RESUMO
Cancer immunotherapies are increasingly combined with targeted therapies to improve therapeutic outcomes. We show that combination of agonistic anti-CD40 with antiangiogenic antibodies targeting 2 proangiogenic factors, vascular endothelial growth factor A (VEGFA) and angiopoietin 2 (Ang2/ANGPT2), induces pleiotropic immune mechanisms that facilitate tumor rejection in several tumor models. On the one hand, VEGFA/Ang2 blockade induced regression of the tumor microvasculature while decreasing the proportion of nonperfused vessels and reducing leakiness of the remaining vessels. On the other hand, both anti-VEGFA/Ang2 and anti-CD40 independently promoted proinflammatory macrophage skewing and increased dendritic cell activation in the tumor microenvironment, which were further amplified upon combination of the 2 treatments. Finally, combined therapy provoked brisk infiltration and intratumoral redistribution of cytotoxic CD8+ T cells in the tumors, which was mainly driven by Ang2 blockade. Overall, these nonredundant synergistic mechanisms endowed T cells with improved effector functions that were conducive to more efficient tumor control, underscoring the therapeutic potential of antiangiogenic immunotherapy in cancer.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Antígenos CD40/agonistas , Neoplasias/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Microambiente Tumoral/efeitos dos fármacos , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Angiopoietina-2/antagonistas & inibidores , Angiopoietina-2/metabolismo , Animais , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígenos CD40/imunologia , Linhagem Celular Tumoral/transplante , Modelos Animais de Doenças , Sinergismo Farmacológico , Feminino , Humanos , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Camundongos , Neoplasias/irrigação sanguínea , Neoplasias/imunologia , Neoplasias/patologia , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Microambiente Tumoral/imunologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Antiangiogenic and cytotoxic effects are considered the principal mechanisms of action of sorafenib, a multitarget kinase inhibitor approved for the treatment of hepatocellular carcinoma (HCC). We report that sorafenib also acts through direct immune modulation, indispensable for its antitumor activity. In vivo cell depletion experiments in two orthotopic HCC mouse models as well as in vitro analysis identified macrophages (MΦ) as the key mediators of the antitumoral effect and demonstrate a strong interdependency of MΦ and natural killer (NK) cells for efficient tumor cell killing. Caspase 1 analysis in sorafenib-treated MΦ revealed an induction of pyroptosis. As a result, cytotoxic NK cells become activated when cocultured with sorafenib-treated MΦ, leading to tumor cell death. In addition, sorafenib was found to down-regulate major histocompatibility complex class I expression of tumor cells, which may reduce the tumor responsiveness to immune checkpoint therapies and favor NK-cell response. In vivo cytokine blocking revealed that sorafenib efficacy is abrogated after inhibition of interleukins 1B and 18. Conclusion: We report an immunomodulatory mechanism of sorafenib involving MΦ pyroptosis and unleashing of an NK-cell response that sets it apart from other spectrum kinase inhibitors as a promising immunotherapy combination partner for the treatment of HCC.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Células Matadoras Naturais/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Piroptose/efeitos dos fármacos , Sorafenibe/farmacologia , Análise de Variância , Animais , Carcinoma Hepatocelular/patologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Humanos , Injeções Intravenosas , Neoplasias Hepáticas/patologia , Macrófagos , Camundongos , Camundongos Transgênicos , Distribuição Aleatória , Carga Tumoral/efeitos dos fármacos , Células Tumorais Cultivadas , Microtomografia por Raio-X/métodos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Immunotherapy with checkpoint inhibitors has proved to be highly effective, with durable responses in a subset of patients. Given their encouraging clinical activity, checkpoint inhibitors are increasingly being tested in clinical trials in combination with chemotherapy. In many instances, there is little understanding of how chemotherapy might influence the quality of the immune response generated by checkpoint inhibitors. In this study, we evaluated the impact of chemotherapy alone or in combination with anti-PD-L1 in a responsive syngeneic tumor model. Although multiple classes of chemotherapy treatment reduced immune cell numbers and activity in peripheral tissues, chemotherapy did not antagonize but in many cases augmented the antitumor activity mediated by anti-PD-L1. This dichotomy between the detrimental effects in peripheral tissues and enhanced antitumor activity was largely explained by the reduced dependence on incoming cells for antitumor efficacy in already established tumors. The effects of the various chemotherapies were also agent specific, and synergy with anti-PD-L1 was achieved by different mechanisms that ultimately helped establish a new threshold for response. These results rationalize the combination of chemotherapy with immunotherapy and suggest that, despite the negative systemic effects of chemotherapy, effective combinations can be obtained through distinct mechanisms acting within the tumor.
