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Protein arginine methyltransferase (PRMT) 5 is over-expressed in a variety of cancers and the master transcription regulator E2F1 is an important methylation target. We have explored the role of PRMT5 and E2F1 in regulating the non-coding genome and report here a striking effect on long non-coding (lnc) RNA gene expression. Moreover, many MHC class I protein-associated peptides were derived from small open reading frames in the lncRNA genes. Pharmacological inhibition of PRMT5 or adjusting E2F1 levels qualitatively altered the repertoire of lncRNA-derived peptide antigens displayed by tumour cells. When presented to the immune system as either ex vivo-loaded dendritic cells or expressed from a viral vector, lncRNA-derived peptides drove a potent antigen-specific CD8 T lymphocyte response, which translated into a significant delay in tumour growth. Thus, lncRNA genes encode immunogenic peptides that can be deployed as a cancer vaccine.

Pancreatic ductal adenocarcinoma (PDAC) is the most lethal type of cancer and the third leading cause of cancer death with the lowest 5-year survival rate. Heterogeneity, difficulty in diagnosis, and rapid metastatic progression are the causes of high mortality in pancreatic cancer. Recent studies have shown that Protein arginine methyltransferase 5 (PRMT5) is overexpressed in pancreatic cancers, and these patients have a worse prognosis. Recently, PRMT5 as an anti-cancer target has gained considerable interest. In this study, we investigated whether inhibition of PRMT5 activity was synergistic with blockade of TGF-β1 signaling, which plays an important role in the construction of the desmoplastic matrix in pancreatic cancer and induces therapeutic vulnerability. Compared with T1-44, a selective inhibitor of PRMT5 activity, the combination of T1-44 with the TGF-β1 signaling inhibitor Vactosertib significantly reduced tumor size and surrounding tissue invasion and significantly improved long-term survival. RNA sequencing analysis of mouse tumors revealed that the combination of T1-44 and Vactosertib significantly altered the expression of genes involved in cancer progression, such as cell migration, extracellular matrix, and apoptotic processes. In particular, the expression of Btg2, known as a tumor suppressor factor in various cancers, was markedly induced by combination treatment. Ectopic overexpression of Btg2 inhibited the EMT response, blocking cell migration, and promoted cancer cell death. These data demonstrate that the combination therapy of T1-44 with Vactosertib is synergistic for pancreatic cancer, suggesting that this novel combination therapy has value in the treatment strategy of patients with pancreatic cancer.

Protein acetylation plays a key role in regulating cellular processes and is subject to aberrant control in diverse pathologies. Although histone deacetylase (HDAC) inhibitors are approved drugs for certain cancers, it is not known whether they can be deployed in other therapeutic contexts. We have explored the clinical HDAC inhibitor, zabadinostat/CXD101, and found that it is a stand-alone regulator of the adaptive immune response. Zabadinostat treatment increased expression of MHC class I and II genes in a variety of cells, including dendritic cells (DCs) and healthy tissue. Remarkably, zabadinostat enhanced the activity of DCs, and CD4 and CD8 T lymphocytes. Using an antigenic peptide presented to the immune system by MHC class I, zabadinostat caused an increase in antigen-specific CD8 T lymphocytes. Further, mice immunised with covid19 spike protein and treated with zabadinostat exhibit enhanced covid19 neutralising antibodies and an increased level of T lymphocytes. The enhanced humoral response reflected increased activity of T follicular helper (Tfh) cells and germinal centre (GC) B cells. Our results argue strongly that zabadinostat has potential to augment diverse therapeutic agents that act through the immune system.

