RESUMO
Radiation therapy is a mainstay of cancer treatment but does not always lead to complete tumor regression. Here we combine radiotherapy with blockade of the 'don't-eat-me' cell-surface molecule CD47 in small cell lung cancer (SCLC), a highly metastatic form of lung cancer. CD47 blockade potently enhances the local antitumor effects of radiotherapy in preclinical models of SCLC. Notably, CD47 blockade also stimulates off-target 'abscopal' effects inhibiting non-irradiated SCLC tumors in mice receiving radiation. These abscopal effects are independent of T cells but require macrophages that migrate into non-irradiated tumor sites in response to inflammatory signals produced by radiation and are locally activated by CD47 blockade to phagocytose cancer cells. Similar abscopal antitumor effects were observed in other cancer models treated with radiation and CD47 blockade. The systemic activation of antitumor macrophages following radiotherapy and CD47 blockade may be particularly important in patients with cancer who suffer from metastatic disease.
Assuntos
Neoplasias Pulmonares , Carcinoma de Pequenas Células do Pulmão , Camundongos , Animais , Antígeno CD47 , Macrófagos , Fagocitose , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológicoRESUMO
The disialoganglioside GD2 is overexpressed on several solid tumors, and monoclonal antibodies targeting GD2 have substantially improved outcomes for children with high-risk neuroblastoma. However, approximately 40% of patients with neuroblastoma still relapse, and anti-GD2 has not mediated significant clinical activity in any other GD2+ malignancy. Macrophages are important mediators of anti-tumor immunity, but tumors resist macrophage phagocytosis through expression of the checkpoint molecule CD47, a so-called 'Don't eat me' signal. In this study, we establish potent synergy for the combination of anti-GD2 and anti-CD47 in syngeneic and xenograft mouse models of neuroblastoma, where the combination eradicates tumors, as well as osteosarcoma and small-cell lung cancer, where the combination significantly reduces tumor burden and extends survival. This synergy is driven by two GD2-specific factors that reorient the balance of macrophage activity. Ligation of GD2 on tumor cells (a) causes upregulation of surface calreticulin, a pro-phagocytic 'Eat me' signal that primes cells for removal and (b) interrupts the interaction of GD2 with its newly identified ligand, the inhibitory immunoreceptor Siglec-7. This work credentials the combination of anti-GD2 and anti-CD47 for clinical translation and suggests that CD47 blockade will be most efficacious in combination with monoclonal antibodies that alter additional pro- and anti-phagocytic signals within the tumor microenvironment.
Assuntos
Neoplasias Ósseas , Antígeno CD47 , Animais , Linhagem Celular Tumoral , Humanos , Imunoterapia , Camundongos , Recidiva Local de Neoplasia , Fagocitose , Microambiente TumoralRESUMO
Monoclonal antibody therapies targeting tumour antigens drive cancer cell elimination in large part by triggering macrophage phagocytosis of cancer cells1-7. However, cancer cells evade phagocytosis using mechanisms that are incompletely understood. Here we develop a platform for unbiased identification of factors that impede antibody-dependent cellular phagocytosis (ADCP) using complementary genome-wide CRISPR knockout and overexpression screens in both cancer cells and macrophages. In cancer cells, beyond known factors such as CD47, we identify many regulators of susceptibility to ADCP, including the poorly characterized enzyme adipocyte plasma membrane-associated protein (APMAP). We find that loss of APMAP synergizes with tumour antigen-targeting monoclonal antibodies and/or CD47-blocking monoclonal antibodies to drive markedly increased phagocytosis across a wide range of cancer cell types, including those that are otherwise resistant to ADCP. Additionally, we show that APMAP loss synergizes with several different tumour-targeting monoclonal antibodies to inhibit tumour growth in mice. Using genome-wide counterscreens in macrophages, we find that the G-protein-coupled receptor GPR84 mediates enhanced phagocytosis of APMAP-deficient cancer cells. This work reveals a cancer-intrinsic regulator of susceptibility to antibody-driven phagocytosis and, more broadly, expands our knowledge of the mechanisms governing cancer resistance to macrophage phagocytosis.