Assuntos
Adenocarcinoma/terapia , Anticorpos Monoclonais/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfócitos T CD8-Positivos/efeitos dos fármacos , Neoplasias do Colo/terapia , Imunoterapia/métodos , Adenocarcinoma/imunologia , Animais , Antígenos de Neoplasias/imunologia , Antígeno B7-H1/imunologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Neoplasias do Colo/imunologia , Feminino , Citometria de Fluxo , Humanos , Imunidade Celular , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais , Carga Tumoral/efeitos dos fármacosRESUMO
BACKGROUND: The treatment of melanoma, the deadliest form of skin cancer, has greatly benefited from immunotherapy. However, many patients do not show a durable response, which is only partially explained by known resistance mechanisms. METHODS: We performed single-cell RNA sequencing of tumor immune infiltrates and matched peripheral blood mononuclear cells of 22 checkpoint inhibitor (CPI)-naive stage III-IV metastatic melanoma patients. After sample collection, the same patients received CPI treatment, and their response was assessed. FINDINGS: CPI responders showed high levels of classical monocytes in peripheral blood, which preferentially transitioned toward CXCL9-expressing macrophages in tumors. Trajectories of tumor-infiltrating CD8+ T cells diverged at the level of effector memory/stem-like T cells, with non-responder cells progressing into a state characterized by cellular stress and apoptosis-related gene expression. Consistently, predicted non-responder-enriched myeloid-T/natural killer cell interactions were primarily immunosuppressive, while responder-enriched interactions were supportive of T cell priming and effector function. CONCLUSIONS: Our study illustrates that the tumor immune microenvironment prior to CPI treatment can be indicative of response. In perspective, modulating the myeloid and/or effector cell compartment by altering the described cell interactions and transitions could improve immunotherapy response. FUNDING: This research was funded by Roche Pharma Research and Early Development.
Assuntos
Inibidores de Checkpoint Imunológico , Melanoma , Neoplasias Cutâneas , Microambiente Tumoral , Humanos , Melanoma/tratamento farmacológico , Melanoma/imunologia , Melanoma/patologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/farmacologia , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Masculino , Feminino , Células Mieloides/imunologia , Células Mieloides/efeitos dos fármacos , Células Mieloides/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/metabolismo , Pessoa de Meia-Idade , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/efeitos dos fármacosRESUMO
Cancer vaccines aim to induce CTL responses against tumors. Challenges for vaccine design are targeting Ag to dendritic cells (DCs) in vivo, facilitating cross-presentation, and conditioning the microenvironment for Th1 type immune responses. In this study, we report that ISCOM vaccines, which consist of ISCOMATRIX adjuvant and protein Ag, meet these challenges. Subcutaneous injection of an ISCOM vaccine in mice led to a substantial influx and activation of innate and adaptive immune effector cells in vaccine site-draining lymph nodes (VDLNs) as well as IFN-γ production by NK and NKT cells. Moreover, an ISCOM vaccine containing the model Ag OVA (OVA/ISCOM vaccine) was efficiently taken up by CD8α(+) DCs in VDLNs and induced their maturation and IL-12 production. Adoptive transfer of transgenic OT-I T cells revealed highly efficient cross-presentation of the OVA/ISCOM vaccine in vivo, whereas cross-presentation of soluble OVA was poor even at a 100-fold higher concentration. Cross-presenting activity was restricted to CD8α(+) DCs in VDLNs, whereas Langerin(+) DCs and CD8α(-) DCs were dispensable. Remarkably, compared with other adjuvant systems, the OVA/ISCOM vaccine induced a high frequency of OVA-specific CTLs capable of tumor cell killing in different tumor models. Thus, ISCOM vaccines combine potent immune activation with Ag delivery to CD8α(+) DCs in vivo for efficient induction of CTL responses.