Telomerase is perceived as an immortality enzyme that might provide longevity to cells and whole organisms. Importantly, it is generally inactive in most somatic cells of healthy, adult men. Consequently, its substrates, i.e. telomeres, get shorter in most human cells with time. Noteworthy, cell life limitation due to telomere attrition during cell divisions, may not be as bad as it looks since longer cell life means longer exposition to harmful factors. Consequently, telomere length (attrition rate) becomes a factor that is responsible for inducing the signaling that leads to the elimination of cells that lived long enough to acquire severe damage. It seems that telomere length that depends on many different factors (including telomerase activity but also genetic factors, a hormonal profile that reflects sex, etc.) might become a useful marker of aging and exposition to stress. Thus in the current paper, we review the factors that affect telomere length in human cells focusing on sex that all together with different environmental and hormonal regulations as well as parental aspect affect telomere attrition rate. We also raise some limitations in the assessment of telomere length that hinders a trustworthy meta-analysis that might lead to acknowledgment of the real value of this parameter.

Aim To present the possibility of non-invasive monitoring of the skin after radiotherapy in regards of epidermal barrier function. Background Radiodermatitis constitutes 95% of all side effects in patients after radiotherapy. The proper assessment of the severity of radiodermatitis can be determined using semi-quantitative clinical scores [Common Terminology Criteria for Adverse Events v 4.0 (CTCAE)].The most accepted way to analyze the epidermal barrier function is to determine Transepidermal Water Loss (TEWL). Material and methods In prospective study, we included 16 patients diagnosed with head and neck cancer treated with radiotherapy or concomitant chemoradiation in whom we performed non-invasive assessments of the skin barrier function, including TEWL measurement. The final analysis included 6 patients (4 treated with adjuvant radiotherapy, 2 with radical chemoradiation). Clinical assessment of irradiated skin was based on target lesion score (TLS) and CTCAE v 4.0 Results The mean TLS score in the middle of irradiation was 1.6 points, after last irradiation it was 2.3 points; 3 months later the mean TLS score was: 0. CTCAE v 4.0 criteria: 2 patients had grade 0, 3 patients - grade 1; 1 patient - grade 2. There were statistically significant differences in TEWL related to irradiated skin in the following time intervals: before vs. in the middle; before vs. day after; in the middle vs. day after; in the middle vs. 3 months after; day after vs. 3 months after. Conclusions The study showed that radiotherapy causes skin barrier dysfunction in all patients independently of clinical radiodermatitis. The biophysical features of this dysfunction can precede clinical symptoms and they can be assessed by non-invasive and objective methods.

Cancer cells, including head and neck cancer cell carcinoma (HNSCC), are characterized by an increased telomerase activity. This enzymatic complex is active in approximately 80–90% of all malignancies, and is regulated by various factors, including methylation status of hTERT gene promoter. hTERT methylation pattern has been thoroughly studied so far. It was proved that hTERT is aberrantly methylated in tumor tissue versus healthy counterparts. However, such effect has not yet been investigated in PBLs (peripheral blood leukocytes) of cancer patients. The aim of this study was to analyze the hTERT gene promoter methylation status in blood leukocytes. DNA was extracted from PBL of 92 patients with histologically diagnosed HNSCC and 53 healthy controls. Methylation status of whole hTERT promoter fragment with independent analysis of each 19 CpG sites was performed using bisulfide conversion technique followed by sequencing of PCR products. Not significant (p = 0.0532) differences in the general frequency of hTERT CpG sites methylation were detected between patients and healthy controls. However, it was discovered that some of analyzed positions (CpG islands: 1 [p = 0.0235], 5 [p = 0.0462], 8 [p = 0.0343]) are significantly more often methylated in HNSCC patients than in controls. The opposite finding was observed in case of CpG position 2 (p = 0.0210). Furthermore, closer analysis of single CpG positions revealed differences in methylation status dependent on anatomical site and TNM classification. To conclude, hTERT promoter methylation status (general or single CpG sites) would be considered as a molecular markers of HNSCC diagnostics.

