Protease-Activated Receptors (PARs): Biology and Therapeutic Potential in Perioperative Stroke.

Perioperative stroke is a devastating complication that occurs during surgery or within 30 days following the surgical procedure. Its prevalence ranges from 0.08 to 10% although it is most likely an underestimation, as sedatives and narcotics can substantially mask symptomatology and clinical presentation. Understanding the underlying pathophysiology and identifying potential therapeutic targets are of paramount importance. Protease-activated receptors (PARs), a unique family of G-protein-coupled receptors, are widely expressed throughout the human body and play essential roles in various physiological and pathological processes. This review elucidates the biology and significance of PARs, outlining their diverse functions in health and disease, and their intricate involvement in cerebrovascular (patho)physiology and neuroprotection. PARs exhibit a dual role in cerebral ischemia, which underscores their potential as therapeutic targets to mitigate the devastating effects of stroke in surgical patients.

Novel insights into voltage-gated ion channels: Translational breakthroughs in medical oncology.

Preclinical evidence suggests that voltage gradients can act as a kind of top-down master regulator during embryogenesis and orchestrate downstream molecular-genetic pathways during organ regeneration or repair. Moreover, electrical stimulation shifts response to injury toward regeneration instead of healing or scarring. Cancer and embryogenesis not only share common phenotypical features but also commonly upregulated molecular pathways. Voltage-gated ion channel activity is directly or indirectly linked to the pathogenesis of cancer hallmarks, while experimental and clinical studies suggest that their modulation, e.g., by anesthetic agents, may exert antitumor effects. A large recent clinical trial served as a proof-of-principle for the benefit of preoperative use of topical sodium channel blockade as a potential anticancer strategy against early human breast cancers. Regardless of whether ion channel aberrations are primary or secondary cancer drivers, understanding the functional consequences of these events may guide us toward the development of novel therapeutic approaches.

The Relation of Calculated Plasma Volume Status to Sublingual Microcirculatory Blood Flow and Organ Injury.

BACKGROUND: The calculated plasma volume status (cPVS) was validated as a surrogate of intravascular filling. The aim of this study is to assess the cPVS in relation to sublingual perfusion and organ injury. METHODS: Pre- and postoperative cPVS were obtained by determining the actual and ideal plasma volume levels in surgical patients. The sublingual microcirculation was assessed using SDF imaging, and we determined the De Backer score, the Consensus Proportion of Perfused Vessels (Consensus PPV), and the Consensus PPV (small). Our primary outcome was the assessment of the distribution of cPVS and its association with intraoperative sublingual microcirculation and postoperative complications. RESULTS: The median pre- and postoperative cPVS were -7.25% (IQR -14.29--1.88) and -0.4% (IQR -5.43-6.06), respectively (p < 0.001). The mean intraoperative administered fluid volume was 2.5 ± 2.5 L (1.14 L h-1). No statistically significant correlation was observed between the pre- or postoperative cPVS and sublingual microcirculation variables. Higher preoperative (OR = 1.04, p = 0.098) and postoperative cPVS (OR = 1.057, p = 0.029) were associated with postoperative organ injury and complications (sepsis (30%), anemia (24%), respiratory failure (13%), acute kidney injury (6%), hypotension (6%), stroke (3%)). CONCLUSIONS: The calculated PVS was associated with an increased risk of organ injury and complications in this cohort.

Effects of Vasopressin Receptor Agonists during the Resuscitation of Hemorrhagic Shock: A Systematic Review and Meta-Analysis of Experimental and Clinical Studies.

