Leukocytes possess the ability to migrate upstream-against the direction of flow-on surfaces of specific chemistry. Upstream migration was first characterized in vitro for T-cells on surfaces comprised of intracellular adhesion molecule-1 (ICAM-1). Upstream migration occurs when the integrin receptor αLβ2 (also known as lymphocyte function-associated antigen-1, or LFA-1) binds to ICAM-1. LFA-1/ICAM-1 interactions are ubiquitous and are widely found in leukocyte trafficking. Upstream migration would be employed after cells come to arrest on the apical surface of the endothelium and might confer an advantage for both trans-endothelial migration and tissue surveillance. It has now been shown that several other motile amoeboid cells which have the responsibility of trafficking from blood vessels into tissues, such as Marginal zone B cells, hematopoietic stem cells, and neutrophils (when macrophage-1 antigen, Mac-1, is blocked), can also migrate upstream on ICAM-1 surfaces. This review will summarize what is known about the basic mechanisms of upstream migration, which cells have displayed this phenomenon, and the possible role of upstream migration in physiology and tissue homeostasis.

BackgroundImmune homeostasis is regulated by a balance of pro- and anti-inflammatory cytokine signals. Dysregulated cytokine expression can cause deleterious immunosuppression or inflammation, which drives disease pathology. In solid tumors, cytokines such as IL10 and TGFβ induce an immunosuppressive tumor microenvironment (TME) that blunts endogenous and therapeutic anti-tumor immunity. Therapeutic strategies to block immunosuppressive cytokines have primarily focused on monoclonal antibodies targeting cytokines/cytokine receptors. While this approach can reduce immunosuppressive signaling, it fails to provide an inflammatory signal that could initiate anti-tumor immunity. Here, we engineered macrophages with synthetic cytokine switch receptors (SR) to develop a cell therapy platform for modulation of pro-/anti-inflammatory signals. Macrophages are homeostatic regulators capable of both initiating inflammation and infiltrating solid tumors, and we leveraged this natural proficiency using SRs that convert tumor-related immunosuppressive (M2) signals into pro-inflammatory (M1) responses for tumor microenvironment (TME) modulation. We termed this engineered myeloid cell platform ’Engineered Microenvironment Converters’ (EM-C) and evaluated its modular ability to target several tumor-associated cytokines.MethodsEM-Cs targeting IL10 or TGFβ were generated by expressing SR in primary human macrophages and monocytes. M2-to-M1 SR were designed to convert IL10 or TGF-β into pro-inflammatory signals based on interferon or toll-like receptor (TLR) signaling pathways. The response of EM-Cs to target cytokines was monitored using phenotypic characterization of surface molecules, measurement of cytokine release, mRNA profiling, and biochemical analysis of downstream signaling. Co-culture assays with bystander immunosuppressive cells were used to assess the ability of EM-Cs to alter their microenvironment. Additionally, combinatorial EM-C were designed to target both IL10 and TGFβ for multiplexed TME conversion.ResultsPro-inflammatory EM-Cs efficiently sequestered IL10 and TGFβ, two prevalent immunosuppressive cytokines in the TME, and converted them into pro-inflammatory signals by upregulating M1 markers, cytokines, and pathways in a dose-dependent manner. EM-Cs furthermore repolarized bystander M2 macrophages towards a pro-inflammatory phenotype following co-culture.ConclusionsWe present a novel immunotherapy platform that harnesses macrophages as ‘living converters’ to locally regulate inflammation in solid tumors. We establish EM-C that convert IL10 or TGFβ into pro-inflammatory signals, showcasing a modular ability to control the inflammatory status of microenvironments without systemic cytokine antagonism. EM-Cs enable the development of target antigen agnostic myeloid cell immunotherapies for solid tumors.

