Experimental Anticancer Drug Research Articles

Enhancing Anti-PD-1 Immunotherapy by Targeting MDSCs via Hepatic Arterial Infusion in Breast Cancer Liver Metastases.

Surgery, chemotherapy, and radiation often have limited utility for advanced metastatic disease in the liver, and despite its promising activity in select cancers, PD-1 blockade therapy similarly has minimal benefit in this setting. Curaxin, CBL0137, is an experimental anti-cancer drug that disrupts the binding of DNA to histones, destabilizes chromatin, and induces Z-DNA formation which may stimulate anti-tumor immune responses. Murine cell lines of colon (CT26) and breast (4T1) cancer were interrogated for survival and CBL0137-associated DNA changes in vitro. Immunocompetent models of liver metastases followed by CBL0137 hepatic arterial infusion (HAI) were used to examine in vivo tumor cell DNA alterations, treatment responses, and the immune contexture associated with CBL0137, both alone and in combination with anti-PD-1 therapy. CBL0137 induced immediate changes to favor tumor cell death in vitro and in vivo with an efficient tumor uptake via the HAI route. Toxicity to CBL0137 was minimal and anti-tumor treatment effects were more efficient with HAI compared to intravenous delivery. Immune effects were pronounced with CBL0137 HAI with concurrent depletion of a specific population of myeloid-derived suppressor cells and maintenance of effector T cell populations. Combination of CBL0137 HAI with PD-1 blockade improved survival in 4T1 tumors but not in CT26 tumors, and therapeutic efficacy relies on the finding of simultaneous and targeted depletion of myeloid-derived suppressor cells and skewing of T cell populations to produce synergy with PD-1 blockade therapy.

Abstract CT105: Early clinical experience with a novel first-in-class G-quadruplex experimental anti-cancer drug

Abstract The human genome contains G-rich regions that are over-represented in many cancer gene promoters. These can form higher-order G-quadruplex arrangements under the influence of an appropriate and selective small molecule compound. The resulting ligand-G-quadruplex complex may be resistant to polymerase read-through and thus results in transcriptional down-regulation of target genes. QN-302, based on a tetra-substituted naphthalene diimide chemotype, is a potent and selective G-quadruplex-binding compound that has been designed and initially developed at University College London. It displays single-digit cell growth inhibition in a panel of pancreatic ductal adenocarcinoma (PDAC) cell lines. RNA-seq analysis shows that it is a pan-quadruplex agent, down-regulating the expression of multiple G-quadruplex-promoter cancer genes and pathways in PDAC. It also shows in vivo anti-tumor activity in several animal models for PDAC, with low toxicity observed at therapeutic doses. QN-302 was licensed to Qualigen Therapeutics Inc in January 2022. It was granted Orphan Drug designation by the FDA in December 2022 for PDAC. This was followed by extensive regulatory toxicological and pharmacological studies together with successful scale-up synthesis, purification and formulation. QN-302 was granted IND clearance in August 2023 by the FDA for phase 1 clinical trials in advanced or metastatic solid tumors. This Phase 1a clinical trial is being conducted in two USA cancer centers (START Midwest, Grand Rapids, MI and Honor Health, Scottsdale, AZ) using a protocol based on 28-day once-weekly dose escalation and evaluation of three potential biomarkers, two of which have been suggested by transcriptomic analysis as being indicative of G-quadruplex-induced drug response and accompanying decrease of anti-tumor activity. So far 1 Cohort (3 patients) has been dosed at the lowest dose of 1.62 mg of QN-302 as part of this Phase 1a clinical trial. Patient #1, a male patient of metastatic pancreatic cancer, is currently in Cycle 4 of therapy showing Stable Disease and no AEs or SAEs reported. Patient #2, a female patient of metastatic colorectal cancer, completed Cycle 2 after which she showed progression upon CT Scan before entering Cycle 3 and was taken off study. Patient #3, a male patient of metastatic PDAC, is currently ending Cycle 2, about to enter Cycle 3, showing Stable Disease upon CT scan. All patients dosed showed no AEs, SAEs or side effects. They were also clinically observed to have improved QOL compared to prior SOC while on therapy with QN-302, This presentation will outline this unique approach and summarize safety, tolerability, PK/PD data and any early clinical data as described above including preliminary biomarker data now that clinical evaluation has commenced. The advantages and challenges of QN-302 will also be discussed. Citation Format: Tariq Arshad, Stephen Neidle, Sreenivasa Chandana, Erkut Borazanci. Early clinical experience with a novel first-in-class G-quadruplex experimental anti-cancer drug [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT105.

