TLR7/8-Agonist-Loaded M1-Macrophage-Derived Nanovesicles Promote the Polarization of Macrophages to Enhance Bladder Cancer Immunotherapy

Abstract Immune checkpoint inhibitors (ICIs), such as PD-1/PD-L1 antibodies, modulate the cancer killing function of immune cells in the tumor microenvironment (TME). However, immunosuppressive M2-type tumor-associated macrophages (TAMs) are abundant in bladder cancer (BC) and able to release substances such as cytokines to promote tumor growth, evade immune cell attack and lead to tumor ICIs treatment resistance. In the present study, we utilized nanovesicles derived from M1 macrophages, which contained constituents of M1 macrophages (M1 NV), and loaded the vesicles with the TLR7/8 agonist R848, a potent driver of M1 macrophages to construct M1 NV-R848 nanovesicles. Compared with M1 NV or R848 treatment alone, M1 NV-R848 was able to induce polarization of M2 macrophages into M1 macrophages more efficient both in vitro and in vivo. Intravenous injection of M1 NV-R848 improved the immunosuppressive TME and inhibited tumor growth and no significantly toxic or immunogenic in MB-49 tumor-bearing mice. In addition, compared with M1 NV-R848 treatment alone, combined injection of M1 NV-R848 and PD-L1 was able to further inhibit MB-49 tumor growth. Thus, our study demonstrates that M1 NV-R848 has the ability to promote the polarization of M2 TAMs to M1 macrophages and to enhance the efficacy of the ICIs PD-L1 in the treatment of UBC with no significantly toxic or immunogenic.

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Reprogrammed M1 macrophages with inhibited STAT3, STAT6 and/or SMAD3 extends lifespan of mice with experimental carcinoma

ZHurnal «Patologicheskaia fiziologiia i eksperimental`naia terapiia» ◽

10.25557/0031-2991.2017.02.4-9 ◽

2017 ◽

pp. 4-9

Author(s):

С.В. Калиш ◽

С.В. Лямина ◽

А.А. Раецкая ◽

И.Ю. Малышев

Keyword(s):

Tumor Growth ◽

Culture Medium ◽

Ehrlich Carcinoma ◽

Macrophage Phenotype ◽

M1 Macrophages ◽

M1 Macrophage ◽

Ec Cells ◽

Experimental Carcinoma

Цель исследования. Репрограммирование М1 фенотипа макрофагов с ингибированными факторами транскрипции М2 фенотипа STAT3, STAТ6 и SMAD и оценка их влияния на развитие карциномы Эрлиха (КЭ) in vitro и in vivo. Методика. Рост опухоли иницировали in vitro путем добавления клеток КЭ в среду культивирования RPMI-1640 и in vivo путем внутрибрюшинной инъекции клеток КЭ мышам. Результаты. Установлено, что M1макрофаги и in vitro, и in vivo оказывают выраженный противоопухолевый эффект, который превосходит антиопухолевые эффекты М1, M1, M1 макрофагов и цисплатина. Заключение. М1 макрофаги с ингибированными STAT3, STAT6 и/или SMAD3 эффективно ограничивают рост опухоли. Полученные данные обосновывают разработку новой технологии противоопухолевой клеточной терапии. Objective. Reprogramming of M1 macrophage phenotype with inhibited M2 phenotype transcription factors, such as STAT3, STAT6 and SMAD and assess their impact on the development of Ehrlich carcinoma (EC) in vitro and in vivo . Methods. Tumor growth in vitro was initiated by addition of EC cells in RPMI-1640 culture medium and in vivo by intraperitoneal of EC cell injection into mice. Results. It was found that M1 macrophages have a pronounced anti-tumor effect in vitro , and in vivo , which was greater than anti-tumor effects of M1, M1, M1 macrophages and cisplatin. Conclusion. M1 macrophages with inhibited STAT3, STAT6 and/or SMAD3 effectively restrict tumor growth. The findings justify the development of new anti-tumor cell therapy technology.

