scholarly journals p53 Protein Isoform Profiles in AML: Correlation with Distinct Differentiation Stages and Response to Epigenetic Differentiation Therapy

Cells ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 833
Author(s):  
Ingvild Haaland ◽  
Sigrun M. Hjelle ◽  
Håkon Reikvam ◽  
André Sulen ◽  
Anita Ryningen ◽  
...  
Keyword(s):  
Cell Lines ◽  
P53 Protein ◽  
Global Gene Expression ◽  
Protein Isoforms ◽  
Chemotherapy Response ◽  
Differentiation Therapy ◽  
Potential Biomarker ◽  
Protein Isoform ◽  
Isoform Expression

p53 protein isoform expression has been found to correlate with prognosis and chemotherapy response in acute myeloid leukemia (AML). We aimed to investigate how p53 protein isoforms are modulated during epigenetic differentiation therapy in AML, and if p53 isoform expression could be a potential biomarker for predicting a response to this treatment. p53 full-length (FL), p53β and p53γ protein isoforms were analyzed by 1D and 2D gel immunoblots in AML cell lines, primary AML cells from untreated patients and AML cells from patients before and after treatment with valproic acid (VPA), all-trans retinoic acid (ATRA) and theophylline. Furthermore, global gene expression profiling analysis was performed on samples from the clinical protocol. Correlation analyses were performed between p53 protein isoform expression and in vitro VPA sensitivity and FAB (French–American–British) class in primary AML cells. The results show downregulation of p53β/γ and upregulation of p53FL in AML cell lines treated with VPA, and in some of the patients treated with differentiation therapy. p53FL positively correlated with in vitro VPA sensitivity and the FAB class of AML, while p53β/γ isoforms negatively correlated with the same. Our results indicate that p53 protein isoforms are modulated by and may predict sensitivity to differentiation therapy in AML.

scholarly journals Truncated WT1 Protein Isoform Expression Is Increased in MCF-7 Cells with Long-Term Estrogen Depletion

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Saavedra-Alonso Santiago ◽  
Zapata-Benavides Pablo ◽  
Mendoza-Gamboa Edgar ◽  
Chavez-Escamilla Ana Karina ◽  
Arellano-Rodríguez Mariela ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Protein Isoforms ◽  
Protein Isoform ◽  
Estrogen Depletion ◽  
Wt1 Protein ◽  
Cells Cultured ◽  
Mcf 7

Background. The wt1 gene codes for a transcription factor that presents several protein isoforms with diverse biological properties, capable of positively and negatively regulating genes involved in proliferation, differentiation, and apoptosis. WT1 protein is overexpressed in more than 90% of breast cancer; however, its role during tumor progression is still unknown. Methodology. In this work, we analyzed the expression of WT1 isoforms in several breast cancer cells with different tumor marker statuses and an in vitro assay using MCF-7 cells cultured with long-term estrogen depletion (MCF-7 LTED cells) with the finality to mimic the process of switching from hormone-dependent to hormone-independent. Moreover, growth kinetics, sensitivity to tamoxifen, and relative expression analysis of ER and Her2/neu were performed. Results. Initially, the expression of 52-54 kDa protein isoform of WT1 in the breast cancer cell line ER (+) was detected by western blot and was absent in ER (-), and the 36-38 kDa protein isoform of WT1 was detected in all cell lines analyzed. The analysis of alternative splicing by RT-PCR shows that the 17AA (+)/KTS (-) isoform of WT1 was the most frequent in the four cell lines analyzed. In vitro, the MCF-7 cells in the estrogen depletion assay show an increase in the expression of the 52-54 kDa isoform of WT1 in the first 48 hours, and this was maintained until week 13, and later, this expression was decreased, and the 36-38 kDa isoform of WT1 did not show change during the first 48 hours but from week 1 showed an increase of expression, and this remained until week 27. Growth kinetic analysis showed that MCF-7 LTED cells presented a 1.4-fold decrease in cellular proliferation compared to MCF-7 cells cultured under normal conditions. In addition, MCF-7 LTED cells showed a decrease in sensitivity to the antiproliferative effect of tamoxifen ( p ≤ 0.05 ). Samples collected until week 57 analyzed by qRT-PCR showed an increase in the relative expression of the Her2/neu and ER. Conclusions. Modulation of protein isoforms showed differential expression of WT1 isoforms dependent on estrogen receptor. The absence of 52-54 kDa and the presence of the 36-38 kDa protein isoform of WT1 were detected in ER-negative breast cancer cell lines classified as advanced stage cells. Long-term estrogen depletion assay in MCF-7 cells increased the expression of the 36-38 kDa isoform and reduced the 52-54 kDa isoform, and these cells show an increase in the expression of tumor markers of ER and Her2/neu. MCF-7 LTED cells showed low proliferation and insensitivity to tamoxifen compared to MCF-7 cells in normal conditions. These results support the theory about the relationship of the 36-38 kDa isoform of WT1 and the absence of ER function in advanced breast cancer.

