Anticancer activity of amphipathic barbiturates

Cancer is one of the leading causes of death worldwide. Challenges related to drug resistance, side effects, and poor response rates makes cancer treatment complicated. However, different forms of immunotherapy have been effective for some cancer patients. Oncolytic therapies which can be administered directly intratumorally represent a promising form of immunotherapy. The aim of oncolytic therapies is to kill cancer cells by inducing immunogenic cell death, which can activate a natural anti-tumor immune response. The current project focused on exploring the potential of amphipathic barbiturates, herein referred to as marine product mimics (MPMs), as novel oncolytic compounds for cancer treatment. The MPMs were developed with inspiration from antimicrobial peptides and the eusynstyelamides, which are a group of natural compounds that have previously been isolated from marine animals. It was demonstrated that the compound MPM-1 could kill a range of different cell types and induced a necrosis like death in the head and neck squamous cell carcinoma cell line HSC-3. Nine additional MPMs were synthesized and included in further studies. In vitro experimentation indicated that the MPMs have an intracellular target and that they accumulate in lysosomes, which could be part of their mechanism of inducing cell death. It was demonstrated that the MPMs could cause the release and exposure of damage-associated molecular patterns which are associated with immunogenic cell death, indicating that they could have the potential to be used in oncolytic cancer therapy. An in vivo study was performed where intratumoral injections of MPM-1 were administered to mice bearing B16F1 melanoma tumors. Complete tumor remission was achieved. However, significant protective effects against a rechallenge with the same cancer cells was not observed. In summary, the MPMs demonstrate potent anticancer activity but whether they have the potential to be used in treatment of human cancer remains to be seen

Exploring the in vitro expansion of CD4 T cells. For improved culturing of CD4 T cells linked to FNAIT

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a rare disease that may cause serious bleedings in the fetus or neonate of a woman who has developed antibodies against the fetus’ platelets. Development of FNAIT has been linked to the presence of platelet reactive CD4 T cells that help B cells to develop into antibody producing plasma cells. To be able to conduct research on such T cells, the Immunology research group must be able to expand and keep them in long term cultures. Recent work revealed that several established T cell clones had started proliferating poorly. In an attempt to understand why and to possibly improve the culturing of future T cell clones, this study looked into some of the conditions that may influence the growth of these cells when expanded in vitro. The established anti- CD3 expansion culture protocol was compared to one that used PHA, but no advantage of using the latter was detected. It was demonstrated that different B-LCLs used as growth promoting feeder cells expressed varying levels of the surface molecules B7-1 and B7-2. This did however not seem to influence their feeder capacity despite the fact that expanding CD4 T cells were shown to express high levels of CD28, which costimulates growth when bound by B7. Expanding CD4 T cells also expressed the inhibitory molecule PD-1, and it was revealed that expression of its ligand, PD-L1, was induced in B-LCLs when used as feeder cells along with PBMCs. Whether this influences the efficiency of an expansion culture is yet to be determined

The marine natural product mimic MPM-1 is cytolytic and induces DAMP release from human cancer cell lines

Bioprospecting contributes to the discovery of new molecules with anticancer properties. Compounds with cytolytic activity and the ability to induce immunogenic cell death can be administered as intratumoral injections with the aim to activate anti-tumor immune responses by causing the release of tumor antigens as well as damage-associated molecular patterns (DAMPs) from dying cancer cells. In the present study, we report the cytolytic and DAMP-releasing efects of a new natural product mimic termed MPM-1 that was inspired by the marine Eusynstyelamides. We found that MPM-1 rapidly killed cancer cells in vitro by inducing a necrosis-like death, which was accompanied by lysosomal swelling and perturbation of autophagy in HSC-3 (human oral squamous cell carcinoma) cells. MPM-1 also induced release of the DAMPs adenosine triphosphate (ATP) and high mobility group box 1 (HMGB1) from Ramos (B-cell lymphoma) and HSC-3 cells, as well as cell surface expression of calreticulin in HSC-3 cells. This indicates that MPM-1 has the ability to induce immunogenic cell death, further suggesting that it may have potential as a novel anticancer compound

Exploring the in vitro expansion of CD4 T cells. For improved culturing of CD4 T cells linked to FNAIT

