Vitamin D Safety and Mineral Homeostasis | Hedvika Hatáková
Reevaluating Vitamin D Safety and Its Influence on Serum Calcium, Magnesium
and PTH Dynamics
Authors: Hedvika Hatáková (1), Pavlína Černá (1), Michal Jirásko (1)
Supervisor: Radek Kučera (1)
(1) Department of Pharmacology and Toxicology, Faculty of Medicine in Pilsen, Charles University
State-of-the-Art: Vitamin D deficiency is a global issue linked to hypocalcemia, bone disorders,
and increased risks of autoimmune, malignant, metabolic, cardiovascular, and infectious diseases.
Limited sun exposure, especially above 33° latitude, and low dietary intake contribute to deficiency. Supplementation is widely used for prevention and overall health. However, chronic high doses may cause severe hypercalcemia. The Endocrine Society sets 4000 IU/day as the upper safe limit, while 3800 IU may already induce hypercalcemia in healthy adults. There is still no clear consensus on vitamin D toxicity, and its link to hypercalcemia is under review. This study investigates how vitamin D affects serum calcium, magnesium, and parathyroid hormone (PTH) levels.
Objective: The aim of the study was to reevaluate the safety of vitamin D and its effect on serum
calcium, magnesium and PTH levels
Material and Methods: Sixty-five healthy volunteers p articipated in t his s tudy between O ctober
2022 and April 2023. Vitamin D was administered at high doses: 4000 IU daily for two months, followed by a 30-day washout period, and then 8000 IU daily for another two months. Blood samples were collected at baseline, after each supplementation phase, and 30 days post-supplementation. Serum levels of vitamin D, parathyroid hormone (PTH), calcium, andmagnesium were analyzed.
Results & Discussion: Statistical analysis revealed a highly significant positive correlation between
serum vitamin D and calcium levels (p < 0,0001). However, this increase was modest, and even at the highest vitamin D concentration observed in this study (208.5 nmol/L), serumcalcium levels remained within the physiological range. The study also confirms an inverse relationship between serum vitamin D and magnesium levels.Well known negative correlation (p< 0,0001) was observed between vitamin D and parathyroid hormone (PTH) levels in the studied cohort, indicating that PTH concentrations declined with increasing vitamin D levels. Model predictions suggest that a PTH value ≤ 1.6 mmol/L would occur only at 285.6 nmol/L, which exceeded the highest vitamin D level recorded.
Conclusion: The study confirmed vitamin D toxicity mechanisms, showing increased calcaemia and decreased parathyroid hormone with higher vitamin D levels. However, both calcaemia and parathyroid hormone remained within physiological limits despite high vitamin D doses. Magnesium decline was modest.
Funding: Supported by “Cooperatio” Program, research area Pharmaceutical Sciences, FNPl, 00669806, BBMRI-CZ: CZ.02.1.01/0.0/0.0/16_013/000167 and LM2015089.
Study program: Doctoral study – Medical Pharmacology | Year of study: 2
ID: 1096
MAPK Signaling Pathway Key Driver of MPNST | Elaheh Mosaieby
Comprehensive Methylation Profiling of Malignant Peripheral Nerve Sheath Tumors Reveals Novel Epigenetic Dysregulation in Key Oncogenic Pathways
Author: Elaheh Mosaieby (1), Petr Martinek (2)
Supervisor: Michael Michal (1, 2)
(1) Department of Pathology, Faculty of Medicine in Pilsen, Charles University and University Hospital, Pilsen (2) Bioptická laboratoř, s.r.o., Pilsen
State-of-the-Art: Malignant Peripheral Nerve Sheath Tumors (MPNST) are extremely aggressive tumors with few treatment options and a poor prognosis. Despite advances in understanding their genetic characteristics, the significance of epigenetic modifications in MPNST development remains unknown. Methylation profiling can help identifying new biomarkers and therapeutic targets by revealing epigenetic dysregulation specific to tumor cells.
Objective: The objective of this study is to conduct a comprehensive methylation profiling of Malignant Peripheral Nerve Sheath Tumors (MPNST) in comparison with normal peripheral nerve tissue. The goal is to identify differentially methylated positions (DMPs) and the associated biological pathways. We aim to explore how these epigenetic alterations may influence the molecular mechanisms and functional pathways that drive MPNST progression.
Material and Methods: A total of 78 MPNST samples and 12 matched normal controls using Infinium Human Methylation BeadChip arrays from public databases were analyzed. Differential methylation analysis was conducted using the LIMMA package, resulting in 168,297 significant DMPs (adj. P.Val < 0.05). Enrichment analysis was performed on genes associated with significant DMPs using Gene Ontology (GO) and KEGG pathway databases. Key pathways were validated using network analysis in Cytoscape.
Results & Discussion: The analysis identified significant hypermethylation in tumor suppressor genes and hypomethylation in oncogenes. The top DMPs included cg19541688 (logFC = 0.61, adj. P.Val = 7.5 × 10^-73), which affects the gene RASSF1, a known tumor suppressor. GO enrichment indicated substantial modifications in small GTPase-mediated signal transduction (p.adjust = 1.09 × 10^-11) and regulation of nervous system development (p.adjust = 7.24 × 10^-11). KEGG pathway analysis revealed dysregulation in the MAPK signaling pathway (p.adjust = 6.1 × 10^-6), which is a key driver of MPNST proliferation and survival. Additional pathways, such as focal adhesion and cellular senescence, were considerably enriched, suggesting a comprehensive epigenetic reprogramming that contributes to tumorigenesis.
