Liposomal Formulation Of Doxorubicin Research Articles

The impact of phospholipids with high transition temperature to enhance redox-sensitive liposomal doxorubicin efficacy in colon carcinoma model

In this study, we have developed a redox-sensitive (RS) liposomal doxorubicin formulation by incorporating 10,10′-diselanediylbis decanoic acid (DDA) organoselenium compound as the RS moiety. Hence, several RS liposomal formulations were prepared by using DOPE, HSPC, DDA, mPEG2000-DSPE, and cholesterol. In situ drug loading using a pH gradient and citrate complex yielded high drug to lipid ratio and encapsulation efficiency (100%) for RS liposomes. Liposomal formulations were characterized in terms of size, surface charge and morphology, drug loading, release properties, cell uptake and cytotoxicity, as well as therapeutic efficacy in BALB/c mice bearing C26 tumor cells. The formulations showed an average particle size of 200 nm with narrow size distributions (PDI < 0.3), and negative surface charges varying from −6 mV to −18.6 mV. Our study confirms that the presence of the DDA compound in liposomes is highly sensitive to hydrogen peroxide at 0.1% w/v, resulting in a significant burst release of up to 40%. The in vivo therapeutic efficacy study in BALB/c mice bearing C26 colon carcinoma confirmed the promising function of RS liposomes in the tumor microenvironment which led to a prolonged median survival time (MST). The addition of hydrogenated soy phosphatidylcholine (HSPC) with a high transition temperature (Tm: 52–53.5°C) extended the MST of our 3-component formulation of F14 (DOPE/HSPC/DDA) to 60 days in comparison to Caelyx (PEGylated liposomal Dox), which is not RS-sensitive (39 days). Overall, HSPC liposomes bearing RS-sensitive moiety enhanced therapeutic efficacy against colon cancer in vitro and in vivo. This achievement unequivocally underscores the criticality of high-TM phospholipids, particularly HSPC, in significantly enhancing liposome stability within the bloodstream. In addition, RS liposomes enable the on-demand release of drugs, leveraging the redox environment of tumor cells, thereby augmenting the efficacy of the formulation.

TXB-001, a newly-developed polymer-conjugated anthracycline: Significantly lower adverse effects in animal models of alopecia and hand-foot syndrome

Anthracycline anti-cancer drugs have been widely used in the treatment of several cancers; however, their use is limited by adverse effects (AEs). Alopecia is a common AE that is minimally invasive, but adversely affects mental health and reduces quality of life (QoL). Hand-foot syndrome (HFS) is a dose-limiting AE of DOXIL, a liposomal formulation of doxorubicin (DOX). Although it is not a life-threatening condition, HFS affects function and reduces QoL. TXB-001 is a new candidate polymer-conjugated anthracycline anti-cancer drug, and modified and optimized polymerized pirarubicin (THP), known as P-THP, is expected to have low toxicity and high efficacy. The anti-cancer effects of TXB-001 were examined using the 4T1 mouse model. An alopecia mouse model and HFS rat model were used to evaluate the alopecia- and HFS-inducing effects of TXB-001 and compare their severity with existing anthracycline anti-cancer drugs. A pharmacokinetic analysis of plasma as well as chest, palmar, and plantar skin samples after the single intravenous administration of DOXIL and TXB-001 to rats was also performed. The results obtained revealed that TXB-001 exerted similar anti-cancer effects to those of DOXIL in mice, weaker alopecia-inducing effects than DOX, DOXIL, and THP in mice, and no or markedly weaker HFS-like changes than DOXIL, which induced significant histopathological changes. The results of the pharmacokinetic analysis showed the accumulation of DOXIL, but not TXB-001, in skin, particularly palmar and plantar skin samples, and these differences were considered to contribute to their HFS-inducing effects.

Abstract 679: TXB-001 is an upgraded conjugated anthracycline that targets tumors with reduced risk of cardiotoxicity, hand-foot syndrome and alopecia

