Preclinical In Vitro Studies with 3D Spheroids to Evaluate Cu(DDC)2 Containing Liposomes for the Treatment of Neuroblastoma

Preclinical in vitro studies of drug candidates for anticancer therapy are generally conducted on well-established 2D cell models. Unfortunately, these models are unable to mimic the properties of in vivo tumors. However, in vitro 3D models (spheroids) have been proven to be superior in reflecting the tumor microenvironment. Diethyldithiocarbamate (DDC−) is the active metabolite of Disulfiram, an approved drug for alcoholism and repurposed for cancer treatment. DDC− binds copper in a molar ratio of 2:1 resulting in a water-insoluble Cu(DDC)2 complex exhibiting anticancer activities. Delivery of the Cu(DDC)2 complex using nanoparticulate carriers provides decisive advantages for a parental application. In this study, an injectable liposomal Cu(DDC)2 formulation was developed and the toxicity was compared with a 2D neuroblastoma and a 3D neuroblastoma cell model. Our results indicate that Cu(DDC)2 liposomes complied with the size requirements of nanoparticles for intravenous injection and demonstrated high drug to lipid ratios as well as colloidal stability upon storage. Furthermore, an efficient cytotoxic effect on neuroblastoma 2D cell cultures and a very promising and even more pronounced effect on 3D cell cultures in terms of neuroblastoma monoculture and neuroblastoma co-culture with primary cell lines was proven, highly encouraging the use of Cu(DDC)2 liposomes for anticancer therapy.

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Chinese Veterinary Medicine B307 Promotes Cardiac Performance and Skeletal Muscle Contraction via Enhancing Intracellular Calcium Levels and Neural Electrical Activity in Animal and Cell Models

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2020/9064824 ◽

2020 ◽

Vol 2020 ◽

pp. 1-10

Author(s):

Chia-Ying Lien ◽

Chen-Wen Lu ◽

Chih-Hsiang Hsu ◽

Tai-Yuan Chuang ◽

Li-Yu Su ◽

...

Keyword(s):

Veterinary Medicine ◽

Electrical Activity ◽

Intracellular Calcium ◽

Cell Model ◽

Color Doppler ◽

Neuroblastoma Cell ◽

Motor Functions ◽

Electrical Activities

The study mainly investigated the effects of Chinese veterinary medicine B307 in cardiac and motor functions in animal models of pigeons and mice. Related cellular mechanisms were also studied in the neuroblastoma cell model of SH-SY5Y. Cardiac functions of pigeons and mice were examined by using moorFLPI Laser color Doppler imager and M-mode echocardiography, and motor functions were examined by using muscle electrical stimulation and force recording in the isolated breast muscle. Intracellular calcium levels and electrical activity of SH-SY5Y cells were examined by using Fura 2-AM fluorescence and MED64 system separately. Our results in vivo found that those pigeons under oral B307 treatment obviously enhanced subcutaneous microcirculation and contractile force and prolonged fatigue time in their breast muscles. Those mice under oral B307 treatment obviously elevated ejection fraction and cardiac output in their hearts. Our results in vitro showed that those SH-SY5Y cells under B307 treatment obviously increased intracellular calcium mobilization and electrical activities. These results revealed that improvement of cardiac and motor functions under B307 treatments may be caused by increasing electrical activities and intracellular calcium levels in neuromuscular cells and a similar mechanism may also occur in muscle cells. Thus, we suggested that B307 can be a functional Chinese veterinary medicine for flying pigeons.

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In vivo and in vitro studies using Clonorchis sinensis adult-derived total protein (CsTP) on cellular function and inflammatory effect in mouse and cell model

Parasitology Research ◽

10.1007/s00436-020-06651-1 ◽

2020 ◽

Vol 119 (5) ◽

pp. 1641-1652

Author(s):

Mei Shang ◽

Hengchang Sun ◽

Yinjuan Wu ◽

Yu Gong ◽

Zeli Tang ◽

...

