The plasticity of pancreatic cancer stem cells: implications in therapeutic resistance

The ever-growing perception of cancer stem cells (CSCs) as a plastic state rather than a hardwired defined entity has evolved our understanding of the functional and biological plasticity of these elusive components in malignancies. Pancreatic cancer (PC), based on its biological features and clinical evolution, is a prototypical example of a CSC-driven disease. Since the discovery of pancreatic CSCs (PCSCs) in 2007, evidence has unraveled their control over many facets of the natural history of PC, including primary tumor growth, metastatic progression, disease recurrence, and acquired drug resistance. Consequently, the current near-ubiquitous treatment regimens for PC using aggressive cytotoxic agents, aimed at ‘‘tumor debulking’’ rather than eradication of CSCs, have proven ineffective in providing clinically convincing improvements in patients with this dreadful disease. Herein, we review the key hallmarks as well as the intrinsic and extrinsic resistance mechanisms of CSCs that mediate treatment failure in PC and enlist the potential CSC-targeting ‘natural agents’ that are gaining popularity in recent years. A better understanding of the molecular and functional landscape of PCSC-intrinsic evasion of chemotherapeutic drugs offers a facile opportunity for treating PC, an intractable cancer with a grim prognosis and in dire need of effective therapeutic advances.Other Information Published in: Cancer and Metastasis Reviews License: https://creativecommons.org/licenses/by/4.0See article on publisher's website: http://dx.doi.org/10.1007/s10555-021-09979-x</p

Cisplatin based therapy: the role of the mitogen activated protein kinase signaling pathway

Cisplatin is a widely used chemotherapeutic agent for treatment of various cancers. However, treatment with cisplatin is associated with drug resistance and several adverse side effects such as nephrotoxicity, reduced immunity towards infections and hearing loss. A Combination of cisplatin with other drugs is an approach to overcome drug resistance and reduce toxicity. The combination therapy also results in increased sensitivity of cisplatin towards cancer cells. The mitogen activated protein kinase (MAPK) pathway in the cell, consisting of extracellular signal regulated kinase, c-Jun N-terminal kinase, p38 kinases, and downstream mediator p90 ribosomal s6 kinase (RSK); is responsible for the regulation of various cellular events including cell survival, cell proliferation, cell cycle progression, cell migration and protein translation. This review article demonstrates the role of MAPK pathway in cisplatin based therapy, illustrates different combination therapy involving cisplatin and also shows the importance of targeting MAPK family, particularly RSK, to achieve increased anticancer effect and overcome drug resistance when combined with cisplatin.Other Information Published in: Journal of Translational Medicine License: http://creativecommons.org/licenses/by/4.0/See article on publisher's website: http://dx.doi.org/10.1186/s12967-018-1471-1</p

Unleashing the immune response to NY-ESO-1 cancer testis antigen as a potential target for cancer immunotherapy

Introduction Cancer Immunotherapy has recently emerged as a promising and effective modality to treat different malignancies. Antigenic profiling of cancer tissues and determination of any pre-existing immune responses to cancer antigens may help predict responses to immune intervention in cancer. NY-ESO-1, a cancer testis antigen is the most immunogenic antigen to date. The promise of NY-ESO-1 as a candidate for specific immune recognition of cancer comes from its restricted expression in normal adult tissue but frequent occurrence in multiple tumors including melanoma and carcinomas of lung, esophageal, liver, gastric, prostrate, ovarian, and bladder. Main body This review summarizes current knowledge of NY-ESO-1 as efficient biomarker and target of immunotherapy. It also addresses limitations and challenges preventing a robust immune response to NY-ESO-1 expressing cancers, and describes pre-clinical and clinical observations relevant to NY-ESO-1 immunity, holding potential therapeutic relevance for cancer treatment. Conclusion NY-ESO-1 induces strong immune responses in cancer patients but has limited objective clinical responses to NY-ESO-1 expressing tumors due to effect of competitive negative signaling from immune-checkpoints and immune-suppressive tumor microenvironment. We propose that combination therapy to increase the efficacy of NY-ESO-1 specific immunotherapeutic interventions should be explored to unleash the immune response against NY-ESO-1 expressing tumors.Other Information Published in: Journal of Translational Medicine License: http://creativecommons.org/licenses/by/4.0/See article on publisher's website: http://dx.doi.org/10.1186/s12967-020-02306-y</p

Tissue microarray based immunohistochemical analysis of p-IκB expression in DLBCL patients: DLBCL array spot showing high expression of p-IκB (A) and low expression of p-IκB (B).

<p>20×/0.70 objective on an Olympus BX 51 microscope. (Olympus America Inc, Center Valley, PA, USA. with the inset showing a 40×0.85 aperture magnified view of the same. (C). Impact of p-IκB expression on prognosis in ABC patients. In patients with activated B cell phenotype(ABC) subgroup, p-IκB over expression showed a poor overall survival of 56.3% at 5 years as compared to 78.2% with low p-IκB expression (p = 0.0724).</p

Correlation of p-IκB expression with clinico-pathological parameters in ABC subtypes of DLBCL samples.