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BTK Inhibition Reverses MDSC-Mediated Immunosuppression and Enhances Response to Anti-PDL1 Therapy in Neuroblastoma

Cancers ◽

10.3390/cancers13040817 ◽

2021 ◽

Vol 13 (4) ◽

pp. 817

Author(s):

Mehreen Ishfaq ◽

Timothy Pham ◽

Cooper Beaman ◽

Pablo Tamayo ◽

Alice L. Yu ◽

...

Keyword(s):

T Cell ◽

Tumor Growth ◽

Checkpoint Inhibitors ◽

No Production ◽

Free Survival ◽

T Cell Infiltration ◽

Car T ◽

Checkpoint Inhibitor Therapy

MDSCs are immune cells of myeloid lineage that plays a key role in promoting tumor growth. The expansion of MDSCs in tumor-bearing hosts reduces the efficacy of checkpoint inhibitors and CAR-T therapies, and hence strategies that deplete or block the recruitment of MDSCs have shown benefit in improving responses to immunotherapy in various cancers, including NB. Ibrutinib, an irreversible molecular inhibitor of BTK, has been widely studied in B cell malignancies, and recently, this drug is repurposed for the treatment of solid tumors. Herein we report that BTK is highly expressed in both granulocytic and monocytic murine MDSCs isolated from mice bearing NB tumors, and its increased expression correlates with a poor relapse-free survival probability of NB patients. Moreover, in vitro treatment of murine MDSCs with ibrutinib altered NO production, decreased mRNA expression of Ido, Arg, Tgfβ, and displayed defects in T-cell suppression. Consistent with these findings, in vivo inhibition of BTK with ibrutinib resulted in reduced MDSC-mediated immune suppression, increased CD8+ T cell infiltration, decreased tumor growth, and improved response to anti-PDL1 checkpoint inhibitor therapy in a murine model of NB. These results demonstrate that ibrutinib modulates immunosuppressive functions of MDSC and can be used either alone or in combination with immunotherapy for augmenting antitumor immune responses in NB.

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P03.20 A murine, myc-driven lymphoma model expressing human CD22 enables testing of targeted therapies and their effects on tumor immune microenvironment

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-itoc7.58 ◽

2020 ◽

Vol 8 (Suppl 2) ◽

pp. A30.1-A30

Author(s):

F Gsottberger ◽

C Brandl ◽

S Petkovic ◽

L Nitschke ◽

A Mackensen ◽

...

Keyword(s):