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Linfócitos T CD8-Positivos/imunologia , Colesterol/administração & dosagem , Apresentação Cruzada/imunologia , Testes Imunológicos de Citotoxicidade/métodos , Células Dendríticas/imunologia , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Fosfolipídeos/administração & dosagem , Saponinas/administração & dosagem , Animais , Linfócitos T CD8-Positivos/metabolismo , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Linhagem Celular Tumoral , Células Cultivadas , Colesterol/imunologia , Células Dendríticas/metabolismo , Combinação de Medicamentos , Feminino , Técnicas de Introdução de Genes , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/patologia , Linfonodos/imunologia , Linfonodos/patologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Melanoma Experimental/prevenção & controle , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células T Matadoras Naturais/imunologia , Células T Matadoras Naturais/metabolismo , Células T Matadoras Naturais/patologia , Fragmentos de Peptídeos/administração & dosagem , Fragmentos de Peptídeos/imunologia , Fosfolipídeos/imunologia , Quillaja/imunologia , Saponinas/imunologiaRESUMO
Granzymes A and B (GrAB) are known principally for their role in mediating perforin-dependent death of virus-infected or malignant cells targeted by CTL. In this study, we show that granzymes also play a critical role as inducers of Ag cross-presentation by dendritic cells (DC). This was demonstrated by the markedly reduced priming of naive CD8(+) T cells specific for the model Ag OVA both in vitro and in vivo in response to tumor cells killed in the absence of granzymes. Reduced cross-priming was due to impairment of phagocytosis of tumor cell corpses by CD8α(+) DC but not CD8α(-) DC, demonstrating the importance of granzymes in inducing the exposure of prophagocytic "eat-me" signals on the dying target cell. Our data reveal a critical and previously unsuspected role for granzymes A and B in dictating immunogenicity by influencing the mode of tumor cell death and indicate that granzymes contribute to the efficient generation of immune effector pathways in addition to their well-known role in apoptosis induction.
Assuntos
Antígenos de Neoplasias/metabolismo , Apresentação Cruzada/imunologia , Granzimas/fisiologia , Fagocitose/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Animais , Antígenos de Neoplasias/imunologia , Morte Celular/imunologia , Linhagem Celular Tumoral , Galinhas , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Granzimas/deficiência , Granzimas/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Ovalbumina/toxicidade , Fragmentos de Peptídeos/toxicidade , Linfócitos T Citotóxicos/enzimologiaRESUMO
Agonistic αCD40 therapy has been shown to inhibit cancer progression in only a fraction of patients. Understanding the cancer cell-intrinsic and microenvironmental determinants of αCD40 therapy response is therefore crucial to identify responsive patient populations and to design efficient combinatorial treatments. Here, we show that the therapeutic efficacy of αCD40 in subcutaneous melanoma relies on preexisting, type 1 classical dendritic cell (cDC1)-primed CD8+ T cells. However, after administration of αCD40, cDC1s were dispensable for antitumor efficacy. Instead, the abundance of activated cDCs, potentially derived from cDC2 cells, increased and further activated antitumor CD8+ T cells. Hence, distinct cDC subsets contributed to the induction of αCD40 responses. In contrast, lung carcinomas, characterized by a high abundance of macrophages, were resistant to αCD40 therapy. Combining αCD40 therapy with macrophage depletion led to tumor growth inhibition only in the presence of strong neoantigens. Accordingly, treatment with immunogenic cell death-inducing chemotherapy sensitized lung tumors to αCD40 therapy in subcutaneous and orthotopic settings. These insights into the microenvironmental regulators of response to αCD40 suggest that different tumor types would benefit from different combinations of therapies to optimize the clinical application of CD40 agonists. SIGNIFICANCE: This work highlights the temporal roles of different dendritic cell subsets in promoting CD8+ T-cell-driven responses to CD40 agonist therapy in cancer.