Machine learning-aided search for ligands of P2Y6 and other P2Y receptors

The P2Y6 receptor, activated by uridine diphosphate (UDP), is a target for antagonists in inflammatory, neurodegenerative, and metabolic disorders, yet few potent and selective antagonists are known to date. This prompted us to use machine learning as a novel approach to aid ligand discovery, with pharmacological evaluation at three P2YR subtypes: initially P2Y6 and subsequently P2Y1 and P2Y14. Relying on extensive published data for P2Y6R agonists, we generated and validated an array of classification machine learning model using the algorithms deep learning (DL), adaboost classifier (ada), Bernoulli NB (bnb), k-nearest neighbors (kNN) classifier, logistic regression (lreg), random forest classifier (rf), support vector classification (SVC), and XGBoost (XGB) classifier models, and the common consensus was applied to molecular selection of 21 diverse structures. Compounds were screened using human P2Y6R-induced functional calcium transients in transfected 1321N1 astrocytoma cells and fluorescent binding inhibition at closely related hP2Y14R expressed in CHO cells. The hit compound ABBV-744, an experimental anticancer drug with a 6-methyl-7-oxo-6,7-dihydro-1H-pyrrolo[2,3-c]pyridine scaffold, had multifaceted interactions with the P2YR family: hP2Y6R inhibition in a non-surmountable fashion, suggesting that noncompetitive antagonism, and hP2Y1R enhancement, but not hP2Y14R binding inhibition. Other machine learning-selected compounds were either weak (experimental anti-asthmatic drug AZD5423 with a phenyl-1H-indazole scaffold) or inactive in inhibiting the hP2Y6R. Experimental drugs TAK-593 and GSK1070916 (100 µM) inhibited P2Y14R fluorescent binding by 50% and 38%, respectively, and all other compounds by < 20%. Thus, machine learning has led the way toward revealing previously unknown modulators of several P2YR subtypes that have varied effects.

Pharmacological Targeting of Senescence with Senolytics as a New Therapeutic Strategy for Neurodegeneration.

Cellular senescence is a state of permanent cell-cycle arrest. Early in life, senescence has a physiologic role in tumor suppression and wound healing. However, gradually, as these senescent cells accumulate over the lifespan of an organism, they contribute to inflammation and the progression of age-related diseases, including neurodegeneration. Targeting senescent cells using a class of drugs known as "senolytics" holds great promise for the management of Alzheimer's and Parkinson's disease. Already, several senolytic compounds have been shown to ameliorate cognitive deficits across several preclinical models of neurodegeneration. Most of these senolytics (e.g., dasatinib) are repurposed clinical or experimental anticancer drugs, which trigger apoptosis of senescent cells by interfering with pro-survival pathways. However, outside of their senolytic function, many first-generation senolytics also have other less appreciated neuroprotective effects, such as potent antioxidant and anti-inflammatory activity. In addition, some senolytic drugs may also have negative dose-limiting toxicities, including thrombocytopenia. In this review, we discuss the various biologic pathways targeted by the leading senolytic drugs, namely dasatinib, quercetin, fisetin, and navitoclax. We further evaluate the clinical transability of these compounds for neurodegeneration, assessing their adverse effects, pharmacokinetic properties, and chemical structure. SIGNIFICANCE STATEMENT: Currently, there are no effective disease-modifying treatments for the most prevalent neurodegenerative disorders, including Alzheimer's and Parkinson's disease. Some of the drugs currently available for treating these diseases are associated with unwanted side-effects and/or become less efficacious with time. Therefore, researchers have begun to explore new innovative treatments for these belligerent diseases, including senolytic drugs. These agents lead to the apoptosis of senescent cells thereby preventing their deleterious role in neurodegeneration.

Molecules promoting circulating clusters of cancer cells suggest novel therapeutic targets for treatment of metastatic cancers.

Treatment of metastatic disease remains among the most challenging tasks in oncology. One of the early events that predicts a poor prognosis and precedes the development of metastasis is the occurrence of clusters of cancer cells in the blood flow. Moreover, the presence of heterogeneous clusters of cancerous and noncancerous cells in the circulation is even more dangerous. Review of pathological mechanisms and biological molecules directly involved in the formation and pathogenesis of the heterotypic circulating tumor cell (CTC) clusters revealed their common properties, which include increased adhesiveness, combined epithelial-mesenchymal phenotype, CTC-white blood cell interaction, and polyploidy. Several molecules involved in the heterotypic CTC interactions and their metastatic properties, including IL6R, CXCR4 and EPCAM, are targets of approved or experimental anticancer drugs. Accordingly, analysis of patient survival data from the published literature and public datasets revealed that the expression of several molecules affecting the formation of CTC clusters predicts patient survival in multiple cancer types. Thus, targeting of molecules involved in CTC heterotypic interactions might be a valuable strategy for the treatment of metastatic cancers.