Tumor Growth ◽

B Cell ◽

Targeted Therapies ◽

Immune Cell ◽

Myeloid Cells ◽

B Cell Lymphoma ◽

Primary Lymphoma ◽

Murine Lymphoma

BackgroundThe tumor microenvironment (TME) is composed of various cell types which closely interact via cell cell contacts and cytokines leading to tumor promotion, immune cell inhibition and drug resistance. TME is increasingly recognized for its role in cancer immunotherapies. In B-cell malignancies, myeloid cells play a central role in supporting tumor growth and immune suppression (Roussel et al., 2017, Cancer Immunol Immunother). Despite the importance of a syngeneic TME, preclinical studies with novel drugs have mainly been performed in models lacking a functional immune system. Therefore, we developed an immune competent murine lymphoma model transgenic to human CD22 to study effects of targeted therapies on TME.Materials and MethodsA chimeric CD22 consisting of human extracellular and murine intracellular CD22 (h/mCD22) was introduced in BL6 mice (BL6h/mCD22). Crossbreeding with BL6λ-myc lead to spontaneous development of murine lymphoma that were serially transplanted. Tumor infiltration and TME was characterized by flow cytometry. Mice were treated with Moxetumomab pasudotox, a CD22 targeted immunotoxin and Doxorubicin.ResultsSpontaneously developed tumors in lymphoid organs from BL6h/mCD22 x λ-myc consist of a monomorphic population of h/mCD22+ murine B cells. Three primary lymphoma subclones were isolated from distinct mice and serially transplanted in syngeneic mice. Stable tumor growth was established after subcutaneous (sc) and intravenous (iv) injection. However, TME of sc tumors was infiltrated by less than 1% immune cells, while myc-driven lymphoma in humans usually show substantial immune infiltration. In contrast to sc tumors, systemically growing lymphoma in murine bone marrow (BM) are infiltrated by 30% myeloid cells and 1% T-cells and in murine spleen by 10% and 30%, respectively. Myeloid cells found in these tumors were shown to suppress T cell proliferation in vitro. To test functionality of the h/mCD22 transgene, lymphoma-bearing mice were treated with Moxetumomab, which reduced BM lymphoma infiltration by 20 to 100-fold and infiltration in spleen by 5 to 20-fold in the three lymphoma models. Effects of treatment on TME were analyzed after treatment with Doxorubicin which is known to activate myeloid cells in vivo. Compared to untreated controls, Doxorubicin increased CD11b+ cells in spleen by 1.5-fold. Among these cells, Ly6G+ granulocytic cells increased most substantially.ConclusionsWe established primary, myc-driven h/mCD22+ B-cell lymphoma which stably engraft in syngeneic mice with a TME mimicking myc-driven lymphoma in men. The model responds well to CD22-targeted therapy and Doxorubicin induces expected immunologic changes. Therefore, our unique model provides a platform to test CD22-targeting treatment strategies in an immune competent background.Disclosure InformationF. Gsottberger: None. C. Brandl: None. S. Petkovic: None. L. Nitschke: None. A. Mackensen: None. F. Müller: None.

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MicroRNA-381 inhibits bladder cancer pathogenesis through directly controlling BMI1 and triggering the Rho/ROCK signaling pathway defect

10.21203/rs.3.rs-23467/v1 ◽

2020 ◽

Author(s):

Dayin Chen ◽

Liang Cheng ◽

Huifeng Cao ◽

Wensi Liu

Keyword(s):

Bladder Cancer ◽

Tumor Growth ◽

Cancer Control ◽

Tumor Formation ◽

Luciferase Assay ◽

Cell Behaviors ◽

Cancer Pathogenesis ◽

Bmi1 Expression

Abstract Background Emerging evidence has noted the important participation of microRNAs (miRNAs) in several human diseases including cancer control. This research was launched to probe the function of miR-381 in bladder cancer (BCs) progression. Methods Aberrantly expressed miRNAs in BCs tissues were analyzed using miRNA microarrays. miR-381 expression in the bladder and paired tumor tissues, and in BCs and normal cell lines was determined. The target relationship between miR-381 and BMI1 was predicted online and validated through a luciferase assay. Gain-of-functions of miR-381 and BMI1 were performed to identify their functions on BCs cell behaviors as well as tumor growth in vivo. Results miR-381 was poor regulated in BCs tissues and cells. A higher miR-381 level indicates a better prognosis of patients with BCs. Artificial up-regulation of miR-381 inhibited proliferation, invasion, migration, resistance to apoptosis, and tumor formation ability of BCs cells. miR-381 directly binds to BMI1 expression. Overexpression of BMI1 partially blocked the tumor suppressing roles of miR-381 in cell malignancy and tumor growth. In addition, miR-381 led to decreased RhoA phosphorylation and ROCK2 activation, which were also reversed by BMI1. Conclusion The study evidenced that miR-381 may act as a beneficiary biomarker in BCs patients. Up-regulation of miR-381 could suppress BCs development both in vivo and in vitro through BMI1 down-regulation and the Rho/ROCK inactivation.

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The M1 aminopeptidase NPEPPS is a novel regulator of cisplatin sensitivity

10.1101/2021.03.04.433676 ◽

2021 ◽

Author(s):

Robert T. Jones ◽

Andrew Goodspeed ◽

Maryam C. Akbarzadeh ◽

Mathijs Scholtes ◽

Hedvig Vekony ◽

...