Abstract TXB-001 is a well-optimized N-(2-hydroxypropyl)methacrylamide copolymer-based conjugated pirarubicin (THP) and a novel candidate for developing anticancer drugs. It exhibits tumor-specific accumulation due to the enhanced permeability and retention effects and prolonged circulation time compared with free THP. Its hydrazone bond is dissociated under lower pH conditions, providing selective and rapid release of THP in acidic tumor environments. Therefore, TXB-001 is expected to have high efficacy and low toxicity. Anthracycline anticancer drugs have been widely used for the treatment of several cancers, but their use is limited by several adverse effects (AEs), including cardiotoxicity, which is a life-threatening AE. Furthermore, the maximum lifetime cumulative dose is decided for each anthracycline in clinical use, which leads to discontinuation of the medication. Hand-foot syndrome (HFS) is a dose-limiting AE of DOXIL®, a liposomal formulation of doxorubicin (DOX), and although it is not a life-threatening condition, it significantly deteriorates quality of life (QOL). Alopecia is another common AE, which is relatively noninvasive but adversely affects mental health and reduces QOL. In this study, we confirmed the efficacy of TXB-001 in a mice triple-negative breast cancer (TNBC) model and evaluated the cardiotoxicity-, HFS- and alopecia-inducing effects of TXB-001 in mice and rats to compare the severity with existing anthracycline anticancer drugs, including DOXIL®. The pharmacokinetic (PK) analysis in several tissues, including plasma, tumor, heart, skin of the chest, and palmar and plantar areas, was also performed after a single intravenous administration in rodents. As a result, TXB-001 showed an equivalent or greater antitumor effects than other anthracyclines in the TNBC model, and the AEs were scarcely observed. TXB-001 did not decrease left ventricular ejection fraction, with no or little HFS-like changes compared with DOXIL®, which showed significant histopathological changes. TXB-001 also showed weaker alopecia-inducing effects than DOX, DOXIL®, and THP. In the PK analysis, TXB-001 was distributed in the tumor at a higher concentration than in other tissues, except for plasma. Hence, most of the THP, the active ingredient, was released in the tumor. The exposure of the heart in the TXB-001-treated group was one-tenth of that in the DOXIL®-treated group. Moreover, accumulation in the skin, especially in palmar and plantar areas, was observed for DOXIL®, but no accumulation in the skin was observed for TXB-001. This PK profile consistently demonstrated the antitumor activity of TXB-001, without AEs. TXB-001 reduces the chance of dose-limiting and QOL-threatening AEs, and thus, cancer patients can continue treatment, making it a more convenient anticancer drug in clinical use with a wide safety margin. Citation Format: Keiyu Oshida, Mikito Hirakata, Emi Tomikawa, Chizuka Sakai, Masashi Uchida, Rieko Shimozono, Akiko Izawa, Yoko Koga, Tsubasa Okano, Hideki Narumi, Lisa Munakata, Ryo Suzuki, Miki Nonaka, Yasuhito Uezono. TXB-001 is an upgraded conjugated anthracycline that targets tumors with reduced risk of cardiotoxicity, hand-foot syndrome and alopecia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 679.

Abstract 3204: Antitumor activity of a coencapsulated liposomal formulation of ceramides and doxorubicin in neuroblastoma models

Abstract Background: Neuroblastoma (NB) represents the most frequent and aggressive form of extracranial solid tumor of infants. Targeted therapies are severely limited by the lack of specific receptors, thus chemotherapy and radiotherapy remain the main treatment choices, which expose NB patients to systemic toxicity. Recently, we have demonstrated that cell surface Nucleolin (NCL) represents a novel cellular target for NB therapy. Methods: The cytotoxic potential of pH-sensitive nanocarriers (NC), decorated with the NCL-recognizing F3 peptide and incorporating a combination of the PI3K/Akt signaling inhibitors, C6-Ceramide (C6-Cer) or C18-ceramide (C18-Cer), and the topoisomerase II inhibitor doxorubicin (DXR) (F3-NC[C6-Cer/DXR] and F3-NC[C18-Cer/DXR], respectively), was tested in vitro on a panel of NB cell lines. Cytotoxicity was evaluated in monolayer and in multicellular tumor spheroid models, by MTS proliferation and Cell Titer Glo 3D cell viability assays, respectively. Apoptotic/necrotic cell death was investigated by Real Time-Glo Annexin VApoptosis and Necrosis assay. Results: F3-NC[C6-Cer/DXR] and F3-NC[C18-Cer/DXR] resulted significantly more effective, in term of reduction of NB cell viability, compared to the corresponding “drugs”-loaded untargeted NC formulations. Both formulations triggered apoptotic cell death, even if in a different kinetic profile. In particular, NB cells treated with F3-NC[C6-Cer/DXR] underwent apoptotic cell death earlier with respect to those treated with F3-NC[C18-Cer/DXR]. Interestingly, F3-NC[C6-Cer/DXR] exerted a significant superior cell killing effect when compared both to F3-NC[C18-Cer/DXR] and to F3-NC encapsulating only DXR, suggesting the importance of the combination treatment to enhance the antitumor effect against NB cells. Conclusion: Our preliminary findings demonstrate that the co-encapsulation and delivery of C6-Ceramide and DXR by NCL-recognizing nanoparticles exert potent antitumor effects against NB models in vitro. Ongoing in vivo experiments on clinically relevant mouse NB models (i.e. recapitulating primary tumor growth, metastases, and minimal residual disease) will allow us to define the possible use of F3-NC[C6-Cer/DXR] as an innovative therapeutic strategy for relapsed/refractory NB. Funding: AIRC IG n. 24397 to PF; CInTech (PRR) C644865576-00000005 to JNM.VB is recipient of AIRC ID 24397 contract. MBC is recipient of PD/BDE/150664/2020 (FCT) fellowship. Citation Format: Veronica Bensa, Mariana Biscaia-Caleiras, Mirco Ponzoni, Joao Nuno Moreira, Chiara Brignole, Fabio Pastorino. Antitumor activity of a coencapsulated liposomal formulation of ceramides and doxorubicin in neuroblastoma models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3204.