Nowotwory nadal stanowią jedną z głównych przyczyn śmiertelności w populacji światowej, a długotrwałe ich leczenie jest trudne i wysoce problematyczne, pomimo znacznego pogłębienia wiedzy z zakresu ich biologii. Obecnie wykorzystywane metody terapeutyczne cechują się wysoką toksycznością, tym samym często udaremniając skuteczne wyleczenie i pogarszając jakość życia chorych. Z tego względu wzrasta zainteresowane środkami, które mogłyby wzmocnić skuteczność konwencjonalnej terapii lub ograniczyć występowanie poważnych skutków ubocznych. Przykładem takiej substancji jest fukoidyna - sulfonowany fukan pochodzenia naturalnego, pozyskiwany z alg i brunatnic. Istnieją liczne doniesienia potwierdzające jej efekty terapeutyczne obejmujące właściwości przeciwnowotworowe, przeciwwirusowe oraz immunomodulujące. Poniższa praca zawiera opis efektów terapeutycznych wywoływanych przez fukoidynę wyekstrahowaną z różnych gatunków alg w poszczególnych modelach nowotworów. Praca uwzględnia poznane dotychczas i proponowane mechanizmy indukcji efektów przeciwnowotworowych oraz krytyczną ocenę ograniczeń fukoidyny jako potencjalnego środka wspomagającego w terapii onkologicznej.

Telomeraza to duży, złożony kompleks enzymatyczny, który odpowiada za proces syntezy telomerów budujących zakończenia eukariotycznych chromosomów. Podwyższona ekspresja genu kodującego katalityczną podjednostkę tego enzymu – hTERT – jest obserwowana w około 88% przypadków nowotworów rejonu głowy i szyi. Komórki rakowe zyskują wówczas wysoki potencjał replikacyjny, a także zdolność do unikania apoptozy. Zahamowanie aktywności telomerazy w terapii przeciwnowotworowej stało się priorytetem badań naukowych już kilkanaście lat temu. Istnieją różne metody wykorzystujące obniżenie ekspresji telomerazy, od terapii genowej do wykorzystania przeciwutleniaczy. Pomimo wielu danych zebranych w ostatnich latach pojawia się coraz więcej pytań w odniesieniu do roli telomerazy w powstawaniu raka. Należy przeprowadzić więcej szczegółowych badań w celu zrozumienia udziału tego enzymu w procesie rozwoju nowotworu w obrębie narządów głowy i szyi. Celem niniejszego artykułu jest zebranie dotychczasowej wiedzy na temat zastosowania telomerazy w terapii onkologicznej.

Arginine methylation widens the mechanism of control by E2F1 from a transcription factor to a regulator of alternative RNA splicing., E2F is a family of master transcription regulators involved in mediating diverse cell fates. Here, we show that residue-specific arginine methylation (meR) by PRMT5 enables E2F1 to regulate many genes at the level of alternative RNA splicing, rather than through its classical transcription-based mechanism. The p100/TSN tudor domain protein reads the meR mark on chromatin-bound E2F1, allowing snRNA components of the splicing machinery to assemble with E2F1. A large set of RNAs including spliced variants associate with E2F1 by virtue of the methyl mark. By focusing on the deSUMOylase SENP7 gene, which we identified as an E2F target gene, we establish that alternative splicing is functionally important for E2F1 activity. Our results reveal an unexpected consequence of arginine methylation, where reader-writer interplay widens the mechanism of control by E2F1, from transcription factor to regulator of alternative RNA splicing, thereby extending the genomic landscape under E2F1 control.