Keyword(s):

Total Protein ◽

Cell Model ◽

Clonorchis Sinensis ◽

Cellular Function ◽

In Vitro Studies ◽

Inflammatory Effect

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An overview of the toxicology of HFA-134a (1,1,1,2-tetrafluoroethane)

Human & Experimental Toxicology ◽

10.1177/096032719501400903 ◽

1995 ◽

Vol 14 (9) ◽

pp. 715-720 ◽

Cited By ~ 37

Author(s):

DJ Alexander ◽

SE Libretto

Keyword(s):

Research And Development ◽

Cell Cultures ◽

In Vitro Studies ◽

In Vivo Studies ◽

Metered Dose Inhalers ◽

Safe Alternative ◽

Local Tolerance ◽

Metered Dose

1 This paper reviews the results of preclinical toxicology studies on HFA-134a carried out by Glaxo Research and Development Ltd. 2 A comprehensive range of studies was conducted in ani mal models suitable for the type of investigation. 3 The inhalation route of administration was used in all in vivo studies (with the exception of local tolerance and sensitisation studies) as patients will be exposed only to vapour during actuation of metered-dose inhalers. Cell cultures used for in vitro studies were also exposed to the vapour. 4 There was no mortality of rodents or dogs at extremely high vapour concentrations (81%v/v). 5 HFA-134a was considered not to be toxic or oncogenic and to provide a safe alternative to chlorofluorocarbons for use in pharmaceutical metered-dose inhalers.

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A New Cell Model for Investigating Prion Strain Selection and Adaptation

Viruses ◽

10.3390/v11100888 ◽

2019 ◽

Vol 11 (10) ◽

pp. 888

Author(s):

Alexandra Philiastides ◽

Juan Manuel Ribes ◽

Daniel Chun-Mun Yip ◽

Christian Schmidt ◽

Iryna Benilova ◽

...

Keyword(s):

Molecular Mechanisms ◽

Cell Model ◽

Prion Diseases ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Strain Selection ◽

Prion Strain ◽

Prion Strains

Prion diseases are fatal neurodegenerative diseases that affect humans and animals. Prion strains, conformational variants of misfolded prion proteins, are associated with distinct clinical and pathological phenotypes. Host-strain interactions result in the selective damage of distinct brain areas and they are responsible for strain selection and/or adaptation, but the underlying molecular mechanisms are unknown. Prion strains can be distinguished by their cell tropism in vivo and in vitro, which suggests that susceptibility to distinct prion strains is determined by cellular factors. The neuroblastoma cell line PK1 is refractory to the prion strain Me7, but highly susceptible to RML. We challenged a large number of clonal PK1 lines with Me7 and successfully selected highly Me7-susceptible subclones (PME) to investigate whether the prion strain repertoire of PK1 can be expanded. Notably, the Me7-infected PME clones were more protease-resistant when compared to RML-infected PME clones, which suggested that cell-adapted Me7 and RML are distinct prion strains. Strikingly, Me7-refractory cells, including PK1 and astrocytes in cortico-hippocampal cultures, are highly susceptible to prions, being derived from homogenates of Me7-infected PME cells, suggesting that the passage of Me7 in PME cells leads to an extended host range. Thus, PME clones represent a compelling cell model for strain selection and adaptation.

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Targeting MDM2 for Neuroblastoma Therapy: In Vitro and In Vivo Anticancer Activity and Mechanism of Action

Cancers ◽

10.3390/cancers12123651 ◽

2020 ◽

Vol 12 (12) ◽

pp. 3651

Author(s):

Wei Wang ◽

Xinjie Wang ◽

Mehrdad Rajaei ◽

Ji Youn Youn ◽

Atif Zafar ◽

...

Keyword(s):

Neuroblastoma Cell ◽

Wild Type ◽

Anticancer Activities ◽

High Risk Disease ◽

M Phase ◽

Mdm2 Expression ◽

Xenograft Models ◽

Underlying Mechanisms

Background: Neuroblastoma is an aggressive pediatric solid tumor with an overall survival rate of <50% for patients with high-risk disease. The majority (>98%) of pathologically-diagnosed neuroblastomas have wild-type p53 with intact functional activity. However, the mouse double minute 2 (MDM2) homolog, an E3 ubiquitin ligase, is overexpressed in neuroblastoma and leads to inhibition of p53. MDM2 also exerts p53-independent oncogenic functions. Thus, MDM2 seems to be an attractive target for the reactivation of p53 and attenuation of oncogenic activity in neuroblastoma. Methods: In this study, we evaluated the anticancer activities and underlying mechanisms of action of SP141, a first-in-class MDM2 inhibitor, in neuroblastoma cell lines with different p53 backgrounds. The findings were confirmed in mouse xenograft models of neuroblastoma. Results: We demonstrate that SP141 reduces neuroblastoma cell viability, induces apoptosis, arrests cells at the G2/M phase, and prevents cell migration, independent of p53. In addition, in neuroblastoma xenograft models, SP141 inhibited MDM2 expression and suppressed tumor growth without any host toxicity at the effective dose. Conclusions: MDM2 inhibition by SP141 results in the inhibition of neuroblastoma growth and metastasis, regardless of the p53 status of the cells and tumors. These findings provide proof-of-concept that SP141 represents a novel treatment option for both p53 wild-type and p53 null neuroblastoma.

