Impact of organotin compounds on the growth of epidermoid Lewis carcinoma

Introduction: Search for new compounds with a broad antitumor and antimetastatic potency due to multiple targeting remains important in medicinal chemistry, pharmacology and oncology. We report the efficacy of hybrid organotin agents bis-(3,5-di-tert-butyl-4-hydroxyphenylthiolate) dimethyltin (Ме3) and (3,5-di-tert-butyl-4-hydroxyphenylthiolate) triphenyltin (Ме5). Materials and methods: The compounds were administered to mice bearing the spontaneously metastatic epidermoid Lewis lung carcinoma (LLC). The efficacy of the treatment was evaluated by mean life span, percentage of tumor growth inhibition, number of lung metastases, frequency of metastasis, tumor weight 21 days after tumor cell inoculation, and a degree of lung damage according to the method of D. Tarin and J.E. Price. Results and discussion: For new organotin compounds containing an antioxidant protective fragment of 2,6-di-tert-butylphenol, moderate antitumor and pronounced antimetastatic effects were revealed in the Lewis model of epidermoid lung carcinoma; more active for Me5. Some features of the development of the process of metastasis were recorded with the introduction of various doses of hybrid organotin compounds. Conclusion: Substances Ме3 and Ме5 possess specific activity on the model under investigation, which allows one to suggest these organotins as promising series of antitumor and antimetastatic agents with multiple targeting mode of action.

Detection and therapy of neuroblastoma minimal residual disease using [64/67Cu]Cu-SARTATE in a preclinical model of hepatic metastases

Background A major challenge to the long-term success of neuroblastoma therapy is widespread metastases that survive initial therapy as minimal residual disease (MRD). The SSTR2 receptor is expressed by most neuroblastoma tumors making it an attractive target for molecularly targeted radionuclide therapy. SARTATE consists of octreotate, which targets the SSTR2 receptor, conjugated to MeCOSar, a bifunctional chelator with high affinity for copper. Cu-SARTATE offers the potential to both detect and treat neuroblastoma MRD by using [64Cu]Cu-SARTATE to detect and monitor the disease and [67Cu]Cu-SARTATE as the companion therapeutic agent. In the present study, we tested this theranostic pair in a preclinical model of neuroblastoma MRD. An intrahepatic model of metastatic neuroblastoma was established using IMR32 cells in nude mice. The biodistribution of [64Cu]Cu-SARTATE was measured using small-animal PET and ex vivo tissue analysis. Survival studies were carried out using the same model: mice (6–8 mice/group) were given single doses of saline, or 9.25 MBq (250 µCi), or 18.5 MBq (500 µCi) of [67Cu]Cu-SARTATE at either 2 or 4 weeks after tumor cell inoculation. Results PET imaging and ex vivo biodistribution confirmed tumor uptake of [64Cu]Cu-SARTATE and rapid clearance from other tissues. The major clearance tissues were the kidneys (15.6 ± 5.8% IA/g at 24 h post-injection, 11.5 ± 2.8% IA/g at 48 h, n = 3/4). Autoradiography and histological analysis confirmed [64Cu]Cu-SARTATE uptake in viable, SSTR2-positive tumor regions with mean tumor uptakes of 14.1–25.0% IA/g at 24 h. [67Cu]Cu-SARTATE therapy was effective when started 2 weeks after tumor cell inoculation, extending survival by an average of 13 days (30%) compared with the untreated group (mean survival of control group 43.0 ± 8.1 days vs. 55.6 ± 9.1 days for the treated group; p = 0.012). No significant therapeutic effect was observed when [67Cu]Cu-SARTATE was started 4 weeks after tumor cell inoculation, when the tumors would have been larger (control group 14.6 ± 8.5 days; 9.25 MBq group 9.5 ± 1.6 days; 18.5 MBq group 15.6 ± 4.1 days; p = 0.064). Conclusions Clinical experiences of peptide-receptor radionuclide therapy for metastatic disease have been encouraging. This study demonstrates the potential for a theranostic approach using [64/67Cu]Cu-SARTATE for the detection and treatment of SSTR2-positive neuroblastoma MRD.