scholarly journals Hsa_circ_0005273 facilitates breast cancer tumorigenesis by regulating YAP1-hippo signaling pathway

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Xuehui Wang ◽  
Changle Ji ◽  
Jiashu Hu ◽  
Xiaochong Deng ◽  
Wenfang Zheng ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Hippo Signaling ◽  
Hippo Signaling Pathway ◽  
Potential Biomarker

Abstract Background Circular RNAs (circRNAs), a novel class of endogenous RNAs, have shown to participate in the development of breast cancer (BC). Hsa_circ_0005273 is a circRNA generated from several exons of PTK2. However, the potential functional role of hsa_circ_0005273 in BC remains largely unknown. Here we aim to evaluate the role of hsa_circ_0005273 in BC. Methods The expression level of hsa_circ_0005273 and miR-200a-3p were examined by RT-qPCR in BC tissues and cell lines. The effect of knocking down hsa_circ_0005273 in BC cell lines were evaluated by examinations of cell proliferation, migration and cell cycle. In addition, xenografts experiment in nude mice were performed to evaluate the effect of hsa_circ_0005273 in BC. RNA immunoprecipitation assay, RNA probe pull-down assay, luciferase reporter assay and fluorescence in situ hybridization were conducted to confirm the relationship between hsa_circ_0005273, miR-200a-3p and YAP1. Results Hsa_circ_0005273 is over-expressed in BC tissues and cell lines, whereas miR-200a-3p expression is repressed. Depletion of hsa_circ_0005273 inhibited the progression of BC cells in vitro and in vivo, while overexpression of hsa_circ_0005273 exhibited the opposite effect. Importantly, hsa_circ_0005273 upregulated YAP1 expression and inactivated Hippo pathway via sponging miR-200a-3p to promote BC progression. Conclusions Hsa_circ_0005273 regulates the miR-200a-3p/YAP1 axis and inactivates Hippo signaling pathway to promote BC progression, which may become a potential biomarker and therapeutic target.

scholarly journals Alterations of p53 and c-myc in the clonal evolution of malignant lymphoma

Blood ◽  
1994 ◽  
Vol 83 (2) ◽  
pp. 452-459 ◽  
Author(s):  
H Chang ◽  
S Benchimol ◽  
MD Minden ◽  
HA Messner
Keyword(s):  
Growth Factors ◽  
T Cell ◽  
Cell Lines ◽  
P53 Protein ◽  
Clonal Evolution ◽  
Cdna Probe ◽  
Cell Phenotype ◽  
Coding Region ◽  
P53 Genes

Abstract We derived the lymphoma cell lines OCI-Ly 13.1 and OCI-Ly 13.2 from a patient with non-Hodgkin's lymphoma at the time of presentation and during chemotherapy-resistant relapse. These lines were of T-cell phenotype and contained the identical T-cell receptor beta-chain rearrangement, indicating that both lines were members of the same malignant clone. The lines differed in their growth characteristics; OCI-Ly 13.1 grew slowly and required growth factors for colony formation, whereas OCI-Ly 13.2 grew rapidly and formed colonies without addition of growth factors. To test whether or not these biologic differences were associated with specific genetic changes, we evaluated the status of the c-myc and p53 genes of both cell lines. The p53 and c- myc genes of OCI-Ly 13.1 were in germline configuration and produced normal-sized transcripts. The p53 protein expressed in OCI-Ly 13.1 was recognized by the anti-p53 monoclonal antibody, PAb240, indicating a conformation typical of p53 proteins expressed by p53 alleles containing a missense mutation. However, sequencing studies of the entire p53 coding region did not reveal any point mutations. In contrast, the cell line OCI-Ly 13.2 contained structural abnormalities of both the c-myc and p53 genes. In addition, one of the p53 alleles was lost as determined by a cDNA probe for the p53 gene (17p 13.1) and the YNZ22.1 probe (17p 13.3). These changes resulted in the absence of p53 protein and mRNA in OCI-Ly 13.2 as detected by immunoprecipitation and Northern blot analysis, respectively. They may be a reflection of disease progression and may be associated with the altered behavior of the malignant cell population within the patient and in vitro.