Combination Treatment with Liposomal Doxorubicin and Inductive Moderate Hyperthermia for Sarcoma Saos-2 Cells.

Despite efforts in osteosarcoma (OS) research, the role of inductive moderate hyperthermia (IMH) in delivering and enhancing the antitumor effect of liposomal doxorubicin formulations (LDOX) remains unresolved. This study investigated the effect of a combination treatment with LDOX and IMH on Saos-2 human OS cells. We compared cell viability using a trypan blue assay, apoptosis and reactive oxygen species (ROS) measured by flow cytometry and pro-apoptotic Bax protein expression examined by immunocytochemistry in response to IMH (42 MHz frequency, 15 W power for 30 min), LDOX (0.4 μg/mL), and LDOX plus IMH. The lower IC50 value of LDOX at 72 h indicated increased accumulation of the drug in the OS cells. LDOX plus IMH resulted in a 61% lower cell viability compared to no treatment. Moreover, IMH potentiated the LDOX action on the Saos-2 cells by promoting ROS production at temperatures of <42 °C. There was a 12% increase in cell populations undergoing early apoptosis with a less heterogeneous distribution of Bax after combination treatment compared to those treated with LDOX (p < 0.05). Therefore, we determined that IMH could enhance LDOX delivery and its antitumor effect via altered membrane permeabilization, ROS generation, and a lower level of visualized Bax heterogeneity in the Saos-2 cells, suggesting the potential translation of these findings into in vivo studies.

An Automated Electroanalytical Method for the Drug Release Profiling of Liposomal Doxorubicin HCl Formulations

Liposomes have emerged as a drug delivery system for various chemotherapeutics providing enhanced bioavailability and reduced toxicity. In vitro drug release profiling of liposomal formulations is one of the essential tests for the premarket approval and post market quality control. We developed an automated electroanalytical method for drug release profiling of liposomal doxorubicin formulation. In this electroanalytical method, square wave voltammetry mode was selected to determine the released drug, the only redox-active analyte, by measuring the current at the pulsed potential ranges. Therefore, no separation from liposomal encapsulated doxorubicin is needed. This electroanalytical method provided a continuous drug release measurement for 24 h. The drug release increased as the release media pH and temperature increased. At 37 °C, the drug release increased from 7 % to 40 % when the pH increased from 5.5 to 7.4, In addition, at pH 6.5, as the temperature increased from 37 °C to 52 °C, total drug release increased by more than two-fold. Complete drug release (more than 80 %) was obtained at pH 6.5 and 52 °C in less than 3 h. The brand name and the two generic formulations showed similar drug release profile in all experimental conditions. This method is an alternative to traditional methods which require separation steps such as dialysis or solid phase extraction to quantitate released doxorubicin. This method may be further applied in the in vitro release testing of other liposomal formulations containing redox-active drug substances, e.g., liposomes encapsulating daunorubicin.

Liposomal Doxorubicin Kinetic Study in an in vitro 2D and 3D Tumor Model for Osteosarcoma in a Perfusion Bioreactor.

In vivo drug screening in animal models is contrary to ethical values, costly and time-consuming. Traditional static in vitro models do not reflect the basic characteristics of bone tumor microenvironments; therefore, perfusion bioreactors, in particular, would be an applicable choice due to their advantages to regenerate versatile bone tumor models for studying in vitro novel drug delivery systems. In this study, an optimal drug formulation of liposomal doxorubicin was prepared, and the release kinetics of the drug and its toxicity effect on MG-63 bone cancer cell line were investigated in two-dimensional, static three-dimensional media on a PLGA/β-TCP scaffold and also in a dynamic media in a perfusion bioreactor. In this assay, the efficacy of the IC50 of this formulation which had been obtained in two-dimensional cell culture (=0.1 μg/ml), was studied in static and dynamic three-dimensional media after 3 and 7 days. Liposomes with good morphology and encapsulation efficiency of 95% had release kinetics of the Korsmeyer-Peppas model. The results of cell growth before treatment and cell viability after treatment in all three environments were compared. Cell growth in 2D was rapid, while it was slow in static 3D conditions. In the dynamic 3D environment, it was significant compared to the static tumor models. Cell viability after 3 and 7 days from treatment was 54.73% and 13.39% in 2D conditions, 72.27% and 26.78% in the static 3D model, while 100% and 78.92% in the dynamic culture indicating the effect of drug toxicity over time, but drug resistance of 3D models compared to 2D culture. In the bioreactor, the formulation used in the mentioned concentration showed very small cytotoxicity demonstrating the dominance of mechanical stimuli on cell growth over drug toxicity. Increasing drug resistance in 3D models compared to 2D models indicates the superiority of liposomal Dox over free form to reduce IC50 concentration.