<p>^@IPI & Stage information was available only in 89 patients. ?GC vs. ABC Germinal Centre (GC) versus Activated B Cell (ABC) phenotype.^$Of the 201 Diffuse Large B cell lymphomas, p-IκB results were available in 151 cases and the remaining 50 spots were non informative. Analysis failure for these IHC markers was attributed to missing or non representative spots. Of these 151 TMA spots with available p-IκB data, range of non available spots for rest of the IHC markers ranged from 1 spot for XIAP to 18 spots for XIAP. The remaining cases were considered for correlation analysis.</p

Thymoquinone-induced generation of ROS in ABC cells.

<p>(<b>A</b>) <b>Thymoquinone increases ROS generation in ABC cells.</b> HBL-1 and RIVA cells were incubated in the absence or presence of 10 µM TQ for indicated time periods. After washing with PBS, cells were incubated with 10 µM H2DCFDA and incubated in the dark for 30 minutes at 37°C as described in material and method. Cells were washed in PBS, re-suspended in PBS and analyzed by flow cytometry. (<b>B</b>) <b>NAC prevents TQ-induced ROS release in ABC cells.</b> HBL-1 and RIVA cell lines were pre-treated with 10 mM NAC for 21hours followed by treatment with 10 µM TQ for various time periods. Cells were washed in PBS and incubated with 10 µM H2DCFDA and incubated in the dark for 30 minutes at 37°C. Cells were washed in PBS, re-suspended in PBS and analyzed by flow cytometry. Bar graph displays the mean +/− SD (standard deviation) of three independent experiments, * p<0.05, statistically significant (Students <i>t</i>-test). (<b>C</b>) (<b>C</b>) <b>NFκB activity is down-regulated by TQ as measured by Luciferase assay.</b> RIVA cells were transiently transfected with p65-LUC plasmid for 48 hours. After transfection, RIVA cells were divided into three groups; one group was left untreated as control, the second group was treated with 10 µM TQ for 24 hours and third group was pretreated with 10 mM NAC for 3 hours followed by treatment with 10 µM TQ for 24 hours. After 24 hours, cells were lysed in luciferase lysis buffer and luciferase activity was measured. pCDNA plasmid was used as transfection control and β-galactosidase was used to normalize the transfection efficiency. <b>(D)</b><b>NAC prevents TQ-induced down-regulation of targets of p65.</b> HBL-1 and RIVA cells were pre-treated with 10 mM NAC for three hours followed by treatment with 10 µM TQ for 24 hours. Cells were lysed and proteins were immunoblotted with antibodies against IκBα, Bcl-2 and Survivin.</p

TQ-induced up-regulation of DR5 does not play a part in TQ-induced apoptosis.

<p>(A) TQ treatment causes up-regulation of DR5 in ABC cells. HBL1 and RIVA cells were treated with 10 µM TQ for indicated time periods. After cell lysis, equal amounts of proteins were immuno-blotted with antibodies against DR5 and beta actin. (B) TQ-induced DR5 up-regulation is ROS dependent. HBL-1 cells were pre-treated with either 10 mM NAC or 80 µM z-VAD for 3 hours and subsequently treated with 10 µM TQ for 24 hours. Cells were lysed and equal amounts of proteins were immunoblotted with antibodies against DR5 and beta-actin. (C and D) TQ-induced apoptosis is not DR5 dependent. (C) HBL-1 and RIVA cells were either transfected with 50 and 100 nM siRNA, specific against DR5 or scrambled siRNA for 48 hours. Cells were then treated with 5 and 10 µM TQ for 24 hours, following which cells were stained with fluorescent-conjugated Annexin V/PI and analyzed by flow cytometry. Bar graph displays the mean +/− SD (standard deviation) of three independent experiments, * p<0.05, statistically significant (Students <i>t</i>-test). (D) HBL-1 and RIVA cells were either transfected with 50 and 100 nM siRNA, specific against DR5 or scrambled siRNA for 48 hours and treated with 5 and 10 µM TQ for 24 hours. Cells were lysed and equal amounts of proteins were immuno-blotted with antibodies against DR5, caspase-8, caspase-3, PARP and beta-actin. (E): Combination treatment with sub-optimal doses of TQ and TRAIL induce synergistic inhibition of cell viability and apoptosis in ABC cells. HBL-1 and RIVA cells were treated with either 1 µM TQ in the presence and absence of 1 ng TRAIL for 24 hours. Following treatment, cells were analyzed for cell viability by MTT assay as described in material and methods. Bar graph displays the mean +/− SD (standard deviation) of three independent experiments, * p<0.05, statistically significant (Students <i>t</i>-test). (F) HBL-1 and RIVA cells were treated with either 1 µM TQ in the presence and absence of 1 ng TRAIL for 24 hours. Following treatment, cells were stained with fluorescent-conjugated Annexin V/PI and analyzed by flow cytometry (G) HBL-1 cells were treated with either 1 µM TQ in the presence and absence of 1 ng TRAIL for 24 hours. Following treatment, cells were lysed and equal amounts of proteins were immuno-blotted with antibodies against caspase-8, caspase-3, PARP, and beta-actin.</p

TQ-induced caspases dependent apoptosis in ABC cells.