IDENTIFICATION OF ANTI-CANCEROUS DRUGS FOR THE MUTATED SNAP25 PROTEIN RELATED TO BRAIN TUMOR THROUGH STRUCTURE-BASED VIRTUAL SCREENING APPROACH

Objective: Brain tumor is a formidable challenge for drug development, and drugs derived from many advanced technologies are being tested in clinical trials. Synaptosomal-associated protein of 25 kDa (SNAP25) is a membrane-binding protein in neurons and it is critical in neurotransmission for the fusion of plasma membrane and synaptic vesicle making it a prime target to address brain tumors. The SNAP-25 gene is responsible for personality disorders, schizophrenia, attention deficit, and hyperactivity disorder in human beings. It is recently discovered, that this protein is responsible for brain cancer as well. Methods: In the present research, 17 investigational and experimental anticancer drugs were selected from the PubChem and DrugBank databases to identify potential inhibitors with high stability to treat mutated SNAP25 protein. For this purpose, we have used the structure-based virtual screening technique wherein, the candidate molecules are computationally docked into the 3D structure of the biological. The docking was achieved in PyRx 0.8 software and the drugs were then ranked based on their predicted binding affinity or complementarity to the binding site. Results: Based on the ligand binding energy, the top six compounds having greater inhibitory effects towards SNAP25 were selected and then visualized with Pymol and Biovia visualizers. The compound Crenolanib has better pharmacological properties and demonstrated higher binding affinities with the target protein. Therefore, this Crenolanib docked confirmations were appraised for molecular dynamic simulations. Conclusion: The study concluded that the anticancer drug Crenolanib exerted inhibitory potential against the mutated protein SNAP-25 and therefore it can be exploited as a cancer modulator to address brain tumors.

Abstract 5380: Anticancer compound ONC201 induces prolonged stress and arrest of proliferation in BT474 human breast cancer cells without induction of cell death

Abstract Background and goals: ONC201 an experimental anticancer drug, targeting mitochondrial caseinolytic peptidase, ClpP is currently under investigation. Current data on the effect of ONC201 along with observations that ONC201 causes cytotoxic effect, there are data demonstrating that ONC201 is rather cytostatic and does not induce cell death. Recently, we demonstrated ONC201-induced arrest of proliferation and a decrease in the number of mitochondrial nucleoids. Here we extended our study to describe the effect of short-and long-term exposure of BT474 cells to ONC201. Observations: Proliferation and cell cycle arrest. ONC201 in a dose-dependent manner (0-50 µM) inhibited the proliferation of BT474 cells without induction of apoptotic cell death. Cells exposed to 10 µM ONC201 accumulated in G0/G1 phase (49.4 ± 11.5% vs 78.7 ± 5.1%) and declined in the S-phase of the cell cycle (39.2 ± 6.6% vs 9.0 ± 2.1%). In parallel, ONC201 exposure induced a decline in Cyclin E and Cdk2 protein levels. Nucleoids and mtDNA. Exposure of BT474 cells, to 10 µM ONC 201, decreased the number of mitochondrial nucleoids from 249 ± 52 to 155 ± 38 per cell after 24 h. Longer 72 h exposure further reduced the number of nucleoids to 84 ± 36 per cell. ONC201 exposure also induced reduction of mitochondrial size from 2.8 ± 0.5 µm2 to 1.5 ± 0.2 µm2 and depletion of mtDNA. Short-term and long-term consequences of cell exposure to ONC201. Short-term treatment (24 h exposure) demonstrated the reversible effect of ONC201 on the expression of stress proteins (ATF4, CHOP, and GDF-15), cell cycle regulatory proteins (cyclin E, Cdk2), and partial restoration of mitochondrial nucleoids (to 214 ± 46 nucleoids per cell). On the contrary, long-term treatment (72 h) resulted in the irreversible and persistent arrest of proliferation, decline of cell cycle regulatory proteins, and activation of stress proteins. Respiration. ONC201 exposure suppressed State 3, State 4, and uncoupled respiration in digitonin-permeabilized BT474 cells. Complex I activity was decreased from 3.3 ± 0.4 ng-atoms O/min/106 cells to 0.5 ± 0.1 and 0.4 ± 0.3 ng-atoms O/min/106 cells, in cells exposed to ONC201 for 24h and 72h, respectively. Similarly, ONC201 decreased the activity of Complex II from 3.5 ± 0.5 ng-atoms O/min/106 cells to 1.5 ± 0.6 and 0.3 ± 0.1 ng-atoms O/min/106. Conclusions: The consequences of ONC201 treatment were dependent on the duration of exposure to the drug. Short-term (24 h) exposure of BT474 cells resulted in reversible induction of stress proteins (ATF4, CHOP, and GDF-15), which were reversed following 120 h incubation in drug-free media. On the contrary, long-term (72 h) exposure of cells to ONC201 induced irreversible and persistent arrest of proliferation and elevation of expression of stress proteins. Thus, ONC201 can induce prolonged and irreversible elevation of stress proteins and persistent G1/S cell cycle arrest. Citation Format: Artem A. Mishukov, Irina V. Odinokova, Serazhutdin A. Abdullaev, Vitaly K. Zhalimov, Emily M. Fennel, Lucas J. Aponte-Collazo, Paul R. Graves, Lee M. Graves, Ekhson L. Holmuhamedov. Anticancer compound ONC201 induces prolonged stress and arrest of proliferation in BT474 human breast cancer cells without induction of cell death [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5380.