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a rare disease that may cause serious bleedings in the fetus or neonate of a woman who has developed antibodies against the fetus’ platelets. Development of FNAIT has been linked to the presence of platelet reactive CD4 T cells that help B cells to develop into antibody producing plasma cells. To be able to conduct research on such T cells, the Immunology research group must be able to expand and keep them in long term cultures. Recent work revealed that several established T cell clones had started proliferating poorly. In an attempt to understand why and to possibly improve the culturing of future T cell clones, this study looked into some of the conditions that may influence the growth of these cells when expanded in vitro. The established anti- CD3 expansion culture protocol was compared to one that used PHA, but no advantage of using the latter was detected. It was demonstrated that different B-LCLs used as growth promoting feeder cells expressed varying levels of the surface molecules B7-1 and B7-2. This did however not seem to influence their feeder capacity despite the fact that expanding CD4 T cells were shown to express high levels of CD28, which costimulates growth when bound by B7. Expanding CD4 T cells also expressed the inhibitory molecule PD-1, and it was revealed that expression of its ligand, PD-L1, was induced in B-LCLs when used as feeder cells along with PBMCs. Whether this influences the efficiency of an expansion culture is yet to be determined

Human and Murine Toll-like Receptor-Driven Disease in Systemic Lupus Erythematosus

The pathogenesis of systemic lupus erythematosus (SLE) is linked to the differential roles of toll-like receptors (TLRs), particularly TLR7, TLR8, and TLR9. TLR7 overexpression or gene duplication, as seen with the Y-linked autoimmune accelerator (Yaa) locus or TLR7 agonist imiquimod, correlates with increased SLE severity, and specific TLR7 polymorphisms and gainof-function variants are associated with enhanced SLE susceptibility and severity. In addition, the X-chromosome location of TLR7 and its escape from X-chromosome inactivation provide a genetic basis for female predominance in SLE. The absence of TLR8 and TLR9 have been shown to exacerbate the detrimental effects of TLR7, leading to upregulated TLR7 activity and increased disease severity in mouse models of SLE. The regulatory functions of TLR8 and TLR9 have been proposed to involve competition for the endosomal trafficking chaperone UNC93B1. However, recent evidence implies more direct, regulatory functions of TLR9 on TLR7 activity. The association between age-associated B cells (ABCs) and autoantibody production positions these cells as potential targets for treatment in SLE, but the lack of specific markers necessitates further research for precise therapeutic intervention. Therapeutically, targeting TLRs is a promising strategy for SLE treatment, with drugs like hydroxychloroquine already in clinical use

The pH-Responsive Liposomes—The Effect of PEGylation on Release Kinetics and Cellular Uptake in Glioblastoma Cells

Nanomedicine has been, to a certain degree, a success story in the development of superior anticancer therapies. However, there are tumors that remain a huge challenge for nanoformulations, for instance, brain tumors such as glioblastoma, the most common and aggressive brain tumor. To utilize the fact that such tumors are characterized by an acidic extracellular environment, we selected pH-responsive liposomes as a potential drug delivery system for superior delivery to GBM. Liposomes comprising PEGylated lipid of two chain lengths with encapsulated fluorescent marker calcein were characterized and challenged against non-PEGylated vesicles. The in vitro calcein release from three liposomal formulations (<200 nm), namely non-PEGylated (pH-Lip) and PEGylated, pH-Lip–PEG750, and pH-Lip–PEG2000, was followed at three pH conditions to prove the pH-responsiveness. The intracellular delivery of a liposomally encapsulated marker was determined in GL261 glioblastoma cell lines in vitro using both flow cytometry and confocal microscopy. The inclusion of PEG2000 within liposomal formulation resulted in reduced in vitro pH-responsiveness compared to pH-Lip and pH-Lip750. All three pH-responsive liposomal formulations improved intracellular uptake in GL261 cells compared to non-pH-responsive liposomes, with negligible differences regarding PEG length. The proposed formulations should be further evaluated in glioblastoma models

The Marine Natural Product Mimic MHP88 Shows Anticancer Activity and has the Potential to Cause Immunogenic Cell Death

Conference web site at https://www.eaa2020.org/.The marine natural product mimic MHP 88 is a novel synthetic molecule based on unique structures found in molecules from an arctic marine bryozoan. Initial studies showed that MHP88 kills cancer cells efficiently, but is not hemolytic. In this study, we look closer at the mode of death induced by MHP88 in oral cancer (HSC-3) and lymphoma (Ramos) cell lines. Immunogenic cell death (ICD) is a mode of death that can be induced by some anticancer molecules and is characterized by the release and expression of certain damage associated molecular patterns (DAMPs) which have immune stimulating effects. Specifically, the release of high mobility group box 1 (HMGB1) and ATP, as well as the translocation of calreticulin from the ER lumen to the outside of the cell membrane, constitute the major hallmarks of ICD. In vivo ICD induced in cancer cells has a vaccination effect, protecting the host from future challenge with the same cancer cells. Animal studies on the molecule LTX-401, which has similar properties as MHP88 have successfully demonstrated this effect

