Concentrations Of Doxorubicin Research Articles

Hybrid Nanoparticle-Assisted Chemo-Photothermal Therapy and Photoacoustic Imaging in a Three-Dimensional Breast Cancer Cell Model.

Bioinspired nanoparticles have recently been gaining attention as promising multifunctional nanoplatforms for therapeutic applications in cancer, including breast cancer. Here, the efficiency of the chemo-photothermal and photoacoustic properties of hybrid albumin-modified nanoparticles (HSA-NPs) loaded with doxorubicin was evaluated in a three-dimensional breast cancer cell model. The HSA-NPs showed a higher uptake and deeper penetration into breast cancer spheroids than healthy breast cell 3D cultures. Confocal microscopy revealed that, in tumour spheroids incubated with doxorubicin-loaded NPs for 16 h, doxorubicin was mainly localised in the cytoplasm, while a strong signal was detectable at the nuclear level after 24 h, suggesting a time-dependent uptake. To evaluate the cytotoxicity of doxorubicin-loaded NPs, tumour spheroids were treated for up to 96 h with increasing concentrations of NPs, showing marked toxicity only at the highest concentration of doxorubicin. When doxorubicin administration was combined with laser photothermal irradiation, enhanced cytotoxicity was observed at lower concentrations and incubation times. Finally, the photoacoustic properties of doxorubicin-loaded NPs were evaluated in tumour spheroids, showing a detectable signal increasing with NP concentration. Overall, our data show that the combined effect of chemo-photothermal therapy results in a shorter exposure time to doxorubicin and a lower drug dose. Furthermore, owing to the photoacoustic properties of the NPs, this nanoplatform may represent a good candidate for theranostic applications.

Self-assembly of maltose-albumin nanoparticles for efficient targeting delivery and therapy in liver cancer

The effective delivery and targeted release of drugs within tumor cells are critical factors in determining the therapeutic efficacy of nanomedicine. To achieve this objective, a conjugate of maltose (Mal) and bovine serum albumin (BSA) was synthesized by the Maillard reaction and self-assembled into nanoparticles with active-targeting capabilities upon pH/heating induction. This nanoparticle could be effectively loaded with doxorubicin (DOX) to form stable nanodrugs (Mal-BSA/DOX) that were sensitive to low pH or high glutathione (GSH), thereby achieving a rapid drug release (96.82 % within 24 h). In vitro cell experiments indicated that maltose-modified BSA particles efficiently enhance cellular internalization via glucose transporters (GLUT)-mediated endocytosis, resulting in increased intracellular DOX levels and heightened expression of γ-H2AX. Consequently, these results ultimately lead to selective tumor cells death, as evidenced by an IC50 value of 3.83 μg/mL in HepG2 cells compared to 5.87 μg/mL in 293t cells. The efficacy of Mal-BSA/DOX in tumor targeting therapy has been further confirmed by in vivo studies, as it effectively delivered a higher concentration of DOX to tumor tissue. This targeted delivery approach not only reduces the systemic toxicity of DOX but also effectively inhibits tumor growth (TGI, 75.95 %). These findings contribute valuable insights into the advancement of targeting-albumin nanomedicine and further support its potential in tumor treatment.

MMP‐2 regulates Src activation via repression of the CHK/MATK tumor suppressor in osteosarcoma

AbstractBackgroundDoxorubicin, a first‐line anticancer drug for osteosarcoma treatment, has been the subject of recent research exploring the mechanisms behind its chemoresistance and its ability to enhance cell migration at sublethal concentrations. Matrix metalloproteinase‐2 (MMP‐2), a type IV collagenase and zinc‐dependent endopeptidase, is well‐known for degrading the extracellular matrix and promoting cancer metastasis. Our previous work demonstrated that nuclear MMP‐2 regulates ribosomal RNA transcription via histone clipping, thereby controlling gene expression. Additionally, MMP‐2 activity is regulated by the non‐receptor tyrosine kinase and oncogene, Src, which plays a crucial role in cell adhesion, invasion, and metastasis. Src kinase is primarily regulated by two endogenous inhibitors: C‐terminal Src kinase (Csk) and Csk homologous kinase (CHK/MATK).AimIn this study, we reveal that the MMP‐2 gene acts as an upstream regulator of Src kinase activity by suppressing its endogenous inhibitor, CHK/MATK, in osteosarcoma cells.Methods and ResultsWe show that enhanced osteosarcoma cell migration which is induced by sublethal concentrations of doxorubicin can be overcome by inactivating the MMP‐2 gene or overexpressing CHK/MATK. Our findings highlight the MMP‐2 gene as a promising additional target for combating cancer cell migration and metastasis. This is due to its role in suppressing on the gene and protein expression of the tumor suppressor CHK/MATK in osteosarcoma.ConclusionBy targeting the MMP‐2 gene, we can potentially enhance the effectiveness of doxorubicin treatment and reduce chemoresistance in osteosarcoma.

