scholarly journals Radiosensitization of malignant gliomas following intracranial delivery of paclitaxel biodegradable polymer microspheres

2014 ◽  
Vol 120 (5) ◽  
pp. 1078-1085 ◽  
Author(s):  
Patrik Gabikian ◽  
Betty M. Tyler ◽  
Irma Zhang ◽  
Khan W. Li ◽  
Henry Brem ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Radiation Treatment ◽  
Malignant Gliomas ◽  
In Vivo Studies ◽  
9L Gliosarcoma ◽  
Fischer 344 ◽  
Improvement In Survival ◽  
Paclitaxel Treatment

Object The aim of this study was to demonstrate that paclitaxel could function as a radiosensitizer for malignant glioma in vitro and in vivo. Methods The radiosensitizing effect of paclitaxel was tested in vitro using the human U373MG and rat 9L glioma cell lines. Cell cycle arrest in response to paclitaxel exposure was quantified by flow cytometry. Cells were subsequently irradiated, and toxicity was measured using the clonogenic assay. In vivo studies were performed in Fischer 344 rats implanted with intracranial 9L gliosarcoma. Rats were treated with control polymer implants, paclitaxel controlled-release polymers, radiotherapy, or a combination of the 2 treatments. The study end point was survival. Results Flow cytometry demonstrated G2-M arrest in both U373MG and 9L cells following 6–12 hours of paclitaxel exposure. The order in which the combination treatment was administered was significant. Exposure to radiation treatment (XRT) during the 6–12 hours after paclitaxel treatment resulted in a synergistic reduction in colony formation. This effect was greater than the effect from either treatment alone and was also greater than the effect of radiation exposure followed by paclitaxel. Rats bearing 9L gliosarcoma tumors treated with paclitaxel polymer administration followed by single-fraction radiotherapy demonstrated a synergistic improvement in survival compared with any other treatment, including radiotherapy followed by paclitaxel treatment. Median survival for control animals was 13 days; for those treated with paclitaxel alone, 21 days; for those treated with XRT alone, 21 days; for those treated with XRT followed by paclitaxel, 45 days; and for those treated with paclitaxel followed by XRT, more than 150 days (p < 0.0001). Conclusions These results indicate that paclitaxel is an effective radiosensitizer for malignant gliomas because it renders glioma cells more sensitive to ionizing radiation by causing G2-M arrest, and induces a synergistic response to chemoradiotherapy.

scholarly journals Efficacy of local polymer-based and systemic delivery of the anti-glutamatergic agents riluzole and memantine in rat glioma models

2014 ◽  
Vol 120 (4) ◽  
pp. 854-863 ◽  
Author(s):  
Kaleb Yohay ◽  
Betty Tyler ◽  
Kyle D. Weaver ◽  
Andrea C. Pardo ◽  
Dan Gincel ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Malignant Gliomas ◽  
In Vitro Assays ◽  
Systemic Delivery ◽  
Organotypic Cultures ◽  
9L Gliosarcoma ◽  
Rat Glioma ◽  
F98 Glioma

Object The poor outcome of malignant gliomas is largely due to local invasiveness. Previous studies suggest that gliomas secrete excess glutamate and destroy surrounding normal peritumoral brain by means of excitotoxic mechanisms. In this study the authors assessed the effect on survival of 2 glutamate modulators (riluzole and memantine) in rodent glioma models. Methods In an in vitro growth inhibition assay, F98 and 9L cells were exposed to riluzole and memantine. Mouse cerebellar organotypic cultures were implanted with F98 glioma cells and treated with radiation, radiation + riluzole, or vehicle and assessed for tumor growth. Safety and tolerability of intracranially implanted riluzole and memantine CPP:SA polymers were tested in F344 rats. The efficacy of these drugs was tested against the 9L model and riluzole was further tested with and without radiation therapy (RT). Results In vitro assays showed effective growth inhibition of both drugs on F98 and 9L cell lines. F98 organotypic cultures showed reduced growth of tumors treated with radiation and riluzole in comparison with untreated cultures or cultures treated with radiation or riluzole alone. Three separate efficacy experiments all showed that localized delivery of riluzole or memantine is efficacious against the 9L gliosarcoma tumor in vivo. Systemic riluzole monotherapy was ineffective; however, riluzole given with RT resulted in improved survival. Conclusions Riluzole and memantine can be safely and effectively delivered intracranially via polymer in rat glioma models. Both drugs demonstrate efficacy against the 9L gliosarcoma and F98 glioma in vitro and in vivo. Although systemic riluzole proved ineffective in increasing survival, riluzole acted synergistically with radiation and increased survival compared with RT or riluzole alone.

