Treatment with decitabine induces the expression of stemness markers, PD-L1 and NY-ESO-1 in colorectal cancer: potential for combined chemoimmunotherapy.

BACKGROUND: The mechanism of tumor immune escape and progression in colorectal cancer (CRC) is widely investigated in-vitro to help understand and identify agents that might play a crucial role in response to treatment and improve the overall survival of CRC patients. Several mechanisms of immune escape and tumor progression, including expression of stemness markers, inactivation of immunoregulatory genes by methylation, and epigenetic silencing, have been reported in CRC, indicating the potential of demethylating agents as anti-cancer drugs. Of these, a chemotherapeutic demethylating agent, Decitabine (DAC), has been reported to induce a dual effect on both DNA demethylation and histone changes leading to an increased expression of target biomarkers, thus making it an attractive anti-tumorigenic drug. METHODS: We compared the effect of DAC in primary 1076 Col and metastatic 1872 Col cell lines isolated and generated from patients' tumor tissues. Both cell lines were treated with DAC, and the expression of the NY-ESO-1 cancer-testis antigen, the PD-L1 immunoinhibitory marker, and the CD44, Nanog, KLF-4, CD133, MSI-1 stemness markers were analyzed using different molecular and immunological assays. RESULTS: DAC treatment significantly upregulated stemness markers in both primary 1076 Col and meta-static 1872 Col cell lines, although a lower effect occurred on the latter: CD44 (7.85 fold; ***p = 0.0001 vs. (4.19 fold; *p = 0.0120), Nanog (4.1 fold; ***p < 0.0001 vs.1.69 fold; ***p = 0.0008), KLF-4 (4.33 fold; ***p < 0.0001 vs.2.48 fold; ***p = 0.0005), CD133 (16.77 fold; ***p = 0.0003 vs.6.36 fold; *p = 0.0166), and MSI-1 (2.33 fold; ***p = 0.0003 vs.2.3 fold; ***p = 0.0004), respectively. Interestingly, in the metastatic 1872 Col cells treated with DAC, the expression of both PD-L1 and NY-ESO-1 was increased tenfold (*p = 0.0128) and fivefold (***p < 0.0001), respectively. CONCLUSIONS: We conclude that the upregulation of both stemness and immune checkpoint markers by DAC treatment on CRC cells might represent a mechanism of immune evasion. In addition, induction of NY-ESO-1 may represent an immuno-therapeutic option in metastatic CRC patients. Finally, the combination of DAC and anti-PD-1/anti-PD-L1 antibodies treatment should represent a potential therapeutic intervention for this group of patients.

TRPV2: A Cancer Biomarker and Potential Therapeutic Target.

The Transient Receptor Potential Vanilloid type-2 (TRPV2) channel exhibits oncogenicity in different types of cancers. TRPV2 is implicated in signaling pathways that mediate cell survival, proliferation, and metastasis. In leukemia and bladder cancer, the oncogenic activity of TRPV2 was linked to alteration of its expression profile. In multiple myeloma patients, TRPV2 overexpression correlated with bone tissue damage and poor prognosis. In prostate cancer, TRPV2 overexpression was associated with the castration-resistant phenotype and metastasis. Loss or inactivation of TRPV2 promoted glioblastoma cell proliferation and increased resistance to CD95-induced apoptotic cell death. TRPV2 overexpression was associated with high relapse-free survival in triple-negative breast cancer, whereas the opposite was found in patients with esophageal squamous cell carcinoma or gastric cancer. Another link was found between TRPV2 expression and either drug-induced cytotoxicity or stemness of liver cancer. Overall, these findings validate TRPV2 as a prime candidate for cancer biomarker and future therapeutic target.

Curcumin-Mediated Apoptotic Cell Death in Papillary Thyroid Cancer and Cancer Stem-Like Cells through Targeting of the JAK/STAT3 Signaling Pathway.

The constitutive activation of Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) signal transduction is well elucidated in STAT3-mediated oncogenesis related to thyroid cancer and is considered to be a plausible therapeutic target. Hence, we investigated whether curcumin, a natural compound, can target the JAK/STAT3 signaling pathway to induce cytotoxic effects in papillary thyroid cancer (PTC) cell lines (BCPAP and TPC-1) and derived thyroid cancer stem-like cells (thyrospheres). Curcumin suppressed PTC cell survival in a dose-dependent manner via the induction of caspase-mediated apoptosis and caused the attenuation of constitutively active STAT3 (the dephosphorylation of Tyr705-STAT3) without affecting STAT3. Gene silencing with STAT3-specific siRNA showed the modulation of genes associated with cell growth and proliferation. The cotreatment of PTC cell lines with curcumin and cisplatin synergistically potentiated cytotoxic effects via the suppression of JAK/STAT3 activity along with the inhibition of antiapoptotic genes and the induction of proapoptotic genes, and it also suppressed the migration of PTC cells by downregulating matrix metalloproteinases and the inhibition of colony formation. Finally, thyrospheres treated with curcumin and cisplatin showed suppressed STAT3 phosphorylation, a reduced formation of thyrospheres, and the downregulated expression of stemness markers, in addition to apoptosis. The current study's findings suggest that curcumin synergistically enhances the anticancer activity of cisplatin in PTC cells as well as in cancer stem-like cells by targeting STAT3, which suggests that curcumin combined with chemotherapeutic agents may provide better therapeutic outcomes.

The Role of Extracellular Vesicles as Modulators of the Tumor Microenvironment, Metastasis and Drug Resistance in Colorectal Cancer.