Radiodermatitis is one of the commonest side effects of radiotherapy. They are usually assessed by semi-quantitative clinical scores, which are not validated and may be subject to inter-observer variability. A few previous studies suggested that high-frequency ultrasonography (HF-USG) is useful in the assessment of the acute phase of radiation dermatitis in breast cancer patients. (a) To monitor skin changes by HF-USG during the course of radiotherapy due to head and neck cancers, and (b) to determine whether there is any connection between skin sonograms and the skin scoring criteria. This prospective, observational study includes patients diagnosed with head and neck cancers, treated with radiotherapy or concomitant chemoradiation. The final analysis includes six patients. In every patient, the HF-USG as well as dermatological assessment (target lesion score-TLS and CACE v. 4.0) were performed 4×: before, in the middle, day after, and 3 months after radiotherapy. There were significant differences between non-irradiated skin thickness and thickness of skin with clinically obvious radiodermatitis (TLS grade 1-4; P < .0001), as well as between irradiated, unchanged skin thickness (TLS grade 0) and thickness of skin with clinically obvious radiodermatitis (TLS grade 1-4; P = .0002). There was no significant difference between non-irradiated and irradiated, unchanged skin thickness (TLS grade 0; P = .9318). In four patients, we demonstrated subepidermal low echogenic band (SLEB). HF-USG can be useful tool to noninvasive and objective assessment of skin changes during radiotherapy.

Telomerase is known to contribute to telomere maintenance and to provide cancer cell immortality. However, numerous reports are showing that the function of the enzyme goes far beyond chromosome ends. The study aimed to explore how telomerase downregulation in MCF7 and MDA-MB-231 breast cancer cells affects their ability to survive. Consequently, sensitivity to drug resistance, proliferation, and adhesion were assessed. The lentiviral-mediated human telomerase reverse transcriptase (hTERT) downregulation efficiency was performed at gene expression and protein level using qPCR and Western blot, respectively. Telomerase activity was evaluated using the Telomeric Repeat Amplification Protocol (TRAP) assay. The study revealed that hTERT downregulation led to an increased sensitivity of breast cancer cells to doxorubicin which was demonstrated in MTT and clonogenic assays. During a long-term doubling time assessment, a decreased population doubling level was observed. Interestingly, it did not dramatically affect cell cycle distribution. hTERT downregulation was accompanied by an alteration in β1-integrin- and by focal adhesion kinase (FAK)-driven pathways together with the reduction of target proteins phosphorylation, i.e., paxillin and c-Src. Additionally, autophagy activation was observed in MDA-MB-231 cells manifested by alternations in Atg5, Beclin 1, LC3II/I ratio, and p62. These results provide new evidence supporting the possible therapeutic potential of telomerase downregulation leading to induction of autophagy and cancer cells elimination.

Cancer cells, including head and neck cancer cell carcinoma (HNSCC), are characterized by an increased telomerase activity. This enzymatic complex is active in approximately 80-90% of all malignancies, and is regulated by various factors, including methylation status of hTERT gene promoter. hTERT methylation pattern has been thoroughly studied so far. It was proved that hTERT is aberrantly methylated in tumor tissue versus healthy counterparts. However, such effect has not yet been investigated in PBLs (peripheral blood leukocytes) of cancer patients. The aim of this study was to analyze the hTERT gene promoter methylation status in blood leukocytes. DNA was extracted from PBL of 92 patients with histologically diagnosed HNSCC and 53 healthy controls. Methylation status of whole hTERT promoter fragment with independent analysis of each 19 CpG sites was performed using bisulfide conversion technique followed by sequencing of PCR products. Not significant (p = 0.0532) differences in the general frequency of hTERT CpG sites methylation were detected between patients and healthy controls. However, it was discovered that some of analyzed positions (CpG islands: 1 [p = 0.0235], 5 [p = 0.0462], 8 [p = 0.0343]) are significantly more often methylated in HNSCC patients than in controls. The opposite finding was observed in case of CpG position 2 (p = 0.0210). Furthermore, closer analysis of single CpG positions revealed differences in methylation status dependent on anatomical site and TNM classification. To conclude, hTERT promoter methylation status (general or single CpG sites) would be considered as a molecular markers of HNSCC diagnostics.