Assuntos
Antígenos CD40 , Células Dendríticas , Macrófagos , Neoplasias , Animais , Antígenos CD40/agonistas , Linfócitos T CD8-Positivos , Células Dendríticas/metabolismo , Humanos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/metabolismoRESUMO
Numerous studies have been performed in vitro and in various animal models to modulate the interaction of dendritic cells (DC) and T cells by Fas (CD95/Apo-1) signalling to delete activated T cells via induction of activation-induced cell death (AICD). Previously, we could demonstrate that Fas ligand (FasL/CD95L)-expressing 'killer-antigen-presenting cells' can be generated from human monocyte-derived mature DC (mDC) using adenoviral gene transfer. To evaluate whether these FasL-expressing mDC (mDC-FasL) could eliminate alloreactive primary human T cells in vitro, co-culture experiments were performed. Proliferation of human T cells was markedly reduced in primary co-cultures with allogeneic mDC-FasL, whereas a strong proliferative T-cell response could be observed in co-cultures with enhanced green fluorescent protein-transduced mDC. Inhibition of T-cell proliferation was related to the transduction efficiency, and the numbers of mDC-FasL present in co-cultures. In addition, proliferation of pre-activated alloreactive CD4(+) and CD8(+) T cells could be almost completely inhibited in secondary co-cultures using mDC-FasL as stimulatory cells, which was the result of induction of apoptosis in the majority of preactivated T cells. The specific deletion of alloreactive T cells by mDC-FasL was confirmed by an unaffected proliferative response of surviving T cells towards allogeneic 'third-party' peripheral blood mononuclear cells in a third stimulation, or upon unspecific stimulation with anti-CD3/CD28 beads. The results of this study demonstrate that allospecifically activated T cells are efficiently eliminated by mDC-FasL, supporting further investigations to apply FasL-expressing 'killer-DC' as a novel strategy for the treatment of allograft rejection.
Assuntos
Células Dendríticas/imunologia , Proteína Ligante Fas/imunologia , Isoantígenos/imunologia , Ativação Linfocitária/imunologia , Linfócitos T/imunologia , Diferenciação Celular , Proliferação de Células , Separação Celular , Técnicas de Cocultura , Células Dendríticas/citologia , Citometria de Fluxo , Humanos , Teste de Cultura Mista de Linfócitos , Monócitos/citologia , Monócitos/imunologia , Transdução GenéticaRESUMO
A sensitive, selective, and comprehensive method for the quantitative determination of tryptophan and 18 of its key metabolites in serum, urine, and cell culture supernatants was developed. The analytes were separated on a C18 silica column by reversed-phase liquid chromatography and detected by electrospray ionization tandem mass spectrometry in positive ion multiple reaction monitoring (MRM) mode, except for indoxyl sulfate which was measured in negative ion MRM mode in a separate run. The limits of detection and lower limits of quantification were in the range of 0.1-50 and 0.5-100 nM, respectively. Fully (13)C isotope-labeled and deuterated internal standards were used to achieve accurate quantification. The applicability of the method to analyze serum, urine, and cell culture supernatants was demonstrated by recovery experiments and the evaluation of matrix effects. Precision for the analysis of serum, urine, and cell culture supernatants ranged between 1.3% and 16.0%, 1.5% and 13.5%, and 1.0% and 17.4%, respectively. The method was applied to analyze changes in tryptophan metabolism in cell culture supernatants from IFN-γ-treated monocytes and immature or mature dendritic cells.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas em Tandem/métodos , Triptofano/sangue , Triptofano/urina , Linhagem Celular , Meios de Cultura/química , Células Dendríticas/química , Células Dendríticas/metabolismo , Humanos , Monócitos/química , Monócitos/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Triptofano/metabolismoRESUMO
Colony-stimulating factor 1 receptor (CSF1R) blockade abates tumor-associated macrophage (TAM) infiltrates and provides marked clinical benefits in diffuse-type tenosynovial giant cell tumors. However, facial edema is a common adverse event associated with TAM elimination in patients. In this study, we examined molecular and cellular events associated with edema formation in mice and human patients with cancer treated with a CSF1R blocking antibody. Extended antibody treatment of mice caused marked body weight gain, an indicator of enhanced body fluid retention. This was associated with an increase of extracellular matrix-remodeling metalloproteinases (MMPs), namely MMP2 and MMP3, and enhanced deposition of hyaluronan (HA) and proteoglycans, leading to skin thickening. Discontinuation of anti-CSF1R treatment or blockade of MMP activity restored unaltered body weight and normal skin morphology in the mice. In patients, edema developed at doses well below the established optimal biological dose for emactuzumab, a CSF1R dimerization inhibitor. Patients who developed edema in response to emactuzumab had elevated HA in peripheral blood. Our findings indicate that an early increase of peripheral HA can serve as a pharmacodynamic marker for edema development and suggest potential interventions based on MMP inhibition for relieving periorbital edema in patients treated with CSF1R inhibitors.
