Open Access
Systemic CD4 immunity and PD-L1/PD-1 blockade immunotherapy
(MDPI, 2022) Escors Murugarren, David; Bocanegra Gondán, Ana Isabel; Chocarro de Erauso, Luisa; Blanco, Ester; Piñeiro Hermida, Sergio; Garnica, Maider; Fernández Rubio, Leticia; Vera García, Ruth; Arasanz Esteban, Hugo; Kochan, Grazyna; Ciencias de la Salud; Osasun Zientziak
PD-L1/PD-1 blockade immunotherapy has changed the therapeutic approaches for the treatment of many cancers. Nevertheless, the mechanisms underlying its efficacy or treatment failure are still unclear. Proficient systemic immunity seems to be a prerequisite for efficacy, as recently shown in patients and in mouse models. It is widely accepted that expansion of anti-tumor CD8 T cell populations is principally responsible for anti-tumor responses. In contrast, the role of CD4 T cells has been less studied. Here we review and discuss the evidence supporting the contribution of CD4 T cells to anti-tumor immunity, especially recent advances linking CD4 T cell subsets to efficacious PD-L1/PD-1 blockade immunotherapy. We also discuss the role of CD4 T cell memory subsets present in peripheral blood before the start of immunotherapies, and their utility as predictors of response.
Open Access
Cutting-edge: preclinical and clinical development of the first approved LAG-3 inhibitor
(MDPI, 2022) Chocarro de Erauso, Luisa; Bocanegra Gondán, Ana Isabel; Blanco, Ester; Fernández Rubio, Leticia; Arasanz Esteban, Hugo; Echaide Górriz, Míriam; Garnica, Maider; Ramos, Pablo; Piñeiro Hermida, Sergio; Vera García, Ruth; Escors Murugarren, David; Kochan, Grazyna; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
Immune checkpoint inhibitors (ICIs) have revolutionized medical practice in oncology since the FDA approval of the first ICI 11 years ago. In light of this, Lymphocyte-Activation Gene 3 (LAG-3) is one of the most important next-generation immune checkpoint molecules, playing a similar role as Programmed cell Death protein 1 (PD-1) and Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4). 19 LAG-3 targeting molecules are being evaluated at 108 clinical trials which are demonstrating positive results, including promising bispecific molecules targeting LAG-3 simultaneously with other ICIs. Recently, a new dual anti-PD-1 (Nivolumab) and anti-LAG-3 (Relatimab) treatment developed by Bristol Myers Squibb (Opdualag), was approved by the Food and Drug Administration (FDA) as the first LAG-3 blocking antibody combination for unresectable or metastatic melanoma. This novel immunotherapy combination more than doubled median progression-free survival (PFS) when compared to nivolumab monotherapy (10.1 months versus 4.6 months). Here, we analyze the large clinical trial responsible for this historical approval (RELATIVITY-047), and discuss the preclinical and clinical developments that led to its jump into clinical practice. We will also summarize results achieved by other LAG-3 targeting molecules with promising anti-tumor activities currently under clinical development in phases I, I/II, II, and III. Opdualag will boost the entry of more LAG-3 targeting molecules into clinical practice, supporting the accumulating evidence highlighting the pivotal role of LAG-3 in cancer.
Open Access
PD-L1 in systemic immunity: unraveling its contribution to PD-1/PD-L1 blockade immunotherapy
(MDPI, 2020) Bocanegra Gondán, Ana Isabel; Blanco, Ester; Fernández Hinojal, Gonzalo; Chocarro de Erauso, Luisa; Zuazo Ibarra, Miren; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
The use of monoclonal antibodies targeting PD-1/PD-L1 axis completely changed anticancer treatment strategies. However, despite the significant improvement in overall survival and progression-free survival of patients undergoing these immunotherapy treatments, the only clinically accepted biomarker with some prediction capabilities for the outcome of the treatment is PD-L1 expression in tumor biopsies. Nevertheless, even when having PD-L1-positive tumors, numerous patients do not respond to these treatments. Considering the high cost of these therapies and the risk of immune-related adverse events during therapy, it is necessary to identify additional biomarkers that would facilitate stratifying patients in potential responders and non-responders before the start of immunotherapies. Here, we review the utility of PD-L1 expression not only in tumor cells but in immune system cells and their influence on the antitumor activity of immune cell subsets.
