Perforin and Granzyme B Expressed by Murine Myeloid-Derived Suppressor Cells: A Study on Their Role in Outgrowth of Cancer Cells

A wide-range of myeloid-derived suppressor cell (MDSC)-mediated immune suppressive functions has previously been described. Nevertheless, potential novel mechanisms by which MDSCs aid tumor progression are, in all likelihood, still unrecognized. Next to its well-known expression in natural killer cells and cytotoxic T lymphocytes (CTLs), granzyme B (GzmB) expression has been found in different cell types. In an MDSC culture model, we demonstrated perforin and GzmB expression. Furthermore, similar observations were made in MDSCs isolated from tumor-bearing mice. Even in MDSCs from humans, GzmB expression was demonstrated. Of note, B16F10 melanoma cells co-cultured with perforin/GzmB knock out mice (KO) MDSCs displayed a remarkable decrease in invasive potential. B16F10 melanoma cells co-injected with KO MDSCs, displayed a significant slower growth curve compared to tumor cells co-injected with wild type (WT) MDSCs. In vivo absence of perforin/GzmB in MDSCs resulted in a higher number of CD8+ T-cells. Despite this change in favor of CD8+ T-cell infiltration, we observed low interferon-¿ (IFN-¿) and high programmed death-ligand 1 (PD-L1) expression, suggesting that other immunosuppressive mechanisms render these CD8+ T-cells dysfunctional. Taken together, our results suggest that GzmB expression in MDSCs is another means to promote tumor growth and warrants further investigation to unravel the exact underlying mechanism

Radioresistance of Human Cancers: Clinical Implications of Genetic Expression Signatures

Although radiotherapy is given to more than 50% of cancer patients, little progress has been made in identifying optimal radiotherapy - drug combinations to improve treatment efficacy. Using molecular data from The Cancer Genome Atlas (TCGA), we extracted a total of 1016 cancer patients that received radiotherapy. The patients were diagnosed with head-and-neck (HNSC - 294 patients), cervical (CESC - 166 patients) and breast (BRCA - 549 patients) cancer. We analyzed mRNA expression patterns of 50 hallmark gene sets of the MSigDB collection, which we divided in eight categories based on a shared biological or functional process. Tumor samples were split into upregulated, neutral or downregulated mRNA expression for all gene sets using a gene set analysis (GSEA) pre-ranked analysis and assessed for their clinical relevance. We found a prognostic association between three of the eight gene set categories (Radiobiological, Metabolism and Proliferation) and overall survival in all three cancer types. Furthermore, multiple single associations were revealed in the other categories considered. To the best of our knowledge, our study is the first report suggesting clinical relevance of molecular characterization based on hallmark gene sets to refine radiation strategies.</jats:p

Ferroptosis: Frenemy of Radiotherapy

Recently, it was established that ferroptosis, a type of iron-dependent regulated cell death, plays a prominent role in radiotherapy-triggered cell death. Accordingly, ferroptosis inducers attracted a lot of interest as potential radio-synergizing drugs, ultimately enhancing radioresponses and patient outcomes. Nevertheless, the tumor microenvironment seems to have a major impact on ferroptosis induction. The influence of hypoxic conditions is an area of interest, as it remains the principal hurdle in the field of radiotherapy. In this review, we focus on the implications of hypoxic conditions on ferroptosis, contemplating the plausibility of using ferroptosis inducers as clinical radiosensitizers. Furthermore, we dive into the prospects of drug repurposing in the domain of ferroptosis inducers and radiosensitizers. Lastly, the potential adverse effects of ferroptosis inducers on normal tissue were discussed in detail. This review will provide an important framework for subsequent ferroptosis research, ascertaining the feasibility of ferroptosis inducers as clinical radiosensitizers

Antidiabetic Biguanides Radiosensitize Hypoxic Colorectal Cancer Cells Through a Decrease in Oxygen Consumption