Assuntos
Edema , Macrófagos , Neoplasias , Peptídeo Hidrolases , Proteoglicanas , Animais , Anticorpos Monoclonais Humanizados/uso terapêutico , Humanos , Camundongos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidoresRESUMO
The pore-forming protein perforin is synthesized as an inactive precursor in natural killer (NK) cells and cytotoxic T lymphocytes (CTLs), and becomes active when a short C-terminal peptide is cleaved within acidic lysosome-like cytotoxic granules. Although it was shown more than a decade ago that this cleavage is pH dependent and can be inhibited by the generic cysteine cathepsin inhibitor E-64d, no protease capable of processing the perforin C terminus has been identified. Neither is it known whether a single protease is responsible or the processing has inbuilt redundancy. Here, we show that incubation of human NK cells and primary antigen-restricted mouse CTLs with the cathepsin L (CatL) inhibitor L1 resulted in a marked inhibition of perforin-dependent target cell death and reduced perforin processing. In vitro, CatL preferentially cleaved a site on full-length recombinant perforin close to its C terminus. The NK cells of mice deficient in CatL showed a reduction but not a complete absence of processed perforin, indicating that cysteine proteases other than CatL are also able to process perforin. We conclude that granule-bound cathepsins are essential for processing perforin to its active form, and that CatL is an important, but not exclusive, participant in this process.
Assuntos
Catepsina L/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Animais , Biocatálise/efeitos dos fármacos , Catepsina L/antagonistas & inibidores , Catepsina L/genética , Linhagem Celular Tumoral , Inibidores de Cisteína Proteinase/farmacologia , Citotoxicidade Imunológica/efeitos dos fármacos , Citotoxicidade Imunológica/genética , Citotoxicidade Imunológica/imunologia , Ácido Egtázico/farmacologia , Granzimas/metabolismo , Humanos , Células K562 , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucina/análogos & derivados , Leucina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Ovalbumina/imunologia , Fragmentos de Peptídeos/imunologia , Peptídeo Hidrolases/metabolismo , Perforina , Proteínas Citotóxicas Formadoras de Poros/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologiaRESUMO
Particular interest to harness the innate immune system for cancer immunotherapy is fueled by limitations of immune checkpoint blockade. Plasmacytoid dendritic cells (pDC) are detected in a variety of solid tumors and correlate with poor clinical outcome. Release of type I interferons in response to toll-like-receptor (TLR)7 and TLR9 activation is the pDC hallmark. Mouse and human pDC differ substantially in their biology concerning surface marker expression and cytokine production. Here, we employed humanized mouse models (HIS) to study pDC function. We performed a comprehensive characterization of transgenic, myeloid-enhanced mouse strains (NOG-EXL and NSG-SGM3) expressing human interleukin-3 (hIL-3) and granulocyte-macrophage colony stimulating factor (GM-CSF) using identical humanization protocols. Only in HIS-NOG-EXL mice sufficient pDC infiltration was detectable. Therefore, we selected this strain for subsequent tumor studies. We analyzed pDC frequency in peripheral blood and tumors by comparing HIS-NOG-EXL with HIS-NOG mice bearing three different ovarian and breast tumors. Despite the substantially increased pDC numbers in peripheral blood of HIS-NOG-EXL mice, we detected TLR7/8 agonist responsive and thus functional pDCs only in certain tumor models independent of the mouse strain employed. However, HIS-NOG-EXL mice showed in general a superior humanization phenotype characterized by reconstitution of different myeloid subsets, NK cells and B cells producing physiologic IgG levels. Hence, we provide first evidence that the tumor milieu but not genetically introduced cytokines defines intratumoral (i.t.) frequencies of the rare pDC subset. This study provides model systems to investigate in vivo pro- and anti-tumoral human pDC functions.