Embargo
Study of PD-1/LAG-3 signaling in T cells and development of therapeutic strategies to counteract PD-1/LAG-3 mediated resistance to cancer immunotherapy
(2025) Chocarro de Erauso, Luisa; Escors Murugarren, David; Kochan, Grazyna; Ciencias de la Salud; Osasun Zientziak
A significant number of cancer patients do not benefit from PD-L1/PD-1 blockade immunotherapies. PD-1 and LAG-3 co-upregulation in T-cells is one of the major mechanisms of resistance by establishing a highly dysfunctional state in T-cells. However, how LAG-3 mechanistically triggers its downstream inhibitory signalling remains largely unknown. Likewise, how LAG-3 cooperates with PD-1 to stablish a highly dysfunctional state in T cells in cancer is also unknown. A better understanding of LAG-3 and PD-1/LAG-3 signaling will uncover the reasons behind intrinsic resistance to PD-1 blockade, develop novel treatments and improve current therapies. This PhD thesis comprises the study of PD-1/LAG-3 signaling in T-cells and the development of therapeutic strategies to counteract PD-1/LAG-3-mediated resistance to cancer immunotherapy. The results of this thesis have been organized into three chapters. In chapter 1, PD-1/LAG-3 molecular co-signaling mechanisms were identified in T-cells, uncovering CBLB ubiquitin ligases as potential targets. The combination of CBL-B inhibitors with anti-PD- 1/anti-LAG-3 immunotherapies overcame PD-1/LAG-3-mediated resistance in models of lung cancer refractory to immunotherapies. In chapter 2, the consequences of PD-1/LAG-3 cosignaling over the proteome and phosphoproteome associated to the TCR signalosome were characterized. MYC was identified as an inhibited upstream regulator of PD-1/LAG-3 proteomes and phosphoproteomes in T-cells. In chapter 3, T-cell lines expressing LAG-3 mutants in its signaling domains were engineered and analyzed by quantitative highthroughput differential proteomics and phosphoproteomics. The pathways and molecules regulated by each of LAG-3 domains and their effect on the TCR signalosome both in terms of protein expression and phosphorylation activity were characterized. These results highlighted the critical role of KIEELE and EP domains to endow LAG-3 with inhibitory functions. Overall, the results will help to identify the mechanisms of intrinsic resistance to PD-1 blockade mediated by LAG-3 co-signaling, and uncover novel therapeutic targets to counteract PD- 1/LAG-3-mediated resistance.
Open Access
A proteomic atlas of lineage and cancer-polarized expression modules in myeloid cells modeling immunosuppressive tumor-infiltrating subsets
(MDPI, 2021) Blanco, Ester; Ibañez Vea, María; Hernández, Carlos; Drici, Lylia; Martínez de Morentin Iribarren, Xabier; Gato Cañas, María; Ausín, Karina; Bocanegra Gondán, Ana Isabel; Zuazo Ibarra, Miren; Chocarro de Erauso, Luisa; Arasanz Esteban, Hugo; Fernández Hinojal, Gonzalo; Fernández Irigoyen, Joaquín; Smerdou, Cristian; Garnica, Maider; Echaide Górriz, Míriam; Fernández Rubio, Leticia; Morente Sancho, Pilar; Ramos-Castellanos, Pablo; Llopiz, Diana; Santamaría Martínez, Enrique; Larsen, Martin R.; Escors Murugarren, David; Kochan, Grazyna; Osasun Zientziak; Institute for Multidisciplinary Research in Applied Biology - IMAB; Ciencias de la Salud; Gobierno de Navarra / Nafarroako Gobernua
Monocytic and granulocytic myeloid-derived suppressor cells together with tumor-infiltrating macrophages constitute the main tumor-infiltrating immunosuppressive myeloid populations. Due to the phenotypic resemblance to conventional myeloid cells, their identification and purification from within the tumors is technically difficult and makes their study a challenge. We differentiated myeloid cells modeling the three main tumor-infiltrating types together with uncommitted macrophages, using ex vivo differentiation methods resembling the tumor microenvironment. The phenotype and proteome of these cells was compared to identify linage-dependent relationships and cancer-specific interactome expression modules. The relationships between monocytic MDSCs and TAMs, monocytic MDSCs and granulocytic MDSCs, and hierarchical relationships of expression networks and transcription factors due to lineage and cancer polarization were mapped. Highly purified immunosuppressive myeloid cell populations that model tumor-infiltrating counterparts were systematically analyzed by quantitative proteomics. Full functional interactome maps have been generated to characterize at high resolution the relationships between the three main myeloid tumor-infiltrating cell types. Our data highlights the biological processes related to each cell type, and uncover novel shared and differential molecular targets. Moreover, the high numbers and fidelity of ex vivo-generated subsets to their natu-ral tumor-shaped counterparts enable their use for validation of new treatments in high-throughput experiments.