Telomere shortening is associated with cancer development, primarily through the induction of genomic instability. The majority of studies have indicated that individuals with shorter blood telomeres may be at a higher risk of developing various types of cancer. There is increasing evidence that the study of the alterations in telomere length may improve cancer prognosis. The aim of the present study was to verify the use of telomere length parameters in the diagnostics of breast cancer stage. Telomere length was analyzed in the blood leukocytes of 52 patients with breast cancer relative to 47 control subjects using quantitative polymerase chain reaction. The effects of stage, grade, estrogen receptor, progesterone receptor and human epidermal growth factor 2 (HER2) status were assessed. The current study demonstrated that the average telomeric sequence length was significantly shorter in leukocytes from individuals diagnosed with a more severe stage of breast cancer (T2N1M0) than in leukocytes in the early stages of the disease (T1N0M0) (P=0.0207). Furthermore, the data indicated that telomeres in leukocytes derived from patients with HER2+ breast cancer were significantly longer compared with those with the HER2− type (P=0.0347). These results suggest that the assessment of telomeres in blood leukocytes may, at least partially, correspond with breast cancer staging and HER2 receptor status.

Head and neck cancer is characterized by malignant tumors arising from the epithelium covering the upper aerodigestive tract, and the majority of these epithelial malignancies are squamous cell carcinomas (SCCs) of the oral cavity (OSCCs). The aim of the current work was to identify miRNAs regulated in OSCC cancerous tissue when compared to a healthy adjacent tissue and to verify the presence of the same miRNAs in the circulation of these patients. For that serum samples and biopsies of healthy and tumor tissues were collected from five patients diagnosed with OSCC of the oral cavity, RNA was extracted from these samples and microRNAs libraries were prepared and sequenced. A total 255 miRNAs were identified in tissue and 381 different miRNAs were identified in serum samples. When comparing the miRNA expression between tumor and healthy tissue we identified 48 miRNAs (25 down- and 23 up-regulated) that were differentially expressed (FDR

Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous group of malignant tumours that affects over 500,000 patients per year. Treatment failure is generally due to the heterogeneity of these tumours and to the serious adverse effects associated with treatment. Immunological system impairment, which is common in HNSCC, further contributes to treatment failure by mediating tumour escape mechanisms. To date, the only clinically approved targeted therapy agent is cetuximab, a monoclonal antibody (mAb) that binds to, and inhibits, epidermal growth factor receptor, which is widely overexpressed in HNSCC. Cetuximab has been proven to induce antibody-dependent cellular cytotoxicity, further magnifying its therapeutic effect. DNA sequencing of HNSCC cells has identified the presence of mutated genes, thus making their protein products potential targets for therapeutic inhibition. Immune mechanisms have been found to have a significant impact on carcinogenesis, thus providing the rationale to support efforts to identify anticancer compounds with immunomodulatory properties. In the context of the rapid development of novel targeted agents, the aim of the present paper is to review our current understanding of HNSCC and to review the novel anticancer agents (mAbs and TKIs) introduced in recent years, including an assessment of their efficacy and mechanisms of action.

Squamous cell carcinoma of the head and neck (HNSCC) is the sixth leading cause of cancer worldwide, representing over half a million incidents every year. Cancer cells, including HNSCC, are characterized by increased telomerase activity. This enzymatic complex is active in ~90% of all cancer types and is responsible for the lengthening of telomeres. Highly recurrent point mutations in the human telomerase reverse transcriptase (hTERT) promoter have recently been reported in a number of human neoplasms. The aim of the present study was to analyze the prevalence of the hTERT promoter C250T mutation and telomere length in the blood leukocytes of 61 patients with HNSCC and 49 healthy individuals. Quantitative polymerase chain reaction identified the hTERT promoter mutation in 36% of patients with HNSCC. To the best of our knowledge this is first report indicating the presence of shorter telomeres in early stage tumors. In addition, the results suggest that the C250T hTERT promoter mutation and telomere length assessment may serve as important molecular markers of HNSCC progression.