Platelet extracellular vesicles are efficient delivery vehicles of doxorubicin, an anti-cancer drug: preparation and in vitro characterization

Platelet extracellular vesicles (PEVs) are an emerging delivery vehi for anticancer drugs due to their ability to target and remain in the tumor microenvironment. However, there is still a lack of understanding regarding yields, safety, drug loading efficiencies, and efficacy of PEVs. In this study, various methods were compared to generate PEVs from clinical-grade platelets, and their properties were examined as vehicles for doxorubicin (DOX). Sonication and extrusion produced the most PEVs, with means of 496 and 493 PEVs per platelet (PLT), respectively, compared to 145 and 33 by freeze/thaw and incubation, respectively. The PEVs were loaded with DOX through incubation and purified by chromatography. The size and concentration of the PEVs and PEV-DOX were analyzed using dynamic light scattering and nanoparticle tracking analysis. The results showed that the population sizes and concentrations of PEVs and PEV-DOX were in the ranges of 120–150 nm and 1.2–6.2 × 1011 particles/mL for all preparations. The loading of DOX determined using fluorospectrometry was found to be 2.1 × 106, 1.7 × 106, and 0.9 × 106 molecules/EV using freeze/thaw, extrusion, and sonication, respectively. The internalization of PEVs was determined to occur through clathrin-mediated endocytosis. PEV-DOX were more efficiently taken up by MDA-MB-231 breast cancer cells compared to MCF7/ADR breast cancer cells and NIH/3T3 cells. DOX-PEVs showed higher anticancer activity against MDA-MB-231 cells than against MCF7/ADR or NIH/3T3 cells and better than acommercial liposomal DOX formulation. In conclusion, this study demonstrates that PEVs generated by PLTs using extrusion, freeze/thaw, or sonication can efficiently load DOX and kill breast cancer cells, providing a promising strategy for further evaluation in preclinical animal models. The study findings suggest that sonication and extrusion are the most efficient methods to generate PEVs and that PEVs loaded with DOX exhibit significant anticancer activity against MDA-MB-231 breast cancer cells.

Delivery of PEGylated liposomal doxorubicin by bispecific antibodies improves treatment in models of high-risk childhood leukemia.

High-risk childhood leukemia has a poor prognosis because of treatment failure and toxic side effects of therapy. Drug encapsulation into liposomal nanocarriers has shown clinical success at improving biodistribution and tolerability of chemotherapy. However, enhancements in drug efficacy have been limited because of a lack of selectivity of the liposomal formulations for the cancer cells. Here, we report on the generation of bispecific antibodies (BsAbs) with dual binding to a leukemic cell receptor, such as CD19, CD20, CD22, or CD38, and methoxy polyethylene glycol (PEG) for the targeted delivery of PEGylated liposomal drugs to leukemia cells. This liposome targeting system follows a "mix-and-match" principle where BsAbs were selected on the specific receptors expressed on leukemia cells. BsAbs improved the targeting and cytotoxic activity of a clinically approved and low-toxic PEGylated liposomal formulation of doxorubicin (Caelyx) toward leukemia cell lines and patient-derived samples that are immunophenotypically heterogeneous and representative of high-risk subtypes of childhood leukemia. BsAb-assisted improvements in leukemia cell targeting and cytotoxic potency of Caelyx correlated with receptor expression and were minimally detrimental in vitro and in vivo toward expansion and functionality of normal peripheral blood mononuclear cells and hematopoietic progenitors. Targeted delivery of Caelyx using BsAbs further enhanced leukemia suppression while reducing drug accumulation in the heart and kidneys and extended overall survival in patient-derived xenograft models of high-risk childhood leukemia. Our methodology using BsAbs therefore represents an attractive targeting platform to potentiate the therapeutic efficacy and safety of liposomal drugs for improved treatment of high-risk leukemia.

Abstract 2420: Interstitial chemo-phototherapy for treatment of locally advanced hepatocellular carcinoma: A preclinical study

