The liver, liver metastasis and liver cancer: A special case for immunotherapy with cytokines and immunostimulatory monoclonal antibodies

"The immunologic outcome of liver transplantations tells us that this organ is somehow specialized as a tolerogenic organ under steady state conditions."Fil: Mazzolini Rizzo, Guillermo Daniel. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ochoa, María C. Universidad de Navarra; EspañaFil: Morales Kastresana, Aizea. Universidad de Navarra; EspañaFil: Sanmamed, Miguel F.. Universidad de Navarra; EspañaFil: Melero, Ignacio. Universidad de Navarra; Españ

Cancer immunotherapy with immunomodulatory anti-CD137 and anti-PD-1 monoclonal antibodies requires Batf3-dependent dendritic cells

Weak and ineffective antitumor cytotoxic T lymphocyte (CTL) responses can be rescued by immunomodulatory mAbs targeting PD-1 or CD137. Using Batf3−/− mice, which are defective for cross-presentation of cell-associated antigens, we show that BATF3-dependent dendritic cells (DC) are essential for the response to therapy with anti-CD137 or anti–PD-1 mAbs. Batf3−/− mice failed to prime an endogenous CTL-mediated immune response toward tumor-associated antigens, including neoantigens. As a result, the immunomodulatory mAbs could not amplify any therapeutically functional immune response in these mice. Moreover, administration of systemic sFLT3L and local poly-ICLC enhanced DC-mediated cross-priming and synergized with anti–CD137- and anti–PD-1–mediated immunostimulation in tumor therapy against B16-ovalbumin–derived melanomas, whereas this function was lost in Batf3−/− mice. These experiments show that cross-priming of tumor antigens by FLT3L- and BATF3-dependent DCs is crucial to the efficacy of immunostimulatory mAbs and represents a very attractive point of intervention to enhance their clinical antitumor effects

Dendritic Cells Take up and Present Antigens from Viable and Apoptotic Polymorphonuclear Leukocytes

Dendritic cells (DC) are endowed with the ability to cross-present antigens from other cell types to cognate T cells. DC are poised to meet polymorphonuclear leukocytes (PMNs) as a result of being co-attracted by interleukin-8 (IL-8), for instance as produced by tumor cells or infected tissue. Human monocyte-derived and mouse bone marrow-derived DC can readily internalize viable or UV-irradiated PMNs. Such internalization was abrogated at 4°C and partly inhibited by anti-CD18 mAb. In mice, DC which had internalized PMNs containing electroporated ovalbumin (OVA) protein, were able to cross-present the antigen to CD8 (OT-1) and CD4 (OT-2) TCR-transgenic T cells. Moreover, in humans, tumor cell debris is internalized by PMNs and the tumor-cell material can be subsequently taken up from the immunomagnetically re-isolated PMNs by DC. Importantly, if human neutrophils had endocytosed bacteria, they were able to trigger the maturation program of the DC. Moreover, when mouse PMNs with E. coli in their interior are co-injected in the foot pad with DC, many DC loaded with fluorescent material from the PMNs reach draining lymph nodes. Using CT26 (H-2d) mouse tumor cells, it was observed that if tumor cells are intracellularly loaded with OVA protein and UV-irradiated, they become phagocytic prey of H-2d PMNs. If such PMNs, that cannot present antigens to OT-1 T cells, are immunomagnetically re-isolated and phagocytosed by H-2b DC, such DC productively cross-present OVA antigen determinants to OT-1 T cells. Cross-presentation to adoptively transferred OT-1 lymphocytes at draining lymph nodes also take place when OVA-loaded PMNs (H-2d) are coinjected in the footpad of mice with autologous DC (H-2b). In summary, our results indicate that antigens phagocytosed by short-lived PMNs can be in turn internalized and productively cross-presented by DC

Minimal information for studies of extracellular vesicles 2018 (MISEV2018):a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points

Molecular pathways: hypoxia response in immune cells fighting or promoting cancer

Both malignant and stromal components in tumors are influenced by the physiologic conditions of the microenvironment. Hypoxia is a prominent feature of solid tumors as a result of defective vascularization and intense metabolic activity. The gene-expression control mechanisms that adapt tissues to hypoxia are exploited by tumors to promote angiogenesis and vasculogenesis. The functions of infiltrating immune cells (macrophages and lymphocytes) and other stromal components are also influenced by a limited O(2) supply. Hypoxia-inducible factors (HIF) are the main molecular transcriptional mediators in the hypoxia response. The degradation and activity of HIF-1α and HIF-2α are tightly controlled by the fine-tuned action of oxygen-sensing prolyl and asparaginyl hydroxylase enzymes. Recent evidence indicates that hypoxia can modulate the differentiation and function of T lymphocytes and myeloid cells, skewing their cytokine-production profiles and modifying the expression of costimulatory receptors. This conceivably includes tumor-infiltrating lymphocytes. Hypoxia not only directly affects tumor-infiltrating leukocytes but also exerts effects on tumor cells and vascular cells that indirectly cause selective chemokine-mediated recruitment of suppressive and proangiogenic T-cell subsets. This review focuses on changes induced by hypoxia in immune cells infiltrating solid malignancies. Such changes may either promote or fight cancer, and thus are important for immunotherapy

