Expósito, F. (Francisco)
- Publications
- item.page.relationships.isContributorAdvisorOfPublication
- item.page.relationships.isContributorOfPublication
10 results
Search Results
Now showing 1 - 10 of 10
- Two cell line models to study multiorganic metastasis and immunotherapy in lung squamous cell carcinoma(2022) Expósito, F. (Francisco); Pio, R. (Rubén); Sainz, C. (Cristina); Agorreta, J. (Jackeline); Bosco, G. (Graziella); Thomas, R. (Roman); Larrayoz, M. (Marta); Azpilicueta, A. (Arantza); Montuenga-Badia, L.M. (Luis M.); Biurrun, G. (Gabriel) de; Bertolo, C. (Cristina); Melero, I. (Ignacio); Redrado, M. (Miriam); Valencia, K. (Karmele); Redín, E. (Esther); Echepare, M. (Mirari); Serrano-Tejero, D. (Diego); Calvo-González, A. (Alfonso); Ajona, D. (Daniel); Zandueta, C. (Carolina)There is a paucity of adequate mouse models and cell lines available to study lung squamous cell carcinoma (LUSC). We have generated and characterized two models of phenotypically different transplantable LUSC cell lines, i.e. UN-SCC679 and UN-SCC680, derived from A/J mice that had been chemically induced with N-nitroso-tris-chloroethylurea (NTCU). Furthermore, we genetically characterized and compared both LUSC cell lines by performing whole-exome and RNA sequencing. These experiments revealed similar genetic and transcriptomic patterns that may correspond to the classic LUSC human subtype. In addition, we compared the immune landscape generated by both tumor cells lines in vivo and assessed their response to immune checkpoint inhibition. The differences between the two cell lines are a good model for the remarkable heterogeneity of human squamous cell carcinoma. Study of the metastatic potential of these models revealed that both cell lines represent the organotropism of LUSC in humans, i.e. affinity to the brain, bones, liver and adrenal glands. In summary, we have generated valuable cell line tools for LUSC research, which recapitulates the complexity of the human disease.
- SRC family kinase (SFK) inhibitor dasatinib improves the antitumor activity of anti-PD-1 in NSCLC models by inhibiting Treg cell conversion and proliferation(2021) Expósito, F. (Francisco); Pio, R. (Rubén); Sainz, C. (Cristina); Senent, Y. (Yaiza); Garmendia, I. (Irati); Remirez, A. (Ana); Agorreta, J. (Jackeline); Lozano-Moreda, T. (Teresa); García-Pedrero, J.M. (Juana M.); Montuenga-Badia, L.M. (Luis M.); Bertolo, C. (Cristina); Villalba-Esparza, M. (María); Redrado, M. (Miriam); Redín, E. (Esther); Andrea, C.E. (Carlos Eduardo) de; Ortiz-Espinosa, S. (Sergio); Serrano-Tejero, D. (Diego); Calvo-González, A. (Alfonso); Ajona, D. (Daniel); Lasarte, J.J. (Juan José)Introduction The use of immune-checkpoint inhibitors has drastically improved the management of patients with non-small cell lung cancer (NSCLC), but innate and acquired resistances are hurdles needed to be solved. Immunomodulatory drugs that can reinvigorate the immune cytotoxic activity, in combination with antiprogrammed cell death 1 (PD-1) antibody, are a great promise to overcome resistance. We evaluated the impact of the SRC family kinases (SFKs) on NSCLC prognosis, and the immunomodulatory effect of the SFK inhibitor dasatinib, in combination with anti-PD-1, in clinically relevant mouse models of NSCLC. Methods A cohort of patients from University Clinic of Navarra (n=116) was used to study immune infiltrates by multiplex immunofluorescence (mIF) and YES1 protein expression in tumor samples. Publicly available resources (TCGA, Km Plotter, and CIBERSORT) were used to study patient's survival based on expression of SFKs and tumor infiltrates. Syngeneic NSCLC mouse models 393P and UNSCC680AJ were used for in vivo drug testing. Results Among the SFK members, YES1 expression showed the highest association with poor prognosis. Patients with high YES1 tumor levels also showed high infiltration of CD4+/FOXP3+ cells (regulatory T cells (Tregs)), suggesting an immunosuppressive phenotype. After testing for YES1 expression in a panel