Background: Immortalized mammalian cell lines are a valuable research tool, though they represent... more Background: Immortalized mammalian cell lines are a valuable research tool, though they represent a highly simplified model. Due to accumulated mutations they may not reflect characteristics of the disease or even the tissue they derive from. Objective: We aim to pinpoint factors distinguishing SeAx cells from two other cutaneous T-cell lymphoma (CTCL) cell lines, namely Hut78 and MyLa2000. Of note, these factors may influence cell sensitivity in an unspecific way and therefore should be taken under consideration. Methods: We evaluated transcriptional levels of drug transporters across cell lines, cell membrane permeability, functionality of pathways related to DNA damage response and activation of G2/M block. Results: Analysis of the transcriptional levels of genes coding drug efflux pumps indicated that they are not consistently down-regulated in SeAx. However, we noted that SeAx cell membrane is markedly more permeable than Hut78 and MyLa2000, which may contribute to increased chemosensitivity in an unspecific way. Moreover, though DNA damage response seemed to be at least partly functional in SeAx cells, they fail to activate G2/M block in response to psoralen + UVA treatment. Any DNA damage should be repaired before cells enter mitosis, in order to uphold genome integrity. Thus, a defective cell cycle block may contribute to cell sensitivity. Conclusions: We believe that factors such as increased membrane permeability or defective cell cycle block should be accounted for when comparing sensitivity of cell line panels to chemotherapeutics of interest. It is worth to exclude a simple, indiscriminative mechanisms of cell resistance or sensitivity before attempting comparisons. Cell lines that are indiscriminately sensitive to a broad range of chemicals may contribute to overestimating the cytotoxic potential of tested compounds if used in cytotoxicity studies.
Rhaponticum carthamoides is an endemic medicinal plant of Siberian origin. Its roots tissue is we... more Rhaponticum carthamoides is an endemic medicinal plant of Siberian origin. Its roots tissue is well known as a rich source of biologically active compounds. R. carthamoides crude extracts were obtained with two different extraction methods (sonication and shaking) and three extraction solvents with different polarity (chloroform, methanol and water). All extract were screened for antioxidative activity (phosphomolybdenum assay, β-carotene bleaching test, DPPH and FRAP tests), cytotoxic (MTT assay) and mutagenic activities (modified Ames test, with the use Vibrio harveyi strains). All tested extracts were able to reduce molybdenum and inhibit about 85% of linoleic acid peroxidation. However, only water and methanol extracts reduced Fe+3 ions and scavenged DPPH free radical. On the other hand chloroform extracts indicated minor toxicity against all tested cell lines. None of the tested extracts showed mutagenic activity. Key words: Antioxidative activity, cytotoxic activity, mutagen...
Rhaponticum carthamoides is a medicinal plant indigenous to Siberia. Although its root tissue is ... more Rhaponticum carthamoides is a medicinal plant indigenous to Siberia. Although its root tissue is a well-known source of biologically active compounds, recent studies have focused on the above-ground part of the plant due to its easier accessibility. An R. carthamoides plantation was recently established in Northern Poland. Composition and antioxidant activity were compared for R. carthamoides leaf extracts obtained from plants grown in two plantations: in Poland (Lubiewice) and in Russia (Koriazma); the latter plantation has growing conditions similar to those in the R. carthamoides natural habitat. Plant secondary compounds were extracted with chloroform, methanol or water. As indicated by gas chromatography-mass spectrometry (GC-MS), the content of most compounds analyzed was higher in extracts from plants grown in Lubiewice (Poland). Antioxidative activity (as measured by diphenyl 1-picrylhydrazyl free radical (DPPH) and ferric reducing antioxidant power (FRAP) assays) and cytoto...
DNA-damaging cancer therapies induce interferon expression and stimulate the immune system, promo... more DNA-damaging cancer therapies induce interferon expression and stimulate the immune system, promoting therapy responses. The immune-activating STING (Stimulator of Interferon Genes) pathway is induced when DNA or double-stranded RNA (dsRNA) is detected in the cell cytoplasm, which can be caused by viral infection or by DNA damage following chemo- or radiotherapy. Here, we investigated the responses of cutaneous T-cell lymphoma (CTCL) cells to the clinically applied DNA crosslinking photochemotherapy (combination of 8–methoxypsoralen and UVA light; 8–MOP + UVA). We showed that this treatment evokes interferon expression and that the type III interferon IFNL1 is the major cytokine induced. IFNL1 upregulation is dependent on STING and on the cytoplasmic DNA sensor cyclic GMP-AMP synthase (cGAS). Furthermore, 8–MOP + UVA treatment induced the expression of genes in pathways involved in response to the tumor necrosis factor, innate immune system and acute inflammatory response. Notably, ...