scholarly journals P06.01 Delta24-ACT oncolytic adenovirus as a therapeutic approach for DIPG

2019 ◽  
Vol 21 (Supplement_3) ◽  
pp. iii36-iii36
Author(s):  
V Laspidea ◽  
M Puigdelloses ◽  
M García-Moure ◽  
I Iñigo-Marco ◽  
J Gallego ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Death ◽  
Cell Lines ◽  
In Vivo Studies ◽  
Immunogenic Cell Death ◽  
Murine Models ◽  
Antitumoral Effect ◽  
Infected Cells

Abstract BACKGROUND Diffuse intrinsic pontine glioma (DIPG) is an aggressive brain tumor, being the leading cause of pediatric death caused by cancer. We previously showed that administration of the oncolytic virus Delta-24-RGD to DIPG murine models was safe and led to an increase in the median survival of these animals. However, not all the animals responded, underscoring the need to improve this therapy. In order to increase the antitumoral effect of the virus, we have engineered Delta-24-RGD with the costimulatory ligand 4-1BBL (Delta24-ACT). 4-1BB is a costimulatory receptor that promotes the survival and expansion of activated T cells, and the generation and maintenance of memory CD8+ T cells. In this project, we evaluated the oncolytic effect of Delta24-ACT and the antitumor immune response in DIPG murine models. MATERIALS AND METHODS We use the NP53 and XFM murine DIPG cell lines. Flow cytometry was used to assess cell infectivity and ligand expression. We analyzed viral replication using a method based in hexon detection, and viral cytotoxic effect using the MTS assay. For immunogenic cell death analysis, we measured ATP secretion by a luminometric assay and calreticulin location by flow cytometry and immunofluorescence. For in vivo studies, cells and virus were injected in the pons of the mice, using the screw-guided system. RESULTS In vitro, Delta24-ACT was able to infect and induce cell death in a dose-dependent manner in murine DIPG cell lines. In addition, Delta24-ACT was able to replicate in these tumor cells and to express viral proteins. Moreover, infected cells expressed 41BBL in their membranes. Delta24-ACT could induce immunogenic cell death due to an increased secretion of ATP and calreticulin translocation to the membrane of infected cells (in no-infected cells it located in the ER), DAMPs that can trigger the immune response activation. In vivo, Delta24-ACT demonstrated to be safe in all the tested doses and was able to induce a significant increase in the median survival of the treated animals. Moreover, long-term survivors display immunological memory. CONCLUSIONS Delta24-ACT treatment led to antitumoral effect in DIPG murine cell lines in vitro. Of significance, we have demonstrated that in vivo administration of Delta24-ACT is safe and results in an enhanced antitumor effect. Future in vivo studies will explore the underlying immune mechanism of the virus.