Reactions of proteins with a few organopalladium compounds of medicinal interest.

Pd compounds form a promising class of experimental anticancer drug candidates whose mechanism of action is still largely unknown; in particular, a few organopalladium compounds seem very attractive. To gain mechanistic insight into medicinal palladium compounds, we have explored here – through ESI MS analysis – the interactions of four organopalladium agents (1–4) – showing remarkable in vitro antiproliferative properties – with a few representative model proteins, i.e., lysozyme (HEWL), ribonuclease A (RNase), and carbonic anhydrase (hCAI). The tested panel included three Pd allyl compounds with one or two carbene ligands and a palladacyclopentadienyl complex. Notably, the Pd allyl compounds turned out to manifest, on the whole, a modest tendency to react with the above proteins. Only complex 3 produced small amounts of characteristic adducts with hCAI bearing either one or two Pd allyl groups. In contrast, the palladacyclopentadienyl complex 4 manifested a greater and peculiar reactivity with all the above proteins generating invariably protein adducts with a mass increase of +256 Da where a butadienyl group – with no associated Pd – is attached to the proteins. Afterwards, we extended our investigations to the C-terminal dodecapeptide of thioredoxin reductase bearing the –Cys–Sec– reactive motif. In this latter case adducts were formed with all tested Pd compounds; however, complex 4 manifested towards this dodecapeptide a type of reactivity deeply different from that observed with HEWL, RNase A and hCAI. The mechanistic implications of these findings are discussed.

The BH3 mimetic compound BH3I-1 impairs mitochondrial dynamics and promotes stress response in addition to its pro-apoptotic key function

BH3 mimetics, such as BH3I-1, act as Bcl-2 antagonists, promote apoptosis and are used in basic research studies on apoptotic signaling and are currently tested as experimental anti-tumor agents. The present study addresses time- and dose-dependent responses of BH3I-1 on apoptosis, cellular stress defense mediated by heme oxygenase-1 (HO-1), and mitochondrial morphology. As expected, treatment of normal human dermal fibroblasts with BH3I-1 induced apoptosis as determined by typical markers including cytochrome c release, loss of procaspase-3, and PARP cleavage. Induction of the cellular stress response marker HO-1 precedes apoptosis induction whereas fragmentation of the mitochondrial network was triggered even more rapidly. No difference in apoptosis induction was found upon depletion of HO-1 by siRNA compared to controls suggesting that apoptosis induction by BH3I-1 is not affected by HO-1. To evaluate the functional interplay between mitochondrial fragmentation and HO-1 induction, murine embryonic fibroblasts lacking the fission factor Drp1 were used. In Drp1 knock out cells, HO-1 levels were low compared to wild type cells, both in untreated controls as well as after BH3I-1 exposure, demonstrating that Drp1 is at least in part required for determining basal and inducible HO-1 levels. Considering the sequence of events, it was shown here that BH3I-1 dependent apoptosis is a rather late event, while effects on mitochondrial morphology and cellular stress response (HO-1 induction) are observed rapidly after exposure of cells to the compound. We propose that BH3I-1 is a valuable tool for studying cellular stress responses as well as mitochondrial dynamics in future studies. Since BH3 mimetics are promising experimental anticancer drugs, our data further imply that additional biological effects such as upregulation of detoxifying systems or changes in mitochondrial dynamics could interfere, in combination therapy, with selective drug toxicity and thus need to be taken into account for drug development.