Overcoming bioprocess bottlenecks in the large-scale expansion of high-quality hiPSC aggregates in vertical-wheel stirred suspension bioreactors

Background Human induced pluripotent stem cells (hiPSCs) hold enormous promise in accelerating breakthroughs in understanding human development, drug screening, disease modeling, and cell and gene therapies. Their potential, however, has been bottlenecked in a mostly laboratory setting due to bioprocess challenges in the scale-up of large quantities of high-quality cells for clinical and manufacturing purposes. While several studies have investigated the production of hiPSCs in bioreactors, the use of conventional horizontal-impeller, paddle, and rocking-wave mixing mechanisms have demonstrated unfavorable hydrodynamic environments for hiPSC growth and quality maintenance. This study focused on using computational fluid dynamics (CFD) modeling to aid in characterizing and optimizing the use of vertical-wheel bioreactors for hiPSC production. Methods The vertical-wheel bioreactor was modeled with CFD simulation software Fluent at agitation rates between 20 and 100 rpm. These models produced fluid flow patterns that mapped out a hydrodynamic environment to guide in the development of hiPSC inoculation and in-vessel aggregate dissociation protocols. The effect of single-cell inoculation on aggregate formation and growth was tested at select CFD-modeled agitation rates and feeding regimes in the vertical-wheel bioreactor. An in-vessel dissociation protocol was developed through the testing of various proteolytic enzymes and agitation exposure times. Results CFD modeling demonstrated the unique flow pattern and homogeneous distribution of hydrodynamic forces produced in the vertical-wheel bioreactor, making it the opportune environment for systematic bioprocess optimization of hiPSC expansion. We developed a scalable, single-cell inoculation protocol for the culture of hiPSCs as aggregates in vertical-wheel bioreactors, achieving over 30-fold expansion in 6 days without sacrificing cell quality. We have also provided the first published protocol for in-vessel hiPSC aggregate dissociation, permitting the entire bioreactor volume to be harvested into single cells for serial passaging into larger scale reactors. Importantly, the cells harvested and re-inoculated into scaled-up vertical-wheel bioreactors not only maintained consistent growth kinetics, they maintained a normal karyotype and pluripotent characterization and function. Conclusions Taken together, these protocols provide a feasible solution for the culture of high-quality hiPSCs at a clinical and manufacturing scale by overcoming some of the major documented bioprocess bottlenecks.

Overcoming Bioprocess Bottlenecks in the Large-Scale Expansion of High Quality hiPSC Aggregates in Vertical-Wheel Stirred Suspension Bioreactors

BackgroundHuman induced pluripotent stem cells (hiPSCs) hold enormous promise in accelerating breakthroughs in understanding human development, drug screening, disease modeling and cell and gene therapies. Their potential, however, has been bottlenecked in a mostly laboratory setting due to bioprocess challenges in the scale-up of large quantities of high-quality cells for clinical and manufacturing purposes. While several studies have investigated the production of hiPSCs in bioreactors, the use of conventional horizontal-impeller, paddle and rocking-wave mixing mechanisms have demonstrated unfavourable hydrodynamic environments for hiPSC growth and quality maintenance. This study focused on using computational fluid dynamics (CFD) modeling to aid in characterizing and optimizing the use of vertical-wheel bioreactors for hiPSC production.MethodsThe vertical-wheel bioreactor was modeled with CFD simulation software Fluent at agitation rates between 20rpm and 100rpm. These models produced fluid flow patterns that mapped out a hydrodynamic environment to guide in the development of hiPSC inoculation and in-vessel aggregate dissociation protocols. The effect of single-cell inoculation on aggregate formation and growth was tested at select CFD modeled agitation rates and feeding regimes in the vertical-wheel bioreactor. An in-vessel dissociation protocol was developed through the testing of various proteolytic enzymes and agitation exposure times.ResultsCFD modeling demonstrated the unique flow pattern and homogeneous distribution of hydrodynamic forces produced in the vertical-wheel bioreactor, making it the opportune environment for systematic bioprocess optimization of hiPSC expansion. We developed a scalable, single-cell inoculation protocol for the culture of hiPSCs as aggregates in vertical-wheel bioreactors, achieving over 30-fold expansion in 6 days without sacrificing cell quality. We have also provided the first published protocol for in-vessel hiPSC aggregate dissociation, permitting the entire bioreactor volume to be harvested into single-cells for serial passaging into larger scale reactors. Importantly, the cells harvested and re-inoculated into scaled-up vertical-wheel bioreactors not only maintained consistent growth kinetics, they maintained a normal karyotype and pluripotent characterization and function.ConclusionsTaken together, these protocols provide a feasible solution for the culture of high quality hiPSCs at a clinical and manufacturing scale by overcoming some of the major documented bioprocess bottlenecks.