of murine cell lines, 393P and UNSCC680AJ were selected for in vivo studies. In the 393P model, dasatinib+anti-PD-1 treatment resulted in synergistic activity, with 87% tumor regressions and development of immunological memory that impeded tumor growth when mice were rechallenged. In vivo depletion experiments further showed that CD8+ and CD4+ cells are necessary for the therapeutic effect of the combination. The antitumor activity was accompanied by a very significant decrease in the number of Tregs, which was validated by mIF in tumor sections. In the UNSCC680AJ model, the antitumor effects of dasatinib+anti-PD-1 were milder but similar to the 393P model. In in vitro assays, we demonstrated that dasatinib blocks proliferation and transforming growth factor beta-driven conversion of effector CD4+ cells into Tregs through targeting of phospholymphocyte-specific protein tyrosine kinase and downstream effectors pSTAT5 and pSMAD3. Conclusions YES1 protein expression is associated with increased numbers of Tregs in patients with NSCLC. Dasatinib synergizes with anti-PD-1 to impair tumor growth in NSCLC experimental models. This study provides the preclinical rationale for the combined use of dasatinib and PD-1/programmed death-ligand 1 blockade to improve outcomes of patients with NSCLC.
- Cancer cell-intrinsic alterations associated with an immunosuppressive tumor microenvironment and resistance to immunotherapy in lung cancer(2023) Expósito, F. (Francisco); Houry, M. (Maeva); Arozarena, I. (Imanol); Leon, S. (Sergio); Montuenga-Badia, L.M. (Luis M.); Otegui, N. (Nerea); Valencia, K. (Karmele); Redín, E. (Esther); Serrano-Tejero, D. (Diego); Calvo-González, A. (Alfonso)Simple Summary Immunotherapy for non-small cell lung cancer (NSCLC) is a clinical reality with impressive efficacy for some patients. However, less than half of them will benefit from this therapeutic regime, and finding indicators of response is necessary for selecting the patients likely to respond. So far, detection of PD-L1 in tumors by immunohistochemistry is the only validated predictive biomarker. In recent years, certain changes in the tumor cells (intrinsic alterations), including mutations, epigenetic changes and metabolic rewiring, have been shown to modify the type of tumor microenvironment (TME) where such a tumor grows. This TME may determine response or refractoriness to immunotherapy. Examples of key mutations are KRAS, SKT11(LKB1), KEAP1 and TP53, as well as co-mutations of these genes. Reshaping the TME with novel strategies in these particular TMEs could increase the efficacy of immunotherapy in lung cancer patients. Despite the great clinical success of immunotherapy in lung cancer patients, only a small percentage of them (<40%) will benefit from this therapy alone or combined with other strategies. Cancer cell-intrinsic and cell-extrinsic mechanisms have been associated with a lack of response to immunotherapy. The present study is focused on cancer cell-intrinsic genetic, epigenetic, transcriptomic and metabolic alterations that reshape the tumor microenvironment (TME) and determine response or refractoriness to immune checkpoint inhibitors (ICIs). Mutations in KRAS, SKT11(LKB1), KEAP1 and TP53 and co-mutations of these genes are the main determinants of ICI response in non-small-cell lung cancer (NSCLC) patients. Recent insights into metabolic changes in cancer cells that impose restrictions on cytotoxic T cells and the efficacy of ICIs indicate that targeting such metabolic restrictions may favor therapeutic responses. Other emerging pathways for therapeutic interventions include epigenetic modulators and DNA damage repair (DDR) pathways, especially in small-cell lung cancer (SCLC). Therefore, the many potential pathways for enhancing the effect of ICIs suggest that, in a few years, we will have much more personalized medicine for lung cancer patients treated with immunotherapy. Such strategies could include vaccines and chimeric antigen receptor (CAR) cells.
