scholarly journals Modulated Electro-Hyperthermia Resolves Radioresistance of Panc1 Pancreas Adenocarcinoma and Promotes DNA Damage and Apoptosis In Vitro

2020 ◽  
Vol 21 (14) ◽  
pp. 5100 ◽  
Author(s):  
Gertrud Forika ◽  
Andrea Balogh ◽  
Tamas Vancsik ◽  
Attila Zalatnai ◽  
Gabor Petovari ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Chromatin Condensation ◽  
Annexin V ◽  
Therapy Resistance ◽  
In Vivo Model ◽  
Tumor Growth Inhibition ◽  
Dna Double Strand Breaks ◽  
Cell Fractions

The poor outcome of pancreas ductal adenocarcinomas (PDAC) is frequently linked to therapy resistance. Modulated electro-hyperthermia (mEHT) generated by 13.56 MHz capacitive radiofrequency can induce direct tumor damage and promote chemo- and radiotherapy. Here, we tested the effect of mEHT either alone or in combination with radiotherapy using an in vivo model of Panc1, a KRAS and TP53 mutant, radioresistant PDAC cell line. A single mEHT shot of 60 min induced ~50% loss of viable cells and morphological signs of apoptosis including chromatin condensation, nuclear shrinkage and apoptotic bodies. Most mEHT treatment related effects exceeded those of radiotherapy, and these were further amplified after combining the two modalities. Treatment related apoptosis was confirmed by a significantly elevated number of annexin V single-positive and cleaved/activated caspase-3 positive tumor cells, as well as sub-G1-phase tumor cell fractions. mEHT and mEHT+radioterapy caused the moderate accumulation of γH2AX positive nuclear foci, indicating DNA double-strand breaks and upregulation of the cyclin dependent kinase inhibitor p21waf1 besides the downregulation of Akt signaling. A clonogenic assay revealed that both mono- and combined treatments affected the tumor progenitor/stem cell populations too. In conclusion, mEHT treatment can contribute to tumor growth inhibition and apoptosis induction and resolve radioresistance of Panc1 PDAC cells.

scholarly journals The Novel Cyclin-Dependent Kinase Inhibitor Flavopiridol Downregulates Bcl-2 and Induces Growth Arrest and Apoptosis in Chronic B-Cell Leukemia Lines

Blood ◽  
1997 ◽  
Vol 90 (11) ◽  
pp. 4307-4312 ◽  
Author(s):  
Andrea König ◽  
Gary K. Schwartz ◽  
Ramzi M. Mohammad ◽  
Ayad Al-Katib ◽  
Janice L. Gabrilove
Keyword(s):  
Cell Line ◽  
Kinase Inhibitor ◽  
Chromatin Condensation ◽  
Human Tumor ◽  
Trypan Blue Exclusion ◽  
Clinical Phase ◽  
Cellular Effects ◽  
Barr Virus

Abstract Flavopiridol is a novel, potent inhibitor of cyclin-dependent kinases (CDK). This synthetic flavone has been reported to exhibit antitumor activity in murine and human tumor cell lines in vitro and in vivo and is currently undergoing clinical phase I evaluation. In the present study, 1 Epstein-Barr virus (EBV)-transformed B-prolymphocytic cell line (JVM-2), 1 EBV-transformed B-CLL cell line (I83CLL), and 1 non-EBV transformed B-CLL cell line (WSU-CLL) were used as targets. Treatment of the cells with flavopiridol (100 nmol/L to 400 nmol/L) led to a marked dose- and time-dependent inhibition of cell growth and survival as determined using trypan blue exclusion. Morphologic analysis showed characteristic apoptotic changes such as chromatin condensation and fragmentation, membrane blebbing, and formation of apoptotic bodies. Furthermore, quantitative assessment of apoptosis-associated DNA strand breaks by in situ TdT labeling showed that a significant number of flavopiridol-treated cells underwent apoptosis. These cellular effects were associated with a significant decrease in bcl-2 expression as observed by Northern and Western blotting. The results showed that flavopiridol downregulates bcl-2 mRNA and bcl-2 protein expression within 24 hours. Genistein and quercetin, two flavonoids that do not inhibit CDKs, did not affect bcl-2 expression. These data suggest an additional mechanism of action of this new flavone which might be useful as an agent in the treatment of chronic lymphoid malignancies.

scholarly journals Enhancing Functional Platelet Release In Vivo from in Vitro-Grown Megakaryocytes Using the Protein Kinase Inhibitor SU6656