Overcoming Bioprocess Bottlenecks in the Large-Scale Expansion of High Quality hiPSC Aggregates in Vertical-Wheel Stirred Suspension Bioreactors

BackgroundHuman induced pluripotent stem cells (hiPSCs) hold enormous promise in accelerating breakthroughs in understanding human development, drug screening, disease modeling and cell and gene therapies. Their potential, however, has been bottlenecked in a mostly laboratory setting due to bioprocess challenges in the scale-up of large quantities of high-quality cells for clinical and manufacturing purposes. While several studies have investigated the production of hiPSCs in bioreactors, the use of conventional horizontal-impeller, paddle and rocking-wave mixing mechanisms have demonstrated unfavourable hydrodynamic environments for hiPSC growth and quality maintenance. This study focused on using computational fluid dynamics (CFD) modeling to aid in characterizing and optimizing the use of vertical-wheel bioreactors for hiPSC production.MethodsThe vertical-wheel bioreactor was modeled with CFD simulation software Fluent at agitation rates between 20rpm and 100rpm. These models produced fluid flow patterns that mapped out a hydrodynamic environment to guide in the development of hiPSC inoculation and in-vessel aggregate dissociation protocols. The effect of single-cell inoculation on aggregate formation and growth was tested at select CFD modeled agitation rates and feeding regimes in the vertical-wheel bioreactor. An in-vessel dissociation protocol was developed through the testing of various proteolytic enzymes and agitation exposure times.ResultsCFD modeling demonstrated the unique flow pattern and homogeneous distribution of hydrodynamic forces produced in the vertical-wheel bioreactor, making it the opportune environment for systematic bioprocess optimization of hiPSC expansion. We developed a scalable, single-cell inoculation protocol for the culture of hiPSCs as aggregates in vertical-wheel bioreactors, achieving over 30-fold expansion in 6 days without sacrificing cell quality. We have also provided the first published protocol for in-vessel hiPSC aggregate dissociation, permitting the entire bioreactor volume to be harvested into single-cells for serial passaging into larger scale reactors. Importantly, the cells harvested and re-inoculated into scaled-up vertical-wheel bioreactors not only maintained consistent growth kinetics, they maintained a normal karyotype and pluripotent characterization and function.ConclusionsTaken together, these protocols provide a feasible solution for the culture of high quality hiPSCs at a clinical and manufacturing scale by overcoming some of the major documented bioprocess bottlenecks.

Hyperbaric oxygen suppresses stemness-associated properties and Nanog and oncostatin M expression, but upregulates β-catenin in orthotopic glioma models

Objective This study aimed to explore whether initial hyperbaric oxygen treatment affects the stemness of glioma stem cells using an in vivo basal ganglia glioma model. Methods A basal ganglia glioma rat model was established. Rats were exposed to normal oxygen or hyperbaric oxygen on days 2, 4, 6, 8, 10, and 12. After 16 days of glioma cell inoculation, western blot, ELISA, and flow cytometry were performed to examine stemness-associated properties by examining the expression of CD133, A2B5, Nanog, oncostatin M, β-catenin, Oct-3/4, Sox2, and Nestin. Results Initial hyperbaric oxygen treatment began to affect glioma stemness-associated properties. The proportion of CD133+A2B5+ cells was significantly reduced after initial hyperbaric oxygen treatment. Additionally, the expression of stemness-related genes such as Nanog and oncostatin M was reduced, while TGF-β and β-catenin were increased. Conclusions Initial hyperbaric oxygen treatment not only alters the hypoxic microenvironment but also affects the stemness-associated properties of cancer stem cells.

Microsurgical Technique of Locoregional Injection into the Hepatic Artery in Tumor-Bearing Mice

Background: Intraarterial injection into the hepatic artery represents an important route for locoregional administration for the treatment of hepatic tumors. In the present work, we describe microsurgical methodology for injection into the hepatic artery in mice. The technique was recently used for analysis of the phenomenon of endothelial capture in liver tumors. Methods: Two different models of hepatic tumors in C57BL/6 mice were used. Tumors were induced by intrahepatic cell inoculation. The preferential blood supply of tumors was studied using blocking of bioavailability of nontumoral endothelial epitope and the subsequent injection of fluorescent endothelium-specific antibody. The selective intraarterial injection of labeled antibody was performed in tumor-bearing mice. The procedure addressed variations of vascular anatomy of the hepatic artery in mice and used direct intraarterial injection with dispensable catheterization. Results: Both experimental tumor models showed preferential blood supply from the hepatic artery. The technique of hepatic arterial injection was adapted and performed according to two major anatomic variations of the hepatic artery. Using this technique, the selective enrichment of labeled antibody to tumor and liver blood vessels, which were perfused during the first intravascular passage, was demonstrated. Conclusions: The experimental hepatic arterial injection in mice is a feasible but demanding microsurgical procedure. The choice of subsequent operation steps is dependent on the vascular anatomy of the hepatic artery which has two major variations in mice.