BACKGROUND: The clinical impact of vasopressin in hemorrhagic shock remains largely unknown. OBJECTIVE: This systematic review and meta-analysis was designed to investigate the effects of vasopressin receptor agonists during the resuscitation of hemorrhagic shock. METHODS: A systematic search of PubMed (MEDLINE), Scopus, and PubMed Central was conducted for relevant articles. Experimental (animal) and clinical studies were included. The primary objective was to investigate the correlation of vasopressin receptor agonist use with mortality and various hemodynamic parameters. RESULTS: Data extraction was possible in thirteen animal studies and two clinical studies. Differences in risk of mortality between patients who received a vasopressin receptor agonist were not statistically significant when compared to those who were not treated with such agents [RR (95% CI): 1.17 (0.67, 2.08); p = 0.562; I2 = 50%]. The available data were insufficient to conduct a meta-analysis assessing the effect of vasopressin receptor agonists on hemodynamics. Drawing safe conclusions from animal studies was challenging, due to significant heterogeneity in terms of species and dosage of vasopressin receptor agonists among studies. CONCLUSIONS: Differences in risk of mortality between patients who received a vasopressin receptor agonist were not statistically significant when compared to those who were not treated with such agents after hemorrhagic shock. More data are needed to deduce certain conclusions.

Why Metformin Should Not Be Used as an Oxidative Phosphorylation Inhibitor in Cancer Patients.

BACKGROUND: Preclinical studies have suggested that metformin exerts antitumor effects on various types of cancers. However, the results of human clinical trials have been inconsistent. SUMMARY: Metformin is widely considered to be a prime example of a clinically relevant compound that inhibits oxidative phosphorylation (OXPHOS). However, the efficacy of metformin in inhibiting OXPHOS in cancer patients remains uncertain. The available evidence suggests that the plasma concentration of metformin remains within the micromolar range when administered at therapeutic doses. While millimolar concentrations are necessary to inhibit complex I activity in isolated mitochondria, there is no evidence supporting the idea that metformin accumulates within the mitochondria. Metformin exerts a modest effect on the adenosine diphosphate to adenosine triphosphate (ATP) ratio, resulting in AMP-activated protein kinase activation, which promotes ATP-generating catabolic pathways and restores cellular energy balance. KEY MESSAGES: The value of metformin as an OXPHOS inhibitor for cancer treatment is debatable, and caution should be exercised while using metformin for this purpose.

Current Status of Tivozanib in the Treatment of Patients With Advanced Renal Cell Carcinoma.

The introduction of tyrosine kinase inhibitors (TKIs) against vascular endothelial growth factor receptors (VEGFRs) has transformed the therapeutic landscape for patients with advanced renal cell carcinoma (RCC). However, dose reductions and interruptions are frequently needed due to limited toxicity, mostly from off-target effects. Tivozanib is a potent, selective VEGFR TKI with weak off-target effects. TIVO-1 and TIVO-3 were randomized controlled phase 3 trials that investigated the efficacy and safety of tivozanib versus sorafenib as initial targeted therapy and after failing two previous lines (including targeted therapy), respectively. Tivozanib did not confer any survival advantage, but it significantly increased progression-free survival, response rates, and the duration of responses with a superior safety profile. Although results from subgroup analysis need to be interpreted cautiously, tivozanib demonstrated superiority after two previous lines of VEGFR TKIs or after axitinib, another selective VEGFR inhibitor. Tivozanib also demonstrated durable activity after therapy with an immune-checkpoint inhibitor, while an ongoing study investigating the combination of tivozanib/nivolumab has shown promising preliminary results regarding efficacy and safety. In conclusion, tivozanib was recently added to our therapeutic armamentarium against advanced RCC. Ongoing rational therapeutic combinations of tivozanib will determine the optimal setting in which the maximum benefit can be derived.

Ursolic Acid against Prostate and Urogenital Cancers: A Review of In Vitro and In Vivo Studies.

Prostate cancer is the second most diagnosed form of cancer in men worldwide and accounted for roughly 1.3 million cases and 359,000 deaths globally in 2018, despite all the available treatment strategies including surgery, radiotherapy, and chemotherapy. Finding novel approaches to prevent and treat prostate and other urogenital cancers effectively is of major importance. Chemicals derived from plants, such as docetaxel and paclitaxel, have been used in cancer treatment, and in recent years, research interest has focused on finding other plant-derived chemicals that can be used in the fight against cancer. Ursolic acid, found in high concentrations in cranberries, is a pentacyclic triterpenoid compound demonstrated to have anti-inflammatory, antioxidant, and anticancer properties. In the present review, we summarize the research studies examining the effects of ursolic acid and its derivatives against prostate and other urogenital cancers. Collectively, the existing data indicate that ursolic acid inhibits human prostate, renal, bladder, and testicular cancer cell proliferation and induces apoptosis. A limited number of studies have shown significant reduction in tumor volume in animals xenografted with human prostate cancer cells and treated with ursolic acid. More animal studies and human clinical studies are required to examine the potential of ursolic acid to inhibit prostate and other urogenital cancers in vivo.