A Transgenic Mouse Model of Aggressive B-Cell Malignancy for Evaluating Anti-Human CD37 Therapeutics

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 188-188
Author(s):  
Kyle A Beckwith ◽  
Frank W Frissora ◽  
Matthew R Stefanovski ◽  
Jutta Deckert ◽  
Carlo M Croce ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
B Cells ◽  
Mouse Model ◽  
B Cell ◽  
Transgenic Mouse ◽  
In Vivo Studies ◽  
B Cell Malignancy ◽  
Cell Malignancy

Abstract Abstract 188 BACKGROUND: Introduction of the anti-CD20 antibody rituximab has led to remarkable progress in the development of targeted therapies for CLL and other B-cell malignancies. Despite prolonging patient survival, therapies targeting CD20 have not been curative. In recent years, alternative targets for therapeutic antibodies have emerged. One of the most promising targets has been CD37, which is highly expressed on malignant B-cells in chronic lymphocytic leukemia (CLL) and non-Hodgkin's lymphoma. The recent interest in this target has led to the generation of novel anti-CD37 therapeutics that could benefit from more extensive preclinical evaluation. However, preclinical development of these agents has been limited by the absence of appropriate leukemia animal models that provide targets expressing human CD37 (hCD37). Here we describe the development and characterization of a transgenic mouse where CLL-like leukemic B-cells express hCD37 and aggressively transplant into syngenic hosts. We demonstrate the utility of this unique mouse model by evaluating the in vivo efficacy of IMGN529, a novel antibody-drug conjugate targeting hCD37 that consists of the CD37-targeting K7153A antibody linked to the maytansinoid DM1 via the thioether SMCC linker. METHODS: The hCD37 transgenic mouse (hCD37-Tg) founder lines were generated by conventional methodology at the OSU Transgenic Facility. B-cell specific expression of hCD37 is driven by immunoglobulin heavy chain promoter and Ig-μ enhancer elements. Founder lines were evaluated by RT-PCR and flow cytometry to confirm RNA and protein expression, respectively. These lines were then crossed with the EμTCL1 mouse model of CLL to generate hCD37xTCL1 mice that develop CD5+CD19+hCD37+ leukemia. For in vivo studies, splenocytes from a leukemic hCD37xTCL1 donor were injected i.v. into healthy hCD37-Tg mice. Mice were randomly assigned to the following treatment groups (n=8–10 per group): IMGN529 conjugate, its K7153A antibody component, or negative controls (isotype antibody-DM1 conjugate or trastuzumab). Upon diagnosis of leukemia, a 10 mg/kg dose was administered i.p. and repeat doses were given 2 times per week for 3 weeks (70 mg/kg total). Peripheral blood disease was monitored by flow cytometry, using counting beads to obtain the absolute number of leukemic CD5+CD19+ B-cells. CD37 expression levels were determined by quantitative flow cytometry. In vitro cytotoxicity was evaluated after 24 hour incubation by flow cytometry with Annexin V and propidium iodide staining. RESULTS: IMGN529 and its K7153A antibody component demonstrated comparable in vitro activity against freshly isolated human CLL cells even in the absence of cross-linking agents (mean IMGN529 cytotoxicity=50.04% vs. 48.85% for K7153A; p=0.175; n=9). Both compounds also demonstrated cytotoxicity against hCD37 Tg B-cells ex vivo in a cross-linking dependent manner, and while expression of hCD37 in hCD37-Tg animals was B-cell specific, the expression levels were substantially lower than those observed in human CLL cells. In vivo studies with transferred hCD37xTCL1 splenocytes demonstrated rapid and complete depletion of CD5+CD19+ leukemic B-cells in response to IMGN529 conjugate, but not K7153A antibody treatment. After 1 week of IMGN529 treatment, peripheral blood leukemia was nearly undetectable and previously detected massive splenomegaly was no longer palpable. In contrast, leukemic counts and spleen sizes continued to increase in control cohorts. CONCLUSIONS: In summary, our group has generated a mouse model that develops a transplantable CD5+CD19+ leukemia expressing hCD37. We demonstrate the utility of this model for both in vitro and in vivo testing of therapeutics targeting hCD37. In addition, preclinical mouse studies expose the robust anti-leukemic effects of IMGN529 in this in vivo model of aggressive B-cell malignancy, despite the relatively low expression of hCD37 on the leukemic B-cells. Our engraftment model shows that IMGN529 is capable of eliminating widespread and highly proliferative mouse leukemia by a mechanism that is both CD37 antigen and conjugate dependent. Therefore, we propose that this novel therapeutic may also exhibit substantial efficacy in a wide range of human B-cell malignancies, even those with relatively low CD37 expression. [This work was supported by NIH (NM, JCB), LLS (NM, JCB) and Pelotonia (KAB)]. Disclosures: Deckert: ImmunoGen Inc.: Employment.