- Cancer epigenetic biomarkers in liquid biopsy for high incidence malignancies(2021) Palanca-Ballester, C. (Cora); Expósito, F. (Francisco); Jantus-Lewintre, E. (Eloisa); Calabuig-Fariñas, S. (Silvia); Rodríguez-Casanova, A. (Aitor); Montuenga-Badia, L.M. (Luis M.); Valencia, K. (Karmele); Redín, E. (Esther); Torres, S. (Susana); Serrano-Tejero, D. (Diego); Calvo-González, A. (Alfonso); Sandoval, J. (Juan); Diaz-Lagares, A. (Ángel)Simple Summary Apart from genetic changes, cancer is characterized by epigenetic alterations, which indicate modifications in the DNA (such as DNA methylation) and histones (such as methylation and acetylation), as well as gene expression regulation by non-coding (nc)RNAs. These changes can be used in biological fluids (liquid biopsies) for diagnosis, prognosis and prediction of cancer drug response. Although these alterations are not widely used as biomarkers in the clinical practice yet, increasing number of commercial kits and clinical trials are expected to prove that epigenetic changes are able to offer valuable information for cancer patients. Early alterations in cancer include the deregulation of epigenetic events such as changes in DNA methylation and abnormal levels of non-coding (nc)RNAs. Although these changes can be identified in tumors, alternative sources of samples may offer advantages over tissue biopsies. Because tumors shed DNA, RNA, and proteins, biological fluids containing these molecules can accurately reflect alterations found in cancer cells, not only coming from the primary tumor, but also from metastasis and from the tumor microenvironment (TME). Depending on the type of cancer, biological fluids encompass blood, urine, cerebrospinal fluid, and saliva, among others. Such samples are named with the general term "liquid biopsy" (LB).
- TMPRSS4: a novel tumor prognostic indicator for the stratification of stage IA tumors and a liquid biopsy biomarker for NSCLC patients(MDPI AG, 2019) Behrens, C. (C.); Pajares, M.J. (María José); Expósito, F. (Francisco); Pio, R. (Rubén); Sainz, C. (Cristina); Lozano, M.D. (María Dolores); Remirez, A. (Ana); Jantus-Lewintre, E. (Eloisa); Camps, C. (Carlos); Montuenga-Badia, L.M. (Luis M.); Villalba-Esparza, M. (María); Redrado, M. (Miriam); Andrea, C.E. (Carlos Eduardo) de; Wistuba, I.I. (Ignacio I.); Calvo-González, A. (Alfonso)Relapse rates in surgically resected non-small-cell lung cancer (NSCLC) patients are between 30% and 45% within five years of diagnosis, which shows the clinical need to identify those patients at high risk of recurrence. The eighth TNM staging system recently refined the classification of NSCLC patients and their associated prognosis, but molecular biomarkers could improve the heterogeneous outcomes found within each stage. Here, using two independent cohorts (MDA and CIMA-CUN) and the eighth TNM classification, we show that TMPRSS4 protein expression is an independent prognostic factor in NSCLC, particularly for patients at stage I: relapse-free survival (RFS) HR, 2.42 (95% CI, 1.47–3.99), p < 0.001; overall survival (OS) HR, 1.99 (95% CI, 1.25–3.16), p = 0.004). In stage IA, high levels of this protein remained associated with worse prognosis (p = 0.002 for RFS and p = 0.001 for OS). As TMPRSS4 expression is epigenetically regulated, methylation status could be used in circulating tumor DNA from liquid biopsies to monitor patients. We developed a digital droplet PCR (ddPCR) method to quantify absolute copy numbers of methylated and unmethylated CpGs within the TMPRSS4 and SHOX2 (as control) promoters in plasma and bronchoalveolar lavage (BAL) samples. In case-control studies, we demonstrated that TMPRSS4 hypomethylation can be used as a diagnostic tool in early stages, with an AUROC of 0.72 (p = 0.008; 91% specificity and 52% sensitivity) for BAL and 0.73 (p = 0.015; 65% specificity and 90% sensitivity) for plasma, in early stages. In conclusion, TMPRSS4 protein expression can be used to stratify patients at high risk of relapse/death in very early stages NSCLC patients. Moreover, analysis of TMPRSS4 methylation status by ddPCR in blood and BAL is feasible and could serve as a non-invasive biomarker to monitor surgically resected patients.