Innovative Methodology in the Discovery of Novel Drug Targets in the Free-Living Amoebae.

Despite advances in drug discovery and modifications in the chemotherapeutic regimens, human infections caused by free-living amoebae (FLA) have high mortality rates (~95%). The FLA that cause fatal human cerebral infections include Naegleria fowleri, Balamuthia mandrillaris and Acanthamoeba spp. Novel drug-target discovery remains the only viable option to tackle these central nervous system (CNS) infection in order to lower the mortality rates caused by the FLA. Of these FLA, N. fowleri causes primary amoebic meningoencephalitis (PAM), while the A. castellanii and B. Mandrillaris are known to cause granulomatous amoebic encephalitis (GAE). The infections caused by the FLA have been treated with drugs like Rifampin, Fluconazole, Amphotericin-B and Miltefosine. Miltefosine is an anti-leishmanial agent and an experimental anti-cancer drug. With only rare incidences of success, these drugs have remained unsuccessful to lower the mortality rates of the cerebral infection caused by FLA. Recently, with the help of bioinformatic computational tools and the discovered genomic data of the FLA, discovery of newer drug targets has become possible. These cellular targets are proteins that are either unique to the FLA or shared between the humans and these unicellular eukaryotes. The latter group of proteins has shown to be targets of some FDA approved drugs prescribed in non-infectious diseases. This review out-lines the bioinformatics methodologies that can be used in the discovery of such novel drug-targets, their chronicle by in-vitro assays done in the past and the translational value of such target discoveries in human diseases caused by FLA.

419TiP - Chemosensitization of carboplatin by NOX66: Pharmacokinetics and safety

Background: The experimental anti-cancer drug, idronoxil, is a selective inhibitor of PI3K/AKT in tumor cells, with studies showing it to be a potent chemosensitizing agent of carboplatin in vitro and in animal studies across a wide range of cancer cell types. These results, however, have not translated to clinical efficacy, with a Phase 3 study of combined oral idronoxil and intravenous carboplatin in patients with late-stage platinum-refractory ovarian cancer discontinued with no efficacy seen. This lack of efficacy is now thought to be due to complete conversion of idronoxil to bio-inactive Phase 2 metabolites. NOX66 is a suppository formulation of idronoxil designed to protect the drug from Phase 2 metabolism. This first-in-human study will look at the ability of NOX66 to deliver relatively high levels of idronoxil in a bio-active form, investigating (a) PK and (b) safety of NOX66 administration both as a monotherapy and in combination with carboplatin. Trial design: This is an open label, Phase 1 PK and safety study of NOX66 as a monotherapy and in combination with carboplatin. Patients included have end stage, refactory solid tumours, and no further therapy options available. A total of 16 patients will be recruited into the study in two cohorts of 8 patients. NOX66 suppositories are formulated using 400mg of idronoxil per 2.2g suppository. Patients are allocated to receive either one or two suppositories per day. Study Part 1: NOX66 PK: Patients receive NOX66 for 14 consecutive days as monotherapy, with a follow up period of 7 days post-dosing. Blood samples will be collected throughout the monotherapy arm to measure levels of idronoxil. If no significant adverse events are noted in this 21 day period, a patient will continue in the study. Study Part 2: NOX66 plus Carboplatin: Patients receive NOX66 at the same dose as in Part 1, for 7 days. Carboplatin is administered on Day 2 of treatment. Up to 6 cycles of chemotherapy are administered, at intervals of 28 days. For Cycles 1-3, low dose (AUC4) carboplatin is administered. Subject to safety review, standard dose (AUC6) carboplatin is administered for cycles 4-6. Safety assessment is continued throughout the study, with measures to identify efficacy signals (CT scan, ECOG) performed at baseline and after Cycles 3 and 6. Clinical trial identification: Trial Protocol Number: NOX66-001A Clinicaltrials.gov NCT02941523 Legal entity responsible for the study: Noxopharm Limited Funding: Noxopharm Limited Disclosure: I. Minns: Employee of Noxopharm Limited. G. Kelly: Member of the board of Directors, employee and a shareholder of Noxopharm Limited.