Abstract Background Patients with hepatocellular carcinoma (HCC) have a poor prognosis with a 5-year survival rate of only 35%. Patients with locally advanced HCC (single or multinodal tumors &amp;gt;3 cm) may not be candidates for curative treatments such as surgical resection, liver transplant or thermal ablation due to tumor size, poor liver function, or other medical co-morbidities. The objective of this study was to investigate the potential use of interstitial chemo-phototherapy (I-CPT) to treat locally advanced HCC with a porphyrin-phospholipid (PoP) liposomal formulation of doxorubicin (Dox) that combines the targeted delivery of doxorubicin with photodynamic therapy to improve local drug delivery. We aimed to identify a safe and effective light dose for the photoactivation of Dox-Pop in the treatment of locally advanced HCC tumors in two pre-clinical models. Methods Sprague-Dawley rats were inoculated with Morris hepatoma cells directly into the left lateral liver lobe. When the tumors reached 2-2.5 cm along the longest axis, they were treated with Dox-Pop followed by light (i.e. I-CPT). For each rat, we applied our image-based treatment planning to guide the light delivery. Following treatment, tumor response was measured through weekly contrast-enhanced magnetic resonance imaging (CE-MRI). Complete response was defined as no evidence of the tumor via MRI or pathological examination. Our image-based treatment planning was used to translate the findings in the rat study to woodchucks with 1.5-4 cm hepatitis virus induced HCC. Weekly blood samples and bi-weekly CE-MRI were performed to assess tumor response up to 70 days. Results Five out of the six rats treated with I-CPT had complete response, with three of the five surviving ≥10 weeks post treatment without tumor recurrence or progression. The other two rats with complete response died between 5-7 weeks post treatment. Death was not attributed to the original treatment. Control, untreated rats were euthanized due to tumor progression within 1-2 weeks after tumors reached 2-2.5 cm. From the rat studies, we identified that a light dose rate and dose of respectively, ≥13.4 mW/cm2 and ≥25 J/cm2 can effectively activate the Dox-Pop liposomes. Of the five woodchucks treated with I-CPT using our effective light regimen, two survived ≥ 68 days post treatment without tumor progression. Another woodchuck had a complete response, but was euthanized 55 days post treatment due to the development of a second tumor. One woodchuck had progressive disease following I-CPT. One woodchuck died unexpectedly 5 days post I-CPT. Death was attributed to high tumor necrosis. Conclusions This study demonstrated the potential use of I-CPT as a treatment option for locally advanced HCC not candidate for standard of care therapies. Ongoing studies are being conducted in woodchucks to optimize the I-CPT light settings and improve tumor response. Citation Format: Emily Oakley, Minhyung Kim, Sanjana Ghosh, Hilliard Kutscher, Jonathan Lovell, Leslie Curtin, Sandra Sexton, Alan Hutson, Gal Shafirstein. Interstitial chemo-phototherapy for treatment of locally advanced hepatocellular carcinoma: A preclinical study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2420.

Gold cluster encapsulated liposomes: theranostic agent with stimulus triggered release capability

Cancer is a major cause of death worldwide. Cancer-resistant to chemo or radiotherapy treatment is a challenge that could be overcome by a nanotechnology approach. Providing a theranostic nano-platform for different cancer treatment strategies could be revolutionary. Here we introduce a multifunctional theranostic nanostructure which has the capacity for improving cancer diagnosis and treatment through better chemo and radiotherapy and current x-ray imaging systems through co-encapsulation of a small gold cluster and anticancer drug doxorubicin. 2nm gold clusters represent good heating under radio frequency electric field (RF-EF) exposure and have been used for in vitro hyperthermia treatment of cancerous cells. Liposomal doxorubicin (169 ± 19.8nm) with gold clusters encapsulation efficiency of 13.2 ± 3.0% and doxorubicin encapsulation efficiency of 64.7 ± 0.7% were prepared and studied as a theranostic agent with a high potential in different cancer treatment modalities. Exposure to a radiofrequency electric field on prepared formulation caused 20.2 ± 2.1% drug release and twice decreasing of IC50 on colorectal carcinoma cells. X-ray attenuation efficiency of the liposomal gold cluster was better than commercial iohexol and free gold clusters in different concentrations. Finally, treatment of gold clusters on cancerous cells results in a significant decrease in the viability of irradiated cells to cobalt-60 beam. Based on these experiments, we concluded that the conventional liposomal formulation of doxorubicin that has been co-encapsulated with small gold clusters could be a suitable theranostic nanostructure for cancer treatment and merits further investigation.

A multicenter randomized trials to compare the bioequivalence and safety of a generic doxorubicin hydrochloride liposome injection with Caelyx ® in advanced breast cancer.

To compare the pharmacokinetic (PK) bioequivalence (BE) and safety of a generic pegylated liposomal doxorubicin (PLD) formulation with the reference product Caelyx®. A multicenter, single-dose, open-label, randomized, two-way crossover study was conducted in patients with breast cancer. For each period, the patients were administered with the test or the reference PLD intravenously at a dose of 50 mg/m2. Cmax, AUC0-t and AUC0-∞ for free, and encapsulated doxorubicin (doxorubicin) and partial AUC (AUC0-48h, AUC48h-t) for encapsulated doxorubicin were evaluated in 17 blood samples taken predose, and increasing time intervals over the following 14 days in each period. A washout period of 28-35 days was observed before crossing over. 48 patients were enrolled and randomised, of which 44 were included and analysed in bioequivalence set (BES). The 90% confidence intervals (CIs) of the geometric mean ratio (GMR) of Cmax, AUC0-t and AUC0-∞ for free doxorubicin and encapsulated doxorubicin all fall within the bioequivalent range of 80% to 125%. The 90% CIs of GMR of partial AUC (AUC0-48h, AUC48h-t) for encapsulated doxorubicin also fall within the bioequivalent range. 48 patients were all included in the safety set (SS). The incidence of treatment-emergent adverse events (TEAEs) related to T and R was 95.8% (46/48) and 97.8% (45/46) respectively. The highest incidence of TEAEs was various laboratory abnormalities. 2 patients withdrew due to T-drug-related AEs. Only one patient experienced serious adverse events and no death occurred in this study. There were no significant differences between the safety profiles of the generic formulation and Caelyx®. Bioequivalence between the test and the reference products was established for free and encapsulated doxorubicin. http://www.chinadrugtrials.org.cn, identifier [CTR20210375].

