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1.
Glycobiology ; 34(7)2024 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-38785323

RESUMO

Aberrant glycosylation is a key mechanism employed by cancer cells to evade immune surveillance, induce angiogenesis and metastasis, among other hallmarks of cancer. Sialic acids, distinctive terminal glycan structures located on glycoproteins or glycolipids, are prominently upregulated across various tumor types, including colorectal cancer (CRC). Sialylated glycans modulate anti-tumor immune responses through their interactions with Siglecs, a family of glycan-binding receptors with specificity for sialic acid-containing glycoconjugates, often resulting in immunosuppression. In this paper, we investigated the immunomodulatory function of ST3Gal5, a sialyltransferase that catalyzes the addition of α2-3 sialic acids to glycosphingolipids, since lower expression of ST3Gal5 is associated with better survival of CRC patients. We employed CRISPR/Cas9 to knock out the ST3Gal5 gene in two murine CRC cell lines MC38 and CT26. Glycomics analysis confirmed the removal of sialic acids on glycolipids, with no discernible impact on glycoprotein sialylation. Although knocking out ST3Gal5 in both cell lines did not affect in vivo tumor growth, we observed enhanced levels of regulatory T cells in CT26 tumors lacking ST3Gal5. Moreover, we demonstrate that the absence of ST3Gal5 affected size and blood vessel density only in MC38 tumors. In summary, we ascertain that sialylation of glycosphingolipids has a limited influence on the anti-tumor immune response in CRC, despite detecting alterations in the tumor microenvironment, possibly due to a shift in ganglioside abundance.


Assuntos
Neoplasias Colorretais , Gangliosídeos , Sialiltransferases , Sialiltransferases/metabolismo , Sialiltransferases/genética , Gangliosídeos/metabolismo , Gangliosídeos/imunologia , Animais , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Camundongos , Linhagem Celular Tumoral , Humanos , beta-Galactosídeo alfa-2,3-Sialiltransferase
2.
J Immunother Cancer ; 11(11)2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37940346

RESUMO

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers. Despite the successful application of immune checkpoint blockade in a range of human cancers, immunotherapy in PDAC remains unsuccessful. PDAC is characterized by a desmoplastic, hypoxic and highly immunosuppressive tumor microenvironment (TME), where T-cell infiltration is often lacking (immune desert), or where T cells are located distant from the tumor islands (immune excluded). Converting the TME to an immune-inflamed state, allowing T-cell infiltration, could increase the success of immunotherapy in PDAC. METHOD: In this study, we use the KPC3 subcutaneous PDAC mouse model to investigate the role of tumor-derived sialic acids in shaping the tumor immune landscape. A sialic acid deficient KPC3 line was generated by genetic knock-out of the CMAS (cytidine monophosphate N-acetylneuraminic acid synthetase) enzyme, a critical enzyme in the synthesis of sialic acid-containing glycans. The effect of sialic acid-deficiency on immunotherapy efficacy was assessed by treatment with anti-programmed cell death protein 1 (PD-1) and agonistic CD40. RESULT: The absence of sialic acids in KPC3 tumors resulted in increased numbers of CD4+ and CD8+ T cells in the TME, and reduced frequencies of CD4+ regulatory T cells (Tregs) within the T-cell population. Importantly, CD8+ T cells were able to infiltrate the tumor islands in sialic acid-deficient tumors. These favorable alterations in the immune landscape sensitized sialic acid-deficient tumors to immunotherapy, which was ineffective in sialic acid-expressing KPC3 tumors. In addition, high expression of sialylation-related genes in human pancreatic cancer correlated with decreased CD8+ T-cell infiltration, increased presence of Tregs, and poorer survival probability. CONCLUSION: Our results demonstrate that tumor-derived sialic acids mediate T-cell exclusion within the PDAC TME, thereby impairing immunotherapy efficacy. Targeting sialic acids represents a potential strategy to enhance T-cell infiltration and improve immunotherapy outcomes in PDAC.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Camundongos , Animais , Humanos , Linfócitos T CD8-Positivos , Ácidos Siálicos/farmacologia , Ácido N-Acetilneuramínico/farmacologia , Neoplasias Pancreáticas/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Imunoterapia/métodos , Microambiente Tumoral
3.
Cancer Metastasis Rev ; 42(3): 941-958, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37266839

