. However, how these two groups of gliomas progress, in a microenvironment-dependent manner, is still a pending question. Here we describe that the expression of Tau is epigenetically induced by mutant IDH1/2, whereas is almost absent from tumors with EGFR/PTEN mutations. Moreover, Tau (MAPT) expression is inversely correlated with overall survival in EGFR-amplified gliomas. Using orthotopic EGFR-related models, we have observed that Tau overexpression or microtubule stabilizers impair the mesenchymal transformation of glioma cells, with profound changes in tumor vasculature and a significant decrease in tumor burden. However, epithelial-to-mesenchymal transformed EGFR-mutant cells, acting as pericytes, induce neo-vasculogenesis and favor aggressive glioma growth, a process that is no longer sensitive to Tau. Altogether our data indicate that the genomic background controls glioma aggressiveness by modifying the vascular microenvironment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/541326v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@1c4b33forg.highwire.dtl.DTLVardef@d3f636org.highwire.dtl.DTLVardef@1c957c1org.highwire.dtl.DTLVardef@1a04a0d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2018

Prostaglandin F2α-induced TGFβ-PMEPA1 pathway is a critical mediator of epithelial plasticity and ovarian carcinoma progression.

Prostaglandin (PG) F2 has been scarcely studied in cancer. We have identified a new role for PGF2 in ovarian cancer, stimulating the production of TGF{beta} and the consequent induction of PMEPA1. We show that this induction increases cell plasticity and proliferation, enhancing tumor growth through PMEPA1. Thus, PMEPA1 overexpression in ovarian carcinoma cells, significantly increased cell proliferation rates, whereas PMEPA1 silencing decreased proliferation. In addition, PMEPA1 overexpression buffered TGF{beta} signaling, via reduction of SMAD-dependent signaling. PMEPA1 overexpressing cells acquired an epithelial morphology, associated to higher E-cadherin expression levels while {beta}-catenin nuclear translocation was inhibited. Interestingly, in mouse xenografts, PMEPA1 overexpressing ovarian cells had a clear survival and proliferative advantage, resulting in higher metastatic capacity, while PMEPA1 silencing had the opposite effect. Furthermore, high PMEPA1 expression in a cohort of advanced ovarian cancer patients was observed, correlating with E-cadherin expression. Most importantly, high PMEPA1 mRNA levels were associated with lower patient survival.\n\nSignificanceThis work identifies Prostaglandin F2 as an inducer, through NFAT, of TGF{beta} and together with this up-regulate PMEPA1, which is identified both as a drug target and as a prognostic biomarker in ovarian tumors, potentially useful in clinical practice.

cancer biology↗

bioRxiv2017

Clusterization in head and neck squamous carcinomas based on lncRNA expression: molecular and clinical correlates

BackgroundLong non-coding RNAs (lncRNAs) have emerged as key players in a remarkably variety of biological processes and pathologic conditions, including cancer. Next-generation sequencing technologies and bioinformatics procedures predict the existence of tens of thousands of lncRNAs, from which we know the functions of only a handful of them, and very little is known in cancer types such as head and neck squamous cell carcinomas (HNSCCs).\n\nResultsHere, we use RNA-seq expression data from The Cancer Genome Atlas (TCGA) and various statistic and software tools in order to get insight about the lncRNome in HNSCC. Based on lncRNAs expression across 426 samples, we discover five distinct tumor clusters that we compare with reported clusters based on various genomic/genetic features. Results demonstrate significant associations between lncRNA-based clustering and DNA-methylation, TP53 mutation, and human papillomavirus infection. Using \"guilt by association\" procedures, we infer the possible biological functions of representative lncRNAs of each cluster. Furthermore, we found that lncRNA clustering is correlated with some important clinical and pathologic features, including patient survival after treatment, tumor grade or sub-anatomical location.\n\nConclusionsWe present a landscape of lncRNAs in HNSCC, and provide associations with important genotypic and phenotypic features that may help to understand the disease.

genomics↗