BackgroundMacrophages expressing chimeric antigen receptors (CAR-M) have been shown to reduce tumor burden, remodel the tumor microenvironment (TME), and coordinate a systemic immune response in pre-clinical solid tumor models. Solid tumors overexpress immunosuppressive molecules, such as CD47, which reduces macrophage tumor phagocytosis. We have previously demonstrated that CD47 is a checkpoint that reduces CAR-M function, and have shown that CRISPR-mediated SIRPα knockout (KO) CAR-M are refractory to the anti-phagocytic checkpoint protein CD47.1 Here, we have enhanced our ability to control gene expression in CAR-M by creating a single vector system that incorporates synthetic shRNA into the CAR intron, enabling simultaneous CAR expression with knockdown of SIRPα. The intronic shRNA platform allows gene-silenced CAR-M and CAR-Monocytes to be manufactured using a single vector in a streamlined single-day process.MethodsTo generate gene-silenced CAR-M, we transduced primary human macrophages or monocytes with a novel adenoviral vector comprising a CAR transgene with custom intronic shRNA expressed under a shared promoter. We characterized shRNA-modified CAR-M using a model anti- human epidermal growth factor receptor 2 (HER2) CAR paired with shRNA targeting SIRPα. CAR-M phenotype was characterized using flow cytometry. The anti-tumor efficacy of CAR-M in vitro was monitored by quantifying killing, phagocytosis, and cytokine production in co-culture assays with HER2+ tumor cell lines. The in vivo efficacy of CAR-M was characterized using metastatic solid tumor xenograft models.ResultsIntronic shRNA enabled concomitant CAR expression with target SIRPα knockdown. Reduction in SIRPα expression by shRNA was comparable to that achieved by CRISPR/Cas9 ribonucleoprotein. Additionally, the inclusion of an intron significantly augmented expression of the neighboring CAR transgene. Compared to unmodified CAR-M, SIRPα-knockdown CAR-M exhibited enhanced killing, phagocytosis, and cytokine production against HER2+ tumor cells in vitro. Furthermore, SIRPα-knockdown CAR-M significantly delayed tumor growth and prolonged survival in vivo.ConclusionsWe show the feasibility of generating gene-silenced primary CAR-M in a single transduction step by integrating CAR delivery with custom intronic shRNA, and we demonstrate that targeted gene knockdown of SIRPα can enhance the anti-tumor activity of CAR-M in vivo. The intronic shRNA design is a generalizable platform that will be valuable for future CAR designs, target tumor antigens, and gene knockout targets.ReferenceSloas C, Gabbasov R, Anderson N, Abramson S, Klichinsky M, Ohtani Y. 144 SIRPα deficient CAR-Macrophages exhibit enhanced anti-tumor function and bypass the CD47 immune checkpoint. J Immunother Cancer. 2021;9:A152-A152.

BackgroundMacrophages are abundant in the solid tumor microenvironment (sTME) and can promote tumor growth (M2) or enhance anti-tumor immunity (M1). CAR expression can redirect macrophage function to selectively target and phagocytose antigen overexpressing cancer cells. CAR-M can reprogram the sTME and present neoantigens to T cells, leading to epitope spreading and anti-tumor immunity. CT-0508 is comprised of autologous monocyte-derived proinflammatory macrophages expressing an anti-HER2 CAR. Pre-clinical studies demonstrated that CT-0508 induced targeted cancer cell phagocytosis while sparing normal cells, decreased tumor burden, prolonged survival, and was safe. Notably, anti-HER2 CAR-M treatment led to activation of the sTME, with infiltration of CD8+ and CD4+ T cells, NK cells, dendritic cells, and increased activated CD8+ tumor infiltrating lymphocytes. In a pre-clinical anti-PD1 resistant solid tumor model, mice that received anti-HER2 CAR-M and anti-PD1 demonstrated improved tumor control, overall survival, and TME activation compared to single treatment alone, indicating synergy and capacity for CAR-M to sensitize solid tumors to checkpoint blockade.1MethodsThis Phase 1, First in Human study evaluates the safety, tolerability, cell manufacturing feasibility, trafficking, TME activation, and preliminary evidence of efficacy of investigational product CT-0508 in 18 participants (pts) with locally advanced (unresectable)/metastatic solid tumors overexpressing HER2. Pts previously treated with anti-HER2 therapies are eligible. Filgrastim mobilized autologous CD14+ monocytes are collected by apheresis, without the need for lymphodepleting chemotherapy, followed by manufacturing and cryopreservation. Group 1 pts (n = 9; enrollment complete) received fractionated doses of CT-0508 over Days 1, 3, and 5. Group 2 pts (n = 9) receive CT-0508 as a single infusion on D1. Additional cohorts include: CT-0508 co-administered with pembrolizumab and CT-0508 monotherapy administered intraperitoneally in pts with peritoneal predominant disease. Correlative assessments include pre- and post-treatment biopsies and blood samples for safety (immunogenicity), trafficking (PCR, RNA scope), CT-0508 persistence in blood and tumor, target antigen engagement, TME modulation (single cell RNA sequencing), immune response (TCR sequencing) and others.ReferenceKlichinsky M, et al. Human chimeric antigen receptor macrophages for cancer immunotherapy. Nature Biotechnology. 2020;38:947–953.