RESUMO

Gangliosides are sialylated glycolipids, mainly present at the cell surface membrane, involved in a variety of cellular signaling events. During malignant transformation, the composition of these glycosphingolipids is altered, leading to structural and functional changes, which are often negatively correlated to patient survival. Cancer cells have the ability to shed gangliosides into the tumor microenvironment, where they have a strong impact on anti-tumor immunity and promote tumor progression. Since most ganglioside species show prominent immunosuppressive activities, they might be considered checkpoint molecules released to counteract ongoing immunosurveillance. In this review, we highlight the current state-of-the-art on the ganglioside-mediated immunomodulation, specified for the different immune cells and individual gangliosides. In addition, we address the dual role that certain gangliosides play in the tumor microenvironment. Even though some ganglioside species have been more extensively studied than others, they are proven to contribute to the defense mechanisms of the tumor and should be regarded as promising therapeutic targets for inclusion in future immunotherapy regimens.


Assuntos
Gangliosídeos , Neoplasias , Humanos , Gangliosídeos/metabolismo , Microambiente Tumoral , Neoplasias/metabolismo , Glicolipídeos , Glicoesfingolipídeos
4.
Cell Rep ; 37(7): 110013, 2021 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-34788605

RESUMO

Autotaxin (ATX; ENPP2) produces lysophosphatidic acid (LPA) that regulates multiple biological functions via cognate G protein-coupled receptors LPAR1-6. ATX/LPA promotes tumor cell migration and metastasis via LPAR1 and T cell motility via LPAR2, yet its actions in the tumor immune microenvironment remain unclear. Here, we show that ATX secreted by melanoma cells is chemorepulsive for tumor-infiltrating lymphocytes (TILs) and circulating CD8+ T cells ex vivo, with ATX functioning as an LPA-producing chaperone. Mechanistically, T cell repulsion predominantly involves Gα12/13-coupled LPAR6. Upon anti-cancer vaccination of tumor-bearing mice, ATX does not affect the induction of systemic T cell responses but, importantly, suppresses tumor infiltration of cytotoxic CD8+ T cells and thereby impairs tumor regression. Moreover, single-cell data from melanoma tumors are consistent with intratumoral ATX acting as a T cell repellent. These findings highlight an unexpected role for the pro-metastatic ATX-LPAR axis in suppressing CD8+ T cell infiltration to impede anti-tumor immunity, suggesting new therapeutic opportunities.


Assuntos
Linfócitos do Interstício Tumoral/metabolismo , Diester Fosfórico Hidrolases/metabolismo , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Quimiotaxia/fisiologia , Feminino , Humanos , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Lisofosfolipídeos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias , Diester Fosfórico Hidrolases/fisiologia , Receptores de Ácidos Lisofosfatídicos/metabolismo , Transdução de Sinais/fisiologia , Microambiente Tumoral
5.
Oncoimmunology ; 9(1): 1786888, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32939320

RESUMO

Blockade of the PD-1 receptor has revolutionized the treatment of metastatic melanoma, with significant increases in overall survival (OS) and a dramatic improvement in patient quality of life. Despite the success of this approach, the number of benefitting patients is limited and there is a need for predictive biomarkers as well as a deeper mechanistic analysis of the cellular populations involved in clinical responses. With the aim to find predictive biomarkers for PD-1 checkpoint blockade, an in-depth immune monitoring study was conducted in 36 advanced melanoma patients receiving pembrolizumab or nivolumab treatment at Karolinska University Hospital. Blood samples were collected before treatment and before administration of the second and fourth doses. Peripheral blood mononuclear cells were isolated and stained for flow cytometric analysis within 2 h of sample collection. Overall survival and progression-free survival (PFS) were inversely correlated with CD69 expression NK cells. In the myeloid compartment, high frequencies of non-classical monocytes and low frequencies of monocytic myeloid derived suppressor cells (MoMDSCs) correlated with response rates and OS. A deeper characterization of monocytic subsets showed that PD-L1 expression in MDSCs, non-classical and intermediate monocytes was significantly increased in patients with shorter PFS in addition to correlating inversely with OS. Our results suggest that cellular populations other than T cells can be critical in the outcome of PD-1 blockade treatment. Specifically, the frequencies of activated NK cells and monocytic subsets are inversely correlated with survival and clinical benefit, suggesting that their role as predictive biomarkers should be further evaluated.