Background and Purpose: The anti-diabetic biguanide drugs metformin and phenformin exhibit antitumor activity in various models. However, their radiomodulatory effect under hypoxic conditions, particularly for phenformin, is largely unknown. This study therefore examines whether metformin and phenformin as mitochondrial complex I blockades could overcome hypoxic radioresistance through inhibition of oxygen consumption. Materials and Methods: A panel of colorectal cancer cells (HCT116, DLD-1, HT29, SW480, and CT26) was exposed to metformin or phenformin for 16 h at indicated concentrations. Afterward, cell viability was measured by MTT and colony formation assays. Apoptosis and reactive oxygen species (ROS) were detected by flow cytometry. Phosphorylation of AMP-activated protein kinase (AMPK) was examined by western blot. Mitochondria complexes activity and oxygen consumption rate (OCR) were measured by seahorse analyzer. The radiosensitivity of tumor cells was assessed by colony formation assay under aerobic and hypoxic conditions. The in vitro findings were further validated in colorectal CT26 tumor model. Results: Metformin and phenformin inhibited mitochondrial complex I activity and subsequently reduced OCR in a dose-dependent manner starting at 3 mM and 30 μM, respectively. As a result, the hypoxic radioresistance of tumor cells was counteracted by metformin and phenformin with an enhancement ratio about 2 at 9 mM and 100 μM, respectively. Regarding intrinsic radioresistance, both of them did not exhibit any effect although there was an increase of phosphorylation of AMPK and ROS production. In tumor-bearing mice, metformin or phenformin alone did not show any anti-tumor effect. While in combination with radiation, both of them substantially delayed tumor growth and enhanced radioresponse, respectively, by 1.3 and 1.5-fold. Conclusion: Our results demonstrate that metformin and phenformin overcome hypoxic radioresistance through inhibition of mitochondrial respiration, and provide a rationale to explore metformin and phenformin as hypoxic radiosensitizers

Table_1_Radioresistance of Human Cancers: Clinical Implications of Genetic Expression Signatures.docx

Although radiotherapy is given to more than 50% of cancer patients, little progress has been made in identifying optimal radiotherapy - drug combinations to improve treatment efficacy. Using molecular data from The Cancer Genome Atlas (TCGA), we extracted a total of 1016 cancer patients that received radiotherapy. The patients were diagnosed with head-and-neck (HNSC - 294 patients), cervical (CESC - 166 patients) and breast (BRCA - 549 patients) cancer. We analyzed mRNA expression patterns of 50 hallmark gene sets of the MSigDB collection, which we divided in eight categories based on a shared biological or functional process. Tumor samples were split into upregulated, neutral or downregulated mRNA expression for all gene sets using a gene set analysis (GSEA) pre-ranked analysis and assessed for their clinical relevance. We found a prognostic association between three of the eight gene set categories (Radiobiological, Metabolism and Proliferation) and overall survival in all three cancer types. Furthermore, multiple single associations were revealed in the other categories considered. To the best of our knowledge, our study is the first report suggesting clinical relevance of molecular characterization based on hallmark gene sets to refine radiation strategies.</p

Exploring the Metabolic Impact of FLASH Radiotherapy

FLASH radiotherapy (FLASH RT) is an innovative modality in cancer treatment that delivers ultrahigh dose rates (UHDRs), distinguishing it from conventional radiotherapy (CRT). FLASH RT has demonstrated the potential to enhance the therapeutic window by reducing radiation-induced damage to normal tissues while maintaining tumor control, a phenomenon termed the FLASH effect. Despite promising outcomes, the precise mechanisms underlying the FLASH effect remain elusive and are a focal point of current research. This review explores the metabolic and cellular responses to FLASH RT compared to CRT, with particular focus on the differential impacts on normal and tumor tissues. Key findings suggest that FLASH RT may mitigate damage in healthy tissues via altered reactive oxygen species (ROS) dynamics, which attenuate downstream oxidative damage. Studies indicate the FLASH RT influences iron metabolism and lipid peroxidation pathways differently than CRT. Additionally, various studies indicate that FLASH RT promotes the preservation of mitochondrial integrity and function, which helps maintain apoptotic pathways in normal tissues, attenuating damage. Current knowledge of the metabolic influences following FLASH RT highlights its potential to minimize toxicity in normal tissues, while also emphasizing the need for further studies in biologically relevant, complex systems to better understand its clinical potential. By targeting distinct metabolic pathways, FLASH RT could represent a transformative advance in RT, ultimately improving the therapeutic window for cancer treatment

Retroviral and Lentiviral Vectors for the Induction of Immunological Tolerance

Retroviral and lentiviral vectors have proven to be particularly efficient systems to deliver genes of interest into target cells, either in vivo or in cell cultures. They have been used for some time for gene therapy and the development of gene vaccines. Recently retroviral and lentiviral vectors have been used to generate tolerogenic dendritic cells, key professional antigen presenting cells that regulate immune responses. Thus, three main approaches have been undertaken to induce immunological tolerance; delivery of potent immunosuppressive cytokines and other molecules, modification of intracellular signalling pathways in dendritic cells, and de-targeting transgene expression from dendritic cells using microRNA technology. In this review we briefly describe retroviral and lentiviral vector biology, and their application to induce immunological tolerance

