Limits...
Enhanced recruitment of genetically modified CX3CR1-positive human T cells into Fractalkine/CX3CL1 expressing tumors: importance of the chemokine gradient.

Siddiqui I, Erreni M, van Brakel M, Debets R, Allavena P - J Immunother Cancer (2016)

Bottom Line: Adoptive T-cell based immunotherapies constitute a promising approach to treat cancer, however, a major problem is to obtain effective and long-lasting anti-tumor responses.Furthermore, significant inhibition of tumor growth was found in mice receiving modified CX3CR1-T cells.Our results also establish that a correct chemokine gradient between the systemic circulation and the tumor is an essential requirement in adoptive T-cell based immunotherapy to efficiently recruit T cell effectors at the correct sites.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Inflammation, Humanitas Clinical and Research Center, 20089 Rozzano, Milan Italy ; Ludwig Center for Cancer Research, Department of Oncology, University of Lausanne, 1066 Epalinges, Switzerland.

ABSTRACT

Background: Adoptive T-cell based immunotherapies constitute a promising approach to treat cancer, however, a major problem is to obtain effective and long-lasting anti-tumor responses. Lack of response may be due to insufficient trafficking of specific T cells to tumors. A key requirement for efficient migration of cytotoxic T cells is that they express chemokine receptors that match the chemokines produced by tumor or tumor-associated cells.

Methods: In this study, we investigated whether the in vivo tumor trafficking of activated T cells could be enhanced by the expression of the chemokine receptor CX3CR1. Two human colorectal cancer cell lines were used to set up a xenograft tumor model in immunodeficient mice; the NCI-H630, constitutively expressing the chemokine ligand CX3CL1 (Fractalkine), and the RKO cell line, transduced to express CX3CL1.

Results: Human primary T cells were transduced with the receptor CX3CR1-eGFP. Upon in vivo adoptive transfer of genetically modified CX3CR1-T cells in mice bearing NCI-H630 tumors, enhanced lymphocyte migration and tumor trafficking were observed, compared to mice receiving Mock-T cells, indicating improved homing ability towards ligand-expressing tumor cells. Furthermore, significant inhibition of tumor growth was found in mice receiving modified CX3CR1-T cells. In contrast, tumors formed by RKO cells transduced with the ligand (RKO-CX3CL1) were not affected, nor more infiltrated upon transfer of CX3CR1-T lymphocytes, likely because high levels of the chemokine were shed by tumor cells in the systemic circulation, thus ifying the blood-tissue chemokine gradient.

Conclusions: This study demonstrates that ectopic expression of CX3CR1 enhanced the homing of adoptively transferred T cells towards CX3CL1-producing tumors, resulting in increased T cell infiltration in tumor tissues and decreased tumor growth. Our results also establish that a correct chemokine gradient between the systemic circulation and the tumor is an essential requirement in adoptive T-cell based immunotherapy to efficiently recruit T cell effectors at the correct sites.

No MeSH data available.


Related in: MedlinePlus

Adoptive transfer of CX3CR1-positive T cells to mice bearing NCI-H630 tumors. a Schematic representation of adoptive transfer of eGFP or CX3CR1-eGFP expressing T cells in NSG mice implanted with NCI-H630 tumors. b Representative flow cytometry plot of explanted and disaggregated tumors for the analysis of infiltrating human T lymphocytes (CD45 + CD3+/CX3CR1+) after adoptive transfer. Cells were gated on live cells, the plots show the population of CD3/CD45 positive cells and CD45/CX3CR1 positive cells within the tumors. c Proportion of CD8+ and CD4+ subpopulation within CD3 positive TILs. d Proportion of CX3CR1+ T cells in gated CD3+ tumor-infiltrating T cells (blue line: mice receiving CX3CR1-expressing T cells, red line: mice receiving eGFP-T cells CD3). e Summary of percentage of total CD3+ (left) and CX3CR1+ (right) T cells in explanted tumors after adoptive transfer, obtained from different mice (each dot represents a single mouse). +/−SEM **P < 0.01 for difference between control (eGFP) and test (CX3CR1) group (Student’s t test)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4836203&req=5