Open Access
PD-1/LAG-3 co-signaling profiling uncovers CBL ubiquitin ligases as key immunotherapy targets
(EMBO Press, 2024-07-19) Chocarro de Erauso, Luisa; Blanco, Ester; Fernández-Rubio, Leticia; Garnica, Maider; Zuazo Ibarra, Miren; García Granda, María Jesús; Bocanegra Gondán, Ana Isabel; Echaide Górriz, Míriam; Johnston, Colette; Edwards, Carolyn J.; Legg, James; Pierce, Andrew J.; Arasanz Esteban, Hugo; Fernández Hinojal, Gonzalo; Vera García, Ruth; Ausín, Karina; Santamaría Martínez, Enrique; Fernández Irigoyen, Joaquín; Kochan, Grazyna; Escors Murugarren, David; Ciencias de la Salud; Osasun Zientziak
Many cancer patients do not benefit from PD-L1/PD-1 blockade immunotherapies. PD-1 and LAG-3 co-upregulation in T-cells is one of the major mechanisms of resistance by establishing a highly dysfunctional state in T-cells. To identify shared features associated to PD-1/LAG-3 dysfunctionality in human cancers and T-cells, multiomic expression profiles were obtained for all TCGA cancers immune infiltrates. A PD-1/LAG-3 dysfunctional signature was found which regulated immune, metabolic, genetic, and epigenetic pathways, but especially a reinforced negative regulation of the TCR signalosome. These results were validated in T-cell lines with constitutively active PD-1, LAG-3 pathways and their combination. A differential analysis of the proteome of PD-1/LAG-3 T-cells showed a specific enrichment in ubiquitin ligases participating in E3 ubiquitination pathways. PD-1/LAG-3 co-blockade inhibited CBL-B expression, while the use of a bispecific drug in clinical development also repressed C-CBL expression, which reverted T-cell dysfunctionality in lung cancer patients resistant to PD-L1/PD-1 blockade. The combination of CBL-B-specific small molecule inhibitors with anti-PD-1/anti-LAG-3 immunotherapies demonstrated notable therapeutic efficacy in models of lung cancer refractory to immunotherapies, overcoming PD-1/LAG-3 mediated resistance. © The Author(s) 2024.
Open Access
Immune profiling uncovers memory T-cell responses with a Th17 signature in cancer patients with previous SARS-CoV-2 infection followed by mRNA vaccination
(MDPI, 2022) Echaide Górriz, Míriam; Labiano, Ibone; Delgado, Marina; Fernández de Lascoiti, Ángela; Ochoa, Patricia; Garnica, Maider; Ramos, Pablo; Chocarro de Erauso, Luisa; Fernández Rubio, Leticia; Arasanz Esteban, Hugo; Bocanegra Gondán, Ana Isabel; Blanco, Ester; Piñeiro Hermida, Sergio; Morente Sancho, Pilar; Vera García, Ruth; Alsina, María; Escors Murugarren, David; Kochan, Grazyna; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
It is unclear whether patients with cancer present inherently impaired responses to COVID-19 and vaccination due to their treatments, neoplastic diseases or both. To address this question, immune profiling was performed in three cohorts of healthy donors and oncologic patients: infected with SARS-CoV-2, BNT162b2-vaccinated, and with previous COVID-19 disease and subsequently vaccinated. Cancer patients showed good antibody responses to vaccination, but poor induction of T-cell responses towards the S protein when compared to infection. Following natural infection, the major targets for T-cells were the SARS-CoV-2 structural proteins M and S, but not the N protein. Similar to antibody titers, the T-cell responses quickly decayed after six months post-vaccination. Significant memory T-cell expansion was observed in vaccinated donors only if previously diagnosed with COVID-19 before undergoing vaccination. Oncologic patients with previous COVID-19 followed by vaccination exhibited potent IL-17+ CD4 and CD8 T-cell responses and elevated numbers of circulating neutrophils in peripheral blood. © 2022 by the authors.