Circulating CD62E Occurs in Soluble Form as Well as Bound to Endothelial Microparticles (EMP): Functional Differences in Platelet Aggregation in Thrombotic Thrombocytopenic Purpura (TTP).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2624-2624
Author(s):  
Joaquin J. Jimenez ◽  
Wenche Jy ◽  
Lucia M. Mauro ◽  
Michael N. Markou ◽  
George W. Burke ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Platelet Aggregation ◽  
Acute Phase ◽  
Critical Role ◽  
In Vivo Studies ◽  
Soluble Form ◽  
Dependent Manner

Abstract Injured endothelial cells (EC) are believed to play a critical role in the pathophysiology of TTP. Soluble markers of endothelial disturbance measured by enzyme-linked immunoassay (ELISA) have been found elevated in TTP. We have recently demonstrated an increase in the release of CD31/42b- EMP, and CD62E+ EMP. Moreover, we have observed that CD62E+ EMP also express vWF. The aim of this study was to quantitate soluble (s) vs. EMP-bound CD62E (bCD62E) in vitro and in vivo, in relation to the functional activity of vWF+ EMP. METHODS: Brain and renal microvascular endothelial cells (MVEC) were cultured and treated with 10ng/mL TNF-α to induce activation, or deprived of serum and growth factors (GFD) to induce apoptosis. Culture supernatants were collected and evaluated in a time-dependent manner. For in vivo studies, platelet-poor plasma was obtained from 4 TTP patients during the acute phase and upon remission. Filtration through 0.1μm, which retains most EMP, was employed to discriminate between (s) and bCD62E. sCD62E was measured by ELISA post-filtration and bCD62E by ELISA pre-filtration. Additionally, CD62E+ and CD62E+/vWF+ EMP were measured by flow cytometry. To assess pro-aggregatory function, EMP were added to washed platelets in the presence of 1 mg/mL ristocetin and aggregates were measured by flow cytometry. RESULTS: In vitro: Activation did not induce release of sCD62E at 3 hours, although bCD62E was present (1.5±0.5X106 EMP/mL). At 6 hours, some sCD62E was detected in the filtrate (0.09±0.02 ng/mL), but most was present in the unfiltered medium (3.5±0.85 ng/mL), signifying that the majority was bCD62E, confirmed by a doubling of CD62E+ EMP (3.0±0.6X106/mL). Subsequently, sCD62E levels were 1.0±0.2 ng/mL at 12 hr, 3.5±0.7 ng/mL at 18 hr, and 5±0.9 ng/mL at 24 hr. In contrast, EMP counts at 12, 18 and 24 hours were 4.6±1, 7±1.3 and 9±1.8 X106/mL (p=0.01, p=0.01, p=0.02, respectively). For all time periods, 40-60% of CD62E were positive for vWF. In control or GFD cultures, there was not a significant increase in sCD62E or CD62E+ EMP at any time period. MVEC from renal gave similar results. In acute TTP plasma samples, CD62E measured by ELISA was significantly increased (65±22 ng/mL) vs. remission (30±6 ng/mL). bCD62E accounted for 50% in acute and 15% in remission. CD62E+/vWF+ EMP were significantly elevated in plasma from acute TTP patients vs. remission (15±4.5 vs. 3±0.5, p=0.01). Sample filtration resulted in a decrease of &gt;95% EMP in both acute and remission TTP plasma. MVEC-derived CD62E+/vWF+ EMP resulted in a dose-dependent increase in platelet aggregation. Additionally, plasma from 4 TTP patients with elevated CD62E+/vWF+ EMP obtained during the acute phase enhanced the formation of platelet aggregates by 48±12% (p=0.02) above remission plasma with low EMP counts. CONCLUSIONS: The results demonstrate that CD62E heretofore regarded as a soluble marker of endothelial dysfunction, in reality exists in both a soluble and EMP-bound form. Indeed, this distinction is highly relevant because CD62E+ EMP also express vWF and are pro-aggregatory to platelets. These EMP have been shown to be elevated during the acute phase of TTP and decrease upon remission. Thus, CD62E+/vWF+ EMP may be active participants in the formation of platelet-rich thrombi in TTP.