Evaluation of anticancer effect of micelle-encapsulated doxorubicin using gamma-distributed delay model

In clinical trials of nanoparticle-based anticancer drug delivery systems, an inexplicable lack of correlation between the function imparted to the nanoparticles and their therapeutic effects is noted. To elucidate the cause of this lack of correlation, we developed a pharmacokinetic-pharmacodynamic (PK-PD) model of tumor growth kinetics in a tumor-bearing rat model injected with polymeric micelle-encapsulated doxorubicin (DOX) using parameters related to tumor accumulation and anticancer activity. DOX was loaded onto a polyethylene glycol-peptide block polymer (PEG-poly(D/F)n: PPDF) via an amide bond to prepare sustained-release polymeric micelles (PPDF-DOX) and folic acid receptor-targeting polymeric micelles (FA-PPDF-DOX). Plasma concentration–time profiles and pharmacokinetic parameters of DOX released from PPDF-DOX into the tumors were examined using microdialysis analysis on the tumors in Walker 256 tumor-bearing rats after PPDF-DOX administration. PPDF-DOX and FA-PPDF-DOX showed delayed tumor accumulation for up to 4 h after injection. Therefore, the analysis was performed using a three-compartment model, incorporating a gamma-distributed delay model between the central and tumor compartments. The PK model was used to simulate eight repeated doses administered every other day. The cumulative area under the curve of FA-PPDF-DOX was 4.2-fold higher than that of PPDF-DOX. The PK-PD model analysis was performed based on intratumoral DOX concentration profiles and changes in tumor diameter. The time efficacy index—a measure of drug efficacy—indicated a 45k-fold higher antitumor effect of FA-PPDF-DOX over that of PPDF-DOX. These results suggest that the active-targeting function versus the effect of sustained-release polymeric micelles may depend on the delayed tumor retention of anticancer drugs.

Doxorubicin/poly(caprolactone)/polyethylene glycol nanoparticles prepared by electrospray

AbstractThe development of sustainable drug delivery systems is very important in the field of pharmaceutical research and has many advantages over the usual methods of drug administration, including improving effectiveness, reducing side effects, and increasing patient compliance. Among various techniques, electrospraying as a voltage‐driven process has emerged as a promising technology for the fabrication of drug‐loaded nanoparticles. In recent years, the application of electrospray approach for drug release has been widely investigated. However, optimization of electrospray process parameters and development of mathematical models to predict drug release kinetics from microparticles are still challenging. To address these issues, a combination of Central Composite Design (CCD) and Response Surface Methodology (RSM) has been used. This paper focuses on the application of CCD‐RSM for modeling and optimization of doxorubicin (DOX)‐poly(caprolactone) (PCL)‐polyethylene glycol (PEG) microparticles prepared using electrospray method. Doxorubicin, a widely used anticancer drug, exhibits potent cytotoxicity against a wide range of cancers. PCL‐PEG microparticles are promising as drug carriers due to their biocompatibility, controlled release properties, and ability to protect the encapsulated drug from degradation. The CCD‐RSM approach allows systematic investigation and optimization of several formulation and process variables, including polymer concentration, PCL‐PEG weight ratio, and applied voltage on drug release behavior. In addition, the effect of rheology and electrical conductivity of polymer fluid on the size of particles obtained from electrospraying was investigated. The effect of adding folic acid and doxorubicin in the optimized sample of nanoparticles and the effect of different concentrations of doxorubicin were investigated, and SEM, DLS, and FTIR techniques were used to characterize the microparticles, and the MMT test was performed to evaluate their cytotoxicity. The results showed that the optimized formulation is effective for sustained drug release over a long period for drug delivery applications.

Doxorubicin induced epigenetic regulation of dendritic cell maturation in association with T cell activation facilitates tumor protective immune response in non-small cell lung cancer (NSCLC)