Outcomes from treatment of metastatic renal-cell carcinoma following failure of first-line anti-VEGF/VEGFR therapy: real-life evidence on the change of the treatment paradigm.

Recently approved agents for post-vascular endothelial growth factor/post-vascular endothelial growth factor receptor (VEGF/VEGFR) inhibitors treatment of metastatic renal-cell carcinomas (mRCC), such as axitinib, nivolumab, and cabozantinib were shown to improve prognosis and substituted everolimus in this setting. We studied practice patterns, efficacy, and tolerability of these agents in a real-world series of Greek patients. We included patients with mRCC who received everolimus, axitinib, or nivolumab after progression on first-line anti-VEGF/VEGFRs therapy. Patients were stratified into three groups. Group A received nivolumab with or without cabozantinib at some point in their disease. Group B received axitinib but without nivolumab or cabozantinib. Group C received only everolimus among the four approved agents. Overall, 131 patients were included in the analysis. Everolimus and nivolumab were mainly used in the second line, while axitinib and cabozantinib were mostly used in the third and fourth lines. Median overall survival (OS) from first-line initiation was 8.7 [95% confidence interval (CI), 4-not reached], 3.6 (95% CI, 2-6), and 2.1 years (95% CI, 1.4-2.6) for Group A, B, and C, respectively ( P < 0.001). Median OS from the initiation of second-line therapy was 3.5, 2.7, and 1.3 years, respectively ( P < 0.001). There was no impact of first-line agent or treatment timing on survival. International Metastatic Renal Cell Carcinoma Database Consortium risk stratification was associated with OS. Toxicities observed were within expected frequencies. Grade ≥3 events were rare. Adoption of modern standards in everyday treatment of mRCC results in prolongation of survival. Real-world datasets are the new landmarks of survival for future research.

The Role οf Ion Channels in the Development and Progression of Prostate Cancer.

Ion channels have major regulatory functions in living cells. Apart from their role in ion transport, they are responsible for cellular electrogenesis and excitability, and may also regulate tissue homeostasis. Although cancer is not officially classified as a channelopathy, it has been increasingly recognized that ion channel aberrations play an important role in virtually all cancer types. Ion channels can exert pro-tumorigenic activities due to genetic or epigenetic alterations, or as a response to molecular signals, such as growth factors, hormones, etc. Increasing evidence suggests that ion channels and pumps play a critical role in the regulation of prostate cancer cell proliferation, apoptosis evasion, migration, epithelial-to-mesenchymal transition, and angiogenesis. There is also evidence suggesting that ion channels might play a role in treatment failure in patients with prostate cancer. Hence, they represent promising targets for diagnosis, staging, and treatment, and their effects may be of particular significance for specific patient populations, including those undergoing anesthesia and surgery. In this article, the role of major types of ion channels involved in the development and progression of prostate cancer are reviewed. Identifying the underlying molecular mechanisms of the pro-tumorigenic effects of ion channels may potentially inform the development of novel therapeutic strategies to counter this malignancy.

Niclosamide in prostate cancer: An inhibitor of AR-V7, a mitochondrial uncoupler, or more?

