Arginase 1 is a key driver of immune suppression in pancreatic cancer
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Evaluation Summary:
Menjivar et al. identify a previously unrecognized role of myeloid cell Arginase1 (Arg1) activity in shaping the anti-tumor immune response in pancreatic ductal adenocarcinoma (PDAC). The proposed therapeutic combination is a new approach for pancreatic cancer, with an enhanced response to immune therapy upon arginase inhibition.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)
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Abstract
An extensive fibroinflammatory stroma rich in macrophages is a hallmark of pancreatic cancer. In this disease, it is well appreciated that macrophages are immunosuppressive and contribute to the poor response to immunotherapy; however, the mechanisms of immune suppression are complex and not fully understood. Immunosuppressive macrophages are classically defined by the expression of the enzyme Arginase 1 (ARG1), which we demonstrated is potently expressed in pancreatic tumor-associated macrophages from both human patients and mouse models. While routinely used as a polarization marker, ARG1 also catabolizes arginine, an amino acid required for T cell activation and proliferation. To investigate this metabolic function, we used a genetic and a pharmacologic approach to target Arg1 in pancreatic cancer. Genetic inactivation of Arg1 in macrophages, using a dual recombinase genetically engineered mouse model of pancreatic cancer, delayed formation of invasive disease, while increasing CD8 + T cell infiltration. Additionally, Arg1 deletion induced compensatory mechanisms, including Arg1 overexpression in epithelial cells, namely Tuft cells, and Arg2 overexpression in a subset of macrophages. To overcome these compensatory mechanisms, we used a pharmacological approach to inhibit arginase. Treatment of established tumors with the arginase inhibitor CB-1158 exhibited further increased CD8 + T cell infiltration, beyond that seen with the macrophage-specific knockout, and sensitized the tumors to anti-PD1 immune checkpoint blockade. Our data demonstrate that Arg1 drives immune suppression in pancreatic cancer by depleting arginine and inhibiting T cell activation.
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Author Response
Reviewer #1 (Public Review):
In this study, Menjivar et al. examine the specific role of the enzyme arginase 1 (Arg1), which is expressed in immunosuppressive macrophages and catabolizes arginine to ornithine, in pancreatic cancer. They use an elegant genetic approach that leverages a dual recombinase-based genetically engineered mouse model of pancreatic cancer, which efficiently deletes Arg1 and recovers extracellular arginine in cultured macrophages. Within the pancreas, macrophage Arg1 deletion increased T cell infiltration and fewer mice developed invasive pancreatic cancer. Interestingly, when tumors did develop, the authors observed that compensatory mechanisms of arginine depletion were induced, including Arg1 overexpression in epithelial cells identified as tuft cells or Arg2 overexpression in macrophages. To …
Author Response
Reviewer #1 (Public Review):
In this study, Menjivar et al. examine the specific role of the enzyme arginase 1 (Arg1), which is expressed in immunosuppressive macrophages and catabolizes arginine to ornithine, in pancreatic cancer. They use an elegant genetic approach that leverages a dual recombinase-based genetically engineered mouse model of pancreatic cancer, which efficiently deletes Arg1 and recovers extracellular arginine in cultured macrophages. Within the pancreas, macrophage Arg1 deletion increased T cell infiltration and fewer mice developed invasive pancreatic cancer. Interestingly, when tumors did develop, the authors observed that compensatory mechanisms of arginine depletion were induced, including Arg1 overexpression in epithelial cells identified as tuft cells or Arg2 overexpression in macrophages. To overcome these compensatory mechanisms, pharmacological targeting of arginase was tested and found to increase T cell infiltration and sensitize to immune checkpoint blockade, suggesting this is a promising approach for pancreatic cancer.
Strengths:
This is a very rigorous, well-designed study and the findings are broadly interesting for the metabolism, immunometabolism, and pancreatic cancer communities. The methods are comprehensive and the experimental details in the legends are complete.
Weaknesses:
The claim that Arg1 deletion in macrophages delayed the formation of invasive disease is not completely justified by the data presented. Only a small number of mice are analyzed, and no statistics are included.
While in the original submission this claim was based on a relatively small number of animals, we have now increased each cohort. The new graph is included in Figure 2E (Response Figure 1); statistical analysis is also included and show the differences to be significant.
Moreover, the abstract does not comprehensively summarize the findings. Many findings, including compensatory upregulation of ARG1 in tuft cells and ARG2 in myeloid cells, are not mentioned, nor was the rationale for the pharmacological approach. Finally, the claim that their data demonstrate that Arg1 is more than simply a marker of macrophage function. While this is the first time this has been examined in pancreatic cancer, a general role for Arg1 and arginine metabolism by myeloid cells in immunosuppression has already been established by multiple studies, including those cited by the authors, in multiple tumor types. This is an overstatement of the findings.
We apologize for the lack of clarity, in the attempt to meet the word limit for the abstract. We have now amended the abstract to better reflect the total of our findings and the context of our work.