TPS2666 Background: Macrophages are abundant in the solid tumor microenvironment (sTME) and can promote tumor growth (M2) or enhance anti-tumor immunity (M1). CAR expression can redirect macrophage function to selectively target and phagocytose antigen overexpressing cancer cells. CAR-M can reprogram the sTME and present neoantigens to T cells, leading to epitope spreading and anti-tumor immunity. CT-0508 is comprised of autologous monocyte-derived proinflammatory macrophages expressing an anti-HER2 CAR. Pre-clinical studies showed that CT-0508 induced targeted cancer cell phagocytosis while sparing normal cells, decreased tumor burden, prolonged survival, and was safe. Notably, anti-HER2 CAR-M treatment led to activation of the sTME, with infiltration of CD8+ and CD4+ T cells, NK cells, dendritic cells, and increased activated CD8+ tumor infiltrating lymphocytes. In a pre-clinical anti-PD1 resistant solid tumor model, mice that received anti-HER2 CAR-M and anti-PD1 demonstrated improved tumor control, overall survival, and TME activation compared to single treatment alone, indicating synergy and capacity for CAR-M to sensitize solid tumors to checkpoint blockade. Methods: This Phase 1, First in Human study evaluates the safety, tolerability, cell manufacturing feasibility, trafficking, TME activation, and preliminary evidence of efficacy of investigational product CT-0508 in 18 participants (pts) with locally advanced (unresectable) /metastatic solid tumors overexpressing HER2. Pts previously treated with anti-HER2 therapies are eligible. Filgrastim mobilized autologous CD14+ monocytes are collected by apheresis, followed by manufacturing and cryopreservation. Group 1 pts (n = 9; enrollment complete) received fractionated doses of CT-0508 over Days 1, 3, and 5. Group 2 pts (n = 9) receive CT-0508 as a single infusion on D1. Additional cohorts include: CT-0508 co-administered with pembrolizumab and CT-0508 monotherapy administered intraperitoneally in pts with peritoneal predominant disease. Correlative assessments include pre- and post-treatment biopsies and blood samples for safety (immunogenicity), trafficking (PCR, RNA scope), CT-0508 persistence in blood and tumor, target antigen engagement, TME modulation (single cell RNA sequencing), immune response (TCR sequencing) and others. Clinical trial information: NCT04660929 .

Abstract Background: Macrophages are abundant in the solid tumor microenvironment (sTME) and can exhibit both pro- and anti-tumor functions. Macrophages can be redirected by CAR expression to phagocytose cancer cells in an antigen-specific manner. CAR-M can reprogram the sTME and present neoantigens to T cells, leading to epitope spreading and anti-tumor immunity. CT-0508 is comprised of autologous monocyte-derived proinflammatory macrophages expressing an anti-HER2 CAR. Pre-clinical studies showed that CT-0508 induced targeted cancer cell phagocytosis while sparing normal cells, decreased tumor burden, prolonged survival, and was safe and effective. Notably, anti-HER2 CAR-M treatment led to activation of the sTME, with infiltration of CD8+ and CD4+ T cells, NK cells, dendritic cells, and increased activated CD8+ tumor infiltrating lymphocytes. In a pre-clinical model of advanced solid tumor resistant to PD1 blockade, mice treated with anti-HER2 CAR-M combined with a PD1 blocking antibody demonstrated improved tumor control, overall survival, and TME activation compared to either treatment alone, indicating synergy and capacity for CAR-M to sensitize solid tumors to checkpoint blockade. Methods: This Phase 1, FIH study is evaluating safety, tolerability, cell manufacturing feasibility, trafficking, TME activation, and preliminary evidence of efficacy of investigational product CT-0508 in 18 pt with locally advanced (unresectable)/metastatic solid tumors overexpressing HER2. Pt previously treated with anti-HER2 therapies are eligible. Filgrastim mobilized autologous CD14+ monocytes are collected by apheresis, followed by manufacturing and cryopreservation. Group 1 pt (n = 9; enrollment complete) received fractionated doses over Days 1, 3, and 5. Group 2 pt (n = 9) receive