Flavopiridol Induces Apoptosis in Chronic Lymphocytic Leukemia Cells Via Activation of Caspase-3 Without Evidence of bcl-2 Modulation or Dependence on Functional p53

Blood ◽  
1998 ◽  
Vol 92 (10) ◽  
pp. 3804-3816 ◽  
Author(s):  
John C. Byrd ◽  
Charlotte Shinn ◽  
Jamie K. Waselenko ◽  
Ephraim J. Fuchs ◽  
Teresa A. Lehman ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Cell Lines ◽  
Caspase 3 ◽  
Mononuclear Cells ◽  
P53 Protein ◽  
Lymphocytic Leukemia ◽  
Vitro Activity ◽  
Interleukin 4 ◽  
In Vitro Activity

Abstract Flavopiridol has been reported to induce apoptosis in lymphoid cell lines via downregulation of bcl-2. The in vitro activity of flavopiridol against human chronic lymphocytic leukemia (CLL) cells and potential mechanisms of action for inducing cytotoxicity were studied. The in vitro viability of mononuclear cells from CLL patients (n = 11) was reduced by 50% at 4 hours, 24 hours, and 4 days at a flavopiridol concentration of 1.15 μmol/L (95% confidence interval [CI] ±0.31), 0.18 μmol/L (95% CI ±0.04), and 0.16 μmol/L (95% CI ±0.04), respectively. Loss of viability in human CLL cells correlated with early induction of apoptosis. Exposure of CLL cells to 0.18 μmol/L of flavopiridol resulted in both decreased expression of p53 protein and cleavage of the caspase-3 zymogen 32-kD protein with the appearance of its 20-kD subunit. Contrasting observations of others in tumor cell lines, flavopiridol cytotoxicity in CLL cells did not correlate with changes in bcl-2 protein expression alterations. We evaluated flavopiridol’s dependence on intact p53 by exposing splenocytes from wild-type (p53+/+) and p53 null (p53−/−) mice that demonstrated no preferential cytotoxicity as compared with a marked differential with F-ara-a and radiation. Incubation of CLL cells with antiapoptotic cytokine interleukin-4 (IL-4) did not alter the LC50 of flavopiridol, as compared with a marked elevation noted with F-ara-a in the majority of patients tested. These data demonstrate that flavopiridol has significant in vitro activity against human CLL cells through activation of caspase-3, which appears to occur independently of bcl-2 modulation, the presence of IL-4, or p53 status. Such findings strongly support the early introduction of flavopiridol into clinical trials for patients with B-CLL.

scholarly journals Article Commentary: Prospects for In Vitro Myofilament Maturation in Stem Cell-Derived Cardiac Myocytes

2015 ◽  
Vol 10s1 ◽  
pp. BMI.S23912 ◽  
Author(s):  
Jonas Schwan ◽  
Stuart G. Campbell
Keyword(s):  
Computational Models ◽  
Disease Modeling ◽  
Heart Tissue ◽  
Human Stem Cells ◽  
In Vitro Systems ◽  
Protein Isoform ◽  
Isoform Expression ◽  
Complex Relationships ◽  
Tissue Specimens

Cardiomyocytes derived from human stem cells are quickly becoming mainstays of cardiac regenerative medicine, in vitro disease modeling, and drug screening. Their suitability for such roles may seem obvious, but assessments of their contractile behavior suggest that they have not achieved a completely mature cardiac muscle phenotype. This could be explained in part by an incomplete transition from fetal to adult myofilament protein isoform expression. In this commentary, we review evidence that supports this hypothesis and discuss prospects for ultimately generating engineered heart tissue specimens that behave similarly to adult human myocardium. We suggest approaches to better characterize myofilament maturation level in these in vitro systems, and illustrate how new computational models could be used to better understand complex relationships between muscle contraction, myofilament protein isoform expression, and maturation.