Rhaponticum carthamoides plants ("maral root") are widely used in Siberian folk medicin... more Rhaponticum carthamoides plants ("maral root") are widely used in Siberian folk medicine. The present study reports for the first time the presence of pentacyclic terpenoid, α-amyrin, in methanol extract from leaves of this plant. α-Amyrin induced proliferation of human keratinocytes (HaCaT) by about 18% while other extract components were ineffective. A panel of biochemical and cell-based assays testing the antioxidative and cytoprotective activites of α-amyrin indicated no antioxidative activity of this compound. α-Amyrin did not protect HaCaT cells against the damage caused by UVB radiation.
Photodermatology, Photoimmunology & Photomedicine, 2017
Background: Photochemotherapy with psoralen and ultraviolet A (PUVA), with or without adjuvant in... more Background: Photochemotherapy with psoralen and ultraviolet A (PUVA), with or without adjuvant interferon-α (IFN-α), is a first line therapy for early stage mycosis fungoides and other forms of cutaneous T-cell lymphoma (CTCL). However, the mechanism by which PUVA with IFN-α work in CTCL is poorly understood. Purpose: To develop a model to investigate the mechanisms of PUVA and PUVA with IFN-α in CTCL cells. Methods: An in vitro model to study the molecular mechanisms of PUVA was created using two different CTCL cell lines, MyLa, which has functional p53, and HuT-78, in which p53 is inactivated due to a homozygous nonsense mutation. Results: PUVA caused G2/M cell cycle block and apoptosis of MyLa and HuT-78 accompanied by increase in the expression of the mitochondrial proapoptotic genes Bax, BAK, and PUMA and a downregulation in antiapoptotic Bcl-2. p53 was induced and c-Myc was repressed by PUVA, but neither were essential for PUVA-induced apoptosis. IFN-α augmented PUVA-induced apoptosis via the JAK1 pathway, and this activity could be inhibited by ruxolitinib. Conclusion: PUVA induces p53-independent apotosis in CTCL cell lines, and this process is augmented by type I interferons via the JAK1 pathway.
Please cite this article as: Biskup Edyta, Naym David Gram, Gniadecki Robert.Smallmolecule inhibi... more Please cite this article as: Biskup Edyta, Naym David Gram, Gniadecki Robert.Smallmolecule inhibitors of Ataxia Telangiectasia and Rad3 related kinase (ATR) sensitize lymphoma cells to UVA radiation.
The extensive involvement of miRNA in the pathophysiology of psoriasis is well-documented. Howeve... more The extensive involvement of miRNA in the pathophysiology of psoriasis is well-documented. However, in order for this information to be useful towards therapeutic manipulation of miRNA-levels, it is essential that detailed functional mechanisms are elucidated. MiR-125b has previously been shown to be strongly associated with psoriasis, and presents as an obvious candidate for further investigation OBJECTIVES: Elucidate the specific pathway and mechanism of interest in this association METHODS: A three-step bioinformatical hypothesis-generation pipeline was performed to identify genes of interest. This pipeline was based on miR-125b-binding, expression in psoriatic lesions and genome-wide association study-based evidence of involvement. The identified candidate gene was then carefully evaluated using luciferase binding assays, in-vitro overexpression, siRNA knock-down and downstream gene read-outs RESULTS: Based on our bioinformatical pipeline, the Ubiquitin Specific Peptidase 2 (USP...
The Notch pathway is important for survival of cutaneous T-cell lymphoma (CTCL) cells. We investi... more The Notch pathway is important for survival of cutaneous T-cell lymphoma (CTCL) cells. We investigated the effect of chemotherapy (doxorubicin, etoposide, and gemcitabine) and radiation modalities on Notch signaling in CTCL cell lines. Chemotherapy induced Notch1 expression at the mRNA and protein level in MyLa2000 and Hut78. Upregulation of well-established Notch targets supported the functional activity of Notch1. Transfection of Notch1 siRNA into MyLa2000 cells was not able to suppress the effects of chemotherapy on Notch1 activation significantly. Notch1 knockdown in combination with doxorubicin, etoposide, or gemcitabine compared to chemotherapy alone decreased cell viability by 12, 20, and 26%, respectively (p < 0.05). Additionally, X-rays (in MyLa2000 but not SeAx) and psoralen plus UVA (PUVA) (in MyLa2000, Hut78, and SeAx) increased the expression of Notch1 family members. Our results indicate that CTCL cells activate the Notch pathway in vitro in response to chemotherapy...
Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant... more Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant to conventional chemotherapy. USP2 has been shown to promote resistance to chemotherapeutic agents in several cancer models.We show here USP2 is expressed in quiescent and activated T-cells and its expression is 50% lower in CTCL cell lines (MyLa2000, SeAx and Hut-78) than in normal T-cells. USP2 is expressed in neoplastic cells in early, plaque-stage mycosis fungoides (MF) and is downregulated in advanced tumor stages. Upon treatment with psoralen with UVA (PUVA) or a p53 activator, nutlin3a, USP2 expression is significantly increased in MyLa2000 (p53wt/wt), but not in SeAx (p53mut) or Hut-78 (p53-/-). USP2 knockdown decreases MyLa2000 cell viability after PUVA by 50% but not Hut-78, suggesting that the function of USP2 in CTCL cells is p53-dependent. Furthermore, USP2 knockdown results in a decreased Mdm2 expression and upregulation of p53. Taken together, our findings suggest that US...
Basal cell carcinoma (BCC) is the most common malignant neoplasm with an annual incidence approac... more Basal cell carcinoma (BCC) is the most common malignant neoplasm with an annual incidence approaching 200/100,000 person-years. Mohs micrographic surgery (MMS) is widely used in North America and in Europe for treatment of BCC. This technique ensures radical tumour removal, sparing of the surrounding healthy skin, and it also offers higher cure rates than standard tumour excision with a predefined margin of healthy skin. The superiority of MMS relies on the fact that the entire (100%) margin of the excised tissue is examined microscopically for residual tumour in contrast to the traditional histopathological examination, in which 2% of the margin is examined. In Denmark, MMS was first introduced by us in 2012. In the present study, we retrospectively included all patients who underwent MMS from May 2012 to June 2015. A total of 231 patients with 263 BCC were included. The mean age was 66.1 years. The most common localisations were the forehead (31.3%), the nose (31.0%) and the cheek...
MicroRNAs (miRNAs) are small noncoding RNA molecules involved in gene regulation. Aberrant expres... more MicroRNAs (miRNAs) are small noncoding RNA molecules involved in gene regulation. Aberrant expression of miRNA has been associated with the development or progression of several diseases, including cancer. In a previous study, we found that the expression of miRNA-125b (miR-125b) was twofold lower in malignant melanoma producing lymph node micrometastases than in nonmetastasizing tumors. To get further insight into the functional role of miR-125b, we assessed whether its overexpression or silencing affects apoptosis, proliferation, or senescence in melanoma cell lines. We showed that overexpression of miR-125b induced typical senescent cell morphology, including increased cytoplasmatic/nucleus ratio and intensive cytoplasmatic b-galactosidase expression. In contrast, inhibition of miR-125b resulted in 30-35% decreased levels of spontaneous apoptosis. We propose that downregulation of miR-125b in an early cutaneous malignant melanoma can contribute to the increased metastatic capability of this tumor. Melanoma Res 21:253-256 c 2011 Wolters Kluwer Health | Lippincott Williams & Wilkins.
We have investigated cell growth dynamics and cyclins B1 and E expression in cell lines derived f... more We have investigated cell growth dynamics and cyclins B1 and E expression in cell lines derived from mycosis fungoides (MyLa), Sézary syndrome (SeAx), and CD30+ lympho-proliferative diseases (Mac1, Mac2a, JK). Mac1 and Mac2a had the highest growth rate (doubling time 18-28 h, >90% cycling cells) whereas SeAx was proliferating slowly (doub-ling time 55 h, approximately 35% cycling cells). Expression of cyclin B1 correlated positively with doubling time whereas expression of cyclin E was unscheduled and constant across the investigated cell lines. All cell lines exhibited high expression of PCNA. Thus, we concluded that cyclin B1 could be used for rapid screening of cell proliferation in malignant lymphocytes derived from cutaneous T-cell lymphoma.
The tumor suppressor p53 is often mutated in human cancers. Restoring its antitumor activity has ... more The tumor suppressor p53 is often mutated in human cancers. Restoring its antitumor activity has been shown to be a promising therapeutic approach for cancer treatment. Here we analyzed the activity and mechanism of a p53 reactivator, ellipticine, in a cellular model of cutaneous T-cell lymphoma (CTCL), a disease that is progressive, chemoresistant and refractory to treatment. We tested the effect of ellipticine in three cell lines with different p53 status: MyLa2000 (p53(wt/wt)), SeAx ((G245S)p53) and Hut-78 ((R196Stop)p53). Ellipticine caused apoptosis in MyLa2000 and SeAx and restored the transcriptional activity of (G245S)p53 in SeAx. However, p53 siRNA knockdown experiments revealed that p53 was not required for ellipticine-induced apoptosis in CTCL. The lipophilic antioxidant α-tocopherol inhibited ellipticine-dependent apoptosis and we linked the apoptotic response to the oxidative DNA damage. Our results provide evidence that ellipticine-induced apoptosis is exerted through DNA damage and does not require p53 activation in T-cell lymphoma.