Assuntos
Citotoxicidade Celular Dependente de Anticorpos/genética , Sistemas CRISPR-Cas , Citofagocitose/genética , Macrófagos/imunologia , Neoplasias/imunologia , Neoplasias/patologia , Animais , Anticorpos Monoclonais/imunologia , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antígenos de Neoplasias/imunologia , Antígeno CD47/antagonistas & inibidores , Linhagem Celular Tumoral , Células Cultivadas , Feminino , Edição de Genes , Técnicas de Inativação de Genes , Humanos , Linfoma de Células T/imunologia , Linfoma de Células T/patologia , Macrófagos/citologia , Macrófagos/metabolismo , Masculino , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/genética , Camundongos , Receptores Acoplados a Proteínas G/metabolismoRESUMO
Ovarian cancer and triple-negative breast cancer are among the most lethal diseases affecting women, with few targeted therapies and high rates of metastasis. Cancer cells are capable of evading clearance by macrophages through the overexpression of anti-phagocytic surface proteins called 'don't eat me' signals-including CD471, programmed cell death ligand 1 (PD-L1)2 and the beta-2 microglobulin subunit of the major histocompatibility class I complex (B2M)3. Monoclonal antibodies that antagonize the interaction of 'don't eat me' signals with their macrophage-expressed receptors have demonstrated therapeutic potential in several cancers4,5. However, variability in the magnitude and durability of the response to these agents has suggested the presence of additional, as yet unknown 'don't eat me' signals. Here we show that CD24 can be the dominant innate immune checkpoint in ovarian cancer and breast cancer, and is a promising target for cancer immunotherapy. We demonstrate a role for tumour-expressed CD24 in promoting immune evasion through its interaction with the inhibitory receptor sialic-acid-binding Ig-like lectin 10 (Siglec-10), which is expressed by tumour-associated macrophages. We find that many tumours overexpress CD24 and that tumour-associated macrophages express high levels of Siglec-10. Genetic ablation of either CD24 or Siglec-10, as well as blockade of the CD24-Siglec-10 interaction using monoclonal antibodies, robustly augment the phagocytosis of all CD24-expressing human tumours that we tested. Genetic ablation and therapeutic blockade of CD24 resulted in a macrophage-dependent reduction of tumour growth in vivo and an increase in survival time. These data reveal CD24 as a highly expressed, anti-phagocytic signal in several cancers and demonstrate the therapeutic potential for CD24 blockade in cancer immunotherapy.
Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Antígeno CD24/antagonistas & inibidores , Imunoterapia/métodos , Lectinas/metabolismo , Macrófagos/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Antineoplásicos Imunológicos/imunologia , Antineoplásicos Imunológicos/farmacologia , Antígeno CD24/deficiência , Antígeno CD24/genética , Antígeno CD24/imunologia , Linhagem Celular Tumoral , Humanos , Lectinas/antagonistas & inibidores , Lectinas/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Neoplasias/imunologia , Neoplasias/patologia , Fagocitose/efeitos dos fármacos , Receptores de Superfície Celular/antagonistas & inibidores , Receptores de Superfície Celular/genética , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Evasão Tumoral/efeitos dos fármacos , Evasão Tumoral/imunologiaRESUMO
Exciting progress in the field of cancer immunotherapy has renewed the urgency of the need for basic studies of immunoregulation in both adaptive cell lineages and innate cell lineages. Here we found a central role for major histocompatibility complex (MHC) class I in controlling the phagocytic function of macrophages. Our results demonstrated that expression of the common MHC class I component ß2-microglobulin (ß2M) by cancer cells directly protected them from phagocytosis. We further showed that this protection was mediated by the inhibitory receptor LILRB1, whose expression was upregulated on the surface of macrophages, including tumor-associated macrophages. Disruption of either MHC class I or LILRB1 potentiated phagocytosis of tumor cells both in vitro and in vivo, which defines the MHC class I-LILRB1 signaling axis as an important regulator of the effector function of innate immune cells, a potential biomarker for therapeutic response to agents directed against the signal-regulatory protein CD47 and a potential target of anti-cancer immunotherapy.
Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Macrófagos/imunologia , Neoplasias/imunologia , Fagocitose/imunologia , Animais , Linhagem Celular Tumoral , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Imunoterapia/métodos , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/metabolismo , Macrófagos/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Neoplasias/metabolismo , Neoplasias/terapia , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/terapiaRESUMO
Cancer immunotherapies hold much promise, but their potential in veterinary settings has not yet been fully appreciated. Canine lymphomas are among the most common tumors of dogs and bear remarkable similarity to human disease. In this study, we examined the combination of CD47 blockade with anti-CD20 passive immunotherapy for canine lymphoma. The CD47/SIRPα axis is an immune checkpoint that regulates macrophage activation. In humans, CD47 is expressed on cancer cells and enables evasion from phagocytosis. CD47-blocking therapies are now under investigation in clinical trials for a variety of human cancers. We found the canine CD47/SIRPα axis to be conserved biochemically and functionally. We identified high-affinity SIRPα variants that antagonize canine CD47 and stimulate phagocytosis of canine cancer cells in vitro When tested as Fc fusion proteins, these therapeutic agents exhibited single-agent efficacy in a mouse xenograft model of canine lymphoma. As robust synergy between CD47 blockade and tumor-specific antibodies has been demonstrated for human cancer, we evaluated the combination of CD47 blockade with 1E4-cIgGB, a canine-specific antibody to CD20. 1E4-cIgGB could elicit a therapeutic response against canine lymphoma in vivo as a single agent. However, augmented responses were observed when combined with CD47-blocking therapies, resulting in synergy in vitro and in vivo and eliciting cures in 100% of mice bearing canine lymphoma. Our findings support further testing of CD47-blocking therapies alone and in combination with CD20 antibodies in the veterinary setting. Cancer Immunol Res; 4(12); 1072-87. ©2016 AACR.
Assuntos
Antígenos CD20/imunologia , Antígeno CD47/imunologia , Imunoterapia , Linfoma Difuso de Grandes Células B/terapia , Animais , Linhagem Celular Tumoral , Cães , Feminino , Imunoglobulina G/uso terapêutico , Linfoma Difuso de Grandes Células B/imunologia , Linfoma Difuso de Grandes Células B/veterinária , Macrófagos/imunologia , Masculino , Camundongos , Fagocitose , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Stem-cell differentiation to desired lineages requires navigating alternating developmental paths that often lead to unwanted cell types. Hence, comprehensive developmental roadmaps are crucial to channel stem-cell differentiation toward desired fates. To this end, here, we map bifurcating lineage choices leading from pluripotency to 12 human mesodermal lineages, including bone, muscle, and heart. We defined the extrinsic signals controlling each binary lineage decision, enabling us to logically block differentiation toward unwanted fates and rapidly steer pluripotent stem cells toward 80%-99% pure human mesodermal lineages at most branchpoints. This strategy enabled the generation of human bone and heart progenitors that could engraft in respective in vivo models. Mapping stepwise chromatin and single-cell gene expression changes in mesoderm development uncovered somite segmentation, a previously unobservable human embryonic event transiently marked by HOPX expression. Collectively, this roadmap enables navigation of mesodermal development to produce transplantable human tissue progenitors and uncover developmental processes. VIDEO ABSTRACT.
Assuntos
Mesoderma/citologia , Transdução de Sinais , Proteínas Morfogenéticas Ósseas/metabolismo , Osso e Ossos/citologia , Osso e Ossos/metabolismo , Coração/crescimento & desenvolvimento , Proteínas de Homeodomínio/metabolismo , Humanos , Mesoderma/metabolismo , Miócitos Cardíacos/metabolismo , Células-Tronco Pluripotentes/metabolismo , Linha Primitiva/citologia , Linha Primitiva/metabolismo , Análise de Célula Única , Somitos/metabolismo , Células-Tronco , Proteínas Supressoras de Tumor/metabolismo , Proteínas Wnt/antagonistas & inibidores , Proteínas Wnt/metabolismoRESUMO
Small-cell lung cancer (SCLC) is a highly aggressive subtype of lung cancer with limited treatment options. CD47 is a cell-surface molecule that promotes immune evasion by engaging signal-regulatory protein alpha (SIRPα), which serves as an inhibitory receptor on macrophages. Here, we found that CD47 is highly expressed on the surface of human SCLC cells; therefore, we investigated CD47-blocking immunotherapies as a potential approach for SCLC treatment. Disruption of the interaction of CD47 with SIRPα using anti-CD47 antibodies induced macrophage-mediated phagocytosis of human SCLC patient cells in culture. In a murine model, administration of CD47-blocking antibodies or targeted inactivation of the Cd47 gene markedly inhibited SCLC tumor growth. Furthermore, using comprehensive antibody arrays, we identified several possible therapeutic targets on the surface of SCLC cells. Antibodies to these targets, including CD56/neural cell adhesion molecule (NCAM), promoted phagocytosis in human SCLC cell lines that was enhanced when combined with CD47-blocking therapies. In light of recent clinical trials for CD47-blocking therapies in cancer treatment, these findings identify disruption of the CD47/SIRPα axis as a potential immunotherapeutic strategy for SCLC. This approach could enable personalized immunotherapeutic regimens in patients with SCLC and other cancers.