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1029-1029
Author(s):  
Danuta Jadwiga Jarocha ◽  
Karen K Vo ◽  
Randolph B Lyde ◽  
Vincent M Hayes ◽  
Mortimer Poncz
Keyword(s):  
Kinase Inhibitor ◽  
Conflicts Of Interest ◽  
Specific Inhibitor ◽  
Annexin V ◽  
Rock Inhibitor ◽  
Medicine Research ◽  
Nsg Mice ◽  
Platelet Release

Abstract The clinical demand for platelet transfusions is increasing, threatening the ability to obtain sufficient healthy donors to provide these platelets. Advances in regenerative medicine research have opened the possibility of generating sufficient in vitro-grown megakaryocytes and consequent platelets to supply a portion of the clinical platelet transfusion demand. We have shown that infusing megakaryocytes for obtaining released, functional platelets is a viable alternative strategy than trying to release platelets in vitro. However, for both approaches, in vitro-cultured megakaryocytes have lower ploidy and release fewer platelets than likely occurs in vivo by primary cells. SU6656 inhibitor, a Src kinase inhibitor, has been shown to influence ploidization in several megakaryocyte-like line with purported increase in proplatelets release. However, in our hands, other agents - such as the ROCK inhibitor Y27632 - while increasing polyploidization markedly, inhibited platelet release per infused megakaryocyte in vivo. We grew megakaryocytes from CD34+ cells for 12 days with or without SU6656 (2.5 µM) supplementation during the last 4 days. We found that the SU6656 inhibitor only increased the number of CD34+-derived megakaryocytes by ~15% at the end of the 12 day growth, but more markedly increase the percent of large megakaryocytes measured by FSC parameter in flow cytometry evaluation from 28 up to 41% and percent of high granular megakaryocytes from 27 to 45%. These changes were accompanied with a shift in average ploidy from 4.9 to 6.9 (p<0.0003, N=6). Notably, SU6656-treated megakaryocytes released ~4-fold more platelets per infused megakaryocytes in immunocompromized NSG mice than untreated similarly in vitro-grown megakaryocytes. By 24 hrs, there were 6.5-fold platelets from the infused SU6656-treated megakaryocytes than control untreated (p<0.037, N=6). Released platelets from the drug-treated and untreated megakaryocytes had similar levels of percent thiazole orange positivity as an indication that they were young platelets. Importantly, baseline annexin V, CD62p and PAC1 binding prior to agonist exposure were also similarly and increased to the same extent after thrombin (1U/ml) stimulation. Additionally, incorporation into a growing cremaster laser injury-induced thrombus in vivo was similar further indicating retained function by the platelets released from the drug-treated megakaryocytes. A number of strategies such as modifying the level of transcription factors have been proposed to increase the size, ploidy or proplatelets release from in vitro-grown megakaryocytes. In none of these cases have these released platelets in vivo biology been examined and demonstrated to replicate high release number per megakaryocyte and retained functionality. We show that terminal exposure of in vitro-grown megakaryocytes to the non-specific inhibitor SU6656 significantly increases in vivo yield while leaving in vivo half-life and functionality intact. The exact pathway affected by SU6656 that leads to these results is now being pursued. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Novel Cyclin-Dependent Kinase Inhibitor Flavopiridol Downregulates Bcl-2 and Induces Growth Arrest and Apoptosis in Chronic B-Cell Leukemia Lines

Blood ◽  
1997 ◽  
Vol 90 (11) ◽  
pp. 4307-4312 ◽  
Author(s):  
Andrea König ◽  
Gary K. Schwartz ◽  
Ramzi M. Mohammad ◽  
Ayad Al-Katib ◽  
Janice L. Gabrilove
Keyword(s):  
Cell Line ◽  
Kinase Inhibitor ◽  
Chromatin Condensation ◽  
Human Tumor ◽  
Trypan Blue Exclusion ◽  
Clinical Phase ◽  
Cellular Effects ◽  
Barr Virus