scholarly journals The alternative splicing repressors hnRNP A1/A2 and PTB influence pyruvate kinase isoform expression and cell metabolism

2010 ◽  
Vol 107 (5) ◽  
pp. 1894-1899 ◽  
Author(s):  
Cynthia V. Clower ◽  
Deblina Chatterjee ◽  
Zhenxun Wang ◽  
Lewis C. Cantley ◽  
Matthew G. Vander Heiden ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Pyruvate Kinase ◽  
Cell Lines ◽  
Cancer Cell ◽  
Aerobic Glycolysis ◽  
Lactate Production ◽  
Cancer Cell Lines ◽  
Protein Isoforms ◽  
Hnrnp A1 ◽  
Isoform Expression

Cancer cells preferentially metabolize glucose by aerobic glycolysis, characterized by increased lactate production. This distinctive metabolism involves expression of the embryonic M2 isozyme of pyruvate kinase, in contrast to the M1 isozyme normally expressed in differentiated cells, and it confers a proliferative advantage to tumor cells. The M1 and M2 pyruvate-kinase isozymes are expressed from a single gene through alternative splicing of a pair of mutually exclusive exons. We measured the expression of M1 and M2 mRNA and protein isoforms in mouse tissues, tumor cell lines, and during terminal differentiation of muscle cells, and show that alternative splicing regulation is sufficient to account for the levels of expressed protein isoforms. We further show that the M1-specific exon is actively repressed in cancer-cell lines—although some M1 mRNA is expressed in cell lines derived from brain tumors—and demonstrate that the related splicing repressors hnRNP A1 and A2, as well as the polypyrimidine-tract-binding protein PTB, contribute to this control. Downregulation of these splicing repressors in cancer-cell lines using shRNAs rescues M1 isoform expression and decreases the extent of lactate production. These findings extend the links between alternative splicing and cancer, and begin to define some of the factors responsible for the switch to aerobic glycolysis.

scholarly journals LncRNA STK4 antisense RNA 1(STK4-AS1) affects the osteosarcoma cell cycle by regulating p53 protein

2021 ◽  
Author(s):  
Weitao Yao ◽  
Jingyu Hou ◽  
Guoqing Liu ◽  
Fangxing Wu ◽  
Qiang Yan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Viability ◽  
Cell Lines ◽  
P53 Protein ◽  
Mg63 Cell ◽  
Cyclin A ◽  
Osteosarcoma Cell ◽  
Osteosarcoma Cells ◽  
P53 Overexpression ◽  
Potential Biomarker

Abstract Background: LncRNA STK4-AS1 has been identified as a potential biomarker associated with multiple cancers. We proposed that STK4-AS1 plays a role in the proliferation of osteosarcoma by regulating its cell cycle. Methods: We compared the expression of STK4-AS1 in osteosarcoma vs normal samples both in clinical tissues and cell lines. We overexpressed and knockdown STK4-AS1 in p53 expressing osteosarcoma cells U2OS and p53 muted expressing osteosarcoma cells MG63 and analyzed cell viability and cell cycle. We overexpressed p53 in STK4-AS1 knockdown cells to explore the association of STK4-AS1 and p53 in the cell cycle. Results: The STK4-AS1 expression was higher in osteosarcoma tissue than adjacent normal bone tissues and was higher in osteosarcoma cell lines (U2OS, MG63, and SAOS-2) than in osteoblast cell lines (hFOB and HOB). Knockdown of STK4-AS1 in U2OS decreased the cell viability, increased cells in the G0/G1 phase, decreased cells in the S and G2/M phase, decreased expression of cyclin A and B, increased p53 and p21, and had no effect on cyclin D and cyclin E, while overexpression did the opposes. MG63 cell viability was not affected by altered STK4-AS1 levels. P53 overexpression in STK4-AS1 knockdown cells recovered cell viability, p21, cyclin A, and cyclin B expression. Conclusion: LncRNA STK4-AS1 affected p53 expressing osteosarcoma cells U2OS cell viability through regulating cell cycle, which is mediated by p53/p21 pathway.

