Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: (A) Schematic representation of pooled shRNA screen performed. SET2 cells were infected with a shRNA library targeting 5,000 genes, and selected with puromycin for 3 days before seeding in 10-cm-plates at a density of 30% for treatment with DMSO control or with JAK2 inhibitor SAR302503 at 0.2 and 0.35 μM. Cells were harvested at 0, 7 and 14 days post drug treatment and cellular genomic DNA was subjected to amplification followed by deep sequencing to determine shRNA copy number. (B) The phenotype of combinational effects was revealed by comparing the quantity of each shRNA in the sample treated with JAK2 inhibitor versus that in the sample treated with DMSO. X-axis: samples treated with 200 nM SAR302503 vs. DMSO; Y-axis: samples treated with 350nM SAR302503 vs. DMSO. 1.5-fold decrease in shRNA abundance is marked with dotted lines. Four MYC shRNAs are marked.

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques: shRNA, Infection, Control, Amplification, Sequencing

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: SET2 (A) and UKE1 cells (B) were treated with three PIM inhibitors for 4 hours and then analyzed by immunoblot using indicated antibodies. (C) UKE1 cells were stably infected with doxycycline-inducible MYC expression vector and MYC protein induction was confirmed by immunoblot analysis after 24 and 72 hour incubation. (D) MYC-UKE1 cells were cultured with or without doxycycline and treated with DMSO or serial dilutions of PIM inhibitor (compound-C). Cell viability measurements were taken 72 hours later.

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques: Western Blot, Stable Transfection, Infection, Expressing, Plasmid Preparation, Incubation, Cell Culture

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: SET2 (A) and UKE1 (B) cells were treated with the combination of JAK2 inhibitor SAR302503 and three PIM inhibitors. Cell viability was assessed 72 hours later. Isobologram analysis was performed as described in Materials and Methods. Combination index (CI) was calculated as the sum of IC50combo/IC50JAK2 and IC50combo/IC50PIM according to Chou-Talay's method. The lowest combination index is shown. Results represent triplicate experiments.

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques:

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: SET2 (A) and UKE1 (B) cells were exposed to the constant amount of JAK2 inhibitor (0.3 μM for SET2 and 1 μM for UKE1), while PIM compounds were titrated in a dose response format. Cell viability was measured using CellTiter-Glo® kit before (CTGbefore) and after 3 days of drug treatment (CTGafter). Effects of drug treatments were calculated as a ratio of (CTGafter-CTGbefore)/ CTGbefore. CTGafter=CTGbefore indicates cell stasis; CTGafter

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques:

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: SET2 (A) and UKE1 (B) cells were exposed to the constant amount of JAK2 inhibitor (0.3 μM for SET2 and 1 μM for UKE1), while PIM compounds were titrated in a dose response format. Apoptosis induction was determined after 3 days of treatment by using Homogeneous Caspase Assay kit as described in Materials and Methods. Results of triplicate experiments are shown.

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques: Caspase Assay

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: SET2 (A) and UKE1 (B) cells were treated with JAK2 and PIM inhibitors for 4 hours, either alone or in combination, at indicated doses. Immunoblot was used to analyze total and phosphorylated forms of 4EBP1, S6 and P70S6K in treated cells.

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques: Western Blot

Journal: Oncotarget

Article Title: Combination of PIM and JAK2 inhibitors synergistically suppresses cell proliferation and overcomes drug resistance of myeloproliferative neoplasms

doi:

Figure Lengend Snippet: (A) SET2 cells were exposed to 2 µM of JAK2 inhibitor for 8 weeks to generate resistant clones. Cell viability of four resulting clones, as well as parental cells, was subsequently evaluated in the presence of a dose response of JAK2 compound. (B) Resistant clones were analyzed by immunoblot using indicated antibodies to detect changes in critical survival nodes. (C) Resistant cells were treated with 3 µM of either JAK2 or PIM inhibitor (compound C) alone or in combination for 3 days. Cell viability was measured using CellTiter-Glo® kit before (CTGbefore) and after 3 days of drug treatment (CTGafter). Effects of drug treatments were calculated as a ratio of (CTGafter-CTGbefore)/ CTGbefore. CTGafter=CTGbefore indicates cell stasis; CTGafter

Article Snippet: SET2 cell line was purchased from DSMZ (Braunschweig, GERMANY).

Techniques: Clone Assay, Western Blot