Keyword(s):

Bladder Cancer ◽

Cisplatin Resistance ◽

Treatment Resistance ◽

Anion Channel ◽

List Type ◽

Cancer Models ◽

Cisplatin Sensitivity ◽

Cancer Types

ABSTRACTDespite routine use of platinum-based chemotherapeutics across diverse cancer types, there remains a need to improve efficacy and patient selection for treatment. A multi-omic assessment of five human bladder cancer cell lines and their chemotherapy resistant derivatives, coupled with whole-genome CRISPR screens were used to identify puromycin- sensitive aminopeptidase, NPEPPS, as a novel functional driver of treatment resistance to cisplatin. Depletion of NPEPPS resulted in enhanced cellular cisplatin import, sensitization of resistant cancer cells to cisplatin in vitro and in vivo. Pharmacologic inhibition of NPEPPS with tosedostat in cells and in chemoresistant, patient-derived tumor organoids improved response to cisplatin. Depletion of LRRC8A and LRRC8D, two subunits of the volume regulated anion channel (VRAC), a known importer of intracellular cisplatin, enhanced resistance to cisplatin. Linking NPEPPS function to VRAC cisplatin import supports NPEPPS as a driver of cisplatin resistance and by virtue of clinically available inhibitors, the potential for rapid clinical translation.HIGHLIGHTS∙CRISPR screens with multi-omics identify NPEPPS as a driver of cisplatin resistance∙NPEPPS depletion in multiple bladder cancer models enhances cisplatin sensitivity∙LRRC8A and LRRC8D loss increase resistance to cisplatin in CRISPR screens∙Unique resource of functional and multi-omic data is provided to the community

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Nanoparticle delivery of immunostimulatory oligonucleotides enhances response to checkpoint inhibitor therapeutics

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2001569117 ◽

2020 ◽

Vol 117 (24) ◽

pp. 13428-13436 ◽

Cited By ~ 6

Author(s):

Colin G. Buss ◽

Sangeeta N. Bhatia

Keyword(s):

Animal Models ◽

Tumor Growth ◽

Immune Checkpoint ◽

Immune Checkpoint Inhibitor ◽

Checkpoint Inhibitor ◽

Checkpoint Inhibitors ◽

Engineered Nanoparticles ◽

Therapeutic Effects

The recent advent of immune checkpoint inhibitor (CPI) antibodies has revolutionized many aspects of cancer therapy, but the efficacy of these breakthrough therapeutics remains limited, as many patients fail to respond for reasons that still largely evade understanding. An array of studies in human patients and animal models has demonstrated that local signaling can generate strongly immunosuppressive microenvironments within tumors, and emerging evidence suggests that delivery of immunostimulatory molecules into tumors can have therapeutic effects. Nanoparticle formulations of these cargoes offer a promising way to maximize their delivery and to enhance the efficacy of checkpoint inhibitors. We developed a modular nanoparticle system capable of encapsulating an array of immunostimulatory oligonucleotides that, in some cases, greatly increase their potency to activate inflammatory signaling within immune cells in vitro. We hypothesized that these immunostimulatory nanoparticles could suppress tumor growth by activating similar signaling in vivo, and thereby also improve responsiveness to immune checkpoint inhibitor antibody therapies. We found that our engineered nanoparticles carrying a CpG DNA ligand of TLR9 can suppress tumor growth in several animal models of various cancers, resulting in an abscopal effect on distant tumors, and improving responsiveness to anti-CTLA4 treatment with combinatorial effects after intratumoral administration. Moreover, by incorporating tumor-homing peptides, immunostimulatory nucleotide-bearing nanoparticles facilitate antitumor efficacy after systemic intravenous (i.v.) administration.

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IMMU-28. TARGETING IMMUNOSUPPRESSIVE MYELOID DERIVED SUPPRESSOR CELLS VIA MIF/CD74 SIGNALING AXIS TO ATTENUATE GBM GROWTH

Neuro-Oncology ◽

10.1093/neuonc/noz175.521 ◽

2019 ◽

Vol 21 (Supplement_6) ◽

pp. vi125-vi125

Author(s):

Tyler Alban ◽

Defne Bayik ◽

Balint Otvos ◽

Matthew Grabowski ◽

Manmeet Ahluwalia ◽

...