Flavopiridol is a novel, potent inhibitor of cyclin-dependent kinases (CDK). This synthetic flavone has been reported to exhibit antitumor activity in murine and human tumor cell lines in vitro and in vivo and is currently undergoing clinical phase I evaluation. In the present study, 1 Epstein-Barr virus (EBV)-transformed B-prolymphocytic cell line (JVM-2), 1 EBV-transformed B-CLL cell line (I83CLL), and 1 non-EBV transformed B-CLL cell line (WSU-CLL) were used as targets. Treatment of the cells with flavopiridol (100 nmol/L to 400 nmol/L) led to a marked dose- and time-dependent inhibition of cell growth and survival as determined using trypan blue exclusion. Morphologic analysis showed characteristic apoptotic changes such as chromatin condensation and fragmentation, membrane blebbing, and formation of apoptotic bodies. Furthermore, quantitative assessment of apoptosis-associated DNA strand breaks by in situ TdT labeling showed that a significant number of flavopiridol-treated cells underwent apoptosis. These cellular effects were associated with a significant decrease in bcl-2 expression as observed by Northern and Western blotting. The results showed that flavopiridol downregulates bcl-2 mRNA and bcl-2 protein expression within 24 hours. Genistein and quercetin, two flavonoids that do not inhibit CDKs, did not affect bcl-2 expression. These data suggest an additional mechanism of action of this new flavone which might be useful as an agent in the treatment of chronic lymphoid malignancies.

scholarly journals CAPN1 (Calpain1)-Mediated Impairment of Autophagic Flux Contributes to Cerebral Ischemia-Induced Neuronal Damage

Stroke ◽  
2021 ◽  
Author(s):  
Yueyang Liu ◽  
Xiaohang Che ◽  
Haotian Zhang ◽  
Xiaoxiao Fu ◽  
Yang Yao ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Kinase Inhibitor ◽  
Neuronal Damage ◽  
Neuronal Cell ◽  
In Vivo Model ◽  
Autophagic Flux ◽  
Autophagosome Formation

Background and Purpose: CAPN1 (calpain1)—an intracellular Ca 2+ -regulated cysteine protease—can be activated under cerebral ischemia. However, the mechanisms by which CAPN1 activation promotes cerebral ischemic injury are not defined. Methods: In the present study, we used adeno-associated virus-mediated genetic knockdown and pharmacological blockade (MDL-28170) of CAPN1 to investigate the role of CAPN1 in the regulation of the autophagy-lysosomal pathway and neuronal damage in 2 models, rat permanent middle cerebral occlusion in vivo model and oxygen-glucose–deprived primary neuron in vitro model. Results: CAPN1 was activated in the cortex of permanent middle cerebral occlusion–operated rats and oxygen-glucose deprivation–exposed neurons. Genetic and pharmacological inhibition of CAPN1 significantly attenuated ischemia-induced lysosomal membrane permeabilization and subsequent accumulation of autophagic substrates in vivo and in vitro. Moreover, inhibition of CAPN1 increased autophagosome formation by decreasing the cleavage of the autophagy regulators BECN1 (Beclin1) and ATG (autophagy-related gene) 5. Importantly, the neuron-protective effect of MDL-28170 on ischemic insult was reversed by cotreatment with either class III-PI3K (phosphatidylinositol 3-kinase) inhibitor 3-methyladenine or lysosomal inhibitor chloroquine (chloroquine), suggesting that CAPN1 activation-mediated impairment of autophagic flux is crucial for cerebral ischemia-induced neuronal damage. Conclusions: The present study demonstrates for the first time that ischemia-induced CAPN1 activation impairs lysosomal function and suppresses autophagosome formation, which contribute to the accumulation of substrates and aggravate the ischemia-induced neuronal cell damage. Our work highlights the vital role of CAPN1 in the regulation of cerebral ischemia–mediated autophagy-lysosomal pathway defects and neuronal damage.

Activity of AT13387, a novel, non-ansamycin inhibitor of heat shock protein 90, against gastrointestinal stromal tumors (GIST).

2013 ◽  
Vol 31 (4_suppl) ◽  
pp. 105-105 ◽  
Author(s):  
Daruka Mahadevan ◽  
Geoffrey Shapiro ◽  
Sandra E. Kurtin ◽  
James M. Cleary ◽  
John F. Lyons ◽  
...  
Keyword(s):  
Phase I ◽  
Kinase Inhibitor ◽  
Heat Shock Protein 90 ◽  
Tumor Growth Inhibition ◽  
Resistance Mutations ◽  
Hsp90 Inhibition ◽  
Secondary Resistance ◽  
Imatinib Resistant