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Endogenous hydrogen sulfide sulfhydrates IKKβ at cysteine 179 to control pulmonary artery endothelial cell inflammation

Clinical Science ◽

10.1042/cs20190514 ◽

2019 ◽

Vol 133 (20) ◽

pp. 2045-2059 ◽

Cited By ~ 4

Author(s):

Da Zhang ◽

Xiuli Wang ◽

Siyao Chen ◽

Selena Chen ◽

Wen Yu ◽

...

Keyword(s):

Hydrogen Sulfide ◽

Endothelial Cell ◽

Pulmonary Artery ◽

Cell Model ◽

Site Directed Mutagenesis ◽

Critical Event ◽

Hypertensive Rats ◽

Pulmonary Artery Endothelial Cell

Abstract Background: Pulmonary artery endothelial cell (PAEC) inflammation is a critical event in the development of pulmonary arterial hypertension (PAH). However, the pathogenesis of PAEC inflammation remains unclear. Methods: Purified recombinant human inhibitor of κB kinase subunit β (IKKβ) protein, human PAECs and monocrotaline-induced pulmonary hypertensive rats were employed in the study. Site-directed mutagenesis, gene knockdown or overexpression were conducted to manipulate the expression or activity of a target protein. Results: We showed that hydrogen sulfide (H2S) inhibited IKKβ activation in the cell model of human PAEC inflammation induced by monocrotaline pyrrole-stimulation or knockdown of cystathionine γ-lyase (CSE), an H2S generating enzyme. Mechanistically, H2S was proved to inhibit IKKβ activity directly via sulfhydrating IKKβ at cysteinyl residue 179 (C179) in purified recombinant IKKβ protein in vitro, whereas thiol reductant dithiothreitol (DTT) reversed H2S-induced IKKβ inactivation. Furthermore, to demonstrate the significance of IKKβ sulfhydration by H2S in the development of PAEC inflammation, we mutated C179 to serine (C179S) in IKKβ. In purified IKKβ protein, C179S mutation of IKKβ abolished H2S-induced IKKβ sulfhydration and the subsequent IKKβ inactivation. In human PAECs, C179S mutation of IKKβ blocked H2S-inhibited IKKβ activation and PAEC inflammatory response. In pulmonary hypertensive rats, C179S mutation of IKKβ abolished the inhibitory effect of H2S on IKKβ activation and pulmonary vascular inflammation and remodeling. Conclusion: Collectively, our in vivo and in vitro findings demonstrated, for the first time, that endogenous H2S directly inactivated IKKβ via sulfhydrating IKKβ at Cys179 to inhibit nuclear factor-κB (NF-κB) pathway activation and thereby control PAEC inflammation in PAH.

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Abnormal Sensitivity of Cortisol-Producing Adrenocortical Adenomas to Serotonin: In Vivo and in Vitro Studies

Yearbook of Endocrinology ◽

10.1016/s0084-3741(08)70169-0 ◽

2007 ◽

Vol 2007 ◽

pp. 358-361

Author(s):

D.E. Schteingart

Keyword(s):

In Vitro Studies ◽

Adrenocortical Adenomas

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The Venom of the Boomslang (Dispholidus Typus): In Vivo and in Vitro Studies

Thrombosis and Haemostasis ◽

10.1055/s-0038-1653532 ◽

1969 ◽

Vol 21 (02) ◽

pp. 234-244 ◽

Cited By ~ 5

Author(s):

N Mackay ◽

J.C Ferguson ◽

Antonia Bagshawe ◽

A.T.T Forrester ◽

G.P Mcnicol

Keyword(s):

In Vitro Studies

SummaryAn account is given of the effects of boomslang venom in man. Evidence was found of a fibrinolytic state apparently secondary to the coagulant action of the venom. These features rapidly responded to the administration of specific antivenom. In vitro studies, using a homogenate of boomslang parotids, confirmed the coagulant properties of the venom and showed them to be of much greater potency than the proteolytic actions.

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