Abstract 2109: Transcriptional analysis of human leukemia cells treated with the experimental anticancer drug laromustine

Abstract Laromustine is an experimental anticancer prodrug that demonstrated significant activity against acute myeloid leukemia in human clinical trials. Upon base-catalyzed activation, laromustine yields two reactive, electrophilic species in situ: 90CE, which can chloroethylate the O6 position of guanine in DNA causing lethal interstrand crosslinks, and methyl isocyanate, which can carbamoylate biochemically relevant nucleophiles, such as cysteine sulfhydryl groups. Previous in vitro and ex vivo investigations into laromustine’s molecular mechanism of action reveal several biochemical consequences of methyl isocyanate, many of which may explain either the acute cytotoxicity of the drug’s carbamoylating activity or the observed synergism with the cytotoxicity of guanine O6 chloroethylation. The combined activities of the electrophilic subspecies of laromustine are believed to induce apoptosis in leukemia cells. Presented herein are extensive flow cytometry experiments using FITC-AnnexinV and propidium iodide suggesting a dose-dependent cytotoxicity that includes apoptotosis. So as to gain a more complete understanding of the drug’s toxicity, an investigation into how laromustine modulates gene transcription in HL60 cells is also carried out and reported here. Total mRNA was harvested from cultured HL60 cells treated with sub-lethal doses of laromustine or analogs lacking either carbamoylating or chloroethylating activity. These samples were subjected to GeneChip analyses against control mRNA. More than 4,000 genes were significantly dysregulated upon the cells’ exposure to laromustine. The analog of laromustine possessing only carbamoylating activity caused the dysregulation of nearly 3,000 genes, the vast majority of which were also dysregulated in response to laromustine. 90CE, which lacks carbamoylating activity but retains chloroethylating activity, understood to be principally responsible for therapeutic cytotoxicity, demonstrated very little effect on gene transcription in HL60 cells. Preliminary analysis of gene pathways affected by laromustine suggests that among the altered pathways, those associated with cancer progression, the G1/S checkpoint, and hematological disorders are particularly stressed. The carbamoylating activity of laromustine is responsible for most of these effects. Identifying genetic pathways disrupted by this potentially useful drug may inform clinical strategies or identify potential targets for co-therapeutic agents. Citation Format: Amanda J. Loya, Dylan J. Cincotta, Abdel Elkahloun, David Bodine, Kevin P. Rice. Transcriptional analysis of human leukemia cells treated with the experimental anticancer drug laromustine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2109. doi:10.1158/1538-7445.AM2017-2109

Mesoporous silica as a support for poorly soluble drug: Influence of pH and amino group on the drug release

Two different structures of mesoporous silica materials (MSMs), MCM-41 and MCM-41-NH2, were studied to investigate their suitability for drug delivery systems (DDS). The synthesized mesoporous materials were used as hosts for encapsulation of artesunate (ART) drug. The artesunate, an experimental anticancer drug, was incorporated in both hosts by two methods, mixing and high pressure methods. These new drug delivery systems were characterized by scanning electron microscope (SEM), N2 sorption isotherm, X-ray diffraction (XRD) and Fourier transform infrared (FT-IR). Loading of ART as models into MCM-41 and MCM-41-NH2 was studied using thermogravimetric analysis (TGA) and UV-visible spectroscopy (UV-Vis). The loading uptake and release behaviors of ART were dependent on the textural properties of MCM-41 and MCM-41-NH2. The release of drug was carefully studied in different pH (7and7.4). The effect of the synthesized hosts and ART@mesoporousdrug deliveries on MCF-7 human breast cancer cell line viability was evaluated. Both hosts alone revealed no toxicity to MCF-7 cancer cells. But, ART@MSMs indicated an inhibition of cell livability, when compared to the non-encapsulated drug. These results indicate the application of the MCM-41 mesoporous material for drug loading, and delivery into cancer cells to impel cell death.

Methods for Elucidation of DNA-Anticancer Drug Interactions and their Applications in the Development of New Drugs.

DNA damaging agents including anthracyclines, camptothecins and platinum drugs are among most frequently used drugs in the chemotherapeutic routine. Due to their relatively low selectivity for cancer cells, administration of these drugs is associated with adverse side effects, inherent genotoxicity with risk of developing secondary cancers. Development of new drugs, which could be spared of these drawbacks and characterize by improved antitumor efficacy, remains challenging yet vitally important task. These properties are in large part dictated by the selectivity of interaction between the drug and DNA and in this way the studies aimed at elucidating the complex interactions between ligand and DNA represent a key step in the drug development. Studies of the drug-DNA interactions encompass determination of DNA sequence specificity and mode of DNA binding as well as kinetic, dynamic and structural parameters of binding. Here, we consider the types of interactions between small molecule ligands and polynucleotides, how they are affected by DNA sequence and structure, and what is their significance for the antitumor activity. Based on this knowledge, we discuss the wide array of experimental techniques available to researchers for studying drug-DNA interactions, which include absorption and emission spectroscopies, NMR, magnetic and optical tweezers or atomic force microscopy. We show, using the clinical and experimental anticancer drugs as examples, how these methods provide various types of information and at the same time complement each other to provide full picture of drug- DNA interaction and aid in the development of new drugs.