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Evaluation Summary:
Menjivar et al. identify a previously unrecognized role of myeloid cell Arginase1 (Arg1) activity in shaping the anti-tumor immune response in pancreatic ductal adenocarcinoma (PDAC). The proposed therapeutic combination is a new approach for pancreatic cancer, with an enhanced response to immune therapy upon arginase inhibition.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)
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Reviewer #1 (Public Review):
In this study, Menjivar et al. examine the specific role of the enzyme arginase 1 (Arg1), which is expressed in immunosuppressive macrophages and catabolizes arginine to ornithine, in pancreatic cancer. They use an elegant genetic approach that leverages a dual recombinase-based genetically engineered mouse model of pancreatic cancer, which efficiently deletes Arg1 and recovers extracellular arginine in cultured macrophages. Within the pancreas, macrophage Arg1 deletion increased T cell infiltration and fewer mice developed invasive pancreatic cancer. Interestingly, when tumors did develop, the authors observed that compensatory mechanisms of arginine depletion were induced, including Arg1 overexpression in epithelial cells identified as tuft cells or Arg2 overexpression in macrophages. To overcome these …
Reviewer #1 (Public Review):
In this study, Menjivar et al. examine the specific role of the enzyme arginase 1 (Arg1), which is expressed in immunosuppressive macrophages and catabolizes arginine to ornithine, in pancreatic cancer. They use an elegant genetic approach that leverages a dual recombinase-based genetically engineered mouse model of pancreatic cancer, which efficiently deletes Arg1 and recovers extracellular arginine in cultured macrophages. Within the pancreas, macrophage Arg1 deletion increased T cell infiltration and fewer mice developed invasive pancreatic cancer. Interestingly, when tumors did develop, the authors observed that compensatory mechanisms of arginine depletion were induced, including Arg1 overexpression in epithelial cells identified as tuft cells or Arg2 overexpression in macrophages. To overcome these compensatory mechanisms, pharmacological targeting of arginase was tested and found to increase T cell infiltration and sensitize to immune checkpoint blockade, suggesting this is a promising approach for pancreatic cancer.
Strengths:
This is a very rigorous, well-designed study and the findings are broadly interesting for the metabolism, immunometabolism, and pancreatic cancer communities. The methods are comprehensive and the experimental details in the legends are complete.
Weaknesses:
The claim that Arg1 deletion in macrophages delayed the formation of invasive disease is not completely justified by the data presented. Only a small number of mice are analyzed, and no statistics are included. Moreover, the abstract does not comprehensively summarize the findings. Many findings, including compensatory upregulation of ARG1 in tuft cells and ARG2 in myeloid cells, are not mentioned, nor was the rationale for the pharmacological approach. Finally, the claim that their data demonstrate that Arg1 is more than simply a marker of macrophage function. While this is the first time this has been examined in pancreatic cancer, a general role for Arg1 and arginine metabolism by myeloid cells in immunosuppression has already been established by multiple studies, including those cited by the authors, in multiple tumor types. This is an overstatement of the findings.
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Reviewer #2 (Public Review):
In this manuscript, Menjivar et al. demonstrate the relevance of myeloid cell Arginase1 (Arg1) activity in shaping the anti-tumor immune response in pancreatic ductal adenocarcinoma (PDAC). They show that inhibition of Arg1 in myeloid cells induces macrophage repolarization, enhances anti-tumor CD8+ cell infiltration and activation, leading to decreased tumor growth; and that systemic inhibition of arginases using CB-1158 enhances the tumor response to anti-PD1 immune checkpoint therapy.
Strengths:
Novelty and translational relevance of the results, i.e. the role of myeloid cell Arg1 in the anti-tumor immune response in PDAC.
Weaknesses:
The authors stress both in the abstract and within the manuscript the fact that Arg1 is widely used as a mere "marker of macrophage function" or "anti-inflammatory M2 state" …
Reviewer #2 (Public Review):
In this manuscript, Menjivar et al. demonstrate the relevance of myeloid cell Arginase1 (Arg1) activity in shaping the anti-tumor immune response in pancreatic ductal adenocarcinoma (PDAC). They show that inhibition of Arg1 in myeloid cells induces macrophage repolarization, enhances anti-tumor CD8+ cell infiltration and activation, leading to decreased tumor growth; and that systemic inhibition of arginases using CB-1158 enhances the tumor response to anti-PD1 immune checkpoint therapy.
Strengths:
Novelty and translational relevance of the results, i.e. the role of myeloid cell Arg1 in the anti-tumor immune response in PDAC.
Weaknesses:
The authors stress both in the abstract and within the manuscript the fact that Arg1 is widely used as a mere "marker of macrophage function" or "anti-inflammatory M2 state" and that their work here provides evidence for its functional role in altering anti-tumor immunity.
However, they do not show what breaking down arginine and generating ornithine and urea does/is needed metabolically to impact immunity, as no such metabolic assays or experiments are performed.
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Reviewer #3 (Public Review):
Menjivar et al. present an analysis of the role of immunosuppressive Arginase 1 in myeloid cells in pancreatic cancer. They show that depletion of Arg1 in macrophages leads to attenuation in progression from PanIN to PDAC and use single cell analysis to understand underlying changes in immune activation, including an increase in cytotoxic T cells. Interestingly, the authors observed what seems to be a compensatory upregulation in Arg1 in epithelial cells and used arginase1 inhibitor to assess the therapeutic potential of targeting Arg1 systemically. This study is overall well performed and generates novel mouse models to study immunosuppression in pancreatic cancer. While the notion that Arginase1 is immunosuppressive is not novel, the observation that Arg1 is upregulated in epithelial cells is interesting. …
Reviewer #3 (Public Review):
Menjivar et al. present an analysis of the role of immunosuppressive Arginase 1 in myeloid cells in pancreatic cancer. They show that depletion of Arg1 in macrophages leads to attenuation in progression from PanIN to PDAC and use single cell analysis to understand underlying changes in immune activation, including an increase in cytotoxic T cells. Interestingly, the authors observed what seems to be a compensatory upregulation in Arg1 in epithelial cells and used arginase1 inhibitor to assess the therapeutic potential of targeting Arg1 systemically. This study is overall well performed and generates novel mouse models to study immunosuppression in pancreatic cancer. While the notion that Arginase1 is immunosuppressive is not novel, the observation that Arg1 is upregulated in epithelial cells is interesting. There are several instances of overstating conclusions that, if addressed in the main text (not just relegated to the discussion section), could significantly strengthen the manuscript.
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