Assuntos
Linfócitos B/imunologia , Carcinoma Epitelial do Ovário/imunologia , Células Dendríticas/imunologia , Células Matadoras Naturais/imunologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Interleucina-3/genética , Camundongos , Camundongos SCID , Camundongos Transgênicos , Microambiente TumoralRESUMO
BACKGROUND: This phase Ib study evaluated the safety, clinical activity, pharmacokinetics, and pharmacodynamics (PD) of emactuzumab (anti-colony stimulating factor 1 receptor monoclonal antibody (mAb)) in combination with selicrelumab (agonistic cluster of differentiation 40 mAb) in patients with advanced solid tumors. METHODS: Both emactuzumab and selicrelumab were administered intravenously every 3 weeks and doses were concomitantly escalated (emactuzumab: 500 to 1000 mg flat; selicrelumab: 2 to 16 mg flat). Dose escalation was conducted using the product of independent beta probabilities dose-escalation design. PD analyzes were performed on peripheral blood samples and tumor/skin biopsies at baseline and on treatment. Clinical activity was evaluated using investigator-based and Response Evaluation Criteria In Solid Tumors V.1.1-based tumor assessments. RESULTS: Three dose-limiting toxicities (all infusion-related reactions (IRRs)) were observed at 8, 12 and 16 mg of selicrelumab together with 1000 mg of emactuzumab. The maximum tolerated dose was not reached at the predefined top doses of emactuzumab (1000 mg) and selicrelumab (16 mg). The most common adverse events were IRRs (75.7%), fatigue (54.1%), facial edema (37.8%), and increase in aspartate aminotransferase and creatinine phosphokinase (35.1% both). PD analyzes demonstrated an increase of Ki67+-activated CD8+ T cells accompanied by a decrease of B cells and the reduction of CD14Dim CD16bright monocytes in peripheral blood. The best objective clinical response was stable disease in 40.5% of patients. CONCLUSION: Emactuzumab in combination with selicrelumab demonstrated a manageable safety profile and evidence of PD activity but did not translate into objective clinical responses. TRIALREGISTRATION NUMBER: NCT02760797.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígenos CD40/metabolismo , Neoplasias/tratamento farmacológico , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Anticorpos Monoclonais Humanizados/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Feminino , Humanos , Masculino , Neoplasias/imunologia , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismoRESUMO
Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide and has a high mortality rate due to limited treatment options. Hence, the response of HCC to different cancer immunotherapies is being intensively investigated in clinical trials. Immune checkpoint blockers (ICB) show promising results, albeit for a minority of HCC patients. Mouse models are commonly used to evaluate new therapeutic agents or regimens. However, to make clinical translation more successful, better characterized preclinical models are required. We therefore extensively investigated two immune-competent orthotopic HCC mouse models, namely transplanted Hep-55.1c and transgenic iAST, with respect to morphological, immunological and genetic traits and evaluated both models' responsiveness to immunotherapies. Hep-55.1c tumors were characterized by rich fibrous stroma, high mutational load and pronounced immune cell infiltrates, all of which are features of immune-responsive tumors. These characteristics were less distinct in iAST tumors, though these were highly vascularized. Cell depletion revealed that CD8+ T cells from iAST mice do not affect tumor growth and are tumor tolerant. This corresponds to the failure of single and combined ICB targeting PD-1 and CTLA-4. In contrast, combining anti-PD-1 and anti-CTLA-4 showed significant antitumor efficacy in the Hep-55.1c mouse model. Collectively, our data comprehensively characterize two immune-competent HCC mouse models representing ICB responsive and refractory characteristics. Our characterization confirms these models to be suitable for preclinical investigation of novel cancer immunotherapy approaches that aim to either deepen preexisting immune responses or generate de novo immunity against the tumor.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Modelos Animais de Doenças , Neoplasias Hepáticas/tratamento farmacológico , Animais , Antígenos Transformantes de Poliomavirus/genética , Antineoplásicos Imunológicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/antagonistas & inibidores , Antígeno CTLA-4/imunologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/imunologia , Linhagem Celular Tumoral/transplante , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Feminino , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Resultado do Tratamento , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologiaRESUMO
Despite decades of research, cancer remains a devastating disease and new treatment options are needed. Today cancer is acknowledged as a multifactorial disease not only comprising of aberrant tumor cells but also the associated stroma including tumor vasculature, fibrotic plaques, and immune cells that interact in a complex heterotypic interplay. Myeloid cells represent one of the most abundant immune cell population within the tumor stroma and are equipped with a broad functional repertoire that promotes tumor growth by suppressing cytotoxic T cell activity, stimulating neoangiogenesis and tissue remodeling. Therefore, myeloid cells have become an attractive target for pharmacological intervention. In this review, we summarize the pharmacological approaches to therapeutically target tumor-associated myeloid cells with a focus on advanced programs that are clinically evaluated. In addition, for each therapeutic strategy, the preclinical rationale as well as advantages and challenges from a drug development perspective are discussed.