Experimental anticancer drug may tackle heart disease, too

Experimental anticancer drug may tackle heart disease, too

Abstract 4777: The role of BAG3 in human leukemia cells’ response to the experimental anticancer drug laromustine

Abstract The experimental anticancer drug laromustine, which demonstrates moderate clinical efficacy against acute myelogenous leukemia and glioblastoma multiforme, yields two reactive electrophiles in situ. Therapeutic cytotoxicity is principally due to a 2-chloroethylating species that can cause lethal DNA crosslinks. However, the second electrophile, methyl isocyanate, triggers acute toxicity in cultured cancer cells and synergizes with the cogenerated 2-chloroethylating activity. The mechanism by which laromustine kills cancer cells likely includes apoptosis. We measured changes in transcription for 88 genes relating to this cell death pathway cultured human promyelocytic (HL-60) cells using quantitative real-time reverse transcriptase PCR. Cultured cells were treated with drug for six hours before harvesting mRNA for analysis. Of these genes, the expression of BAG3, whose encoded protein is also known as CAIR-1 or Bis, was increased by more than 300 fold in cells exposed to laromustine. By forming a complex with heat shock protein 70 (Hsp70), it is thought that BAG3 promotes anti-apoptotic activity by interfering with protein chaperones and release of cytochrome-c. We are also examining effects on the entire transcriptome upon treatment with laromustine using GeneChip Microarrays. Finally, we have created a stable BAG3 knockdown cell line using an shRNA construct via lentiviral transfection. Substantial differences in laromustine-induced cytotoxicity in differing BAG3 backgrounds would suggest a mechanistic role in a cell's defense against laromustine and possibly an improved clinical approach. Citation Format: Kevin P. Rice, Amanda J. Loya, Kayla M. Gross. The role of BAG3 in human leukemia cells’ response to the experimental anticancer drug laromustine. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4777.

Anticancer metal drugs and immunogenic cell death

Conventional chemotherapeutics, but also innovative precision anticancer compounds, are commonly perceived to target primarily the cancer cell compartment. However, recently it was discovered that some of these compounds can also exert immunomodulatory activities which might be exploited to synergistically enhance their anticancer effects. One specific phenomenon of the interplay between chemotherapy and the anticancer immune response is the so-called “immunogenic cell death” (ICD). ICD was discovered based on a vaccination effect exerted by cancer cells dying from pretreatment with certain chemotherapeutics, termed ICD inducers, in syngeneic transplantation mouse models. Interestingly, only a minority of drugs is able to trigger ICD without a clear-cut relation to chemical structures or their primary modes-of-action. Nevertheless, generation of reactive oxygen species (ROS) and induction of endoplasmic reticulum (ER) stress are clearly linked to ICD. With regard to metal drugs, oxaliplatin but not cisplatin is considered a bona fide ICD inducer. Taken into account that several experimental metal compounds are efficient ROS and ER stress mediators, presence of potent ICD inducers within the plethora of novel metal complexes seems feasible and has occasionally been reported. In the light of recent successes in cancer immunotherapy, here we review existing literature regarding anticancer metal drugs and ICD induction. We recommend a more profound investigation of the immunogenic features of experimental anticancer metal drugs.

Abstract PL08-04: High-throughput drug screens in cancer cell models to enable precision cancer medicine

Abstract Alterations in cancer genomes strongly influence clinical responses to therapy and can be used as molecular biomarkers of response to targeted agents. However, the identification of biomarkers for effective patient stratification remains a considerable challenge for the development of effective therapies. We are performing large-scale drug sensitivity screens using genetically characterised in vitro cellular models of human cancer. Cancer cells are profiled for differential sensitivity to clinical, pre-clinical and experimental anti-cancer compounds and drug response is linked with genomic features. We aim to identify the molecular features of cancer cells which impact on drug sensitivity and resistance, and investigate the molecular rationale for the observed sensitivity. This provides insights into mechanism of drug action, identifies opportunities for re-purposing existing drugs for new indications, and enables the identification of biomarkers to facilitate the development of targeted cancer therapies. I will describe results from our recent analysis of how cancer-driving alterations identified from the analysis of 11,000 patient tumours correlate with response to 265 compounds profiled in a panel of 1,000 human cancer cell-lines. This comprehensive analysis provides an initial glimpse of the landscape of pharmacogenomic interactions operative in human cancer, and is a resource to identify novel therapeutic possibilities for selected cancer sub-populations. Citation Format: Mathew Garnett. High-throughput drug screens in cancer cell models to enable precision cancer medicine. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr PL08-04.