A liposomal formulation of simvastatin and doxorubicin for improved cardioprotective and anti-cancer effect

Anthracyclines such as doxorubicin (Dox) are the preferred chemotherapeutics for several cancers. However, Dox-induced cardiotoxicity limits its therapeutic potential. Liposomal encapsulation of Dox has been used for patients with risk to develop Dox induced cardiotoxicity but does not surpass the efficacy of the unencapsulated drug. Statins are widely used as cholesterol lowering drugs and have also demonstrated cardioprotective activity in cancer patients undergoing Dox therapy. We developed a liposome loaded with Dox and simvastatin (Sim) and investigated their effect on cardiomyocytes and zebrafish larvae. Furthermore, we investigated if the doses required for cardioprotection compromised the cytotoxicity of Dox in mammary and prostate cancer cells. Combination of Sim and Dox reduced ROS generation in cardiomyocytes, both given as free drugs, or co-encapsulated in liposomes. In contrast, Sim potentiated ROS-generation and cytotoxic activity of Dox towards cancer cells also when co-encapsulated in liposomes. In zebrafish larvae, Sim treatment reduced Dox-induced cardiac affection, and the liposomes did not induce any sign of Dox-induced cardiotoxicity. Our results show that liposomal co-encapsulation of Sim and Dox can be an efficient way of further reducing the risk of cardiotoxic events of liposomal Dox, while retaining, or even potentiating the anti-cancer effect of Dox.

The role of size in PEGylated liposomal doxorubicin biodistribution and anti-tumour activity.

The size of nanoliposome‐encapsulated drugs significantly affects their therapeutic efficacy, biodistribution, targeting ability, and toxicity profile for the cancer treatment. In the present study, the biodistribution and anti‐tumoral activity of PEGylated liposomal Doxorubicin (PLD) formulations with different sizes were investigated. First, 100, 200, and 400 nm PLDs were prepared by remote loading procedure and characterised for their size, zeta potential, encapsulation efficacy, and release properties. Then, in vitro cellular uptake and cytotoxicity were studied by flow cytometry and MTT assay, and compared with commercially available PLD Caelyx®. In vivo studies were applied on BALB/c mice bearing C26 colon carcinoma. The cytotoxicity and cellular uptake tests did not demonstrate any statistically significant differences between PLDs. The biodistribution results showed that Caelyx® and 100 nm liposomal formulations had the most doxorubicin (Dox) accumulation in the tumour tissue and, as a result, considerably suppressed tumour growth compared with 200 and 400 nm PLDs. In contrast, larger nanoparticles (200 and 400 nm formulations) had more accumulation in the liver and spleen. This study revealed that 90 nm Caelyx® biodistribution profile led to the stronger anti‐tumour activity of the drug and hence significant survival extension, and showed the importance of vesicle size in the targeting of nanoparticles to the tumour microenvironment for the treatment of cancer.

TLD-1, a novel liposomal doxorubicin, in patients (pts) with advanced solid tumors: Dose escalation and expansion part of a multicenter open-label phase I trial (SAKK 65/16).