BackgroundNSCLC is one of the leading causes of death and is often diagnosed at late stages with no alternative therapeutic approach. DCs are professional antigen-presenting cells and DC-based immunotherapy has been under the spotlight for its anti-cancer properties. Epigenetic modifications including DNA methylation and histone modification in DCs play a crucial role in regulating their functions such as maturation and activation,innate immune responses, T cell priming, antigen presentation, and cytokine production. In the current study, we investigated the anti-cancer properties of Doxorubicin at a noncytotoxic concentration that could be extrapolated as an epigenetic regulator for DC maturation to elicit anti-tumor activity. MethodologiesPBMCs from normal and NSCLC blood samples were isolated and treated with growth factors. DCs were matured with low dose Doxorubicin and the DC maturation markers were checked by using flow-cytometry. Further, ELISA was performed and low dose Doxorubicin-induced DCs were pulsed with LCA (Lung Cancer Antigen) and primed with CD4 +T helper (Th) cells for cytotoxicity assessment. Further, epigenetic markers of T: DC conjugation were immunofluorescently visualized under a microscope. ChIP-qPCR and Invitro assays such as histone methylation, DNA methylation, and m6A methylation were performed to study the epigenetic changes under low dose Dox treatment. IL-12 neutralization assay was performed to check for the IL-12 dependency of DCs and their effect under Dox at low dose treatment. This was further followed by a Western Blotting analysis for histone and non-histone proteins. ResultsLow dose Doxorubicin induces epigenetic changes in DCs to elicit an anti-tumor response in NSCLC through the generation of CTLs with a concomitant increase in the extracellular secretions of anti-inflammatory cytokines. We also found that low dosage of Doxorubicin matured DCs when pulsed with LCA and primed with CD4 +T helper cells, secrete IFN-γ which is important in orchestrating adaptive immunity by activating CD8 + cytotoxic T-lymphocytes. Also, the secretions of IL-12 help us infer that protective immunity is also induced via Th1 response which triggered selectively the translocation of PKCθ to immunological synapse in between DC and Th. Further, methylation and acetylation markers H3K4me3 and H3K14Ac respectively upregulated whereas levels of STAT5, NFkB, NOTCH1, and DNAPKcs were downregulated. DNA and RNA methylation assays then lead to confirmations about the epigenetic changes caused by low dose Dox treatment. DNA methylation was reduced which resulted in the activation of tumor suppressor gene p53 and Th1-associated transcription factor TBX21. On the other hand, both absolute and relative RNA methylation quantification increased in the presence of Dox at a low dose. ConclusionFrom this study, we understand that non-cytotoxic concentration of Doxorubicin increases the Ag-presenting ability of DCs via an IL-12-dependent mechanism and causes epigenetic modifications in NSCLC.

Lower cardiotoxicity of CPX-351 relative to daunorubicin plus cytarabine free-drug combination in hiPSC-derived cardiomyocytes in vitro

Liposomal formulations are hypothesized to alleviate anthracycline cardiotoxicity, although this has only been documented clinically for doxorubicin. We developed an in vitro multiparametric model using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) to assess the relative toxicity of anthracyclines across formulations. Proof of concept was established by treating hiPSC-CM with equivalent concentrations of free and liposomal doxorubicin. The study was then repeated with free daunorubicin plus cytarabine and CPX-351, a dual-drug liposomal encapsulation of daunorubicin/cytarabine. hiPSC-CM were treated with free-drug or liposomal formulations for 24 h on Days 1, 3, and 5 at equivalent concentrations ranging from 0 to 1000 ng/mL and assessed on subsequent days. Free-drug treatment resulted in concentration-dependent cumulative cytotoxicity (microscopy), more profound decrease in ATP levels, and significant time- and concentration-dependent decreases in oxygen consumption versus liposomal formulations (p < 0.01). Repeated free-drug exposure also resulted in greater release of biomarkers (cardiac troponin I, FABP3) and lactate dehydrogenase, as well as in a biphasic rhythmicity response (initial increase followed by slowing/quiescence of beating) indicating significant injury, which was not observed after repeated exposure to liposomal formulations. Overall, liposomal formulations were considerably less toxic to hiPSC-CM than their free-drug counterparts. Clinical data will be needed to confirm findings for CPX-351.

Microdialysis as a sampling tool for the chemotherapeutic agent Doxorubicin

Doxorubicin is a chemotherapeutic agent used for more than fifty years to treat a great variety of cancers in both children and adults. Despite hereof, pharmacokinetic knowledge is almost solely based on systemic plasma concentrations. Microdialysis is a catheter-based pharmacokinetic sampling tool enabling simultaneous target site sampling of unbound molecules of interest. The aim of this study was to thoroughly evaluate the feasibility of applying microdialysis for sampling of Doxorubicin in both in vitro experiments and an in vivo setting. Doxorubicin relative recovery by gain and by loss was tested for different catheter types, perfusion fluids, concentrations and collection vials. Adsorption tests revealed polystyrene/santoprene vials to be the biggest contributor of unwanted adsorption between Doxorubicin and the microdialysis equipment, and confirmed LoBind Eppendorf tubes to be a suitable alternative. The methodological combination of polyamide membranes, saline as perfusion fluid and LoBind Eppendorf sampling tubes demonstrated no statistically significant differences for relative recovery by gain and by loss, and the relative recovery was also found to be concentration independent. We conclude, that a proper microdialysis set-up can be used to collect samples containing concentrations of the chemotherapeutic drug Doxorubicin in vitro and in vivo, which encourage future pharmacokinetic studies to evaluate current treatment regimens to find the most effective and least toxic anti-neoplastic treatment for the patients.