A recent phase Ib study investigating the use of reformulated niclosamide in combination with abiraterone and prednisone in patients with castration-resistant prostate cancer (CRPC) demonstrated encouraging preliminary efficacy with low toxicity. Preclinical studies have reported that niclosamide at clinically relevant concentrations inhibits androgen receptor splice variant 7 (AR-V7), a known tumor driver in CRPC. However, the magnitude of anti-tumor effects of niclosamide either used alone or in combination with abiraterone in these experimental models, far exceeded what could have been explained as a simple AR-V7 inhibition. Niclosamide at clinically relevant concentrations also acts as an oxidative phosphorylation (OxPhos) uncoupler in mitochondria. This raises the question whether the observed effects of niclosamide were partly mediated by OxPhos inhibition. Most OxPhos inhibitors did not demonstrate selectivity towards cancer cells and failed to enter clinical practice due to unacceptable toxicity. However, some mitochondrial uncouplers have greater cytotoxicity against cancerous cells compared to non-cancerous. Hyperpolarization of cancer cell mitochondria, or the more alkaline mitochondrial matrix of cancer cells could be potential reasons for this. Niclosamide can also alter Wnt/ß-catenin, mTOR, Notch, NF-kB and STAT3 signaling pathways. Hence, the mechanism of action of reformulated niclosamide in CRPC patients requires further investigation. This will potentially lead to new opportunities to develop and investigate even more selective and effective treatments against prostate cancer.

Characterization of prostate cancer adrenal metastases: dependence upon androgen receptor signaling and steroid hormones.

BACKGROUND: Prostate cancer (PCa) typically spreads to the bone, and this distribution is attributed to the central role of the microenvironment in progression. However, metastasis to the adrenal glands, while not as common, does occur. The biology that accounts for adrenal metastases may be attributed to the unique local steroid metabolome and co-clinical characterization may elucidate the role steroid biosynthesis plays in PCa progression. METHODS: Three patients with metastatic PCa who had archived tumor tissue from an adrenalectomy were retrospectively identified, and one adrenal metastasis was developed into a xenograft (MDA-PCa-250). The adrenal metastases were characterized by performing somatic DNA whole exome sequencing (WES), RNA-Seq, immunohistochemistry (IHC), and steroid metabolite quantitation. The influence of steroid metabolites on adrenal metastasis cells and tumor growth was tested in vitro and in vivo. RESULTS: Clinically, adrenalectomy was performed during castration-resistant oligometastatic disease, and two men experienced resensitization to leuprolide. Somatic DNA WES revealed heterogeneous alterations in tumor suppressor and DNA damage repair pathway genes. Adrenal metastases had active androgen receptor (AR) signaling by IHC, and RNA-Seq supported a potential role for adrenal androgen precursor metabolism in activating the AR. Steroid quantitation suggested the adrenal androgen precursors were converted into testosterone in these metastases, and stable isotope tracing of an organoid from MDA-PCa-250 confirmed the capability of adrenal metastases to biosynthesize testosterone from adrenal precursors. In vitro testing of a cell line derived from MDA-PCa-250 showed that testosterone and cortisol stimulated tumor cell growth. In vivo experiments demonstrated that MDA-PCa-250 grew in intact mice with circulating testosterone, but not in castrated mice. CONCLUSIONS: PCa adrenal metastases depend upon AR signaling driven by androgen precursors, androstenedione and dehydroepiandrosterone, available in the microenvironment, despite the presence of heterogeneous somatic DNA alterations. Moreover, MDA-PCa-250 provides a preclinical model that can recapitulate the unique androgen-dependence of adrenal metastases. CLINICAL TRIAL REGISTRATION: This study does not report the clinical results of a clinical trial, but it does use samples from a completed clinical trial that is registered with clinicaltrials.gov (NCT01254864).

Burnout Syndrome among Doctors in Greek Oncology Departments.