- Identification of novel synthetic lethal vulnerability in non small cell lung cancer by co targeting TMPRSS4 and DDR1(Springer Science and Business Media LLC, 2019) Pajares, M.J. (María José); Expósito, F. (Francisco); Pio, R. (Rubén); Sainz, C. (Cristina); Jantus-Lewintre, E. (Eloisa); Camps, C. (Carlos); Montuenga-Badia, L.M. (Luis M.); Villalba-Esparza, M. (María); Guruceaga, E. (Elizabeth); López-López, R. (Rafael); Redrado, M. (Miriam); Valencia, K. (Karmele); Redín, E. (Esther); Lahoz, A. (Agustín); Andrea, C.E. (Carlos Eduardo) de; Calvo-González, A. (Alfonso); Sandoval, J. (Juan); Cirauqui, C. (Cristina); Hervas, D. (D.); Diaz-Lagares, A. (Ángel)Finding novel targets in non-small cell lung cancer (NSCLC) is highly needed and identification of synthetic lethality between two genes is a new approach to target NSCLC. We previously found that TMPRSS4 promotes NSCLC growth and constitutes a prognostic biomarker. Here, through large-scale analyses across 5 public databases we identified consistent co-expression between TMPRSS4 and DDR1. Similar to TMPRSS4, DDR1 promoter was hypomethylated in NSCLC in 3 independent cohorts and hypomethylation was an independent prognostic factor of disease-free survival. Treatment with 5-azacitidine increased DDR1 levels in cell lines, suggesting an epigenetic regulation. Cells lacking TMPRSS4 were highly sensitive to the cytotoxic effect of the DDR1 inhibitor dasatinib. TMPRSS4/DDR1 double knock-down (KD) cells, but not single KD cells suffered a G0/G1 cell cycle arrest with loss of E2F1 and cyclins A and B, increased p21 levels and a larger number of cells in apoptosis. Moreover, double KD cells were highly sensitized to cisplatin, which caused massive apoptosis (~40%). In vivo studies demonstrated tumor regression in double KD-injected mice. In conclusion, we have identified a novel vulnerability in NSCLC resulting from a synthetic lethal interaction between DDR1 and TMPRSS4.
- YES1 is a druggable oncogenic target in SCLC(Elsevier, 2022) Redín, E. (Esther); Garrido-Martín, E. (Eva); Valencia, K. (Karmele); Redrado, M. (Miriam); Solórzano-Rendón, J.L. (José Luis); Carias, R. (Rafael); Echepare, M. (Mirari); Expósito, F. (Francisco); Serrano-Tejero, D. (Diego); Ferrer, I. (Irene); Nunez-Buiza, A. (Angel); Garmendia, I. (Irati); García-Pedrero, J.M. (Juana M.); Gurpide, A. (Alfonso); Paz-Ares, L. (Luis); Politi, K. (Katerina); Montuenga-Badia, L.M. (Luis M.); Calvo-González, A. (Alfonso)Introduction: SCLC is an extremely aggressive subtype of lung cancer without approved targeted therapies. Here we identified YES1 as a novel targetable oncogene driving SCLC maintenance and metastasis. Methods: Association between YES1 levels and prognosis was evaluated in SCLC clinical samples. In vitro functional experiments for proliferation, apoptosis, cell cycle, and cytotoxicity were performed. Genetic and pharmacologic inhibition of YES1 was evaluated in vivo in cell- and patientderived xenografts and metastasis. YES1 levels were evaluated in mouse and patient plasma-derived exosomes. Results: Overexpression or gain/amplification of YES1 was identified in 31% and 26% of cases, respectively, across molecular subgroups, and was found as an independent predictor of poor prognosis. Genetic depletion of YES1 dramatically reduced cell proliferation, three-dimensional organoid formation, tumor growth, and distant metastasis, leading to extensive apoptosis and tumor regressions. Mechanistically, YES1-inhibited cells revealed alterations in the replisome and DNA repair processes, that conferred sensitivity to irradiation. Pharmacologic blockade with the novel YES1 inhibitor CH6953755 or dasatinib induced marked antitumor activity in organoid models and cell- and patient-derived xenografts. YES1 protein was detected in plasma exosomes from patients and mouse models, with levels matching those of tumors, suggesting that circulating YES1 could represent a biomarker for patient selection/ monitoring. Conclusions: Our results provide evidence that YES1 is a new druggable oncogenic target and biomarker to advance the clinical management of a subpopulation of patients with SCLC.