PODO/TERT256 – A promising human immortalized podocyte cell line and its potential use for in vitro research at different oxygen levels

Podocytes are of key interest for the prediction of nephrotoxicity as they are especially sensitive to toxic insults due to their central role in the glomerular filtration apparatus. However, currently, prediction of nephrotoxicity in humans remains insufficiently reliable, thus highlighting the need for advanced in vitro model systems using human cells with improved prediction capacity. Recent approaches for refining in vitro model systems focus on closely replicating physiological conditions as observed under the in vivo situation typical of the respective nephron section of interest. PODO/TERT256, a human immortalized podocyte cell line, were employed in a semi-static transwell system to evaluate its potential use as a human podocyte in vitro system for modelling potential human glomerular toxicity. Furthermore, the impact of routinely employed excessive oxygen tension (21 % - AtmOx), when compared to the physiological oxygen tensions (10 % - PhysOx) observed in vivo, was analyzed. Generally, cultured PODO/TERT256 formed a stable, contact-inhibited monolayer with typical podocyte morphology (large cell body, apical microvilli, finger-like cytoplasmic projections (reminiscent of foot processes), and interdigitating cell-cell junctions) and developed a size-selective filtration barrier. PhysOx, however, induced a more pronounced in vivo like phenotype, comprised of significantly larger cell bodies, significantly enhanced filtration barrier size-selectivity, and a remarkable re-localization of nephrin to the cell membrane, thus suggesting an improved in vitro replication of in vivo characteristics. Preliminary toxicity characterization with the known glomerulotoxin doxorubicin (DOX) suggested an increasing change in filtration permeability, already at the lowest DOX concentrations tested (0.01 μM) under PhysOx, whereas obvious changes under AtmOx were observed as of 0.16 μM and higher with a near all or nothing effect. The latter findings suggested that PODO/TERT256 could serve as an in vitro human podocyte model for studying glomerulotoxicity, whereby culturing at PhyOx tension appeared critical for an improved in vivo-like phenotype and functionality. Moreover, PODO/TERT256 could be incorporated into advanced human glomerulus systems in vitro, recapitulating microfluidic conditions and multiple cell types (endothelial and mesenchymal cells) that can even better predict human glomerular toxicity.

Induced pluripotent stem cell-derived cardiac myocytes to assess anthracycline cardiotoxicity

Abstract Background Current oncological treatments improved survival but also increased awareness of cardiovascular side-effects following cancer therapy. Anthracyclines are a class of highly effective chemotherapeutics, yet their use is regularly complicated by anthracycline-induced cardiotoxicity (AIC). The advent of induced pluripotent stem cell-derived cardiac myocytes (iPSC-CMs) offers the potential for disease modeling and prediction of genetic cardiomyopathies. Purpose We aim to use iPSC-CMs to assess susceptibility to cardiotoxicity induced by anti-cancer treatments on an individual patient level. Furthermore, the assay could be exploited to unravel cellular processes and pathways involved in AIC. Methods Five heart transplant patients following AIC and five anthracycline treated patients without AIC were included in our study. We reprogrammed patient-derived lymphocytes from both groups toward iPSCs using Sendai Virus; afterwards, patient derived iPSCs were differentiated into CM-like cells. Cytotoxicity following chemotherapy exposure was assessed using flow cytometry (FC), in which iPSC-CM-specific cells were first selected based on the expression of signal-regulatory protein alpha (SIRPA) surface marker; thereafter the percentage of apoptotic versus viable cells following doxorubicin exposure was quantitatively assessed. Ultrastructural changes were evaluated using electron microscopy (EM) images, comparing doxorubicin treated and untreated iPSC-CMs. Reactive Oxygen Species (ROS) generation following doxorubicin exposure was assessed using a luciferin precursor-based plate reader assay (ROS-GLO) and FC. Furthermore, whole genome sequencing (WGS) was performed and variants were filtered based on allele frequency, pathogenicity using HumVar trained variant effect predictors and presence in patients versus controls. Results The cytotoxic effect of increasing concentrations of doxorubicin was quantifiable using FC on SIRPA positive iPSC-CMs (Figure 1 panel A). On the ultrastructural level, doxorubicin treated iPSC-CMs manifested darker and more condensed mitochondria as seen with EM (Figure 1 panel B). Increased ROS production was observed following doxorubicin exposure as assessed using ROS-GLO plate reader assay and FC (Figure 1 panel C). We used iPSC-CMs of transplant patients and matched controls to assess inter-individual susceptibility to anthracycline-induced cardiotoxicity. Included patient profiles can be found in Figure 2 panel A. There was a higher rate of cell death at any concentration of doxorubicin in patients with proven AIC as opposed to controls (Figure 2 panel B). WGS analysis showed 2 pathogenic variants in the PRAMEF6 gene present in cases and not in controls (Figure 3 panel C). Conclusion iPSC-CMs represent a robust disease model to study cancer therapy-induced cardiotoxicity. Furthermore, iPSC-CMs allow to study inter-individual differences in susceptibility to cancer therapy-induced cardiotoxicity.Mechanism of AICSusceptibility to AIC

Ultrasound-Induced Tumor Perfusion Changes and Doxorubicin Delivery: A Study on Pulse Length and Pulse Repetition Frequency.