Objective: Economic crisis and austerity has dramatic consequences for health care professionals' mental health. This study aimed to investigate the prevalence of burnout syndrome among doctors working in public or private oncology departments in Greece and its association with factors determined by economic crisis. Method : Medical or radiation oncologists who are members of the National Oncology Societies and practicing oncology at least for one year were enrolled in this study. An On-Line questionnaire consisting of the Maslach Burnout Inventory and an informative questionnaire was utilized. Results: Eighty-six On-Line questionnaires waere analyzed. Radiation oncologists had significantly higher Depersonalization (DP), whereas medical oncologists had significantly higher Emotional Exhaustion (EE) scores. Registrars had higher scores compared to consultants in all subscales of the burnout syndrome. Factors such as ''hospitals are not safe due to the lack or reductions in medical supplies and personnel shortage'' and ''receiving less than 50% of annual leave'' were associated with significantly higher levels of EE. Factors such as ''not afraid of moving abroad'' and ''receiving 100% of annual leave'' were associated with significantly higher levels of low Personal Accomplishments (PA). The principal component analysis yielded three principal components: 'future insecurity', 'feeling secure while working' and 'working conditions associated with burnout syndrome. Conclusion: Several factors associated with austerity resulting from economic crisis significantly influenced prevalence of burnout syndrome among oncologists in Greece. Further studies need to be conducted to mobilize policy makers to develop and implement policies to improve oncologists' mental health.

Risk for Arterial Thromboembolic Events (ATEs) in Patients with Advanced Urinary Tract Cancer (aUTC) Treated with First-Line Chemotherapy: Single-Center, Observational Study.

Arterial thromboembolism has been associated with cancer or its treatment. Unlike venous thromboembolism, the incidence and risk factors have not been extensively studied. Here, we investigated the incidence of arterial thromboembolic events (ATEs) in an institutional series of advanced urinary tract cancer (aUTC) treated with cytotoxic chemotherapy. The ATE definition included peripheral arterial embolism/thrombosis, ischemic stroke and coronary events. A total of 354 aUTC patients were analyzed. Most patients (95.2%) received platinum-based chemotherapy. A total of 12 patients (3.4%) suffered an ATE within a median time of 3.6 months from the start of chemotherapy. The most frequent ATE was ischemic stroke (n = 7). Two ATEs were fatal. The 6-month and 24-month incidence were 2.1% (95% confidence interval [CI]: 0.9-4.1) and 3.6% (95% CI: 1.9-6.2), respectively. Perioperative chemotherapy increased the risk for ATE by 5.55-fold. Tumors other than UTC and pure non-transitional cell carcinoma histology were also independent risk factors. No association with the type of chemotherapy was found. Overall, ATEs occur in 4.6% of aUTC patients treated with chemotherapy and represent a clinically relevant manifestation. Perioperative chemotherapy significantly increases the risk for ATE. The role of prophylaxis in high-risk groups should be prospectively studied.

Potential Ophthalmological Side Effects Induced by Anti-Neoplastic Regimens for the Treatment of Genitourinary Cancers: A Review.

The outcomes of patients with genitourinary (GU) cancers have been steadily improving in recent years. Novel therapies have entered our armamentarium, while several other regimens are currently being studied in clinical trials. This recent explosion of new agents has improved patient survival and the quality of life for patients, but has also significantly increased the frequency of several side effects. The current review will focus on the potential ocular adverse reactions of GU neoplastic treatments. The broad spectrum of manifestations of ocular toxicity underscores the uniqueness and complexity of the anatomic, physiologic, and metabolic features of the human eye. Most side effects are mild in severity and transient, but some can be severe, disabling, and irreversible. Clinicians should be aware of complications that might be vision threatening and impact the patient's quality of life. In this review, we focused on the ocular toxicity of the antineoplastic regimens that are currently used for the treatment of GU, including prostate cancer, bladder cancer, renal cell carcinoma, testicular cancer, pheochromocytoma, adrenocortical carcinoma, and penile cancer.

Is the glucocorticoid receptor a key player in prostate cancer?: A literature review.

Glucocorticoids act through the glucocorticoid receptor (GR) and exert pleiotropic effects in different cancer types. In prostate cancer cells, GR and androgen receptor (AR) share overlapping transcriptomes and cistromes. Under enzalutamide treatment, GR signaling can bypass AR activation and promote castration resistance via the expression of a subset of AR-target genes. However, GR-dependent growth under enhanced antiandrogen inhibition occurs only in a subset of primed cells. On the other hand, glucocorticoids have been used successfully in the treatment of prostate cancer for many years. In the context of AR signaling, GR competes with AR for DNA-binding and has the potential to halt the proliferation rate of prostate cancer cells. Their target genes overlap by <50% and they execute unique functions in vivo. In addition, even when AR and GR upregulate the same transcriptional target gene, the effect might not be identical in magnitude. Besides being able to drive tumor proliferation, GR is also a key player in prostate cancer cell survival. Stimulation of GR activity can undermine the effects of enhanced antiandrogen treatment, chemotherapy and radiotherapy. GR activation in prostate cancer can increase prosurvival gene expression. Identifying the full spectrum of GR activity will inform the optimal use of glucocorticosteroids in prostate cancer. It will also determine the best strategies to target the protumorigenic effects of GR.