scholarly journals The Plant-Derived Compound Resveratrol in Brain Cancer: A Review

Biomolecules ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 161 ◽  
Author(s):  
Terezia Kiskova ◽  
Peter Kubatka ◽  
Dietrich Büsselberg ◽  
Monika Kassayova
Keyword(s):  
Brain Cancer ◽  
Standard Treatment ◽  
Alkylating Agents ◽  
Malignant Gliomas ◽  
In Vivo Studies ◽  
Anticancer Effects ◽  
The Central Nervous System ◽  
Methylguanine Methyltransferase

Despite intensive research, malignant brain tumors are among the most difficult to treat due to high resistance to conventional therapeutic approaches. High-grade malignant gliomas, including glioblastoma and anaplastic astrocytoma, are among the most devastating and rapidly growing cancers. Despite the ability of standard treatment agents to achieve therapeutic concentrations in the brain, malignant gliomas are often resistant to alkylating agents. Resveratrol is a plant polyphenol occurring in nuts, berries, grapes, and red wine. Resveratrol crosses the blood‒brain barrier and may influence the central nervous system. Moreover, it influences the enzyme isocitrate dehydrogenase and, more importantly, the resistance to standard treatment via various mechanisms, such as O6-methylguanine methyltransferase. This review summarizes the anticancer effects of resveratrol in various types of brain cancer. Several in vitro and in vivo studies have presented promising results; however, further clinical research is necessary to prove the therapeutic efficacy of resveratrol in brain cancer treatment.

scholarly journals EXTH-18. ENGINEERED EXOSOMES AS GENE DELIVERY TOOLS FOR THE TREATMENT OF GLIOBLASTOMA

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii90-ii90
Author(s):  
Malcolm McDonald ◽  
Anwar Hossain ◽  
Eric Momin ◽  
Irtiza Hasan ◽  
Sanjay Singh ◽  
...  
Keyword(s):  
Cell Entry ◽  
In Vivo Studies ◽  
In Vitro Proliferation ◽  
Stem Cell Lines ◽  
Improvement In Survival ◽  
U87 Cells ◽  
Delivery Efficiency ◽  
Significant Opportunity

Abstract Although we previously showed that exosomes are capable of delivering anti-glioma microRNAs (miRs) to brain tumors (Lang et al. 2018), our studies revealed significant opportunity to 1) improve packaging and delivery efficiency of exosomes and 2) expand the repertoire of anti-glioma miRs. We hypothesized that incorporation of viral proteins into exosomes would enhance miR packaging and cell entry. To test this hypothesis, we engineered exosomes that express retroviral Gag and VSVg proteins (eExos). Specifically, HEK293T cells were transfected with Gag, VSVg, and with Cre-recombinase containing plasmid (pCre) to generate eExos-pCre. After 48hrs eExos-pCre were isolated by differential ultracentrifugation. Western analyses verified Gag and VSVg in eExos-pCre, and PCR documented pCre in these exosomes. Next, U87 cells harboring a dsRed-eGFP-loxP reporter-gene were treated with eExos-pCre or control exosomes. Flow cytometry demonstrated that eExos-pCre resulted in 82% conversion of red cells to green, compared with controls (2% conversion), verifying the effectiveness of eExos to deliver plasmids containing anti-glioma agents. To identify effective anti-glioma miRs, we conducted a high-throughput screen of 539 miRs against 7 glioma stem cell lines (GSCs) and identified miR-124-2, miR-135-a-2, and Let7i as the most potent anti-glioma miRs. We then studied the ability of eExos to package and deliver plasmids of these miRs either singly (eExos-miR-124, eExos-miR-135, eExos-miRLet7i) or as a tri-cistronic plasmid (eExos-miR-124-135-Let7i). Although eExos-miR-124, eExos-miR-135, and eExos-miRLet7i significantly decreased in vitro proliferation in all three GSCs (p&lt; 0.01), eExos-miR-124-135-Let7i were most effective (p&lt; 0.001). In in vivo studies, mice harboring GSC231 gliomas were injected with each of the eExos-miRs. Most significant improvement in survival was seen with eExos-miR-124-135-Let7i (median 75 versus 32.5 days for controls, p&lt; 0.001). We conclude that eExos are a novel delivery strategy for human gliomas and that a tri-cistronic plasmid of miR-124-135-Let7i is highly effective against GBM and worthy of clinical translation.