Keyword(s):

Tumor Microenvironment ◽

Tumor Growth ◽

Immune Cell ◽

Suppressor Cells ◽

Myeloid Derived Suppressor Cells ◽

Signaling Axis ◽

Immunosuppressive Microenvironment ◽

And Function

Abstract The immunosuppressive microenvironment in glioblastoma (GBM) enables persistent tumor growth and evasion from tumoricidal immune cell recognition. Despite a large accumulation of immune cells in the GBM microenvironment, tumor growth continues, and evidence for potent immunosuppression via myeloid derived suppressor cells (MDSCs) is now emerging. In agreement with these observations, we have recently established that increased MDSCs over time correlates with poor prognosis in GBM, making these cells of interest for therapeutic targeting. In seeking to reduce MDSCs in GBM, we previously identified the cytokine macrophage migration inhibitory factor (MIF) as a possible activator of MDSC function in GBM. Here, using a novel in vitro co-culture system to reproducibly and rapidly create GBM-educated MDSCs, we observed that MIF was essential in the generation of MDSCs and that MDSCs generated via this approach express a repertoire of MIF receptors. CD74 was the primary MIF receptor in monocytic MDSCs (M-MDSC), which penetrate the tumor microenvironment in preclinical models and patient samples. A screen of MIF/CD74 interaction inhibitors revealed that MN-166, a clinically relevant blood brain barrier penetrant drug, which is currently fast tracked for FDA approval, reduced MDSC generation and function in vitro. This effect was specific to M-MDSC subsets expressing CD74, and appeared as reduced downstream pERK signaling and MCP-1 secretion. In vivo, MN-166 was able reduce tumor-infiltrating MDSCs, while conferring a significant increase in survival in the syngeneic glioma model GL261. These data provide proof of concept that M-MDSCs can be targeted in the tumor microenvironment via MN-166 to reduce tumor growth and provide a rationale for future clinical assessment of MN-166 to reduce M-MDSCs in the tumor microenvironment. Ongoing studies are assessing the effects of MDSC inhibition in combination with immune activating approaches, in order to inhibit immune suppression while simultaneously activating the immune system.

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The Potential of T Cell Immunoglobulin and Mucin-Domain containing-3 (Tim-3) in Designing Novel Immunotherapy for Bladder Cancer

Endocrine Metabolic & Immune Disorders - Drug Targets ◽

10.2174/1871530321666210310142141 ◽

2021 ◽

Vol 21 ◽

Author(s):

Monireh Mohsenzadegan ◽

Parizad Bavandpour ◽

Mohammad Reza Nowroozi ◽

Erfan Amini ◽

Masoumeh Kourosh-Arami ◽

...

Keyword(s):

Bladder Cancer ◽

T Cells ◽

T Cell ◽

Tumor Immunity ◽

Checkpoint Inhibitors ◽

Number Of Patients ◽

Anti Cancer ◽

Domain 3

: Targeting inhibitory receptors on T cells in the tumor sites can promote effective anti-tumor immunity in bladder cancer. Unfortunately, the main dilemma is that a large number of patients remain refractory to CTLA-4, PD-1, and PD-L1 blockade therapies. T-cell immunoglobulin and mucin domain 3 (Tim-3) is an inhibitory receptor expressed on T cells and innate immune cells. Both in vivo and in vitro data from patients with advanced cancers support the role of Tim-3 inhibition in satisfactory anti-tumor immunity. In bladder cancer, the expression level of Tim-3 significantly increases with advanced pathological grade and T stage. Therefore, rationality implies that designing novel monoclonal antibodies reactive with Tim-3 alone or in combination with other checkpoint inhibitors may indicate a favorable response in bladder cancer. Here, we aimed to investigate the possibility of targeting Tim-3 as a novel anti-cancer treatment for bladder cancer.

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Immunotherapy Combined with Metronomic Dosing: An Effective Approach for the Treatment of NSCLC

Cancers ◽

10.3390/cancers13081901 ◽

2021 ◽

Vol 13 (8) ◽

pp. 1901

Author(s):

Eleni Skavatsou ◽

Maria Semitekolou ◽

Ioannis Morianos ◽

Theodoros Karampelas ◽

Nikolaos Lougiakis ◽

...