Patterns of indolence in prostate cancer (Review).

Although prostate cancer is a major cause of cancer-related mortality worldwide, most patients will have a relatively indolent clinical course. Contrary to most other types of cancer, even the diagnosis of locally advanced or metastatic disease is not always lethal. The present review aimed to summarize what is known regarding the underlying mechanisms related to the indolent course of subsets of prostate cancer, at various stages. The data suggested that no specific gene alteration by itself was responsible for carcinogenesis or disease aggressiveness. However, pathway analysis identified genetic aberrations in multiple critical pathways that tend to accumulate over the course of the disease. The progression from indolence into aggressive disease is associated with a complex interplay in which genetic and epigenetic factors are involved. The effect of the immune tumor microenvironment is also very important. Emerging evidence has suggested that the upregulation of pathways related to cellular aging and senescence can identify patients with indolent disease. In addition, a number of tumors enter a long-lasting quiescent state. Further research will determine whether halting tumor evolution is a feasible option, and whether the life of patients can be markedly prolonged by inducing tumor senescence or long-term dormancy.

Orphan receptors in prostate cancer.

BACKGROUND: The identification of new cellular receptors has been increasing rapidly. A receptor is called "orphan" if an endogenous ligand has not been identified yet. METHODS: Here we review receptors that contribute to prostate cancer and are considered orphan or partially orphan. This means that the full spectrum of their endogenous ligands remains unknown. RESULTS: The orphan receptors are divided into two major families. The first group includes G protein-coupled receptors. Most are orphan olfactory receptors. OR51E1 inhibits cell proliferation and induces senescence in prostate cancer. OR51E2 inhibits prostate cancer growth, but promotes invasiveness and metastasis. GPR158, GPR110, and GPCR-X play significant roles in prostate cancer development and progression. However, GPR160 induces cell cycle arrest and apoptosis. The other major subset of orphan receptors are nuclear receptors. Receptor-related orphan receptor α (RORα) inhibits tumor growth, but RORγ stimulates androgen receptor signaling. PXR contributes to metabolic deactivation of androgens and inhibits cell proliferation. TLX has protumorigenic effects in prostate cancer, while its knockdown triggers cellular senescence and growth arrest. Estrogen-related receptor ERRγ can inhibit tumor growth but ERRα is protumorigenic. Dax1 and short heterodimeric partner are also inhibitory in prostate cancer. CONCLUSION: There is a "zoo" of relatively underappreciated orphan receptors that play key roles in prostate cancer.

Androgen receptor signaling-mitochondrial DNA-oxidative phosphorylation: A critical triangle in early prostate cancer.

Mitochondria are more than just the cellular powerhouse. They also play key roles in vital functions such as apoptosis, metabolism regulation, and other intracellular interactions. The mitochondrial DNA (mtDNA) encodes for 12 subunits of the oxidative phosphorylation (OXPHOS) system. Depletion of mtDNA in androgen-dependent prostate cancer (PCa) cell lines renders them androgen-independent and more aggressive. Paradoxically, pharmaceutical inhibition of OXPHOS is lethal for subsets of PCa cells, whereas others become dependent on androgen receptor (AR) signaling for survival. Given that the AR-mitochondria interaction is critical for early PCa, it is crucial to understand the details of this interaction. Technical hurdles have made mitochondria traditionally difficult to study, with many techniques used for isolation masking the properties of given individual mitochondria. Although the isolation of mitochondria enables us to study OXPHOS, we miss the context in which mitochondria interact with the rest of the cell. Both AR signaling and mtDNA affect apoptosis, metabolism regulation, cellular calcium storage and homeostasis, intracellular calcium signaling, and redox homeostasis. In this review, we will attempt to understand how the crosstalk between AR-mtDNA-OXPHOS is responsible for "life or death" decisions inside the cells. Our aim is to point toward potential vulnerabilities that can lead to the discovery of novel therapeutic targets.