Fig3: Adoptive transfer of CX3CR1-positive T cells to mice bearing NCI-H630 tumors. a Schematic representation of adoptive transfer of eGFP or CX3CR1-eGFP expressing T cells in NSG mice implanted with NCI-H630 tumors. b Representative flow cytometry plot of explanted and disaggregated tumors for the analysis of infiltrating human T lymphocytes (CD45 + CD3+/CX3CR1+) after adoptive transfer. Cells were gated on live cells, the plots show the population of CD3/CD45 positive cells and CD45/CX3CR1 positive cells within the tumors. c Proportion of CD8+ and CD4+ subpopulation within CD3 positive TILs. d Proportion of CX3CR1+ T cells in gated CD3+ tumor-infiltrating T cells (blue line: mice receiving CX3CR1-expressing T cells, red line: mice receiving eGFP-T cells CD3). e Summary of percentage of total CD3+ (left) and CX3CR1+ (right) T cells in explanted tumors after adoptive transfer, obtained from different mice (each dot represents a single mouse). +/−SEM **P < 0.01 for difference between control (eGFP) and test (CX3CR1) group (Student’s t test)

Mentions: Mice were subcutaneously inoculated with NCI-H630 tumor cells and checked for tumor development; after 18 days, eGFP or CX3CR1-eGFP lymphocytes (3 × 106) were transferred through tail vein injection. Mice received transduced T lymphocytes every three-four days (Fig. 3a). After three consecutive adoptive transfers, mice were sacrificed and tumors harvested, digested enzymatically and tumor-infiltrating lymphocytes (TIL) were analyzed by quantitative real-time PCR and by flow cytometry. The representative cytometry analysis, illustrated in Fig. 3b, shows high CD3+ and CX3CR1+ cells (within CD45+ gated population) of mice receiving CX3CR1- transduced lymphocytes, compared to mice receiving eGFP-lymphocytes.Fig. 3


Enhanced recruitment of genetically modified CX3CR1-positive human T cells into Fractalkine/CX3CL1 expressing tumors: importance of the chemokine gradient.

Siddiqui I, Erreni M, van Brakel M, Debets R, Allavena P - J Immunother Cancer (2016)

Adoptive transfer of CX3CR1-positive T cells to mice bearing NCI-H630 tumors. a Schematic representation of adoptive transfer of eGFP or CX3CR1-eGFP expressing T cells in NSG mice implanted with NCI-H630 tumors. b Representative flow cytometry plot of explanted and disaggregated tumors for the analysis of infiltrating human T lymphocytes (CD45 + CD3+/CX3CR1+) after adoptive transfer. Cells were gated on live cells, the plots show the population of CD3/CD45 positive cells and CD45/CX3CR1 positive cells within the tumors. c Proportion of CD8+ and CD4+ subpopulation within CD3 positive TILs. d Proportion of CX3CR1+ T cells in gated CD3+ tumor-infiltrating T cells (blue line: mice receiving CX3CR1-expressing T cells, red line: mice receiving eGFP-T cells CD3). e Summary of percentage of total CD3+ (left) and CX3CR1+ (right) T cells in explanted tumors after adoptive transfer, obtained from different mice (each dot represents a single mouse). +/−SEM **P < 0.01 for difference between control (eGFP) and test (CX3CR1) group (Student’s t test)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4836203&req=5