Abstract B18: Non-thermal plasma, tirapazamine, and gap junctions: A novel approach to melanoma therapy through ROS induction

Abstract Melanoma cells typically have high levels of superoxide anion which may increase their susceptibility to cell death by reactive oxygen species (ROS). We have used this information to establish a system whereby melanoma cells are saturated with ROS through treatment with non-thermal plasma (NTP). NTP is formed from a mixture of highly charged helium and atmospheric gases to emit a total of 45 molecular species, many of which are ROS. We have recently demonstrated that NTP causes selective apoptotic cell death of melanoma cells as compared to normal skin keratinocytes when plated in co-culture. Since tumors flourish under low oxygen (hypoxic) conditions, we tested the effects of NTP under hypoxia. In order to enhance the effects of NTP, we combined this therapy with the experimental anticancer drug, tirapazamine, which promotes cell toxicity through DNA damage only under hypoxic conditions. The question then remains as to how to increase the target area of melanoma therapy. Our previous studies have demonstrated that gap junctions act via the bystander effect to enable the passage of cell-death promoting signals by NTP. We utilized stably transfected 1205Lu metastatic melanoma cells that express three different levels of gap junctions: endogenous low levels, increased levels through expression of connexin 43, or an absence of gap junctions due to expression of a dominant negative mutant connexin 43. We have shown that the cells with the mutant connexin 43 showed increased invasive potential as well as enhanced expression of HIF1α. The combination therapy with NTP and tirapazamine under hypoxic conditions caused a synergistic effect which increased cell death in all cells, yet was most apparent in the 1205Lu cells overexpressing the functional gap junctions. In addition, the target area of the NTP torch was dramatically increased under these conditions. This suggests that the ROS generated by NTP and tirapazamine induced the bystander effect and promoted the spreading of cell death signals well beyond the area of the plasma treatment. These results are indicative a novel future approach to melanoma therapy through selective ROS induction and modification of gap junction phenotype. Citation Format: Shoshanna N. Zucker, Cory Higley, Zethan Koch, Haneesha Goli, Peter Casey, Akeem Francis, Kevin Burke, Jennifer Zirnheld. Non-thermal plasma, tirapazamine, and gap junctions: A novel approach to melanoma therapy through ROS induction. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Melanoma: From Biology to Therapy; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(14 Suppl):Abstract nr B18.

Modeling of human uveal melanoma in zebrafish xenograft embryos.

Uveal melanoma (UM) is fatal in up to 50% of patients because of liver metastases that are refractory to therapies currently available. While murine xenograft models for human uveal melanoma are available, they have limited utility for screening large compound libraries in drug discovery studies. Therefore, new robust preclinical models are needed that can efficiently evaluate drug efficacy for treatment of this malignancy. Uveal melanoma cell lines generated from primary tumors (92.1, Mel270) and metastases (OMM2.3, OMM2.5, OMM1) were injected into the yolk of 2-day-old zebrafish embryos. After 6 days, proliferation and active migration was quantified via automated confocal image analysis. To determine the suitability of this xenotransplantation model for drug testing, drugs with three different activities (dasatinib, quisinostat, and MLN-4924) were added to the water of uveal melanoma-engrafted embryos. All tested UM cell lines proliferated and migrated in the embryos; significant differences could be discerned between cell lines: Cells derived from metastases showed more migration and proliferation than cells derived from the primary tumors, and provided preclinical models for drug testing. Addition of the Src-inhibitor dasatinib in the water of engrafted embryos reduced proliferation and migration of high Src-expressing 92.1 cells, but did not affect low Src-expressing metastatic OMM2.3 cells. Two experimental anticancer drugs, quisinostat (a histone deacetylase inhibitor) and MLN-4924 (neddylation pathway inhibitor), blocked migration and proliferation of 92.1 and OMM2.3. We established a zebrafish xenograft model of human uveal melanoma with demonstrated applicability for screening large libraries of compounds in drug discovery studies.