Perforin and Granzyme B Expressed by Murine Myeloid-Derived Suppressor Cells: A Study on Their Role in Outgrowth of Cancer Cells

A wide-range of myeloid-derived suppressor cell (MDSC)-mediated immune suppressive functions has previously been described. Nevertheless, potential novel mechanisms by which MDSCs aid tumor progression are, in all likelihood, still unrecognized. Next to its well-known expression in natural killer cells and cytotoxic T lymphocytes (CTLs), granzyme B (GzmB) expression has been found in different cell types. In an MDSC culture model, we demonstrated perforin and GzmB expression. Furthermore, similar observations were made in MDSCs isolated from tumor-bearing mice. Even in MDSCs from humans, GzmB expression was demonstrated. Of note, B16F10 melanoma cells co-cultured with perforin/GzmB knock out mice (KO) MDSCs displayed a remarkable decrease in invasive potential. B16F10 melanoma cells co-injected with KO MDSCs, displayed a significant slower growth curve compared to tumor cells co-injected with wild type (WT) MDSCs. In vivo absence of perforin/GzmB in MDSCs resulted in a higher number of CD8+ T-cells. Despite this change in favor of CD8+ T-cell infiltration, we observed low interferon-γ (IFN-γ) and high programmed death-ligand 1 (PD-L1) expression, suggesting that other immunosuppressive mechanisms render these CD8+ T-cells dysfunctional. Taken together, our results suggest that GzmB expression in MDSCs is another means to promote tumor growth and warrants further investigation to unravel the exact underlying mechanism.</jats:p

Intratumoral Combinatorial Administration of CD1c (BDCA-1)+ Myeloid Dendritic Cells Plus Ipilimumab and Avelumab in Combination with Intravenous Low-Dose Nivolumab in Patients with Advanced Solid Tumors: A Phase IB Clinical Trial

Intratumoral (IT) myeloid dendritic cells (myDCs) play a pivotal role in re-licensing antitumor cytotoxic T lymphocytes. IT injection of the IgG1 monoclonal antibodies ipilimumab and avelumab may induce antibody-dependent cellular cytotoxicity, thereby enhancing the release of tumor antigens that can be captured and processed by CD1c (BDCA-1)+ myDCs. Patients with advanced solid tumors after standard care were eligible for IT injections of &ge;1 lesion with ipilimumab (10 mg) and avelumab (40 mg) and intravenous (IV) nivolumab (10 mg) on day 1, followed by IT injection of autologous CD1c (BDCA-1)+ myDCs on day 2. IT/IV administration of ipilimumab, avelumab, and nivolumab was repeated bi-weekly. Primary objectives were safety and feasibility. Nine patients were treated with a median of 21 &times; 106 CD1c (BDCA-1)+ myDCs, and a median of 4 IT/IV administrations of ipilimumab, avelumab, and nivolumab. The treatment was safe with mainly injection-site reactions, but also immune-related pneumonitis (n = 2), colitis (n = 1), and bullous pemphigoid (n = 1). The best response was a durable partial response in a patient with stage IV melanoma who previously progressed on checkpoint inhibitors. Our combinatorial therapeutic approach, including IT injection of CD1c (BDCA-1)+ myDCs, is feasible and safe, and it resulted in encouraging signs of antitumor activity in patients with advanced solid tumors

Potential of memory T cells in bridging preoperative chemoradiation and immunotherapy in rectal cancer.

The management of locally advanced rectal cancer has passed a long way of developments, where total mesorectal excision and preoperative radiotherapy are crucial to secure clinical outcome. These and other aspects of multidisciplinary strategies are in-depth summarized in the literature, while our mini-review pursues a different goal. From an ethical and medical standpoint, we witness a delayed implementation of novel therapies given the cost/time consuming process of organizing randomized trials that would bridge an already excellent local control in cT3-4 node-positive disease with long-term survival. This unfortunate separation of clinical research and medical care provides a strong motivation to repurpose known pharmaceuticals that suit for treatment intensification with a focus on distant control. In the framework of on-going phase II-III IG/IMRT-SIB trials, we came across an intriguing translational observation that the ratio of circulating (protumor) myeloid-derived suppressor cells to (antitumor) central memory CD8+ T cells is drastically increased, a possible mechanism of tumor immuno-escape and spread. This finding prompts that restoring the CD45RO memory T-cell pool could be a part of integrated adjuvant interventions. Therefore, the immunocorrective potentials of modified IL-2 and the anti-diabetic drug metformin are thoroughly discussed in the context of tumor immunobiology, mTOR pathways and revised Warburg effect