To investigate the appropriate combination of pulse length (PL) and pulse repetition frequency (PRF) when performing ultrasound stimulated microbubble (USMB) to enhance doxorubicin (DOX) delivery to tumors. A total of 48 tumor-bearing mice were divided into four groups, namely groups A-D. The mice in groups B-D were treated with chemotherapy and USMB treatment with different combinations of PL and PRF, and group A was control. Contrast-enhanced ultrasound imaging was conducted to analyze tumor blood perfusion. Fluorescence microscopy and high-performance liquid chromatography were used to qualitatively and quantitatively analyse DOX release. The structural changes of tumors were observed under light microscope and transmission electron microscope. Furthermore, another 24 tumor-bearing mice were treated with sonochemotherapy and some related inflammatory factors were measured to explore the underlying mechanism. With PL of three cycles and PRF of 2 kHz, the tumor perfusion area ratio increased by 26.67%, and the DOX concentration was 4.69 times higher than the control (P < .001). With PL of 34.5 cycles and PRF of 200 Hz, the tumor perfusion area ratio decreased by 12.7% and DOX did not exhibit increased extravasation compared with the control. Microvascular rupture and hemorrhage were observed after long PL and low PRF treatment. While vasodilation and higher levels of some vasodilator inflammatory factors were found after treatment with short PL and high PRF. USMB treatment using short PL and high PRF could enhance tumor blood perfusion and increase DOX delivery, whereas long PL and low PRF could not serve the same purpose.

Quantitative imaging of doxorubicin diffusion and cellular uptake in biomimetic gels with human liver tumor cells

Novel tumor-on-a-chip approaches are increasingly used to investigate tumor progression and potential treatment options. To improve the effect of any cancer treatment it is important to have an in depth understanding of drug diffusion, penetration through the tumor extracellular matrix and cellular uptake. In this study, we have developed a miniaturized chip where drug diffusion and cellular uptake in different hydrogel environments can be quantified at high resolution using live imaging. Diffusion of doxorubicin was reduced in a biomimetic hydrogel mimicking tissue properties of cirrhotic liver and early stage hepatocellular carcinoma (373 ± 108 µm2/s) as compared to an agarose gel (501 ± 77 µm2/s, p = 0.019). The diffusion was further lowered to 256 ± 30 µm2/s (p = 0.028) by preparing the biomimetic gel in cell media instead of phosphate buffered saline. The addition of liver tumor cells (Huh7 or HepG2) to the gel, at two different densities, did not significantly influence drug diffusion. Clinically relevant and quantifiable doxorubicin concentration gradients (1–20 µM) were established in the chip within one hour. Intracellular increases in doxorubicin fluorescence correlated with decreasing fluorescence of the DNA-binding stain Hoechst 33342 and based on the quantified intracellular uptake of doxorubicin an apparent cell permeability (9.00 ± 0.74 × 10–4 µm/s for HepG2) was determined. Finally, the data derived from the in vitro model were applied to a spatio-temporal tissue concentration model to evaluate the potential clinical impact of a cirrhotic extracellular matrix on doxorubicin diffusion and tumor cell uptake. Graphical abstract

A physiologically-based pharmacokinetic model for predicting doxorubicin disposition in multiple tissue levels and quantitative toxicity assessment

Doxorubicin is a widely-used chemotherapeutic drug, however its high toxicity poses a significant challenge for its clinical use. To address this issue, a physiologically-based pharmacokinetic (PBPK) model was implemented to quantitatively assess doxorubicin toxicity at cellular scale. Due to its unique pharmacokinetic behavior (e.g. high volume of distribution and affinity to extra-plasma tissue compartments), we proposed a modified PBPK model structure and developed the model with multispecies extrapolation to compensate for the limitation of obtaining clinical tissue data. Our model predicted the disposition of doxorubicin in multiple tissues including clinical tissue data with an overall absolute average fold error (AAFE) of 2.12. The model’s performance was further validated with 8 clinical datasets in combined with intracellular doxorubicin concentration with an average AAFE of 1.98. To assess the potential cellular toxicity, toxicity levels and area under curve (AUC) were defined for different dosing regimens in toxic and non-toxic scenarios. The cellular concentrations of doxorubicin in multiple organ sites associated with commonly observed adverse effects (AEs) were simulated and calculated the AUC for quantitative assessments. Our findings supported the clinical dosing regimen of 75 mg/m2 with a 21-day interval and suggest that slow infusion and separated single high doses may lower the risk of developing AEs from a cellular level, providing valuable insights for the risk assessment of doxorubicin chemotherapy. In conclusion, our work highlights the potential of PBPK modelling to provide quantitative assessments of cellular toxicity and supports the use of clinical dosing regimens to mitigate the risk of adverse effects.

Individualized chemotherapy drug dose escalation in dogs with multicentric lymphoma.