105 Background: AT13387 is a second-generation potent, novel non-ansamycin HSP90 inhibitor (Kd 0.71nM). The majority of GIST tumors are characterized by activating mutations of c-KIT, an HSP90 client protein. Secondary resistance mutations within c-KIT limit clinical responses to TKIs. The dependence of c-KIT and its mutated forms on HSP90 suggests that HSP90 inhibition may be a valuable treatment option for imatinib-sensitive and resistant clones. In vitro, AT13387 inhibited the proliferation of imatinib-sensitive (GIST882, GIST-T1) and imatinib-resistant (GIST430, GIST48) cell lines. In vivo, AT13387 demonstrated anti-tumor activity in the imatinib-sensitive (GIST-PSW) and imatinib-resistant (GIST430) xenograft models. Induction of HSP70, depletion of phospho-c-KIT and inhibition of c-KIT signaling were observed in both models. Combination treatment of imatinib and AT13387 in the GIST430 model was well tolerated and significantly enhanced tumor growth inhibition over either monotherapies. Methods: In a completed phase I study, AT13387 was administered IV over 1 hour twice weekly or weekly of a 28-day cycle in a standard 3+3 dose-escalation design. The primary endpoint was to determine the MTD; secondary endpoints included PK, PD, safety and tolerability. Results: The PK exposures were dose-dependent and linear. AT13387 was well tolerated on both schedules. DLTs included primarily G2 AEs of GI toxicities, fatigue and infusion site reactions. The once weekly RP2D was determined to be 260 mg/m2. HSP70 induction was 2–7 fold at higher doses. A total of 7 GIST subjects were enrolled. An objective and durable PR was observed in one subject and 2 SDs at 8, 7 and 11 months, respectively. The PR subject demonstrated molecular resistance to kinase inhibitor treatment in the c-KIT gene prior to initiation of AT13387 therapy in two resected lesions by harboring the same activating c-KIT deletion in exon 11 and two separate TKI resistance mutations in exon 17. Conclusions: Overall, these results suggest AT13387 is a promising agent in GIST, including TKI-resistant c-Kit positive GIST. AT13387 is currently being evaluated in combination with imatinib in an ongoing phase I/II study. Clinical trial information: NCT01294202.

scholarly journals Clarithromycin synergizes with cisplatin to inhibit ovarian cancer growth in vitro and in vivo

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Bo Zhou ◽  
Meng Xia ◽  
Bin Wang ◽  
Niresh Thapa ◽  
Lijuan Gan ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Acquired Resistance ◽  
Xenograft Model ◽  
Annexin V ◽  
Therapy Resistance ◽  
Tumor Xenograft ◽  
Cancer Cell Growth ◽  
Cisplatin Therapy

Abstract Background Cisplatin-based chemotherapy is the first-line treatment for ovarian cancer. However, acquired resistance to cisplatin treatment often occurs in epithelial ovarian cancer, and effective and practical methods for overcoming this obstacle are urgently needed. The study aimed to demonstrate the synergistic effect of clarithromycin (CAM) with cisplatin to inhibit ovarian carcinoma cells growth in vitro and in vivo. Results We performed CCK-8 assay to detect apoptosis rates in response to CAM alone or in combination with cisplatin, which were further confirmed by Annexin V and PI staining methods and western blotting. Mechanistically, CAM could reduce endogenous antioxidant enzyme expression and increase the levels of reactive oxygen species (ROS) to augment the cytotoxic effect of cisplatin. Meanwhile, a tumor xenograft model in athymic BALB/c-nude mice demonstrated that CAM combined with cisplatin resulted in reduced tumor growth and weight compared with cisplatin alone. Conclusion Collectively, our results indicate that CAM works synergistically with cisplatin to inhibit ovarian cancer cell growth, which may be manipulated by a ROS-mediated mechanism that enhances cisplatin therapy, and offers a novel strategy for overcoming cisplatin therapy resistance.