Open Access
Systemic blood immune cell populations as biomarkers for the outcome of immune checkpoint inhibitor therapies
(MDPI, 2020) Hernández, Carlos; Arasanz Esteban, Hugo; Chocarro de Erauso, Luisa; Bocanegra Gondán, Ana Isabel; Zuazo Ibarra, Miren; Ciencias de la Salud; Osasun Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa; Gobierno de Navarra / Nafarroako Gobernua
The development of cancer immunotherapy in the last decade has followed a vertiginous rhythm. Nowadays, immune checkpoint inhibitors (ICI) which include anti-CTLA4, anti-PD-1 and anti-PD-L1 antibodies are in clinical use for the treatment of numerous cancers. However, approximately only a third of the patients benefit from ICI therapies. Many efforts have been made for the identification of biomarkers allowing patient stratification into potential responders and progressors before the start of ICI therapies or for monitoring responses during treatment. While much attention is centered on biomarkers from the tumor microenvironment, in many cases biopsies are not available. The identification of systemic immune cell subsets that correlate with responses could provide promising biomarkers. Some of them have been reported to influence the response to ICI therapies, such as proliferation and activation status of CD8 and CD4 T cells, the expression of immune checkpoints in peripheral blood cells and the relative numbers of immunosuppressive cells such as regulatory T cells and myeloid-derived suppressor cells. In addition, the profile of soluble factors in plasma samples could be associated to response or tumor progression. Here we will review the cellular subsets associated to response or progression in different studies and discuss their accuracy in diagnosis.
Open Access
Understanding LAG-3 Signaling
(MDPI, 2021) Chocarro de Erauso, Luisa; Blanco, Ester; Zuazo Ibarra, Miren; Arasanz Esteban, Hugo; Bocanegra Gondán, Ana Isabel; Fernández Rubio, Leticia; Morente Sancho, Pilar; Fernández Hinojal, Gonzalo; Echaide Górriz, Míriam; Garnica, Maider; Ramos, Pablo; Vera García, Ruth; Kochan, Grazyna; Escors Murugarren, David; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
Lymphocyte activation gene 3 (LAG-3) is a cell surface inhibitory receptor with multiple biological activities over T cell activation and effector functions. LAG-3 plays a regulatory role in immunity and emerged some time ago as an inhibitory immune checkpoint molecule comparable to PD-1 and CTLA-4 and a potential target for enhancing anti-cancer immune responses. LAG-3 is the third inhibitory receptor to be exploited in human anti-cancer immunotherapies, and it is considered a potential next-generation cancer immunotherapy target in human therapy, right next to PD-1 and CTLA-4. Unlike PD-1 and CTLA-4, the exact mechanisms of action of LAG-3 and its relationship with other immune checkpoint molecules remain poorly understood. This is partly caused by the presence of non-conventional signaling motifs in its intracellular domain that are different from other conventional immunoregulatory signaling motifs but with similar inhibitory activities. Here we summarize the current understanding of LAG-3 signaling and its role in LAG-3 functions, from its mechanisms of action to clinical applications.
Open Access
Leading edge: intratumor delivery of monoclonal antibodies for the treatment of solid tumors
(MDPI, 2023) Blanco, Ester; Chocarro de Erauso, Luisa; Fernández Rubio, Leticia; Bocanegra Gondán, Ana Isabel; Arasanz Esteban, Hugo; Echaide Górriz, Míriam; Garnica, Maider; Piñeiro Hermida, Sergio; Kochan, Grazyna; Escors Murugarren, David; Ciencias de la Salud; Osasun Zientziak
Immunotherapies based on immune checkpoint blockade have shown remarkable clinical outcomes and durable responses in patients with many tumor types. Nevertheless, these therapies lack efficacy in most cancer patients, even causing severe adverse events in a small subset of patients, such as inflammatory disorders and hyper-progressive disease. To diminish the risk of developing serious toxicities, intratumor delivery of monoclonal antibodies could be a solution. Encouraging results have been shown in both preclinical and clinical studies. Thus, intratumor immunotherapy as a new strategy may retain efficacy while increasing safety. This approach is still an exploratory frontier in cancer research and opens up new possibilities for next-generation personalized medicine. Local intratumor delivery can be achieved through many means, but an attractive approach is the use of gene therapy vectors expressing mAbs inside the tumor mass. Here, we summarize basic, translational, and clinical results of intratumor mAb delivery, together with descriptions of non-viral and viral strategies for mAb delivery in preclinical and clinical development. Currently, this is an expanding research subject that will surely play a key role in the future of oncology.