This study was performed to determine the ability to escalate drug doses in a 15-week CHOP protocol in dogs with multicentric lymphoma. We hypothesized that at least 50% of dogs could successfully be escalated in at least 1 drug. Secondary aims were to establish objective response rate (ORR), progression-free interval (PFI), and overall survival time (OST). Thirty dogs with newly diagnosed multicentric lymphoma were prospectively treated with a 15-week CHOP protocol. This was a prospective cohort study. Drug doses that did not cause dose-limiting adverse effects (AEs) were increased using a standardized escalation protocol. AEs and response were assessed using VCOG criteria. Serial blood samples were collected after the first dose of each drug for pharmacokinetic analysis. Of the 23 dogs with the opportunity to dose escalate, at least 1 drug was successfully escalated in 18 (78%). Vincristine was successfully escalated to 0.8 mg/m2 or higher in 11 dogs, cyclophosphamide to 300 mg/m2 or higher in 16 dogs, and doxorubicin to 35 mg/m2 or 1.4 mg/kg or higher in 9 dogs. Three of the 23 dogs (13%) were hospitalized at least once because of drug-induced AEs. Neutropenia was the most common dose-limiting toxicosis for all drugs. Peak doxorubicin concentrations were significantly lower in dogs where doxorubicin was successfully escalated. The objective response rate was 100%. The median progression free interval was 171 days. The median overall survival time was 254 days. Drugs in the CHOP protocol can often be escalated safely with manageable AEs.

Exposure to anticancer drugs modulates the expression of ACSL4 and ABCG2 proteins in adrenocortical carcinoma cells

Adrenocortical carcinoma (ACC) is a rare and malignant disease, with more than 50 % of patients developing hormone-secreting tumors. These tumors are genetically heterogeneous and potentially lethal, as metastasis is often underway at the time of diagnosis. While chemoresistance can be multifactorial, Acyl CoA synthetase 4 (ACSL4) is known to contribute to the generation of highly aggressive cellular phenotypes, while increased expression and activity of multidrug transporters such as ATP-binding cassette subfamily G member 2 (ABCG2) are known to play a key role. Therefore, the objective of this work was to determine changes in the expression of ACSL4 and ABCG2 in ACC cell lines after exposure to antitumor drugs. Bioinformatics analysis of public database GSE140818 revealed higher ACSL4 and ABCG2 expression in HAC15 cells resistant to mitotane when compared to wild type cells. In addition, our studies revealed an increase in ACSL4 and ABCG2 expression in lowly aggressive H295R cells undergoing early treatment with non-lethal concentrations of mitotane, doxorubicin and cisplatin. Comparable results were obtained in lowly aggressive breast cancer cells MCF-7. The increase in ACSL4 and ABCG2 expression favored tumor cell viability, proliferation and compound efflux, an effect partially offset by ACSL4 and ABCG2 inhibitors. These results provide relevant data on the undesired molecular effects of antitumor drugs and may fuel future studies on patients’ early response to antitumor treatment.

Clinical trial protocol for PanDox: a phase I study of targeted chemotherapy delivery to non-resectable primary pancreatic tumours using thermosensitive liposomal doxorubicin (ThermoDox®) and focused ultrasound

BackgroundThe dense stroma of pancreatic ductal adenocarcinomas is a major barrier to drug delivery. To increase the local drug diffusion gradient, high doses of chemotherapeutic agent doxorubicin can be released from thermally-sensitive liposomes (ThermoDox®) using ultrasound-mediated hyperthermia at the tumour target. PanDox is designed as a Phase 1 single centre study to investigate enhancing drug delivery to adult patients with non-operable pancreatic ductal adenocarcinomas. The study compares a single cycle of either conventional doxorubicin alone or ThermoDox® with focused ultrasound-induced hyperthermia for targeted drug release.MethodsAdults with non-resectable pancreatic ductal adenocarcinoma are allocated to receive a single cycle of either doxorubicin alone (Arm A) or ThermoDox® with focused ultrasound-induced hyperthermia (Arm B), based on patient- and tumour-specific safety conditions. Participants in Arm B will undergo a general anaesthetic and pre-heating of the tumour by extra-corporal focused ultrasound (FUS). Rather than employing invasive thermometry, ultrasound parameters are derived from a patient-specific treatment planning model to reach the 41 °C target temperature for drug release. ThermoDox® is then concurrently infused with further ultrasound exposure. Tumour biopsies at the targeted site from all patients are analysed post-treatment using high performance liquid chromatography to quantify doxorubicin delivered to the tumour. The primary endpoint is defined as a statistically significant enhancement in concentration of total intra-tumoural doxorubicin, comparing samples from patients receiving liposomal drug with FUS to free drug alone. Participants are followed for 21 days post-treatment to assess secondary endpoints, including radiological assessment to measure changes in tumour activity by Positron Emission Tomography Response Criteria in Solid Tumours (PERCIST) criteria, adverse events and patient-reported symptoms.DiscussionThis early phase study builds on previous work targeting tumours in the liver to investigate whether enhancement of chemotherapy delivery using ultrasound-mediated hyperthermia can be translated to the stroma-dense environment of pancreatic ductal adenocarcinoma. If successful, it could herald a new approach towards managing these difficult-to-treat tumours.Trial registrationClinicalTrials.gov Identifier: NCT04852367. Registered 21st April 2022.EudraCT number: 2019–003950-10 (Registered 2019)Iras Project ID: 272253 (Registered 2019)Ethics Number: 20/EE/0284.