Successful/effective cancer therapy in low grade lymphoma is often hampered by cell resistance to... more Successful/effective cancer therapy in low grade lymphoma is often hampered by cell resistance to anti-neoplastic agents. The crucial mechanisms responsible for this phenomenon are poorly understood. Overcoming resistance of tumor cells to anticancer agents, such as proteasome inhibitors, could improve their clinical efficacy. Using cutaneous T-cell lymphoma (CTCL) as a model of the chemotherapy-resistant peripheral lymphoid malignancy, we demonstrated that resistance to proteasome inhibition involved a signaling between the oncogene cMyc and miR-125b-5p. Bortezomib repressed cMyc and simultaneously induced miR-125b-5p that exerted a cytoprotective effect through the downmodulation of MAD4. Overexpression of cMyc repressed miR-125b-5p transcription and sensitized lymphoma cells to bortezomib. The central role of miR-125b-5p was further confirmed in a mouse model of T-cell lymphoma, where xenotransplantation of human CTCL cells overexpressing miR-125b-5p resulted in enhanced tumor growth and a shorter median survival. Our findings describe a novel mechanism through which miR-125b-5p not only regulates tumor growth in vivo, but also increases cellular resistance to proteasome inhibitors via modulation of MAD4.
ABSTRACT We report the antibacterial activity of the plant extracts and three previously isolates... more ABSTRACT We report the antibacterial activity of the plant extracts and three previously isolates known compounds viz., nonacosane-10-ol (alcohol), 23a-homostigmast-5-en-3ß-ol (homolog of β-sterol) and cis- and trans-prtopinium (alkaloid) of Fumaria parviflora Lam (Fumariaceae) for the first time. Plant extracts and pure compounds were in vitro assessed against seven clinical Gram (-) and Gram (+) bacteria viz. Staphylococcus aureus, Staphylococcus epidermis, Escherichia coli, Salmonella typhimurium, Klebsiella pneumonia, Bacillus subtilis, Salmonella typhi. Bacterial culture was maintained on Mueller-Hinton agar nutrient slants and the tested strains were evaluated following well diffusion test, agar dilution method and minimum inhibitory concentration. The zone of inhibition (IZ) and the activity index (AI) were maximum for Gram (-) Escherichia coli (IZ = 28 ± 0.9; AI = 0.93 ± 0.3), Klebsiella pneumonia (IZ = 22 ± 0.4; AI = 0.73 ± 0.4) and Salmonella typhi (IZ = 22 ± 0.4; AI = 0.73 ± 0.9) and these bacteria were strongly sensitive (SS) to the nhexane root extracts. The minimum inhibitory concentration value for the plant extracts was found to be in the range of 3.12 to 50.0 mg ml-1. The three compounds displayed strong antibacterial activity at a conc. of 100, 200 and 300 μg ml-1 against the tested strains. The zone of inhibition of these compounds ranged from minimum (9 ± 0.9, AI = 0.3 ± 0.5) for Salmonella typhimurium to maximum (46 ± 0.9, AI = 1.53 ± 0.3) for E. coli. The cis- and trans-protopinum was the most potent antibacterial compound against all the strains tested at the highest conc. of 300 μg ml-1. The three compounds were completely bactericidal as measured by the viable cell count studies. Fumaria parviflora derived extracts and the phytochemicals particularly the cis- and trans- prtopinium possess antibiotic properties and these compounds could be used in the development of novel chemotherapuric agents.
Plant Cell, Tissue and Organ Culture (PCTOC), 2009
Ruta graveolens L. is a source of pharmacologically active compounds such as coumarins, furanocou... more Ruta graveolens L. is a source of pharmacologically active compounds such as coumarins, furanocoumarins and furoquinolone alkaloids. Hypocotyls, callus and shoots of R. graveolens were inoculated with bacteria from two Agrobacterium rhizogenes strains. Hairy root cultures were established after inoculation of hypocotyls with wild A. rhizogenes strain LBA 9402. The transgenic nature of the regenerated tissue was confirmed by PCR amplification. Coumarins, furanocoumarins and alkaloids present in the hairy root tissue were identified by GC and GC-MS and compared with those present in in vitro shoot cultures. The level of pinnarin and rutacultin, bergapten, isopimpinelin and xanthotoxin was approximately twofold higher in hairy root than in shoot cultures. Two additional coumarins: osthole and osthenol, never been found in R. graveolens, were identified in hairy root tissue. Besides coumarins, alkaloids were identified: dictamnine, skimmianine, kokusaginine, rybalinine and an isomer of rybalinine. The levels of nearly all coumarins and alkaloids in hairy roots cultured in the darkness were higher than those accumulated under a photoperiod mode.