Colorectal cancer (CRC) is one of the most common cancers worldwide, with high morbidity and mortality rates. A number of factors including modulation of the tumor microenvironment, high metastatic capability, and resistance to treatment have been associated with CRC disease progression. Recent studies have documented that tumor-derived extracellular vesicles (EVs) play a significant role in intercellular communication in CRC via transfer of cargo lipids, proteins, DNA and RNAs to the recipient tumor cells. This transfer influences a number of immune-related pathways leading to activation/differentiation/expression of immune cells and modulation of the tumor microenvironment that plays a significant role in CRC progression, metastasis, and drug resistance. Furthermore, tumor-derived EVs are secreted in large amounts in biological fluids of CRC patients and as such the expression analysis of EV cargoes have been associated with prognosis or response to therapy and may be a source of therapeutic targets. This review aims to provide a comprehensive insight into the role of EVs in the modulation of the tumor microenvironment and its effects on CRC progression, metastasis, and drug resistance. On the other hand, the potential role of CRC derived EVs as a source of biomarkers of response and therapeutic targets will be discussed in detail to understand the dynamic role of EVs in CRC diagnosis, treatment, and management.

Evaluation of cationic channel TRPV2 as a novel biomarker and therapeutic target in Leukemia-Implications concerning the resolution of pulmonary inflammation.

Patients treated during leukemia face the risk of complications including pulmonary dysfunction that may result from infiltration of leukemic blast cells (LBCs) into lung parenchyma and interstitium. In LBCs, we demonstrated that transient receptor potential vanilloid type 2 channel (TRPV2), reputed for its role in inflammatory processes, exhibited oncogenic activity associated with alteration of its molecular expression profile. TRPV2 was overexpressed in LBCs compared to normal human peripheral blood mononuclear cells (PBMCs). Additionally, functional full length isoform and nonfunctional short form pore-less variant of TRPV2 protein were up-regulated and down-regulated respectively in LBCs. However, the opposite was found in PBMCs. TRPV2 silencing or pharmacological targeting by Tranilast (TL) or SKF96365 (SKF) triggered caspace-mediated apoptosis and cell cycle arrest. TL and SKF inhibited chemotactic peptide fMLP-induced response linked to TRPV2 Ca2+ activity, and down-regulated expression of surface marker CD38 involved in leukemia and lung airway inflammation. Challenging lung airway epithelial cells (AECs) with LBCs decreased (by more than 50%) transepithelial resistance (TER) denoting barrier function alteration. Importantly, TL prevented such loss in TER. Therefore, TRPV2 merits further exploration as a pharmacodynamic biomarker for leukemia patients (with pulmonary inflammation) who might be suitable for a novel [adjuvant] therapeutic strategy based on TL.

Anticancer Activity of Camel Milk via Induction of Autophagic Death in Human Colorectal and Breast Cancer Cells

Background/ Objective: Camel milk is traditionally known for its human health benefits and believed to be a remedy for various human ailments including cancer. The study was aimed to evaluate the inhibitory effects of commercially available camel milk on cancer cells and its underlying mechanism(s). Materials and Methods: Two cell lines: colorectal cancer HCT 116 and breast cancer MCF-7 were cultured with different doses of camel milk. The effects of camel milk on cell death were determined by MTT assay, viability by trypan blue exclusion assay and migration by in vitro scratch assay. The mechanism was elucidated by western blotting and confocal microscopy was used to confirm autophagy. Results: Camel milk significantly reduced proliferation, viability as well as migration of both the cells. The accumulation of LC3-II protein along with reduction in expression of p62 and Atg 5-12, the autophagy proteins implied induction of autophagy. The (GFP)-LC3 puncta detected by confocal microscopy confirmed the autophagosome formation in response to camel milk treatment. Conclusion: Camel milk exerted antiproliferative effects on human colorectal HCT 116 and breast MCF-7 cancer cells by inducing autophagy.

Rab2 utilizes glyceraldehyde-3-phosphate dehydrogenase and protein kinase C{iota} to associate with microtubules and to recruit dynein.

Rab2 requires glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and atypical protein kinase Ciota (aPKCiota) for retrograde vesicle formation from vesicular tubular clusters that sort secretory cargo from recycling proteins returned to the endoplasmic reticulum. However, the precise role of GAPDH and aPKCiota in the early secretory pathway is unclear. GAPDH was the first glycolytic enzyme reported to co-purify with microtubules (MTs). Similarly, aPKC associates directly with MTs. To learn whether Rab2 also binds directly to MTs, a MT binding assay was performed. Purified Rab2 was found in a MT-enriched pellet only when both GAPDH and aPKCiota were present, and Rab2-MT binding could be prevented by a recombinant fragment made to the Rab2 amino terminus (residues 2-70), which directly interacts with GAPDH and aPKCiota. Because GAPDH binds to the carboxyl terminus of alpha-tubulin, we characterized the distribution of tyrosinated/detyrosinated alpha-tubulin that is recruited by Rab2 in a quantitative membrane binding assay. Rab2-treated membranes contained predominantly tyrosinated alpha-tubulin; however, aPKCiota was the limiting and essential factor. Tyrosination/detyrosination influences MT motor protein binding; therefore, we determined whether Rab2 stimulated kinesin or dynein membrane binding. Although kinesin was not detected on membranes incubated with Rab2, dynein was recruited in a dose-dependent manner, and binding was aPKCiota-dependent. These combined results suggest a mechanism by which Rab2 controls MT and motor recruitment to vesicular tubular clusters.