scholarly journals Liposomal Lapatinib in Combination with Low-Dose Photodynamic Therapy for the Treatment of Glioma

2019 ◽  
Vol 8 (12) ◽  
pp. 2214 ◽  
Author(s):  
Carl Fisher ◽  
Girgis Obaid ◽  
Carolyn Niu ◽  
Warren Foltz ◽  
Alyssa Goldstein ◽  
...  
Keyword(s):  
Photodynamic Therapy ◽  
Cell Lines ◽  
Malignant Gliomas ◽  
Protoporphyrin Ix ◽  
Growth Factor Receptor ◽  
In Vivo Studies ◽  
Epidermal Growth

Background: Malignant gliomas are highly invasive and extremely difficult to treat tumours with poor prognosis and outcomes. Photodynamic therapy (PDT), mediated by Gleolan®, has been studied previously with partial success in treating these tumours and extending lifetime. We aim to determine whether combining PDT using ALA-protoporphyrin IX (PpIX) with a liposomal formulation of the clinical epidermal growth factor receptor (EGFR) inhibitor, lapatinib, would increase the anti-tumour PDT efficacy. Methods: Lapatinib was given in vitro and in vivo 24 h prior to PDT and for 3–5 days following PDT to elicit whether the combination provided any benefits to PDT therapy. Live-cell imaging, in vitro PDT, and in vivo studies were performed to elucidate the effect lapatinib had on PDT for a variety of glioma cell lines and as well as GSC-30 neurospheres in vivo. Results: PDT combined with lapatinib led to a significant increase in PpIX accumulation, and reductions in the LD50 of PpIX mediated PDT in two EGFR-driven cell lines, U87 and U87vIII, tested (p < 0.05). PDT + lapatinib elicited stronger MRI-quantified glioma responses following PDT for two human glioma-derived tumours (U87 and GSC-30) in vivo (p < 0.05). Furthermore, PDT leads to enhanced survival in rats following treatment with lapatinib compared to lapatinib alone and PDT alone (p < 0.05). Conclusions: As lapatinib is approved for other oncological indications, a realization of its potential combination with PDT and in fluorescence-guided resection could be readily tested clinically. Furthermore, as its use would only be in acute settings, long-term resistance should not pose an issue as compared to its use as monotherapy.

Effective conversion of irinotecan to SN-38 after intratumoral drug delivery to an intracranial murine glioma model in vivo

2011 ◽  
Vol 114 (3) ◽  
pp. 689-694 ◽  
Author(s):  
Weijun Wang ◽  
Alex Ghandi ◽  
Leonard Liebes ◽  
Stan G. Louie ◽  
Florence M. Hofman ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Survival Time ◽  
Topoisomerase I ◽  
Malignant Gliomas ◽  
In Vivo Studies ◽  
Dependent Manner ◽  
Systemic Delivery ◽  
Series Of Experiments