CT-0508 as a single infusion on D1. Additional cohorts include: CT-0508 co-administered with pembrolizumab and CT-0508 monotherapy administered intraperitoneally in pt with peritoneal predominant disease. Correlative assessments include pre- and post-treatment biopsies and blood samples for safety (immunogenicity), trafficking (qPCR, RNA in situ hybridization), CT-0508 persistence in blood and tumor, target antigen engagement, TME modulation (single cell RNA sequencing), immune response (TCR sequencing) and others. Citation Format: Yara Abdou, E. Claire Dees, Joanne Mortimer, Naoto Ueno, Melissa Johnson, Richard Maziarz, Jennifer Specht, Yuan Yuan, Paula Puhlman, Mathew Angelos, Saar Gill, Amy Ronczka, Thomas Condamine, Daniel J. Cushing, Michael Klichinsky, Debora Barton, Ramona F. Swaby, Kim Reiss Binder. A phase 1, first-in-human (FIH) study of autologous anti-HER2 chimeric antigen receptor macrophage (CAR-M) in participants (pt) with HER2 overexpressing solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT241.

Abstract Motivation: Cytokines in tissue microenvironments regulate the balance between pro- and anti-inflammatory signals. Dysregulated cytokines cause deleterious immunosuppression or inflammation, which underpins the pathophysiology of solid tumors, chronic kidney disease, and more. Rebalancing inflammation/immunosuppression by rectifying cytokine signals offers a generalizable approach for treating numerous diseases. While doing so through cytokine blockade carries risks due to systemic administration, cellular immunotherapies offer a localized approach that could detect pathogenic cytokines then proportionately rebalance inflammation as needed. Specifically, macrophages are homeostatic regulators responsible for initiating and resolving inflammation. Here, we leveraged macrophages’ ability to regulate inflammation by equipping them with synthetic cytokine switch receptors (SR) that convert immunosuppressive M2 signals into pro-inflammatory M1 responses for solid tumor microenvironment conversion, or vice versa for inflammatory disease. We termed this platform “Engineered Microenvironment Converters” (EM-C) and evaluated its modular ability to target disease-associated cytokines. Methods: EM-Cs targeting IL10, TGFβ, IFNγ and IL17A were generated by expressing SR in primary human macrophages. M2-to-M1 SR were designed to convert IL10 or TGF-β into pro-inflammatory stimuli, and M1-to-M2 SR were designed to IFNγ or IL17A into immunosuppressive signals. The in vitro response of EM-Cs to their target cytokine was monitored using phenotypic characterization of surface molecules, measurement of cytokine production, mRNA sequencing, and biochemical analysis of downstream signaling. Co-culture assays with bystander cells were used to assess the ability of EM-Cs to alter their microenvironment. Results: Pro-inflammatory EM-Cs converted IL10 and TGFβ, two prevalent immunosuppressive cytokines in the TME, into pro-inflammatory signals. Unlike wildtype macrophages, these EM-Cs responded to IL10 or TGFβ with upregulated M1 markers and cytokines in a dose-dependent manner. Furthermore, EM-Cs repolarized bystander M2 macrophages towards a pro-inflammatory phenotype following co-culture. Similarly, anti-inflammatory EM-Cs responded IFNγ and IL17A, two cytokines canonically overexpressed in inflammatory disease, by upregulating M2 markers and inducing an anti-inflammatory environment. Conclusion: We present for the first time a novel immunotherapy platform that harnesses macrophages as “living converters” to locally regulate inflammation for oncology and inflammatory applications. By demonstrating EM-Cs in the M2-to-M1 and M1-to-M2 direction, this platform offers modularity in controlling the inflammatory status of tissue microenvironments without systemic cytokine antagonism. Citation Format: Chris Sloas, Yuhao Huangfu, Rehman Qureshi, Michael Ball, Thomas Condamine, Michael Klichinsky, Yumi Ohtani. Macrophages engineered with cytokine switch receptors: Development of a modular platform for rebalancing inflammation in microenvironments. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4054.