Background: Immortalized mammalian cell lines are a valuable research tool, though they represent... more Background: Immortalized mammalian cell lines are a valuable research tool, though they represent a highly simplified model. Due to accumulated mutations they may not reflect characteristics of the disease or even the tissue they derive from. Objective: We aim to pinpoint factors distinguishing SeAx cells from two other cutaneous T-cell lymphoma (CTCL) cell lines, namely Hut78 and MyLa2000. Of note, these factors may influence cell sensitivity in an unspecific way and therefore should be taken under consideration. Methods: We evaluated transcriptional levels of drug transporters across cell lines, cell membrane permeability, functionality of pathways related to DNA damage response and activation of G2/M block. Results: Analysis of the transcriptional levels of genes coding drug efflux pumps indicated that they are not consistently down-regulated in SeAx. However, we noted that SeAx cell membrane is markedly more permeable than Hut78 and MyLa2000, which may contribute to increased chemosensitivity in an unspecific way. Moreover, though DNA damage response seemed to be at least partly functional in SeAx cells, they fail to activate G2/M block in response to psoralen + UVA treatment. Any DNA damage should be repaired before cells enter mitosis, in order to uphold genome integrity. Thus, a defective cell cycle block may contribute to cell sensitivity. Conclusions: We believe that factors such as increased membrane permeability or defective cell cycle block should be accounted for when comparing sensitivity of cell line panels to chemotherapeutics of interest. It is worth to exclude a simple, indiscriminative mechanisms of cell resistance or sensitivity before attempting comparisons. Cell lines that are indiscriminately sensitive to a broad range of chemicals may contribute to overestimating the cytotoxic potential of tested compounds if used in cytotoxicity studies.
Rhaponticum carthamoides is an endemic medicinal plant of Siberian origin. Its roots tissue is we... more Rhaponticum carthamoides is an endemic medicinal plant of Siberian origin. Its roots tissue is well known as a rich source of biologically active compounds. R. carthamoides crude extracts were obtained with two different extraction methods (sonication and shaking) and three extraction solvents with different polarity (chloroform, methanol and water). All extract were screened for antioxidative activity (phosphomolybdenum assay, β-carotene bleaching test, DPPH and FRAP tests), cytotoxic (MTT assay) and mutagenic activities (modified Ames test, with the use Vibrio harveyi strains). All tested extracts were able to reduce molybdenum and inhibit about 85% of linoleic acid peroxidation. However, only water and methanol extracts reduced Fe+3 ions and scavenged DPPH free radical. On the other hand chloroform extracts indicated minor toxicity against all tested cell lines. None of the tested extracts showed mutagenic activity. Key words: Antioxidative activity, cytotoxic activity, mutagen...
Rhaponticum carthamoides is a medicinal plant indigenous to Siberia. Although its root tissue is ... more Rhaponticum carthamoides is a medicinal plant indigenous to Siberia. Although its root tissue is a well-known source of biologically active compounds, recent studies have focused on the above-ground part of the plant due to its easier accessibility. An R. carthamoides plantation was recently established in Northern Poland. Composition and antioxidant activity were compared for R. carthamoides leaf extracts obtained from plants grown in two plantations: in Poland (Lubiewice) and in Russia (Koriazma); the latter plantation has growing conditions similar to those in the R. carthamoides natural habitat. Plant secondary compounds were extracted with chloroform, methanol or water. As indicated by gas chromatography-mass spectrometry (GC-MS), the content of most compounds analyzed was higher in extracts from plants grown in Lubiewice (Poland). Antioxidative activity (as measured by diphenyl 1-picrylhydrazyl free radical (DPPH) and ferric reducing antioxidant power (FRAP) assays) and cytoto...
DNA-damaging cancer therapies induce interferon expression and stimulate the immune system, promo... more DNA-damaging cancer therapies induce interferon expression and stimulate the immune system, promoting therapy responses. The immune-activating STING (Stimulator of Interferon Genes) pathway is induced when DNA or double-stranded RNA (dsRNA) is detected in the cell cytoplasm, which can be caused by viral infection or by DNA damage following chemo- or radiotherapy. Here, we investigated the responses of cutaneous T-cell lymphoma (CTCL) cells to the clinically applied DNA crosslinking photochemotherapy (combination of 8–methoxypsoralen and UVA light; 8–MOP + UVA). We showed that this treatment evokes interferon expression and that the type III interferon IFNL1 is the major cytokine induced. IFNL1 upregulation is dependent on STING and on the cytoplasmic DNA sensor cyclic GMP-AMP synthase (cGAS). Furthermore, 8–MOP + UVA treatment induced the expression of genes in pathways involved in response to the tumor necrosis factor, innate immune system and acute inflammatory response. Notably, ...