Flavopiridol Induces Apoptosis in Chronic Lymphocytic Leukemia Cells Via Activation of Caspase-3 Without Evidence of bcl-2 Modulation or Dependence on Functional p53

Blood ◽  
1998 ◽  
Vol 92 (10) ◽  
pp. 3804-3816 ◽  
Author(s):  
John C. Byrd ◽  
Charlotte Shinn ◽  
Jamie K. Waselenko ◽  
Ephraim J. Fuchs ◽  
Teresa A. Lehman ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Cell Lines ◽  
Caspase 3 ◽  
Mononuclear Cells ◽  
P53 Protein ◽  
Lymphocytic Leukemia ◽  
Vitro Activity ◽  
Interleukin 4 ◽  
In Vitro Activity

Flavopiridol has been reported to induce apoptosis in lymphoid cell lines via downregulation of bcl-2. The in vitro activity of flavopiridol against human chronic lymphocytic leukemia (CLL) cells and potential mechanisms of action for inducing cytotoxicity were studied. The in vitro viability of mononuclear cells from CLL patients (n = 11) was reduced by 50% at 4 hours, 24 hours, and 4 days at a flavopiridol concentration of 1.15 μmol/L (95% confidence interval [CI] ±0.31), 0.18 μmol/L (95% CI ±0.04), and 0.16 μmol/L (95% CI ±0.04), respectively. Loss of viability in human CLL cells correlated with early induction of apoptosis. Exposure of CLL cells to 0.18 μmol/L of flavopiridol resulted in both decreased expression of p53 protein and cleavage of the caspase-3 zymogen 32-kD protein with the appearance of its 20-kD subunit. Contrasting observations of others in tumor cell lines, flavopiridol cytotoxicity in CLL cells did not correlate with changes in bcl-2 protein expression alterations. We evaluated flavopiridol’s dependence on intact p53 by exposing splenocytes from wild-type (p53+/+) and p53 null (p53−/−) mice that demonstrated no preferential cytotoxicity as compared with a marked differential with F-ara-a and radiation. Incubation of CLL cells with antiapoptotic cytokine interleukin-4 (IL-4) did not alter the LC50 of flavopiridol, as compared with a marked elevation noted with F-ara-a in the majority of patients tested. These data demonstrate that flavopiridol has significant in vitro activity against human CLL cells through activation of caspase-3, which appears to occur independently of bcl-2 modulation, the presence of IL-4, or p53 status. Such findings strongly support the early introduction of flavopiridol into clinical trials for patients with B-CLL.

scholarly journals HLA-G and HLA-F protein isoform expression in breast cancer patients receiving neoadjuvant treatment

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Franziska M. Wuerfel ◽  
Hanna Huebner ◽  
Lothar Häberle ◽  
Paul Gass ◽  
Alexander Hein ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Protein Expression ◽  
Expression Patterns ◽  
Neoadjuvant Treatment ◽  
Protein Isoforms ◽  
F Protein ◽  
Her2 Breast Cancer ◽  
Soluble Hla ◽  
Protein Isoform ◽  
Isoform Expression

Abstract The immunosuppressive human leukocyte antigens HLA-G and HLA-F are expressed on trophoblast and malignant cells. Four membrane-bound and three soluble HLA-G protein isoforms have been described, which have different immunosuppressive potentials. HLA-F has three transcript variants, resulting in three different protein isoforms. The aim of this study was to evaluate the prognostic and predictive value of HLA-G and HLA-F protein isoform expression patterns in patients with breast cancer. Core biopsies were taken at diagnosis in patients with HER2+ (n = 28), luminal B-like (n = 49) and triple-negative (n = 38) breast cancers who received neoadjuvant chemotherapy. Expression levels of HLA-F and -G were correlated with the pathological complete response (pCR). Protein expression was determined by Western blot analysis, using two antibodies for each HLA, specific for different isoforms. The protein expression of HLA isoforms did not significantly differ between breast cancer subtypes. However, some initial indications were found for an association between the soluble HLA-G6 protein isoform and pCR in HER2+ breast cancer. The study provides preliminary evidence for the evaluation of HLA-G isoform expression, in particular HLA-G6, as a possible new marker for pCR in HER2+ breast cancer.

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