Keyword(s):

Immune Cell ◽

Checkpoint Inhibitors ◽

Vascular Endothelial ◽

Dose Administration ◽

In Vivo Models ◽

Pharmacokinetic Properties ◽

Strong Candidate ◽

Endothelial Growth Factor

Pioneering studies on tumor and immune cell interactions have highlighted immune checkpoint inhibitors (ICIs) as revolutionizing interventions for the management of NSCLC, typically combined with traditional MTD chemotherapies, which usually lead to toxicities and resistance to treatment. Alternatively, MTR chemotherapy is based on the daily low dose administration of chemotherapeutics, preventing tumor growth indirectly by targeting the tumor microenvironment. The effects of MTR administration of an oral prodrug of gemcitabine (OralGem), alone or with anti-PD1, were evaluated. Relevant in vitro and in vivo models were developed to investigate the efficacy of MTR alone or with immunotherapy and the potential toxicities associated with each dosing scheme. MTR OralGem restricted tumor angiogenesis by regulating thrombospondin-1 (TSP-1) and vascular endothelial growth factor A (VEGFA) expression. MTR OralGem enhanced antitumor immunity by increasing T effector responses and cytokine release, concomitant with dampening regulatory T cell populations. Promising pharmacokinetic properties afforded minimized blood and thymus toxicity and favorable bioavailability upon MTR administration compared to MTD. The combination of MTR OralGem with immunotherapy was shown to be highly efficacious and tolerable, illuminating it as a strong candidate therapeutic scheme for the treatment of NSCLC.

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Angiotensin-(1-7) and Alamandine Promote Anti-inflammatory Response in Macrophages In Vitro and In Vivo

Mediators of Inflammation ◽

10.1155/2019/2401081 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 11

Author(s):

Melissa de Carvalho Santuchi ◽

Miriane Fernandes Dutra ◽

Juliana Priscila Vago ◽

Kátia Maciel Lima ◽

Izabela Galvão ◽

...

Keyword(s):

Immune Cell ◽

Inflammatory Diseases ◽

Renin Angiotensin System ◽

M1 Macrophages ◽

Macrophage Phenotypes ◽

Anti Inflammatory ◽

Inflammation Resolution ◽

Macrophage Subsets

The renin-angiotensin system (RAS) peptides play an important role in inflammation. Resolution of inflammation contributes to restore tissue homeostasis, and it is characterized by neutrophil apoptosis and their subsequent removal by macrophages, which are remarkable plastic cells involved in the pathophysiology of diverse inflammatory diseases. However, the effects of RAS peptides on different macrophage phenotypes are still emerging. Here, we evaluated the effects of angiotensin-(1-7) (Ang-(1-7)) and the most novel RAS peptide, alamandine, on resting (M0), proinflammatory M(LPS+IFN-γ), and anti-inflammatory M(IL-4) macrophage phenotypes in vitro, as well as on specific immune cell populations and macrophage subsets into the pleural cavity of LPS-induced pleurisy in mice. Our results showed that Ang-(1-7) and alamandine, through Mas and MrgD receptors, respectively, do not affect M0 macrophages but reduce the proinflammatory TNF-α, CCL2, and IL-1β transcript expression levels in LPS+IFN-γ-stimulated macrophages. Therapeutic administration of these peptides in LPS-induced inflammation in mice decreased the number of neutrophils and M1 (F4/80lowGr1+CD11bmed) macrophage frequency without affecting the other investigated macrophage subsets. Our data suggested that both Ang-(1-7) and alamandine, through their respective receptors Mas and MrgD, promote an anti-inflammatory reprogramming of M(LPS+IFN-γ)/M1 macrophages under inflammatory circumstances and potentiate the reprogramming induced by IL-4. In conclusion, our work sheds light on the emerging proresolving properties of Ang-(1-7) and alamandine, opening new avenues for the treatment of inflammatory diseases.

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