3027 Background: TLD-1 is a novel liposomal doxorubicin that compared favorably to conventional liposomal formulations of doxorubicin in preclinical in vivo mouse breast cancer models. This phase I first-in-human trial is aiming to determine the recommended phase II dose (RP2D), toxicity profile, pharmacokinetics and preliminary activity. Methods: Patients with a maximum of 3 prior lines of systemic chemotherapy and preferably anthracycline-sensitive disease were eligible. TLD-1 was administered on day 1 iv over 60-90 minutes (depending on individual dose) q 21 days, for up to 6 or 9 cycles (according to prior anthracycline-exposure) with premedication of 8mg dexamethasone. Dose escalation with dose levels (DL) 1-7 of 10, 16, 23, 30, 35, 40 and 45mg/m2 started with an accelerated titration design, treating one pt at each DL up to DL6 (40mg/m2) followed by a modified continual reassessment method at DL7 due to observed toxicity. Results: 30 pts (F:M = 24:6) have been treated, one each at DLs 1-6, 15 pts at DL7 and an additional 9 pts at DL6. Most frequent tumor types included breast (n = 13), ovarian (n = 6), cervical cancer (n = 2) and cholangiocarcinoma (n = 2). Median age was 67.5 years (range:38-83), 13 pts were exposed to prior anthracyclines. The median number of cycles was 4 (range:1-9). No dose-limiting toxicities (DLT) occurred during cycle 1. At DLs 1 to 5, no treatment-related G3 AEs (TRAE) were observed. At DL6, there was one case of mucositis G3, one of palmar-plantar-erythrodysesthesia (PPE) G3 and one of anemia and neutropenia G3 each. One patient with pre-existing valvular cardiopathy developed symptoms of heart-failure G3 after 8 cycles. Echocardiography showed severe mitral regurgitation with normal LV-EF. In addition one case of urinary-tract infection G3 was seen. Dose-modifications or -delays due to AEs occurred in 7/50 cycles. At DL7, one case of mucositis G3, 3 events of PPE G3 and one case of fatigue G3 were reported. In addition, one case of infection with shingles occurred. Dose-modifications or -delays due to AEs occurred in 12/61 cycles. Shingles and heart failure were reported as SAEs. All toxicities listed above were categorized as TRAE. 29/30 pts were evaluable for response. Three breast cancer pts had a partial response, 2 at DL7 and 1 at DL6, 14 pts had stable disease. Conclusions: No DLT was observed up to DL7. RP2D was defined at 40mg/m2 due to cumulative PPE G3 at DL7. The trial is ongoing with a comparative PK-part evaluating the two iv liposomal formulations of doxorubicin TLD-1 and Caelyx. Clinical trial information: NCT03387917.

Frequency of Cardiac-Toxicity in Patient Treated with Anthracycline, An Institutional Perspective

Introduction: Cancer now affects more than one in three people in their lifetime. Cancer remains the leading cause of death in many countries. Anthracycline-based chemotherapy has played an important part in the current age of cancer treatment, with earlier diagnosis and novel medicines. Objective: To find out the frequency of cardiac-toxicity (cardiomyopathy) in patient treated with anthracycline Methodology: This Descriptive Study was carried out at the Oncology department of Hayatabad Medical Complex, Peshawar for duration of six months from 20 Dec, 2019 to 20 Jun, 2020. Patients attending the medical oncology department of Hayatabad Medical Complex, Peshawar were included in the study. Demographic data like age, sex was collected in the pre-designed Performa. The patients were sent for echocardiography which was done by cardiologist and ejection fraction was noted. The patients suitable for treatment on anthracycline were started with standard doses for his/her carcinoma which were for leukemia 25mg/m2 weekly for 1 months, for lymphoma (NHL) 50mg/m2 after every 21 days for 4.2 months, for breast carcinoma 60mg/m2 after every 21 days for 2.8 months. Results: As per frequencies and percentages for anthracycline induced cardiomyopathy, 31 (15.0%) patients had anthracycline induced cardiomyopathy. Conclusion: Our research shows that although preventing chronic cardiotoxicity may be challenging, prolong infusion methods for anthracyclines offer a reduced risk, and novel liposomal formulations of doxorubicin may provide less cardiotoxicity as compared to conventional doxorubicin. Key words: Anthracyclines; Cardiotoxicity; Treatment

Imaging Tumor Heterogeneity and the Variations in Nanoparticle Accumulation Using Perfluorooctyl Bromide Nanocapsule X‐Ray Computed Tomography Contrast

AbstractSystemically administered nanoparticles can extravasate into solid tumors through active or passive processes, showing high inter‐ and intraindividual variability, leading to inconsistent nanomedicine disposition in tumor lesions and variable therapeutic outcomes. Radio‐opaque Perfluorooctyl bromide‐loaded PEGylated cholesteryl‐functionalized polycaprolactone (PEG‐b‐PCL(Ch)) nanocapsules (PFOB NC) are assessed as imaging probes via X‐ray Computed Tomography (CT) as means to estimate nanoparticle disposition in solid tumors. This imaging platform can enable precision medicine by screening tumors for likelihood of nanoparticle accumulation prior to treatment. Single‐ and double‐lateral xenograft models are employed for imaging‐based nanoparticle accumulation assessment using a preclinical multimodal imaging system. Differences in tumor accumulation and spatial distribution are verified as a function of xenograft type via X‐ray CT and further corroborated by optical imaging. Slow growing OVCAR‐8 xenografts display granular distribution pattern and higher levels of PFOB NC disposition on a volume‐normalized basis. A549 xenografts, displaying faster growth, are less susceptible to PFOB NC accumulation and show only a diffuse‐like spatial distribution of the nanocapsule contrast. A direct correlation between tumor dispositions of PFOB NC and a commercial PEGylated liposomal doxorubicin formulation is found. This study highlights the potential of using nanoparticle‐based radio‐opaque contrasts to better understand nanoparticle disposition and foster improved therapeutic outcomes in cancer nanomedicine.