Fig3: Adoptive transfer of CX3CR1-positive T cells to mice bearing NCI-H630 tumors. a Schematic representation of adoptive transfer of eGFP or CX3CR1-eGFP expressing T cells in NSG mice implanted with NCI-H630 tumors. b Representative flow cytometry plot of explanted and disaggregated tumors for the analysis of infiltrating human T lymphocytes (CD45 + CD3+/CX3CR1+) after adoptive transfer. Cells were gated on live cells, the plots show the population of CD3/CD45 positive cells and CD45/CX3CR1 positive cells within the tumors. c Proportion of CD8+ and CD4+ subpopulation within CD3 positive TILs. d Proportion of CX3CR1+ T cells in gated CD3+ tumor-infiltrating T cells (blue line: mice receiving CX3CR1-expressing T cells, red line: mice receiving eGFP-T cells CD3). e Summary of percentage of total CD3+ (left) and CX3CR1+ (right) T cells in explanted tumors after adoptive transfer, obtained from different mice (each dot represents a single mouse). +/−SEM **P < 0.01 for difference between control (eGFP) and test (CX3CR1) group (Student’s t test)
Mentions: Mice were subcutaneously inoculated with NCI-H630 tumor cells and checked for tumor development; after 18 days, eGFP or CX3CR1-eGFP lymphocytes (3 × 106) were transferred through tail vein injection. Mice received transduced T lymphocytes every three-four days (Fig. 3a). After three consecutive adoptive transfers, mice were sacrificed and tumors harvested, digested enzymatically and tumor-infiltrating lymphocytes (TIL) were analyzed by quantitative real-time PCR and by flow cytometry. The representative cytometry analysis, illustrated in Fig. 3b, shows high CD3+ and CX3CR1+ cells (within CD45+ gated population) of mice receiving CX3CR1- transduced lymphocytes, compared to mice receiving eGFP-lymphocytes.Fig. 3

Bottom Line: Adoptive T-cell based immunotherapies constitute a promising approach to treat cancer, however, a major problem is to obtain effective and long-lasting anti-tumor responses.Furthermore, significant inhibition of tumor growth was found in mice receiving modified CX3CR1-T cells.Our results also establish that a correct chemokine gradient between the systemic circulation and the tumor is an essential requirement in adoptive T-cell based immunotherapy to efficiently recruit T cell effectors at the correct sites.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Inflammation, Humanitas Clinical and Research Center, 20089 Rozzano, Milan Italy ; Ludwig Center for Cancer Research, Department of Oncology, University of Lausanne, 1066 Epalinges, Switzerland.

ABSTRACT

Background: Adoptive T-cell based immunotherapies constitute a promising approach to treat cancer, however, a major problem is to obtain effective and long-lasting anti-tumor responses. Lack of response may be due to insufficient trafficking of specific T cells to tumors. A key requirement for efficient migration of cytotoxic T cells is that they express chemokine receptors that match the chemokines produced by tumor or tumor-associated cells.

Methods: In this study, we investigated whether the in vivo tumor trafficking of activated T cells could be enhanced by the expression of the chemokine receptor CX3CR1. Two human colorectal cancer cell lines were used to set up a xenograft tumor model in immunodeficient mice; the NCI-H630, constitutively expressing the chemokine ligand CX3CL1 (Fractalkine), and the RKO cell line, transduced to express CX3CL1.

Results: Human primary T cells were transduced with the receptor CX3CR1-eGFP. Upon in vivo adoptive transfer of genetically modified CX3CR1-T cells in mice bearing NCI-H630 tumors, enhanced lymphocyte migration and tumor trafficking were observed, compared to mice receiving Mock-T cells, indicating improved homing ability towards ligand-expressing tumor cells. Furthermore, significant inhibition of tumor growth was found in mice receiving modified CX3CR1-T cells. In contrast, tumors formed by RKO cells transduced with the ligand (RKO-CX3CL1) were not affected, nor more infiltrated upon transfer of CX3CR1-T lymphocytes, likely because high levels of the chemokine were shed by tumor cells in the systemic circulation, thus ifying the blood-tissue chemokine gradient.

Conclusions: This study demonstrates that ectopic expression of CX3CR1 enhanced the homing of adoptively transferred T cells towards CX3CL1-producing tumors, resulting in increased T cell infiltration in tumor tissues and decreased tumor growth. Our results also establish that a correct chemokine gradient between the systemic circulation and the tumor is an essential requirement in adoptive T-cell based immunotherapy to efficiently recruit T cell effectors at the correct sites.

No MeSH data available.


Related in: MedlinePlus