Abstract While adoptive cell therapies have seen significant success in the treatment of hematological malignancies, solid tumors remain challenging for the field. A significant obstacle is the exclusion of T cells from the tumor microenvironment (TME). In contrast, monocytes/macrophages are naturally recruited to the TME. These cells then have the potential to phagocytose tumor cells, activate the TME, and prime a broad anti-tumor adaptive immune response via T cell recruitment and activation. We have previously developed CT-0508, a chimeric antigen receptor macrophage (CAR-M) targeting HER2 which showed efficacy in a variety of pre-clinical models and is currently in a Phase I clinical trial for patients with HER2+ solid tumors. Mesothelin is overexpressed in a variety of solid tumors, including mesothelioma, lung, pancreatic, and ovarian cancers. To leverage tumor biology with myeloid cells, we engineered primary human macrophages using the chimeric adenoviral vector Ad5f35 to express a CAR containing a human scFv against human mesothelin. We used both in vitro cell based assays and in vivo xenograft models to assess the activity of CT-1119. CAR-M engineered with an Ad5f35 vector demonstrated high CAR expression, high viability, upregulated M1 (anti-tumor) macrophage markers, and downregulated M2 (pro-tumor) macrophage markers. CT-1119 specifically phagocytosed multiple mesothelin expressing tumor cell lines in a CAR-dependent and antigen-dependent manner. CT-1119 demonstrated robust in vitro killing of the relevant tumor cell lines A549 and MES-OV expressing mesothelin. CAR engagement also induced the release of pro-inflammatory cytokines such as TNFα following stimulation with mesothelin in both cell-free and cell-based contexts in a dose-dependent manner. In vivo, CT-1119 significantly reduced tumor burden in a murine xenograft model of lung cancer. Similarly, human monocytes targeting mesothelin were successfully generated using the same Ad5f35 vector and demonstrated specific activity against mesothelin positive tumor cells. The presented results demonstrate that CT-1119, an autologous human anti-mesothelin CAR-M, can cause phagocytosis, tumor cell killing, and pro-inflammatory cytokine release in response to stimulation with mesothelin. These results show that CAR-M is a feasible approach for the treatment of mesothelin expressing sold tumors via the potential for induction of a systemic anti-tumor response. Citation Format: Nicholas R. Anderson, Brinda Shah, Alison Worth, Rashid Gabbasov, Brett Menchel, Kerri Ciccaglione, Daniel Blumenthal, Stefano Pierini, Sabrina Ceeraz DeLong, Sascha Abramson, Thomas Condamine, Michael Klichinsky. A mesothelin targeting chimeric antigen receptor macrophage (CAR-M) for solid tumor immunotherapy: pre-clinical development of CT-1119. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4053.

BackgroundAdoptive T cell therapies have led to remarkable advances among patients with hematologic malignancies but have had less success in those with solid tumors. Macrophages are actively recruited and abundantly present in the solid tumor microenvironment (sTME). Tumor associated macrophages are predominantly immunosuppressive and support tumor growth (M2), while a subset of proinflammatory macrophages enhance anti-tumor immunogenicity (M1). M1 macrophage function can be augmented by CAR expression to selectively recognize and phagocytose antigen overexpressing cancer cells. Moreover, CAR macrophages can reprogram the sTME and present neoantigens to T cells, leading to epitope spreading and anti-tumor immune memory. Human Epidermal Growth Factor Receptor 2 (HER2) overexpression promotes tumorigenesis in many solid tumors. CT-0508 is a cell product comprised of autologous monocyte-derived proinflammatory macrophages expressing an anti-HER2 CAR and is being investigated in a currently ongoing first in human clinical trial (NCT04660929).MethodsUsing the chimeric adenoviral vector Ad5f35, we manufactured CT-0508 product from apheresis material collected from patients enrolled in group 1 of the CT-0508 phase 1 clinical trial. To assess the activity of pre-infusion CT-0508 products, in vitro cell based assays were utilized including killing, phagocytosis, cytokine secretion, and transcriptomics analysis. Donor-matched untransduced (UTD) macrophages served as controls. Additionally, healthy donor derived products were used in some experiments.ResultsCT-0508 was successfully manufactured with high viability, purity and CAR expression for patients enrolled in group 1. CT-0508 products demonstrated enhanced killing and phagocytosis of HER2-expressing tumor cells over autologous UTD macrophages. CITE-Seq analysis of CT-0508 confirmed an M1 macrophage transcriptional signature compared with pre-manufacturing monocyte and apheresis populations. Using healthy donor derived CT-0508 products, cell product activation was demonstrated by ex vivo CAR engagement and led to an increase in secreted immunostimulatory cytokines and chemokines, consistent with pro-inflammatory macrophage activation including TNF, IL-6, MIP-1α, and MIP-1β. Secreted factor analysis also showed HER2 responsive changes in extracellular matrix remodeling factors and growth factors. Additionally, transcriptomic and proteomic analysis revealed that CAR engagement further enhanced CAR-M M1 polarization. CAR engagement mediated effects are also being investigated in patient derived products.ConclusionsTogether, these results demonstrate that functional CT-0508 CAR-M were successfully manufactured with an M1 phenotype and that CAR-antigen interaction drives cell product activation and amplifies the M1 polarization status of CT-0508 CAR-M.