Assuntos
Antineoplásicos/farmacologia , Desenvolvimento de Medicamentos , Células Mieloides/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Antineoplásicos/síntese química , Antineoplásicos/química , Humanos , Neoplasias/metabolismo , Neoplasias/patologiaRESUMO
Colony-stimulating factor 1 (CSF1) is a key regulator of monocyte/macrophage differentiation that sustains the protumorigenic functions of tumor-associated macrophages (TAMs). We show that CSF1 is expressed in human melanoma, and patients with metastatic melanoma have increased CSF1 in blood compared to healthy subjects. In tumors, CSF1 expression correlated with the abundance of CD8+ T cells and CD163+ TAMs. Human melanoma cell lines consistently produced CSF1 after exposure to melanoma-specific CD8+ T cells or T cell-derived cytokines in vitro, reflecting a broadly conserved mechanism of CSF1 induction by activated CD8+ T cells. Mining of publicly available transcriptomic data sets suggested co-enrichment of CD8+ T cells with CSF1 or various TAM-specific markers in human melanoma, which was associated with nonresponsiveness to programmed cell death protein 1 (PD1) checkpoint blockade in a smaller patient cohort. Combination of anti-PD1 and anti-CSF1 receptor (CSF1R) antibodies induced the regression of BRAFV600E -driven, transplant mouse melanomas, a result that was dependent on the effective elimination of TAMs. Collectively, these data implicate CSF1 induction as a CD8+ T cell-dependent adaptive resistance mechanism and show that simultaneous CSF1R targeting may be beneficial in melanomas refractory to immune checkpoint blockade and, possibly, other T cell-based therapies.
Assuntos
Fator Estimulador de Colônias de Macrófagos/sangue , Melanoma/sangue , Melanoma/patologia , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Humanos , Macrófagos/metabolismo , Camundongos , Proteínas Proto-Oncogênicas B-raf/metabolismo , Receptores de Superfície Celular/metabolismo , Transdução de SinaisRESUMO
Depletion of immunosuppressive tumor-associated macrophages (TAMs) or reprogramming toward a proinflammatory activation state represent different strategies to therapeutically target this abundant myeloid population. In this study, we report that inhibition of colony-stimulating factor-1 receptor (CSF-1R) signaling sensitizes TAMs to profound and rapid reprogramming in the presence of a CD40 agonist before their depletion. Despite the short-lived nature of macrophage hyperactivation, combined CSF-1R+CD40 stimulation of macrophages is sufficient to create a proinflammatory tumor milieu that reinvigorates an effective T cell response in transplanted tumors that are either responsive or insensitive to immune checkpoint blockade. The central role of macrophages in regulating preexisting immunity is substantiated by depletion experiments, transcriptome analysis of ex vivo sorted TAMs, and gene expression profiling of whole tumor lysates at an early treatment time point. This approach enabled the identification of specific combination-induced changes among the pleiotropic activation spectrum of the CD40 agonist. In patients, CD40 expression on human TAMs was detected in mesothelioma and colorectal adenocarcinoma.