Rhaponticum carthamoides plants ("maral root") are widely used in Siberian folk medicin... more Rhaponticum carthamoides plants ("maral root") are widely used in Siberian folk medicine. The present study reports for the first time the presence of pentacyclic terpenoid, α-amyrin, in methanol extract from leaves of this plant. α-Amyrin induced proliferation of human keratinocytes (HaCaT) by about 18% while other extract components were ineffective. A panel of biochemical and cell-based assays testing the antioxidative and cytoprotective activites of α-amyrin indicated no antioxidative activity of this compound. α-Amyrin did not protect HaCaT cells against the damage caused by UVB radiation.
Photodermatology, Photoimmunology & Photomedicine, 2017
Background: Photochemotherapy with psoralen and ultraviolet A (PUVA), with or without adjuvant in... more Background: Photochemotherapy with psoralen and ultraviolet A (PUVA), with or without adjuvant interferon-α (IFN-α), is a first line therapy for early stage mycosis fungoides and other forms of cutaneous T-cell lymphoma (CTCL). However, the mechanism by which PUVA with IFN-α work in CTCL is poorly understood. Purpose: To develop a model to investigate the mechanisms of PUVA and PUVA with IFN-α in CTCL cells. Methods: An in vitro model to study the molecular mechanisms of PUVA was created using two different CTCL cell lines, MyLa, which has functional p53, and HuT-78, in which p53 is inactivated due to a homozygous nonsense mutation. Results: PUVA caused G2/M cell cycle block and apoptosis of MyLa and HuT-78 accompanied by increase in the expression of the mitochondrial proapoptotic genes Bax, BAK, and PUMA and a downregulation in antiapoptotic Bcl-2. p53 was induced and c-Myc was repressed by PUVA, but neither were essential for PUVA-induced apoptosis. IFN-α augmented PUVA-induced apoptosis via the JAK1 pathway, and this activity could be inhibited by ruxolitinib. Conclusion: PUVA induces p53-independent apotosis in CTCL cell lines, and this process is augmented by type I interferons via the JAK1 pathway.
Please cite this article as: Biskup Edyta, Naym David Gram, Gniadecki Robert.Smallmolecule inhibi... more Please cite this article as: Biskup Edyta, Naym David Gram, Gniadecki Robert.Smallmolecule inhibitors of Ataxia Telangiectasia and Rad3 related kinase (ATR) sensitize lymphoma cells to UVA radiation.
The extensive involvement of miRNA in the pathophysiology of psoriasis is well-documented. Howeve... more The extensive involvement of miRNA in the pathophysiology of psoriasis is well-documented. However, in order for this information to be useful towards therapeutic manipulation of miRNA-levels, it is essential that detailed functional mechanisms are elucidated. MiR-125b has previously been shown to be strongly associated with psoriasis, and presents as an obvious candidate for further investigation OBJECTIVES: Elucidate the specific pathway and mechanism of interest in this association METHODS: A three-step bioinformatical hypothesis-generation pipeline was performed to identify genes of interest. This pipeline was based on miR-125b-binding, expression in psoriatic lesions and genome-wide association study-based evidence of involvement. The identified candidate gene was then carefully evaluated using luciferase binding assays, in-vitro overexpression, siRNA knock-down and downstream gene read-outs RESULTS: Based on our bioinformatical pipeline, the Ubiquitin Specific Peptidase 2 (USP...
The Notch pathway is important for survival of cutaneous T-cell lymphoma (CTCL) cells. We investi... more The Notch pathway is important for survival of cutaneous T-cell lymphoma (CTCL) cells. We investigated the effect of chemotherapy (doxorubicin, etoposide, and gemcitabine) and radiation modalities on Notch signaling in CTCL cell lines. Chemotherapy induced Notch1 expression at the mRNA and protein level in MyLa2000 and Hut78. Upregulation of well-established Notch targets supported the functional activity of Notch1. Transfection of Notch1 siRNA into MyLa2000 cells was not able to suppress the effects of chemotherapy on Notch1 activation significantly. Notch1 knockdown in combination with doxorubicin, etoposide, or gemcitabine compared to chemotherapy alone decreased cell viability by 12, 20, and 26%, respectively (p < 0.05). Additionally, X-rays (in MyLa2000 but not SeAx) and psoralen plus UVA (PUVA) (in MyLa2000, Hut78, and SeAx) increased the expression of Notch1 family members. Our results indicate that CTCL cells activate the Notch pathway in vitro in response to chemotherapy...
Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant... more Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant to conventional chemotherapy. USP2 has been shown to promote resistance to chemotherapeutic agents in several cancer models.We show here USP2 is expressed in quiescent and activated T-cells and its expression is 50% lower in CTCL cell lines (MyLa2000, SeAx and Hut-78) than in normal T-cells. USP2 is expressed in neoplastic cells in early, plaque-stage mycosis fungoides (MF) and is downregulated in advanced tumor stages. Upon treatment with psoralen with UVA (PUVA) or a p53 activator, nutlin3a, USP2 expression is significantly increased in MyLa2000 (p53wt/wt), but not in SeAx (p53mut) or Hut-78 (p53-/-). USP2 knockdown decreases MyLa2000 cell viability after PUVA by 50% but not Hut-78, suggesting that the function of USP2 in CTCL cells is p53-dependent. Furthermore, USP2 knockdown results in a decreased Mdm2 expression and upregulation of p53. Taken together, our findings suggest that US...
Basal cell carcinoma (BCC) is the most common malignant neoplasm with an annual incidence approac... more Basal cell carcinoma (BCC) is the most common malignant neoplasm with an annual incidence approaching 200/100,000 person-years. Mohs micrographic surgery (MMS) is widely used in North America and in Europe for treatment of BCC. This technique ensures radical tumour removal, sparing of the surrounding healthy skin, and it also offers higher cure rates than standard tumour excision with a predefined margin of healthy skin. The superiority of MMS relies on the fact that the entire (100%) margin of the excised tissue is examined microscopically for residual tumour in contrast to the traditional histopathological examination, in which 2% of the margin is examined. In Denmark, MMS was first introduced by us in 2012. In the present study, we retrospectively included all patients who underwent MMS from May 2012 to June 2015. A total of 231 patients with 263 BCC were included. The mean age was 66.1 years. The most common localisations were the forehead (31.3%), the nose (31.0%) and the cheek...
MicroRNAs (miRNAs) are small noncoding RNA molecules involved in gene regulation. Aberrant expres... more MicroRNAs (miRNAs) are small noncoding RNA molecules involved in gene regulation. Aberrant expression of miRNA has been associated with the development or progression of several diseases, including cancer. In a previous study, we found that the expression of miRNA-125b (miR-125b) was twofold lower in malignant melanoma producing lymph node micrometastases than in nonmetastasizing tumors. To get further insight into the functional role of miR-125b, we assessed whether its overexpression or silencing affects apoptosis, proliferation, or senescence in melanoma cell lines. We showed that overexpression of miR-125b induced typical senescent cell morphology, including increased cytoplasmatic/nucleus ratio and intensive cytoplasmatic b-galactosidase expression. In contrast, inhibition of miR-125b resulted in 30-35% decreased levels of spontaneous apoptosis. We propose that downregulation of miR-125b in an early cutaneous malignant melanoma can contribute to the increased metastatic capability of this tumor. Melanoma Res 21:253-256 c 2011 Wolters Kluwer Health | Lippincott Williams & Wilkins.
We have investigated cell growth dynamics and cyclins B1 and E expression in cell lines derived f... more We have investigated cell growth dynamics and cyclins B1 and E expression in cell lines derived from mycosis fungoides (MyLa), Sézary syndrome (SeAx), and CD30+ lympho-proliferative diseases (Mac1, Mac2a, JK). Mac1 and Mac2a had the highest growth rate (doubling time 18-28 h, >90% cycling cells) whereas SeAx was proliferating slowly (doub-ling time 55 h, approximately 35% cycling cells). Expression of cyclin B1 correlated positively with doubling time whereas expression of cyclin E was unscheduled and constant across the investigated cell lines. All cell lines exhibited high expression of PCNA. Thus, we concluded that cyclin B1 could be used for rapid screening of cell proliferation in malignant lymphocytes derived from cutaneous T-cell lymphoma.
The tumor suppressor p53 is often mutated in human cancers. Restoring its antitumor activity has ... more The tumor suppressor p53 is often mutated in human cancers. Restoring its antitumor activity has been shown to be a promising therapeutic approach for cancer treatment. Here we analyzed the activity and mechanism of a p53 reactivator, ellipticine, in a cellular model of cutaneous T-cell lymphoma (CTCL), a disease that is progressive, chemoresistant and refractory to treatment. We tested the effect of ellipticine in three cell lines with different p53 status: MyLa2000 (p53(wt/wt)), SeAx ((G245S)p53) and Hut-78 ((R196Stop)p53). Ellipticine caused apoptosis in MyLa2000 and SeAx and restored the transcriptional activity of (G245S)p53 in SeAx. However, p53 siRNA knockdown experiments revealed that p53 was not required for ellipticine-induced apoptosis in CTCL. The lipophilic antioxidant α-tocopherol inhibited ellipticine-dependent apoptosis and we linked the apoptotic response to the oxidative DNA damage. Our results provide evidence that ellipticine-induced apoptosis is exerted through DNA damage and does not require p53 activation in T-cell lymphoma.