BackgroundIn pre-clinical studies, CAR macrophages (CAR-M) phagocytose tumor cells, activate the tumor micro-environment (TME), recruit T cells, and induce anti-tumor T cell immunity. CT-0508 is a first-in-class CAR-M product comprised of autologous monocyte-derived macrophages expressing an anti-HER2 CAR. Here we present preliminary clinical results from the CT-0508 Phase 1 FIH study.MethodsThis multi-center, open-label study is evaluating CT-0508’s safety, tolerability, and manufacturing feasibility in 18 participants with advanced solid tumors overexpressing HER2 with progression on prior therapies. Monocytes are isolated from mobilized apheresis products, differentiated into macrophages, and engineered with an anti-HER2 CAR. Group 1 participants (n = 9) receive a fractionated dose on days 1, 3, 5 and Group 2 participants (n = 9) receive the full dose on day 1. CT-0508 is administered without preparative chemotherapy. Serial blood samples and biopsies (baseline and 2 post-treatment) are collected to investigate safety, pharmacokinetics, and mechanism of action.ResultsNine participants (6F/3M) have been treated, comprising breast (4), esophageal (2), cholangiocarcinoma, ovarian, and parotid gland cancers, with a median age of 58. Participants had received a median of 3 (range, 2-11) prior lines of therapy; 8 had received prior anti-HER2 therapy. CT-0508 was successfully manufactured and well tolerated with no dose-limiting toxicities. Three related SAEs occurred in 2 participants: grade 1 CRS with hospitalization for monitoring and grade 2 infusion reaction that resolved within 1 hour were reported in one participant. Grade 2 CRS with fever and hypoxia occurred in another participant and resolved within ~ 72 hours. Five additional participants experienced Grade 1-2 CRS and/or infusion reactions with rapid resolution. There were no major organ toxicities and no on-target off-tumor toxicities. Post-infusion cytokines were transiently elevated in most participants enrolled in group 1 and were self-limiting. Four of 7 participants evaluated had stable disease. CT-0508 was transiently detectable in the blood and was detected in the TME 8 days and 4 weeks after infusion. CT-0508 modulated the TME, leading to myeloid cell activation, T cell infiltration, activation, and proliferation. TCR sequencing demonstrated newly expanding T cell clones in the blood post-treatment that accumulated within the TME, suggesting expansion of tumor-reactive T cells upon CT-0508 infusion. Data from participants enrolled in group 1 will be presented.ConclusionsCT-0508 was safe and feasible to manufacture.Early correlative data demonstrate trafficking, TME modulation,and induction of anti-tumor T cell immunity. The study is actively enrolling.Trial RegistrationNCT04660929ReferenceKlichinsky M, Ruella M, Shestova O. et al. Human chimeric antigen receptor macrophages for cancer immunotherapy. Nat Biotechnol. 2020;38:947–953.Ethics ApprovalThis study has been approved by each participating site: University of Pennsylvania Abramson Cancer Center IRB approval #844106, University of North Carolina Lineberger Cancer Center IRB approval #20201732 , City of Hope Cancer Center IRB approval #20201732, The University of Texas MD Anderson Cancer Center IRB approval # 2021-0327, Sarah Cannon Research Institute IRB #20201732. All participants gave their informed consent before taking part in the study.