Drug delivery by sonosensitive liposome and microbubble with acoustic-lens attached ultrasound: an in vivo feasibility study in a murine melanoma model

Conventional chemotherapy methods have adverse off-target effects and low therapeutic efficiencies of drug release in target tumors. In this study, we proposed a combination therapy of doxorubicin (DOX)-loaded ultrasound (US)-sensitive liposomal nanocarriers (IMP301), microbubbles (MBs) under focused US exposure using convex acoustic lens-attached US (LENS) to tumor treatment. The therapeutic effects of each treatment in a murine melanoma model were evaluated using contrast-enhanced US (CEUS) and micro-computed tomography (micro-CT) imaging, bioluminescence and confocal microscopy imaging, and liquid chromatography–mass spectroscopy (LC/MS) analysis. Tumor-bearing mice were randomly assigned to one of the following groups: (1) G1: IMP301 only (n = 9); (2) G2: IMP301 + LENS (n = 9); (3) G3: IMP301 + MB + LENS (n = 9); (4) G4: DOXIL only (n = 9); and (5) G5: IMP301 without DOXIL group as a control group (n = 4). Ten days after tumor injection, tumor-bearing mice were treated according to each treatment strategy on 10th, 12th, and 14th days from the day of tumor injection. The CEUS images of the tumors in the murine melanoma model clearly showed increased echo signal intensity from MBs as resonant US scattering. The relative tumor volume of the G2 and G3 groups on the micro-CT imaging showed inhibited tumor growth than the reference baseline of the G5 group. DOX signals on bioluminescence and confocal microscopy imaging were mainly located at the tumor sites. LC/MS showed prominently higher intratumoral DOX concentration in the G3 group than in other treated groups. Therefore, this study effectively demonstrates the feasibility of the synergistic combination of IMP301, MBs, and LENS-application for tumor-targeted treatment. Thus, this study can enable efficient tumor-targeted treatment by combining therapy such as IMP301 + MBs + LENS-application.

Doxorubicin-induced senescence promotes resistance to cell death by modulating genes associated with apoptotic and necrotic pathways in prostate cancer DU145 CD133+/CD44+ cells

Cancer stem cells (CSCs) are the most important cause of cancer treatment failure. Traditional cancer treatments, such as chemotherapy and radiotherapy, damage healthy cells alongside malignant cells, leading to severe adverse effects. Therefore, inducing cellular senescence without triggering apoptosis, which further damages healthy cells, may be an alternative strategy. However, there is insufficient knowledge regarding senescence induction in CSCs that show resistance to treatment and stemness properties. The present study aims to elucidate the effects of senescence induction on proliferation, cell cycle, and apoptosis in prostate CSCs and non-CSCs. Prostate CSCs were isolated from DU145 cancer cells using the FACS method. Subsequently, senescence induction was performed in RWPE-1, DU145, prostate CSCs, and non-CSCs by using different concentrations of Doxorubicin (DOX). Cellular senescence was detected using the senescence markers SA-β-gal, Ki67, and senescence-associated heterochromatin foci (SAHF). The effects of senescence on cell cycle and apoptosis were evaluated using the Muse Cell Analyzer, and genes in signaling pathways associated with the apoptotic/necrotic pathway were analyzed by real-time PCR. Prostate CSCs were isolated with 95.6 ± 1.4% purity according to CD133+/CD44+ characteristics, and spheroid formation belonging to stem cells was observed. After DOX-induced senescence, we observed morphological changes, SA-β-gal positivity, SAHF, and the lack of Ki67 in senescent cells. Furthermore; we detected G2/M cell cycle arrest and downregulation of various apoptosis-related genes in senescent prostate CSCs. Our results showed that DOX is a potent inducer of senescence for prostate CSCs, inhibits proliferation by arresting the cell cycle, and senescent prostate CSCs develop resistance to apoptosis.

CT-guided paravertebral injection of doxorubicin for treatment of postherpetic neuralgia: a database-based retrospective stratified study.