Assuntos
Imunidade , Macrófagos/imunologia , Neoplasias/imunologia , Neoplasias/patologia , Animais , Antígenos CD40/agonistas , Antígenos CD40/metabolismo , Linfócitos T CD8-Positivos/imunologia , Feminino , Humanos , Inflamação/patologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Modelos Biológicos , Fenótipo , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismoRESUMO
Tumour-associated macrophages (TAMs) largely express an alternatively activated (or M2) phenotype, which entails immunosuppressive and tumour-promoting capabilities. Reprogramming TAMs towards a classically activated (M1) phenotype may thwart tumour-associated immunosuppression and unleash anti-tumour immunity. Here we show that conditional deletion of the microRNA (miRNA)-processing enzyme DICER in macrophages prompts M1-like TAM programming, characterized by hyperactive IFN-γ/STAT1 signalling. This rewiring abated the immunosuppressive capacity of TAMs and fostered the recruitment of activated cytotoxic T lymphocytes (CTLs) to the tumours. CTL-derived IFN-γ exacerbated M1 polarization of Dicer1-deficient TAMs and inhibited tumour growth. Remarkably, DICER deficiency in TAMs negated the anti-tumoral effects of macrophage depletion by anti-CSF1R antibodies, and enabled complete tumour eradication by PD1 checkpoint blockade or CD40 agonistic antibodies. Finally, genetic rescue of Let-7 miRNA activity in Dicer1-deficient TAMs partly restored their M2-like phenotype and decreased tumour-infiltrating CTLs. These findings suggest that DICER/Let-7 activity opposes IFN-γ-induced, immunostimulatory M1-like TAM activation, with potential therapeutic implications.
Assuntos
RNA Helicases DEAD-box/metabolismo , Interferon gama/metabolismo , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , MicroRNAs/genética , Ribonuclease III/metabolismo , Microambiente Tumoral/genética , Animais , Células Cultivadas , RNA Helicases DEAD-box/deficiência , Humanos , Camundongos , Neoplasias/genética , Neoplasias/imunologia , Ribonuclease III/deficiênciaRESUMO
Effective cancer immunotherapy requires overcoming immunosuppressive tumor microenvironments. We found that local nitric oxide (NO) production by tumor-infiltrating myeloid cells is important for adoptively transferred CD8(+) cytotoxic T cells to destroy tumors. These myeloid cells are phenotypically similar to inducible nitric oxide synthase (NOS2)- and tumor necrosis factor (TNF)-producing dendritic cells (DC), or Tip-DCs. Depletion of immunosuppressive, colony stimulating factor 1 receptor (CSF-1R)-dependent arginase 1(+) myeloid cells enhanced NO-dependent tumor killing. Tumor elimination via NOS2 required the CD40-CD40L pathway. We also uncovered a strong correlation between survival of colorectal cancer patients and NOS2, CD40, and TNF expression in their tumors. Our results identify a network of pro-tumor factors that can be targeted to boost cancer immunotherapies.
Assuntos
Células Dendríticas/imunologia , Imunoterapia Adotiva/métodos , Neoplasias/imunologia , Neoplasias/terapia , Óxido Nítrico Sintase Tipo II/imunologia , Linfócitos T Citotóxicos/imunologia , Fator de Necrose Tumoral alfa/imunologia , Animais , Arginase/biossíntese , Arginase/imunologia , Antígenos CD40/imunologia , Ligante de CD40/imunologia , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/biossíntese , Microambiente Tumoral , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Macrophage infiltration has been identified as an independent poor prognostic factor for several cancer entities. In mouse tumor models macrophages orchestrate various tumor-promoting processes. This observation sparked an interest to therapeutically target these plastic innate immune cells. To date, blockade of colony stimulating factor-1 or its receptor represents the only truly selective approach to manipulate macrophages in cancer patients. Here, we discuss the currently available information on efficacy and safety of various CSF-1/CSF-1R inhibitors in cancer patients and highlight potential combination partners emerging from preclinical studies while considering the differences between mouse and human macrophage biology.