Imaged-guided focused ultrasound in combination with various formulations of doxorubicin for the treatment of diffuse intrinsic pontine glioma

BackgroundDiffuse intrinsic pontine glioma (DIPG) is a notoriously difficult tumor to treat, with an overall survival of DIPG patients being only 11 months. One of the major obstacles for the effective treatment of DIPG is the blood–brain barrier (BBB). In order to circumvent the BBB, drug delivery methods are needed that target the pontine area. One such approach is microbubble-mediated focused ultrasound (FUS)—a non-invasive method that can temporarily and locally open the BBB. Previously, it was shown that FUS is safe with minimal side effects and rapid recovery times in preclinical animal models with different DIPG tumors. However, recent studies have shown that combining FUS with a single treatment of the chemotherapeutic drug doxorubicin did not improve survival in a DIPG xenograft model. As the duration of doxorubicin exposure might play a role in tumor response, we hypothesized that the use of a long-circulation (PEGylated) liposomal formulation of doxorubicin could lead to improved overall survival through a longer exposure time to the tumor.MethodDIPG xenograft models were established with orthotopic injections of HSJD-DIPG-07 tumor cells into the pontine area of female athymic nude-foxn1nu mice. Tumor engraftment was confirmed with bioluminescence imaging (BLI) 40 days post-inoculation. Mice were randomized into groups receiving either liposomal formulations of doxorubicin (2B3-101 or Caelyx®) or free doxorubicin in combination with or without FUS treatment. Treatment groups received 5 mg/kg 2B3-101 or Caelyx® 1 h before FUS treatment or 5 mg/kg free doxorubicin immediately after FUS.ResultsHistological analysis, however, revealed liposome extravasation in healthy controls but not in HSJD-DIPG-07 xenograft 24 h after treatment. Furthermore, BLI monitoring did not show reduced signal after treatment, which was further illustrated with a survival analysis, showing no significant difference between treated and control animals (p = 0.3).ConclusionWe did not observe a treatment effect after a single dose of free doxorubicin or the liposomal formulations 2B3-101 or Caelyx® in combination with FUS in DIPG-bearing mice.

In vitro and in vivo activity of thermosensitive liposomes loaded with doxorubicin and cisplatin

Thermosensitive liposomes loaded with cisplatin and doxorubicin composed of DPPC, DSPC, and DPPE-PEG5000 with different ratios were prepared by thin film hydration method. The Differential Scanning Calorimetry (DSC) curves showed that the liposomes composed of DPPC-DSPC-DPPE-PEG5000 with phospholipid ratio 95:5:0.05 w/w were a suitable formulation as thermosensitive liposomes with a DSC peak at 42.1 °C. The effect of doxorubicin and cisplatin encapsulated non-thermosensitive and thermosensitive liposomes on cellular proliferation and IC50 in SKBR3 & MDA-MB-231 breast cancer and PC-3 & LNcaP prostate cancer cell lines was investigated. The results showed that doxorubicin loaded into thermosensitive liposomes showed 20-fold decrease in the IC50 at 42 °C while comparing it with the same at 37 °C. Also, the results showed a more than 35-fold and 12-fold decrease in the IC50 of cisplatin thermosensitive liposomes at 42 °C, while compared with free cisplatin and cisplatin thermosensitive liposomes at any temperature. The in vivo results showed that the effect of doxorubicin encapsulated thermosensitive liposomes at hyperthermic conditions during the treatment as the tumor growth inhibition was measured 1.5-fold higher than any of the liposomal formulations of doxorubicin. It was also noticed that the tumor volume reduced to 150 mm3 in doxorubicin thermosensitive liposomes (G8) after 3 weeks during the treatment, but increased to 196 mm3 after 4 weeks. The Kaplan-Meir curve showed the 100% survival of the animals from G8 (thermosensitive liposomes containing doxorubicin plus hyperthermia) after 12 weeks. The flow cytometry data revealed more than 25% apoptotic cells and 6.25% necrotic cells in the tumor cells from the tissues of the G8 group of the animals. The results clearly indicate the superior efficacy of doxorubicin and cisplatin containing thermosensitive liposomes treatment during hyperthermia.

Dual-Loaded Liposomes Tagged with Hyaluronic Acid Have Synergistic Effects in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is the most lethal subtypes of breast cancer. Although chemotherapy is considered the most effective strategy for TNBC, most chemotherapeutics in current use are cytotoxic, meaning they target antiproliferative activity but do not inhibit tumor cell metastasis. Here, a TNBC-specific targeted liposomal formulation of epalrestat (EPS) and doxorubicin (DOX) with synergistic effects on both tumor cell proliferation and metastasis is described. These liposomes are biocompatible and effectively target tumor cells owing to hyaluronic acid (HA) modification on their surface. This active targeting, mediated by CD44-HA interaction, allows DOX and EPS to be delivered simultaneously to tumor cells in vivo, where they suppress not only TNBC tumor growth and the epithelial-mesenchymal transition, but also cancer stem cells, which collectively suppress tumor growth and metastasis of TNBC and may also act to prevent relapse of TNBC.