Object Irinotecan (CPT-11), a topoisomerase I inhibitor, is a cytotoxic agent with activity against malignant gliomas and other tumors. After systemic delivery, CPT-11 is converted to its active metabolite, SN-38, which displays significantly higher cytotoxic potency. However, the achievement of therapeutically effective plasma levels of CPT-11 and SN-38 is seriously complicated by variables that affect drug metabolism in the liver. Thus the capacity of CPT-11 to be converted to the active SN38 intratumorally in gliomas was addressed. Methods For in vitro studies, 2 glioma cell lines, U87 and U251, were tested to determine the cytotoxic effects of CPT-11 and SN-38 in a dose-dependent manner. In vivo studies were performed by implanting U87 intracranially into athymic/nude mice. For a period of 2 weeks, SN-38, CPT-11, or vehicle was administered intratumorally by means of an osmotic minipump. One series of experiments measured the presence of SN-38 or CPT-11 in the tumor and surrounding brain tissues after 2 weeks' exposure to the drug. In a second series of experiments, after 2 weeks' exposure to the drug, the animals were maintained, in the absence of drug, until death. The survival curves were then calculated. Results The results show that the animals that had CPT-11 delivered intratumorally by the minipump expressed SN-38 in vivo. Furthermore, both CPT-11 and SN-38 accumulated at higher levels in tumor tissues compared with uninvolved brain. Intratumoral delivery of CPT-11 or SN-38 extended the average survival time of tumor-bearing animals from 22 days to 46 and 65 days, respectively. Conclusions These results demonstrate that intratumorally administered CPT-11 can be effectively converted to SN-38 and this method of drug delivery is effective in extending the survival time of animals bearing malignant gliomas.

560 Alpha-tocopheryloxyacetic acid induces apoptosis of murine rhabdomyosarcoma in vitro while modulating innate and adaptive immune responses in vivo

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A594-A594
Author(s):  
Fernanda Szewc ◽  
Longzhen Song ◽  
Sean Rinella ◽  
Christopher Dubay ◽  
Emmanuel Akporiaye ◽  
...  
Keyword(s):  
Gene Expression ◽  
Flow Cytometry ◽  
T Cells ◽  
Tumor Growth ◽  
Live Cell Imaging ◽  
Control Diet ◽  
Annexin V ◽  
In Vivo Studies