This study explored the impact of different doeses of doxorubicin in CT-guided transvertebral foraminal injections for postherpetic neuralgia (PHN) treatment and the impact of 0.5% doxorubicin treatment on patients with different disease courses and lesion locations. This retrospective study included 291 patients with PHN who received CT-guided doxorubicin injection at West China Hospital between April 2014 and February 2020. A total of 228 patients received 0.5% doxorubicin treatment and 63 received 0.33% doxorubicin. Both groups showed significantly improvement in visual analogue scale (VAS) and Brief Pain Inventory (BPI) scores. The 0.5% doxorubicin group demonstrated significant lower VAS scores at 6 and 12 months after surgery (all p < 0.001) and a significant lower score on the BPI at 6 and 12 months than the 0.33% doxorubicin group (all p < 0.05). Stratified analysis of 0.5% doxorubicin demonstrated a significant reduction in VAS score at 1 week, 3 months, 6 months, and 12 months after treatment compared to baseline (all p < 0.05) and significant improvements in BPI score after treatment compared to baseline (p < 0.05). The VAS score of the chest group was significant higher than facial, neck and upper limbs and abdomen groupsin groups 1 week after surgery (all p < 0.05). Various aspects of quality of life, including daily life, enjoyment of life, sleep, relationships, work, walking ability, and emotions, significantly decreased after surgery (p < 0.05). Especially in sleep duration, there was an increase in patients reporting intermediate sleep (4-7 h) and a proportion achieving a normal sleep duration of ≥7 h. And no significant differences of BPI were observed among different affected locations. The incidence of adverse events in the 0.5% doxorubicin group and 0.33% doxorubicin group was 8.78 and 6.34%, respectively. CT-guided doxorubicin injection therapy has the potential to alleviate pain and enhance the quality of life in patients with PHN. Notably, the use of a 0.5% doxorubicin concentration yields more pronounced pain relief compared to a 0.33% concentration. While longer durations of PHN and specific affected sites may influence the response to treatment, the overall improvements in quality of life remain consistent.

Establishment and Characterization of Multi-Drug Resistant p53-Negative Osteosarcoma SaOS-2 Subline.

To establish a p53-negative osteosarcoma (OS) SaOS-2 cellular subline exhibiting resistance to specific chemotherapeutic agents, including topoisomerase II inhibitors, taxanes, and vinca alkaloids. The OS subline exhibiting resistance to the chemotherapeutic agents indicated above was generated by the stepwise treatment of the parental SaOS-2 cell line with increasing concentrations of doxorubicin (Dox) for 5 months. Half-inhibitory concentrations (IC50) for Dox, vinblastine (Vin), and paclitaxel (PTX) were calculated by a colorimetric MTS-based assay. Crystal violet staining was used to assess cellular viability, whereas the proliferation capacities of cancer cells were monitored in real-time by the i-Celligence system. Expression of apoptotic markers (e.g., cleaved PARP and caspase-3), DNA repair proteins (e.g., ATM, DNA-PK, Nbs1, Rad51, MSH2, etc.), and certain ABC transporters (P-glycoprotein, MRP1, ABCG2, etc.) was assessed by western blotting and real-time PCR. Flow cytometry was used to examine the fluorescence intensity of Dox and ABC-transporter substrates (e.g., Calcein AM and CMFDA) and to assess their excretion to define the activity of specific ABC-transporters. To confirm OS resistance to Dox in vivo, xenograft experiments were performed. An OS subline generated by a stepwise treatment of the parental SaOS-2 cell line with increasing concentrations of Dox resulted in an increase in the IC50 for Dox, Vin, and PTX (~6-, 4-, and 30-fold, respectively). The acquisition of chemoresistance in vitro was also evidenced by the lack of apoptotic markers (e.g., cleaved PARP and caspase-3) in resistant OS cells treated with the chemotherapeutic agents indicated above. The development of the multidrug resistance (MDR) phenotype in this OS subline was due to the overexpression of ABCB1 (i.e., P-glycoprotein) and ABCC1 (i.e., multidrug resistance protein-1, MRP-1), which was evidenced on both mRNA and protein levels. Due to increased expression of MDR-related proteins, resistant OS exhibited an excessive efflux of Dox. Moreover, decreased accumulation of calcein AM, a well-known fluorescent substrate for both ABCB1 and ABCC1, was observed for resistant OS cells compared to their parental SaOS-2 cell line. Importantly, tariquidar and cyclosporin, well-known ABC inhibitors, retained the intensity of Dox-induced fluorescence in resistant SAOS-2 cells. Furthermore, in addition to the increased efflux of the chemotherapeutic agents from Dox-resistant OS cells, we found higher expression of several DNA repair proteins (e.g., Rad51 recombinase, Mre11, and Nbs1, activated forms of ATM, DNA-PK, Chk1, and Chk2, etc.), contributing to the chemoresistance due to the excessive DNA repair. Lastly, the in vivo study indicated that Dox has no impact on the SaOS-2 Dox-R xenograft tumor growth in a nude mouse model. An acquired resistance of OS to the chemotherapeutic agents might be due to the several mechanisms undergoing simultaneously on the single-cell level. This reveals the complexity of the mechanisms involved in the secondary resistance of OS to chemotherapies.