Tamarix articulata (T. articulata) - An Important Halophytic Medicinal Plant with Potential Pharmacological Properties

2019 ◽  
Vol 20 (4) ◽  
pp. 285-292 ◽  
Author(s):  
Abdullah M. Alnuqaydan ◽  
Bilal Rah
Keyword(s):  
Medicinal Plant ◽  
Biological Activities ◽  
Wide Spectrum ◽  
Biological Properties ◽  
In Vivo Model ◽  
Drug Candidate ◽  
Phytochemical Analysis ◽  
Pharmacological Properties

Background:Tamarix Articulata (T. articulata), commonly known as Tamarisk or Athal in Arabic region, belongs to the Tamaricaece species. It is an important halophytic medicinal plant and a good source of polyphenolic phytochemical(s). In traditional medicines, T. articulata extract is commonly used, either singly or in combination with other plant extracts against different ailments since ancient times.Methods:Electronic database survey via Pubmed, Google Scholar, Researchgate, Scopus and Science Direct were used to review the scientific inputs until October 2018, by searching appropriate keywords. Literature related to pharmacological activities of T. articulata, Tamarix species, phytochemical analysis of T. articulata, biological activities of T. articulata extracts. All of these terms were used to search the scientific literature associated with T. articulata; the dosage of extract, route of administration, extract type, and in-vitro and in-vivo model.Results:Numerous reports revealed that T. articulata contains a wide spectrum of phytochemical(s), which enables it to have a wide window of biological properties. Owing to the presence of high content of phytochemical compounds like polyphenolics and flavonoids, T. articulata is a potential source of antioxidant, anti-inflammatory and antiproliferative properties. In view of these pharmacological properties, T. articulata could be a potential drug candidate to treat various clinical conditions including cancer in the near future.Conclusion:In this review, the spectrum of phytochemical(s) has been summarized for their pharmacological properties and the mechanisms of action, and the possible potential therapeutic applications of this plant against various diseases discussed.

Derivatives of Deoxypodophyllotoxin Induce Apoptosis Through Bcl-2/Bax Proteins Expression

2020 ◽  
Vol 20 ◽  
Author(s):  
Ya-Nan Li ◽  
Ni Ning ◽  
Lei Song ◽  
Yun Geng ◽  
Jun-Ting Fan ◽  
...  
Keyword(s):  
Annexin V ◽  
Mtt Method ◽  
Anthriscus Sylvestris ◽  
Anti Tumor Activity ◽  
Derivatives Of ◽  
Toxicity In Vitro ◽  
Ht 29 ◽  
Mcf 7

Background: Deoxypodophyllotoxin, isolated from theTraditional Chinese Medicine Anthriscus sylvestris, is well-known because of its significant antitumor activity with strong toxicity in vitro and in vivo. Objective: In this article, we synthesized a series of deoxypodophyllotoxin derivatives, and evaluated their antitumor effectiveness.Methods:The anti tumor activity of deoxypodophyllotoxin derivatives was investigated by the MTT method. Apoptosis percentage was measured by flow cytometer analysis using Annexin-V-FITC. Results: The derivatives revealed obvious cytotoxicity in the MTT assay by decreasing the number of late cancer cells. The decrease of Bcl-2/Bax could be observed in MCF-7, HepG2, HT-29 andMG-63 using Annexin V-FITC. The ratio of Bcl-2/Bax in the administration group was decreased, which was determined by the ELISA kit. Conclusion: The derivatives of deoxypodophyllotoxin could induce apoptosis in tumor cell lines by influencing Bcl-2/Bax.

scholarly journals Radiofrequency Irradiation Modulates TRPV1-Related Burning Sensation in Rosacea

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1424
Author(s):  
Seyeon Oh ◽  
Myeongjoo Son ◽  
Joonhong Park ◽  
Donghwan Kang ◽  
Kyunghee Byun
Keyword(s):  
Burning Sensation ◽  
In Vivo Model ◽  
Molecular Evidence ◽  
Exposed Skin ◽  
Heat Treated ◽  
Vitro Model ◽  
Effective Use ◽  
Inflammatory Molecules

Rosacea is a skin inflammatory condition that is accompanied by not only redness and flushing but also unseen symptoms, such as burning, stinging, and itching. TRPV1 expression in UVB-exposed skin can lead to a painful burning sensation. Upregulated TRPV1 expression helps release neuropeptides, including calcitonin gene-related peptide, pituitary adenylate cyclase-activating polypeptide, and vasoactive intestinal peptide, which can activate macrophage and inflammatory molecules. In this study, we found that radiofrequency (RF) irradiation reduced TRPV1 activation and neuropeptide expression in a UVB-exposed in vivo model and UVB- or heat-treated in an in vitro model. RF irradiation attenuated neuropeptide-induced macrophage activation and inflammatory molecule expression. Interestingly, the burning sensation in the skin of UVB-exposed mice and patients with rosacea was significantly decreased by RF irradiation. These results can provide experimental and molecular evidence on the effective use of RF irradiation for the burning sensation in patients with rosacea.

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