BackgroundRelapsed pediatric sarcomas have a poor prognosis with no available curative options. Alpha-Tocopheryloxyacetic acid (a-TEA) is a redox-silent analog of alpha-tocopherol that induces apoptotic and immunogenic cell death of tumor cells at doses that are not harmful to healthy normal cells. In a first-in-human clinical trial, a-TEA was well tolerated in adults with advanced solid tumors (NCT02192346), but has not yet been studied in pediatric sarcoma. We used a murine model of rhabdomyosarcoma (M3-9-M RMS) to assess the in vitro and in vivo anti-tumor effects of a-TEA.MethodsIn vitro studies were performed on the M3-9-M RMS cell line to measure a-TEA-mediated apoptosis using flow cytometry (Annexin V+/7AAD+ cells) and live cell imaging (Annexin V+ cells). In vivo studies involved orthotopic implantation of luciferase+ M3-9-M tumor cells into syngeneic C57BL/6 recipients. Once tumors were palpable, mice were randomized to a control diet or a-TEA-supplemented chow for 21 days and evaluated for bioluminescence, tumor growth and overall survival. Gene expression of tumor-infiltrating and splenic T cells were analyzed by bulk RNA-Seq and flow cytometry respectively.ResultsM3-9-M RMS treatment with 2.5–100 uM a-TEA induced apoptosis in a dose-dependent manner within 24 hours (p < 0.05) as measured by flow cytometry and live cell imaging. In-vivo studies with the M3-9-M RMS mouse model showed that recipients of a-TEA chow had 30–40% reduced tumor growth (p<0.01) and bioluminescence (p<0.05), leading to prolonged survival (> 4 weeks) compared to recipients of matched control chow (p<0.05). Spleen cells isolated from a-TEA-fed tumor-bearing mice demonstrated increased levels of IFN??+ cells, CD4+ T-cells, Ki-67 proliferation, and decrease in splenic CD11b+ arginase-1+ (p<0.01) and PD-L1+ cells (p<0.05) compared to their counterparts on the control diet. Gene set enrichment analyses of excised RMS tumors after a-TEA treatment revealed increased gene expression of CD24, EP300, CXCR4, and c-Jun as compared to tumors from mice fed control chow.ConclusionsThese data indicate that a-TEA mediates apoptosis of RMS in vitro and suppresses in vivo tumor growth, leading to prolonged survival likely via enhanced activation of adaptive immunity through CD4+ T cells and suppression of innate immunity through regulation of myeloid cell subsets. Furthermore, a-TEA may have direct effects on tumor cell proliferation through EP300 and c-Jun as well as indirect effects on tumor growth by regulation of immune cell recruitment through CD24 and CXCR4 gene expression. Administration of a-TEA as a potential salvage treatment for RMS is warranted.AcknowledgementsThe study was supported by NIH TL1 TR002375 (FS), St. Baldrick’s Stand up to Cancer (SU2C) Pediatric Dream Team Translational Research Grant SU2C-AACR-DT-27-17, NIH/NCI R01 CA215461, American Cancer Society Research Scholar Grant RSG- 18-104-01-LIB, and the Midwest Athletes Against Childhood Cancer (MACC) Fund (CMC). SU2C is a division of the Entertainment Industry Foundation. Research grants are administered by the American Association for Cancer Research, the scientific partner of SU2C. The contents of this article do not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US government.Ethics ApprovalThe University of Wisconsin-Madison Animal Care and Use Committee approved all protocols (M005915).

scholarly journals MicroRNAs Expression Induces Apoptosis of Macrophages in Response to Leishmania major (MRHO/IR/75/ER): An In-Vitro and In-Vivo Study

2020 ◽  
Author(s):  
Mostafa GHOLAMREZAEI ◽  
Soheila ROUHANI ◽  
Mehdi MOHEBALI ◽  
Samira MOHAMMADI-YEGANEH ◽  
Mostafa HAJI MOLLA HOSEINI ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Leishmania Major ◽  
Parasite Burden ◽  
Parasite Load ◽  
Lesion Size ◽  
In Vivo Studies ◽  
Raw 264.7 Cells ◽  
Control Group

Background: We aimed to investigate the effect of miR-15a mimic and inhibitor of miR-155 expression on apoptosis induction in macrophages infected with Iranian strain of Leishmania major in-vitro and in-vivo. Methods: RAW 264.7 cells were infected with L. major promastigotes (MRHO/IR/75/ER), and then were treated with miRNAs. For in-vivo experiment, BALB/c mice were inoculated with L. major promastigotes, and then they were treated with miRNAs. For evaluation of miRNA therapeutic effect, in-vitro and in-vivo studies were performed using quantitative Real-time PCR, Flow cytometry, lesion size measurement, and Limiting Dilution Assay (LDA). This study was performed in Shahid Beheshti University of Medical Sciences in 2019. Results: In-vitro results of flow cytometry showed that using miR-15a mimic, miR-155 inhibitor or both of them increased apoptosis of macrophages. In in-vivo, size of lesion increased during experiment in control groups (P<0.05) while application of both miR-155 inhibitor and miR-15a mimic inhibited the increase in the size of lesions within 6 wk of experiment (P=0.85). LDA results showed that microRNA therapy could significantly decrease parasite load in mimic or inhibitor receiving groups compared to the control group (P<0.05). Conclusion: miR-155 inhibitor and miR-15a mimic in L. major infected macrophages can induce apoptosis and reduce parasite burden. Therefore, miRNA-based therapy can be proposed as new treatment for cutaneous leishmaniasis.

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