Epigenetics and prostate cancer: defining the timing of DNA methyltransferase deregulation during prostate cancer progression.

DNA methyltransferases (DNMTs) regulate gene expression by methylating cytosine residues within CpG dinucleotides. Aberrant methylation patterns have been shown in a variety of human tumours including prostate cancer. However, the expression of DNMTs in clinical samples across the spectrum of prostate cancer progression has not been studied before. Tissue microarrays were constructed from the prostatectomy specimens of 309 patients across the spectrum of prostate cancer progression: hormone-naïve low-grade prostate cancer (n=49), hormone-naïve high-grade prostate cancer (n=151), hormonally treated high-grade prostate cancer (n=65), and castrate-resistant prostate cancer (CRPC) including neuroendocrine carcinoma (n=44). Adjacent non-neoplastic parenchyma was also available in 100 patients. In 71 patients with high-grade carcinoma and lymph node metastasis, tissue from the metastasis was also available for analysis. Immunohistochemical staining was performed with antibodies against DNMT1, DNMT2, DNMT3A, DNMT3B, and DNMT3L. Our results showed that DNMT1 and DNMT3L were upregulated early in prostate cancer progression, whereas DNMT2 was upregulated as a response to androgen ablation. DNMT1, DNMT3A, and DNMT3B were higher in the late stages of prostate cancer progression, i.e., the emergence of castrate resistance and androgen-independent growth. Lastly, DNMT1, DNMT2, and DNMT3L were upregulated in lymph node metastases compared to primary carcinomas. Our results highlight a cascade of epigenetic events in prostate cancer progression.

A Standardized Evaluation Method for FOXP3+ Tregs and CD8+ T-cells in Breast Carcinoma: Association With Breast Carcinoma Subtypes, Stage and Prognosis.

BACKGROUND/AIM: The role of FOXP3+ Tregs and CD8+ T-cells in different stages and subtypes of breast carcinoma (BC) is yet to be fully defined, mainly because of methodological variations between studies. The aim of this study was to assess FOXP3+ and CD8+ intratumoral stromal TILs (sTILs) by a standardized method, in order to discern differences between the histological subtypes and BC stage and evaluate the applicability of the method. PATIENTS AND METHODS: FOXP3+ and CD8+ sTILs were studied immunohistochemically in 207 BCs and counted on digital images, amounting to a standard stromal area of a 10×10 grid on ×40 magnification. The results were correlated with clinicopathological features and outcomes. RESULTS: Tregs and CD8+ TILs were more abundant in HER2+ BCs (p=0.02, p=0.007, respectively), estrogen receptor (ER)-BCs (p<0.001, for both cell types), and triple-negative BCs (TNBCs) (p=0.01, p=0.006, respectively). Tregs and CD8+ TILs were associated with high grade (p<0.001 and p=0.002, respectively) and high Ki67 index (p<0.001, for both cell types). Lower CD8/FOXP3 ratio was associated with node metastases (p=0.007). Node metastases and advanced stage paralleled with decreased CD8+ sTILs (p=0.023, p=0.019, respectively). In the entire group and in ER- BCs, CD8+ TILs were associated with favorable distant metastasis-free survival (p=0.021, p<0.001, respectively), disease-free survival (p=0.022, p<0.001, respectively) and breast cancer specific survival (BCSS) (p=0.022, p=0.005). In ER-BCs, Tregs were associated with favorable BCSS (p=0.02). CONCLUSION: Tregs and CD8+ TILs are higher in early-stage TNBCs and HER2+ BCs and diminish with progression to advanced stages. The findings provide support for immunotherapeutic manipulation of TILs, particularly in early stages of these BC subtypes. The evaluation methodology can be easily implemented for standardization of immunohistochemically-detected TILs.