The Marine Natural Product Mimic MHP88 Shows Anticancer Activity and has the Potential to Cause Immunogenic Cell Death

The marine natural product mimic MHP 88 is a novel synthetic molecule based on unique structures found in molecules from an arctic marine bryozoan. Initial studies showed that MHP88 kills cancer cells efficiently, but is not hemolytic. In this study, we look closer at the mode of death induced by MHP88 in oral cancer (HSC-3) and lymphoma (Ramos) cell lines. Immunogenic cell death (ICD) is a mode of death that can be induced by some anticancer molecules and is characterized by the release and expression of certain damage associated molecular patterns (DAMPs) which have immune stimulating effects. Specifically, the release of high mobility group box 1 (HMGB1) and ATP, as well as the translocation of calreticulin from the ER lumen to the outside of the cell membrane, constitute the major hallmarks of ICD. In vivo ICD induced in cancer cells has a vaccination effect, protecting the host from future challenge with the same cancer cells. Animal studies on the molecule LTX-401, which has similar properties as MHP88 have successfully demonstrated this effect

Amphipathic barbiturates as marine product mimics with cytolytic and immunogenic effects on head and neck squamous cell carcinoma cell lines

Publisher Copyright: Copyright © 2023 von Hofsten, Langer, Korelin, Magnussen, Ausbacher, Anderssen, Salo, Strøm, Bayer, Al-Samadi and Berge.The incidence of head and neck squamous cell carcinoma (HNSCC) is increasing and the conventional treatments for this form of cancer can be tough. Despite the success of existing immunotherapies in some HNSCC patients, many do not respond to this type of treatment. Thus, the development of novel anti-cancer therapies should be prioritized. In the current study, the anticancer activity of a panel of novel compounds, herein termed marine product mimics (MPMs), against HNSCC cell lines is explored. The previously reported compound MPM-1, which is structurally related to the novel MPMs, was shown to have promising effects on the HNSCC cell line HSC-3. The results from the current study indicate that the novel MPMs are more potent than MPM-1 but cause a similar type of cell death. The results indicated that the MPMs must cross through the cell membrane to exert their action and that they are lysosomotropic. Further experiments showed that some of the MPMs could induce phosphorylation of eukaryotic initiation factor 2α (eIF2α) in HSC-3 and UT-SCC-24A cells, which indicates that they can activate the integrated stress response that is strongly associated with immunogenic cell death. Cell surface expression of calreticulin and release of HMGB1 and ATP, which are all hallmarks of immunogenic cell death, was also demonstrated in HSC-3 and UT-SCC-24A cells treated with MPMs. This suggests that the MPMs are interesting candidates for future HNSCC cancer therapies.Peer reviewe

The selectivity of galardin and an azasugar-based hydroxamate compound for human matrix metalloproteases and bacterial metalloproteases

<div><p>Inhibitors targeting bacterial enzymes should not interfere with enzymes of the host, and knowledge about structural determinants for selectivity is important for designing inhibitors with a therapeutic potential. We have determined the binding strengths of two hydroxamate compounds, galardin and compound <b>1b</b> for the bacterial zinc metalloproteases, thermolysin, pseudolysin and auerolysin, known to be bacterial virulence factors, and the two human zinc metalloproteases MMP-9 and MMP-14. The active sites of the bacterial and human enzymes have huge similarities. In addition, we also studied the enzyme-inhibitor interactions by molecular modelling. The obtained <i>K</i>_i values of galardin for MMP-9 and MMP-14 and compound <b>1b</b> for MMP-9 are approximately ten times lower than previously reported. Compound <b>1b</b> binds stronger than galardin to both MMP-9 and MMP-14, and docking studies indicated that the diphenyl ether moiety of compound <b>1b</b> obtains more favourable interactions within the S´₁-subpocket than the 4-methylpentanoyl moiety of galardin. Both compounds bind stronger to MMP-9 than to MMP-14, which appears to be due to a larger S´₁-subpocket in the former enzyme. Galardin, but not <b>1b</b>, inhibits the bacterial enzymes, but the galardin <i>K</i>_i values were much larger than for the MMPs. The docking indicates that the S´₁-subpockets of the bacterial proteases are too small to accommodate the diphenyl ether moiety of <b>1b</b>, while the 4-methylpentanoyl moiety of galardin enters the pocket. The present study indicates that the size and shape of the ligand structural moiety entering the S´₁-subpocket is an important determinant for selectivity between the studied MMPs and bacterial MPs.</p></div