Assuntos
Antígeno B7-H1 , Melanoma , Biomarcadores , Humanos , Células Matadoras Naturais , Leucócitos Mononucleares , Melanoma/tratamento farmacológico , Monócitos , Receptor de Morte Celular Programada 1 , Qualidade de Vida
6.
Cancers (Basel) ; 12(9)2020 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-32927726

RESUMO

Cancer stem cells (CSCs) are located in dedicated niches, where they remain inert to chemotherapeutic drugs and drive metastasis. Although plasticity in the CSC pool is well appreciated, the molecular mechanisms implicated in the regulation of cancer stemness are still elusive. Here, we define a fucosylation-dependent reprogramming of colon cancer cells towards a stem cell-like phenotype and function. De novo transcriptional activation of Fut9 in the murine colon adenocarcinoma cell line, MC38, followed by RNA seq-based regulon analysis, revealed major gene regulatory networks related to stemness. Lewisx, Sox2, ALDH and CD44 expression, tumorsphere formation, resistance to 5-FU treatment and in vivo tumor growth were increased in FUT9-expressing MC38 cells compared to the control cells. Likewise, human CRC cell lines highly expressing FUT9 displayed phenotypic features of CSCs, which were significantly impaired upon FUT9 knock-out. Finally, in primary CRC FUT9+ tumor cells pathways related to cancer stemness were enriched, providing a clinically meaningful annotation of the complicity of FUT9 in stemness regulation and may open new avenues for therapeutic intervention.

7.
Methods Mol Biol ; 1913: 167-179, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30666606

RESUMO

Antibody-dependent cell-mediated cytotoxicity (ADCC) is a mechanism in which immune cell activation is induced by the cross-linking of CD16 with the Fc region of antibodies that at the same time bind specifically to cell surface antigens. ADCC stimulates the secretion of perforin, granzymes, and cytokines leading to lysis of the malignant cells. Natural killer (NK) cells express the CD16 receptor and can therefore be activated by ADCC to kill tumor cells. To study the cytotoxicity of NK cells against cancer cells, an ADCC-based assay is described: the chromium release assay. In this method, the antibody trastuzumab, which binds specifically to HER2-positive malignant cells, is used to trigger ADCC.


Assuntos
Radioisótopos de Cromo/metabolismo , Testes Imunológicos de Citotoxicidade/métodos , Células Matadoras Naturais/imunologia , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Neoplasias da Mama/sangue , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Separação Celular/instrumentação , Separação Celular/métodos , Testes Imunológicos de Citotoxicidade/instrumentação , Feminino , Citometria de Fluxo/instrumentação , Citometria de Fluxo/métodos , Proteínas Ligadas por GPI/imunologia , Proteínas Ligadas por GPI/metabolismo , Humanos , Células Matadoras Naturais/metabolismo , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/metabolismo , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Trastuzumab/farmacologia
8.
Methods Mol Biol ; 1913: 181-194, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30666607

RESUMO

Antibody-dependent cell-mediated cytotoxicity (ADCC) is a mechanism in which immune cell activation is induced by the cross-linking of CD16 with the Fc region of antibodies that at the same time bind specifically to cell surface antigens. ADCC stimulates the secretion of perforin, granzymes, and cytokines leading to lysis of the malignant cells. Natural killer (NK) cells express the CD16 receptor and can therefore be activated by ADCC to kill tumor cells. To study the cytotoxicity of NK cells against cancer cells, an ADCC-based assay is described: the flow cytometry-based cytotoxicity assay. In this method, the antibody trastuzumab, which binds specifically to HER2-positive malignant cells, is used to trigger ADCC.


Assuntos
Testes Imunológicos de Citotoxicidade/métodos , Citometria de Fluxo/métodos , Células Matadoras Naturais/imunologia , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Neoplasias da Mama/sangue , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Separação Celular/instrumentação , Separação Celular/métodos , Testes Imunológicos de Citotoxicidade/instrumentação , Feminino , Citometria de Fluxo/instrumentação , Proteínas Ligadas por GPI/imunologia , Proteínas Ligadas por GPI/metabolismo , Humanos , Células Matadoras Naturais/metabolismo , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/metabolismo , Receptores de IgG/imunologia , Receptores de IgG/metabolismo , Trastuzumab/farmacologia
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