Ascorbic acid (vitamin C) has been gaining attention as a potential treatment for human malignancies. Various experimental studies have shown the ability of pharmacological doses of vitamin C alone or in combinations with clinically used drugs to exert beneficial effects in various models of human cancers. Cytotoxicity of high doses of vitamin C in cancer cells appears to be related to excessive reactive oxygen species generation and the resulting suppression of the energy production via glycolysis. A hallmark of cancer cells is a strongly upregulated aerobic glycolysis, which elevates its relative importance as a source of ATP (Adenosine 5′-triphosphate). Aerobic glycolysis is maintained by a highly increased uptake of glucose, which is made possible by the upregulated expression of its transporters, such as GLUT-1, GLUT-3, and GLUT-4. These proteins can also transport the oxidized form of vitamin C, dehydroascorbate, permitting its preferential uptake by cancer cells with the subsequent depletion of critical cellular reducers as a result of ascorbate formation. Ascorbate also has a potential to affect other aspects of cancer cell metabolism due to its ability to promote reduction of iron(III) to iron(II) in numerous cellular metalloenzymes. Among iron-dependent dioxygenases, important targets for stimulation by vitamin C in cancer include prolyl hydroxylases targeting the hypoxia-inducible factors HIF-1/HIF-2 and histone and DNA demethylases. Altered metabolism of cancer cells by vitamin C can be beneficial by itself and promote activity of specific drugs.

Down syndrome (DS) is one of the most common aneuploidy. In general population, its prevalence is 1:600-1:800 live births. It is caused by a trisomy of chromosome 21. DS is phenotypically manifested by premature aging, upward slant to the eyes, epicanthus, flattened face, and poor muscle tone. In addition to physical changes, this syndrome is characterized by early onset of diseases specific to old age, such as Alzheimer's disease, vision and hearing problems, and precocious menopause. Since DS symptoms include premature aging, the shortening of telomeres might be one of the markers of cellular aging. Consequently, the aim of the study was to determine the length of the telomeres in leukocytes from the blood of juvenile patients with DS (n=68) compared to an age-matched control group (n=56) and also to determine the diagnostic or predictive value for this parameter. We show that, for the first time, in juveniles, the average relative telomere length in studied subjects is significantly longer than in the control group (50.46 vs. 40.56, respectively arbitrary units [AU]; p=0.0026). The results provide interesting basis for further research to determine the causes and consequences of telomere maintaining and the dynamics of this process in patients with DS.

Background The accumulation of mutagenic substances in the human body may result in DNA metabolism disruption followed by carcinogenesis. As a consequence of mutations in the genes coding for transmembrane protein pumps, the intracellular concentration of xenobiotics may significantly increase. This, in turn, may provoke altered risk for cancer development. The gene known to be the most relevant in the transport of numerous compounds is ABCB1 (also known as MDR1 ). Numerous mutations and polymorphisms that affect the encoded protein's (PgP) function were identified in this gene. The aim of the study was to define the frequency of 2677G > A,T and 3435C > T polymorphisms in a population of Polish breast cancer patients and to estimate their contribution to cancer development. Methods The polymorphism frequency analysis (209 patients vs. 202 control subjects) was performed either by allele-specific amplification (2677G > A,T) or by restriction fragment length polymorphism (RFLP) using the SAU3AI restriction enzyme (3435C > T) followed by verification with hybridization probe assays in a Real-Time system and sequencing. Results In the control group the frequency of individual 2677 genotypes was: wild homozygous GG = 34%, heterozygous G/T or G/A = 52.5% and variant homozygous AA or TT = 13.5%, while the genotype frequency in the group of studied patients was 43.5, 44.5 and 12%, respectively. In the control group, the frequency of individual 3435 genotypes was: CC = 25.4%, CT = 50.2%, TT = 24.4%, while the genotype frequency in the group of studied patients was 23, 46 and 31%, respectively. Conclusions Thus, no significant differences in the studied polymorphism frequencies were observed. It is then suggested that the studied polymorphisms, although probably good candidates in other tissue cancer types, might not be good predictive factors in breast cancer risk or development in Caucasians.