<p>(A) TQ treatment causes activation and cleavage of caspases in ABC cells. HBL-1 and RIVA cells were treated with and without 5 and 10 µM TQ for 24 hours. Cells were lysed and equal amounts of proteins were immunoblotted with antibodies against caspase-9, caspase-3, PARP and Beta-actin. (B) TQ-induced caspases activation and cleavage is blocked by NAC and caspases inhibitor. HBL-1 cells were pretreated with either 10 mM NAC or 80 µM z-VAD for 3 hours and subsequently treated with 10 µM TQ for 24 hours. Cells were lysed and equal amounts of proteins were immunoblotted with antibodies against caspase-9, caspase-3 cleaved caspase-3, PARP and beta-actin. (C) TQ suppresses growth of ABC cells. ABC cell lines were incubated with 0–50 µM TQ for 24 hours. Cell viability was measured by MTT assays as described in Materials and Methods. The graph displays the mean +/− SD (standard deviation) of three independent experiments, * p<0.05, statistically significant (Students <i>t</i>-test). (D) TQ treatment induces apoptosis in ABC cell lines. ABC cells were treated with 5 and 10 µM TQ (as indicated) for 24 hours and cells were subsequently stained with flourescein-conjugated annexin-V and propidium iodide (PI) and analyzed by flow cytometry.</p

TQ-induced mitochondrial signaling pathway in ABC cells.

<p>(A) TQ treatment causes Bax conformational changes in ABC cells. After treating with 10 µM TQ for indicated time periods, HBL-1 cells were lysed and immuno-precipitated with anti-Bax 6A7 antibody for detection of conformationally changed Bax protein. In addition, the total cell lysates were immuno-blotted with specific anti-Bax polyclonal antibody. (B) NAC prevents TQ-induced Bax conformational changes in ABC cells. HBL-1 cells were pre-treated with either, 10 mM NAC and 80 µM z-VAD/fmk for 3 hours and subsequently treated with 10 µM TQ for 8 hours. Cells were lysed and immunoprecipitated with anti-Bax 6A7 antibody and proteins were immunoblotted with Bax rabbit polyclonal antibody. (C) TQ treatment causes change in mitochondrial membrane potential in ABC cells. ABC cells were treated with and without 5 and 10 µM TQ for 24 hours. Live cells with intact mitochondrial membrane potential and dead cells with lost mitochondrial membrane potential was measured by JC-1 staining and analyzed by flow cytometry as described in Materials and Methods. Bar graph displays the mean +/− SD (standard deviation) of three independent experiments, * p<0.05, statistically significant (Students <i>t</i>-test). (D) TQ treatment causes release of cytochrome c from mitochondria into cytosole. HBL-1 and RIVA cells were treated with 5 and 10 µM TQ for 24 hours. Mitochondrial free cytosolic fractions were isolated and immunoblotted with antibody against cytochrome c and Beta-actin.</p

The cross-talk between miRNAs and JAK/STAT pathway in cutaneous T cell lymphoma: Emphasis on therapeutic opportunities

Mycosis Fungoides (MF) and Sézary Syndrome (SS) belong to a wide spectrum of T cell lymphoproliferative disorders collectively termed cutaneous T cell lymphomas (CTCL). CTCLs represent an archetype of heterogeneous and dynamically variable lymphoproliferative neoplasms typified by distinct clinical, histological, immunophenotypic, and genetic features. Owing to its complex dynamics, the pathogenesis of CTCL remains elusive. However, in recent years, progress in CTCL classification combined with next-generation sequencing analyses has broadened the genetic and epigenetic spectrum of clearly defined CTCL entities such as MF and SS. Several large-scale genome studies have identified the polygenic nature of CTCL and unveiled an idiosyncratic mutational landscape involving genetic aberrations, epigenetic alterations, cell cycle dysregulation, apoptosis, and the constitutive activation of T cell/NF-κB/JAK-STAT signaling pathways. In this review, we summarize the evolving insights on how the intrinsic epigenetic events driven by dysregulated miRNAs, including the oncogenic and tumor-suppressive miRNAs, influence the pathogenesis of MF and SS. We also focus on the interplay between the JAK/STAT pathway and miRNAs in CTCL as well as the significance of the miRNA/STAT axis as a relevant pathogenetic mechanism underlying CTCL initiation and progression. Based on these biologic insights, the current status and recent progress on novel therapies with a strong biological rationale, including miRNA-targeted molecules and JAK/STAT-targeted therapy for CTCL management, are discussed.Other InformationPublished in: Seminars in Cell & Developmental BiologyLicense: http://creativecommons.org/licenses/by/4.0/See article on publisher's website: https://dx.doi.org/10.1016/j.semcdb.2022.09.015</p