- Short-term starvation reduces IGF-1 levels to sensitize lung tumors to PD-1 immune checkpoint blockade(Nature, 2020) Ajona, D. (Daniel); Ortiz-Espinosa, S. (Sergio); Lozano-Moreda, T. (Teresa); Expósito, F. (Francisco); Calvo-González, A. (Alfonso); Valencia, K. (Karmele); Redrado, M. (Miriam); Remirez, A. (Ana); Lecanda, F. (Fernando); Alignani, D. (Diego); Lasarte, J.J. (Juan José); Macaya, I. (Irati); Senent, Y. (Yaiza); Bertolo, C. (Cristina); Sainz, C. (Cristina); Gil-Bazo, I. (Ignacio); Eguren-Santamaría, I. (Iñaki); Lopez-Picazo, J.M. (José M.); Perez-Gracia, J.L. (Jose Luis); Andrea, C.E. (Carlos Eduardo) de; Vicent, S. (Silvestre); Fernandez-Sanmamed, M. (Miguel); Montuenga-Badia, L.M. (Luis M.); Pio, R. (Rubén); González, Á. (Álvaro)Harnessing the immune system by blocking the programmed cell death protein 1 (PD-1) pathway has been a major breakthrough in non-small-cell lung cancer treatment. Nonetheless, many patients fail to respond to PD-1 inhibition. Using three syngeneic models, we demonstrate that short-term starvation synergizes with PD-1 blockade to inhibit lung cancer progression and metastasis. This antitumor activity was linked to a reduction in circulating insulin-like growth factor 1 (IGF-1) and a downregulation of IGF-1 receptor (IGF-1R) signaling in tumor cells. A combined inhibition of IGF-1R and PD-1 synergistically reduced tumor growth in mice. This effect required CD8 cells, boosted the intratumoral CD8/Treg ratio and led to the development of tumor-specific immunity. In patients with non-small-cell lung cancer, high plasma levels of IGF-1 or high IGF-1R expression in tumors was associated with resistance to anti-PD-1–programmed death-ligand 1 immunotherapy. In conclusion, our data strongly support the clinical evaluation of IGF-1 modulators in combination with PD-1 blockade.
- Intratumoral combination therapy with poly(I:C) and resiquimod synergistically triggers tumor-associated macrophages for effective systemic antitumoral immunity(BMJ, 2021) Anfray, C. (Clément); Mainini, F. (Francesco); Digifico, E. (Elisabeth); Maeda, A. (Akihiro); Sironi, M. (Marina); Erreni, M. (Marco); Anselmo, A. (Achille); Ummarino, A. (Aldo); Gandoy, S. (Sara); Expósito, F. (Francisco); Redrado, M. (Miriam); Serrano, D. (Diego); Calvo-González, A. (Alfonso); Martens, M. (Marvin); Bravo, S. (Susana B.); Mantovani, A. (Alberto); Allavena, P. (Paola); Torres-Andón, F. (Fernando)Background: Tumor-associated macrophages (TAMs) play a key immunosuppressive role that limits the ability of the immune system to fight cancer and hinder the antitumoral efficacy of most treatments currently applied in the clinic. Previous studies have evaluated the antitumoral immune response triggered by (TLR) agonists, such as poly(I:C), imiquimod (R837) or resiquimod (R848) as monotherapies; however, their combination for the treatment of cancer has not been explored. This study investigates the antitumoral efficacy and the macrophage reprogramming triggered by poly(I:C) combined with R848 or with R837, versus single treatments. Methods: TLR agonist treatments were evaluated in vitro for toxicity and immunostimulatory activity by Alamar Blue, ELISA and flow cytometry using primary human and murine M-CSF-differentiated macrophages. Cytotoxic activity of TLR-treated macrophages toward cancer cells was evaluated with an in vitro functional assay by flow cytometry. For in vivo experiments, the CMT167 lung cancer model and the MN/MCA1 fibrosarcoma model metastasizing to lungs were used; tumor-infiltrating leukocytes were evaluated by flow cytometry, RT-qPCR, multispectral immunophenotyping, quantitative proteomic experiments, and protein-protein interaction