Molecular pathways: hypoxia response in immune cells fighting or promoting cancer

Both malignant and stromal components in tumors are influenced by the physiologic conditions of the microenvironment. Hypoxia is a prominent feature of solid tumors as a result of defective vascularization and intense metabolic activity. The gene-expression control mechanisms that adapt tissues to hypoxia are exploited by tumors to promote angiogenesis and vasculogenesis. The functions of infiltrating immune cells (macrophages and lymphocytes) and other stromal components are also influenced by a limited O(2) supply. Hypoxia-inducible factors (HIF) are the main molecular transcriptional mediators in the hypoxia response. The degradation and activity of HIF-1α and HIF-2α are tightly controlled by the fine-tuned action of oxygen-sensing prolyl and asparaginyl hydroxylase enzymes. Recent evidence indicates that hypoxia can modulate the differentiation and function of T lymphocytes and myeloid cells, skewing their cytokine-production profiles and modifying the expression of costimulatory receptors. This conceivably includes tumor-infiltrating lymphocytes. Hypoxia not only directly affects tumor-infiltrating leukocytes but also exerts effects on tumor cells and vascular cells that indirectly cause selective chemokine-mediated recruitment of suppressive and proangiogenic T-cell subsets. This review focuses on changes induced by hypoxia in immune cells infiltrating solid malignancies. Such changes may either promote or fight cancer, and thus are important for immunotherapy

Innate functions of immunoglobulin M lessen liver gene transfer with helper-dependent adenovirus.

The immune system poses obstacles to viral vectors, even in the first administration to preimmunized hosts. We have observed that the livers of B cell-deficient mice were more effectively transduced by a helper-dependent adenovirus serotype-5 (HDA) vector than those of WT mice. This effect was T-cell independent as shown in athymic mice. Passive transfer of the serum from adenovirus-naïve WT to Rag1KO mice resulted in a reduction in gene transfer that was traced to IgM purified from serum of adenovirus-naïve mice. To ascribe the gene transfer inhibition activity to either adenoviral antigen-specific or antigen-unspecific functions of IgM, we used a monoclonal IgM antibody of unrelated specificity. Both the polyclonal and the irrelevant monoclonal IgM inhibited gene transfer by the HDA vector to either cultured hepatocellular carcinoma cells or to the liver of mice in vivo. Adsorption of polyclonal or monoclonal IgMs to viral capsids was revealed by ELISAs on adenovirus-coated plates. These observations indicate the existence of an inborn IgM mechanism deployed against a prevalent virus to reduce early post-infection viremia. In conclusion, innate IgM binding to adenovirus serotype-5 capsids restrains gene-transfer and offers a mechanism to be targeted for optimization of vector dosage in gene therapy with HDA vectors

Cancer Immunotherapy with Immunomodulatory Anti-CD137 and Anti-PD-1 Monoclonal Antibodies Requires BATF3-Dependent Dendritic Cells

UNLABELLED: Weak and ineffective antitumor cytotoxic T lymphocyte (CTL) responses can be rescued by immunomodulatory mAbs targeting PD-1 or CD137. Using Batf3(-/-) mice, which are defective for cross-presentation of cell-associated antigens, we show that BATF3-dependent dendritic cells (DC) are essential for the response to therapy with anti-CD137 or anti-PD-1 mAbs. Batf3(-/-) mice failed to prime an endogenous CTL-mediated immune response toward tumor-associated antigens, including neoantigens. As a result, the immunomodulatory mAbs could not amplify any therapeutically functional immune response in these mice. Moreover, administration of systemic sFLT3L and local poly-ICLC enhanced DC-mediated cross-priming and synergized with anti-CD137- and anti-PD-1-mediated immunostimulation in tumor therapy against B16-ovalbumin-derived melanomas, whereas this function was lost in Batf3(-/-) mice. These experiments show that cross-priming of tumor antigens by FLT3L- and BATF3-dependent DCs is crucial to the efficacy of immunostimulatory mAbs and represents a very attractive point of intervention to enhance their clinical antitumor effects. SIGNIFICANCE: Immunotherapy with immunostimulatory mAbs is currently achieving durable clinical responses in different types of cancer. We show that cross-priming of tumor antigens by BATF3-dependent DCs is a key limiting factor that can be exploited to enhance the antitumor efficacy of anti-PD-1 and anti-CD137 immunostimulatory mAbs.Work at the I. Melero lab is funded by MICINN (SAF200803294 and SAF2011-22831), Departamento de salud del Gobierno de Navarra, Redes temáticas de investigación cooperativa RETIC (RD06/0020/0065), and the European commission 7th framework program (ENCITE and IACT). Work in the D. Sancho laboratory is funded by the CNIC and grants from the Spanish Ministry of Economy and Competitiveness (SAF-2013-42920R) and the European Research Council (ERC Starting Independent Researcher Grant 2010, ERC-2010-StG 260414). The CNIC is supported by the Spanish Ministry of Economy and Competitiveness and the Pro-CNIC Foundation. I. Melero and D. Sancho are funded by the European Commission (635122-PROCROP H2020).S