Successful/effective cancer therapy in low grade lymphoma is often hampered by cell resistance to... more Successful/effective cancer therapy in low grade lymphoma is often hampered by cell resistance to anti-neoplastic agents. The crucial mechanisms responsible for this phenomenon are poorly understood. Overcoming resistance of tumor cells to anticancer agents, such as proteasome inhibitors, could improve their clinical efficacy. Using cutaneous T-cell lymphoma (CTCL) as a model of the chemotherapy-resistant peripheral lymphoid malignancy, we demonstrated that resistance to proteasome inhibition involved a signaling between the oncogene cMyc and miR-125b-5p. Bortezomib repressed cMyc and simultaneously induced miR-125b-5p that exerted a cytoprotective effect through the downmodulation of MAD4. Overexpression of cMyc repressed miR-125b-5p transcription and sensitized lymphoma cells to bortezomib. The central role of miR-125b-5p was further confirmed in a mouse model of T-cell lymphoma, where xenotransplantation of human CTCL cells overexpressing miR-125b-5p resulted in enhanced tumor growth and a shorter median survival. Our findings describe a novel mechanism through which miR-125b-5p not only regulates tumor growth in vivo, but also increases cellular resistance to proteasome inhibitors via modulation of MAD4.
ABSTRACT We report the antibacterial activity of the plant extracts and three previously isolates... more ABSTRACT We report the antibacterial activity of the plant extracts and three previously isolates known compounds viz., nonacosane-10-ol (alcohol), 23a-homostigmast-5-en-3ß-ol (homolog of β-sterol) and cis- and trans-prtopinium (alkaloid) of Fumaria parviflora Lam (Fumariaceae) for the first time. Plant extracts and pure compounds were in vitro assessed against seven clinical Gram (-) and Gram (+) bacteria viz. Staphylococcus aureus, Staphylococcus epidermis, Escherichia coli, Salmonella typhimurium, Klebsiella pneumonia, Bacillus subtilis, Salmonella typhi. Bacterial culture was maintained on Mueller-Hinton agar nutrient slants and the tested strains were evaluated following well diffusion test, agar dilution method and minimum inhibitory concentration. The zone of inhibition (IZ) and the activity index (AI) were maximum for Gram (-) Escherichia coli (IZ = 28 ± 0.9; AI = 0.93 ± 0.3), Klebsiella pneumonia (IZ = 22 ± 0.4; AI = 0.73 ± 0.4) and Salmonella typhi (IZ = 22 ± 0.4; AI = 0.73 ± 0.9) and these bacteria were strongly sensitive (SS) to the nhexane root extracts. The minimum inhibitory concentration value for the plant extracts was found to be in the range of 3.12 to 50.0 mg ml-1. The three compounds displayed strong antibacterial activity at a conc. of 100, 200 and 300 μg ml-1 against the tested strains. The zone of inhibition of these compounds ranged from minimum (9 ± 0.9, AI = 0.3 ± 0.5) for Salmonella typhimurium to maximum (46 ± 0.9, AI = 1.53 ± 0.3) for E. coli. The cis- and trans-protopinum was the most potent antibacterial compound against all the strains tested at the highest conc. of 300 μg ml-1. The three compounds were completely bactericidal as measured by the viable cell count studies. Fumaria parviflora derived extracts and the phytochemicals particularly the cis- and trans- prtopinium possess antibiotic properties and these compounds could be used in the development of novel chemotherapuric agents.
Plant Cell, Tissue and Organ Culture (PCTOC), 2009
Ruta graveolens L. is a source of pharmacologically active compounds such as coumarins, furanocou... more Ruta graveolens L. is a source of pharmacologically active compounds such as coumarins, furanocoumarins and furoquinolone alkaloids. Hypocotyls, callus and shoots of R. graveolens were inoculated with bacteria from two Agrobacterium rhizogenes strains. Hairy root cultures were established after inoculation of hypocotyls with wild A. rhizogenes strain LBA 9402. The transgenic nature of the regenerated tissue was confirmed by PCR amplification. Coumarins, furanocoumarins and alkaloids present in the hairy root tissue were identified by GC and GC-MS and compared with those present in in vitro shoot cultures. The level of pinnarin and rutacultin, bergapten, isopimpinelin and xanthotoxin was approximately twofold higher in hairy root than in shoot cultures. Two additional coumarins: osthole and osthenol, never been found in R. graveolens, were identified in hairy root tissue. Besides coumarins, alkaloids were identified: dictamnine, skimmianine, kokusaginine, rybalinine and an isomer of rybalinine. The levels of nearly all coumarins and alkaloids in hairy roots cultured in the darkness were higher than those accumulated under a photoperiod mode.
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Papers by Edyta Biskup