analysis. Results: Results demonstrated the higher efficacy of poly(I:C) combined with R848 versus single treatments or combined with R837 to polarize macrophages toward M1-like antitumor effectors in vitro. In vivo, the intratumoral synergistic combination of poly(I:C)+R848 significantly prevented tumor growth and metastasis in lung cancer and fibrosarcoma immunocompetent murine models. Regressing tumors showed increased infiltration of macrophages with a higher M1:M2 ratio, recruitment of CD4+ and CD8+ T cells, accompanied by a reduction of immunosuppressive CD206+ TAMs and FOXP3+/CD4+ T cells. The depletion of both CD4+ and CD8+ T cells resulted in complete loss of treatment efficacy. Treated mice acquired systemic antitumoral response and resistance to tumor rechallenge mediated by boosted macrophage cytotoxic activity and T-cell proliferation. Proteomic experiments validate the superior activation of innate immunity by poly(I:C)+R848 combination versus single treatments or poly(I:C)+R837, and protein-protein-interaction network analysis reveal the key activation of the STAT1 pathway. Discussion: These findings demonstrate the antitumor immune responses mediated by macrophage activation on local administration of poly(I:C)+R848 combination and support the intratumoral application of this therapy to patients with solid tumors in the clinic.
- PTEN loss confers resistance to anti-PD-1 therapy in non-small cell lung cancer by increasing tumor infiltration of regulatory T cells(American Association for Cancer Research, 2023) Expósito, F. (Francisco); Redrado, M. (Miriam); Houry, M. (Maeva); Hastings, K. (Katherine); Molero, M. (Magdalena); Lozano-Moreda, T. (Teresa); Solórzano-Rendón, J.L. (José Luis); Sanz, J. (Julián); Adradas, V. (Vera); Amat, R. (Ramon); Redín, E. (Esther); Leon, S. (Sergio); Legarra, N. (Naroa); Garcia, J. (Javier); Serrano-Tejero, D. (Diego); Valencia, K. (Karmele); Robles-Oteiza, C. (Camila); Foggetti, G. (Giorgia); Otegui, N. (Nerea); Felip, E. (Enriqueta); Lasarte, J.J. (Juan José); Paz-Ares, L. (Luis); Zugazagoitia, J. (Jon); Politi, K. (Katerina); Montuenga-Badia, L.M. (Luis M.); Calvo-González, A. (Alfonso)Immunotherapy resistance in non-small cell lung cancer (NSCLC) may be mediated by an immunosuppressive microenvironment, which can be shaped by the mutational landscape of the tumor. Here, we observed genetic alterations in the PTEN/PI3K/AKT/mTOR pathway and/or loss of PTEN expression in >25% of patients with NSCLC, with higher frequency in lung squamous carcinomas (LUSC). Patients with PTEN-low tumors had higher levels of PD-L1 and PD-L2 and showed worse progression-free survival when treated with immunotherapy. Development of a Pten-null LUSC mouse model revealed that tumors with PTEN loss were refractory to antiprogrammed cell death protein 1 (anti-PD-1), highly metastatic and fibrotic, and secreted TGFβ/CXCL10 to promote conversion of CD4+ lymphocytes into regulatory T cells (Treg). Human and mouse PTEN-low tumors were enriched in Tregs and expressed higher levels of immunosuppressive genes. Importantly, treatment of mice bearing Pten-null tumors with TLR agonists and anti-TGFβ antibody aimed to alter this immunosuppressive microenvironment and led to tumor rejection and immunologic memory in 100% of mice. These results demonstrate that lack of PTEN causes immunotherapy resistance in LUSCs by establishing an immunosuppressive tumor microenvironment that can be reversed therapeutically. Significance: PTEN loss leads to the development of an immunosuppressive microenvironment in lung cancer that confers resistance to anti-PD-1 therapy, which can be overcome by targeting PTEN loss-mediated immunosuppression.