Conclusion: With the use of this study‘s thorough methylation profile of MPNST, significant epigenetic changes affecting important carcinogenic pathways were found. The identified DMPs and associated pathways present potential biomarkers for early diagnosis and novel therapeutic targets. Our findings lay the groundwork for future functional studies to validate these epigenetic changes and their role in MPNST development.
Study program: Doctoral study – Pathology | Year of study: 4
ID: 1080
Bladder Cancer: NAC Prior to Cystectomy | Kseniia Khomenko
Neoadjuvant Chemotherapy in Muscle-Invasive Urothelial Carcinoma of the Bladder Prior to Radical Cystectomy: A Retrospective Evaluation
Author: Kseniia Khomenko (1)
Supervisor: Tomáš Pitra (1)
(1) Department of Urology, Faculty of Medicine in Pilsen, Charles University and University Hospital, Pilsen
State-of-the-Art: Urothelial carcinoma of the bladder is the seventh most common cancer globally. Radical cystectomy remains the standard treatment for muscle-invasive bladder cancer (MIBC) and for high- and very high-risk non–muscle-invasive bladder cancer (NMIBC). Neoadjuvant chemotherapy (NAC) is a recommended part of treatment according to European Association of Urology (EAU) guidelines. Studies and meta-analyses have shown a survival benefit with NAC, especially with cisplatin-based regimens. The most commonly used protocols are M-VAC (methotrexate, vinblastine, doxorubicin, and cisplatin) and gemcitabine plus cisplatin (GC), though real-world responses vary with tumor biology, patient selection, and regimen choice.
Objective: To evaluate the pathological response and oncological outcomes of patients with MIBC and high- and very high-risk NMIBC treated with neoadjuvant chemotherapy (M-VAC or gemcitabine/cisplatin) followed by radical cystectomy.
Material and Methods: A retrospective review was conducted on 24 patients with histologically confirmed MIBC or high- and very high-risk NMIBC treated between 2018 and 2023 at our clinic. All patients received neoadjuvant chemotherapy with either M-VAC or gemcitabine plus cisplatin prior to undergoing radical cystectomy. Pathological response was assessed on cystectomy specimens and categorized as complete response (ypT0), partial pathological response (≤ypT1), or no pathological response (≥ypT1-2). Follow-up data were collected to assess recurrence and metastasis rates.
Results & Discussion: Between January 2018 and December 2023, 99 patients underwent radical cystectomy for urothelial bladder carcinoma at our department. Of these, 24 received neoadjuvant chemotherapy with either M-VAC or gemcitabine-cisplatin (GC). Twelve achieved complete pathological response (pT0), two had a partial response (≤pT1), and ten showed no pathological downstaging (≥pT2). During follow-up, three patients developed metastases, while data were unavailable for five. Complete responders had significantly improved disease-free survival, supporting the prognostic value of pathological response. While not statistically significant, pT0 rates were higher with M-VAC. Study limitations include small sample size and retrospective design.
Conclusion: NAC before radical cystectomy in MIBC led to a 50% complete and 8.3% partial pathological response. Response correlated with better outcomes, suggesting that, in select cases, post-NAC surveillance may be feasible. Larger studies are needed to guide treatment choices.
Funding: Charles University Prague, Faculty of Medicine in Pilsen (Cooperatio Program, SURG), Institutional Research of the University Hospital Pilsen (FNPl 00669806).
Study program: Doctoral study – Surgery | Year of study: 2
ID: 1115
Anti-androgenic Effect on Prostate Cancer | Roman Viták
Testing boron derivatives of anti-androgenic drugs on prostate cancer cell lines
Author: Roman Viták (1,2)
Supervisor: Radek Kučera (1)
(1) Department of Pharmacology and Toxicology, Faculty of Medicine in Pilsen, Charles University
(2) Department of Pharmaceutical Chemistry and Pharmaceutical Analysis, Faculty of Pharmacy
in Hradec Králové, Charles University
State-of-the-Art: Prostate cancer is one of the most common malignancies in men and its
treatment involves androgen deprivation therapy. Current treatments for prostate cancer include
hormonal therapy, chemotherapy, and targeted biological therapies. Antiandrogens such
as flutamide, bicalutamide, and enzalutamide play a key role in the treatment of advanced
disease, but often lead to the development of resistance. Boron derivatives have become the
subject of intense research in recent years due to their unique pharmacokinetic properties and
ability to selectively interact with target proteins. Studies suggest that modification of antiandrogens
with a boron substitution compound could lead to improved affinity for the androgen
receptor while reducing toxicity to healthy cells.
Objective: The aim of the study was to test the antiproliferative activity of boron derivatives
of nonsteroidal antiandrogens on androgen-dependent and androgen-independent prostate
cancer cell lines. And compare the efficacy and toxicity of the new derivatives with commercially
available antiandrogens.
Material and Methods: We synthesized a series of boron derivatives of nonsteroidal antiandrogens
that were tested on androgen-dependent and androgen-independent prostate cancer
cell lines. Non-carcinogenic cell lines were used as controls. In vitro toxicity was assessed using
standard WST-1 assays, which allow quantification of cell viability based on metabolic activity.
Cells were exposed to test compounds for 24, 48 or 72 hours and IC50 values were determined
by comparison with control drugs from the group of nonsteroidal antiandrogens.
Results & Discussion: Several of the most active newly synthesized structural derivatives of
nonsteroidal antiandrogens showed higher antiproliferative activity against androgen-dependent
prostate cancer lines compared to the standards flutamide and bicalutamide, while at the
same time having lower toxicity against non-carcinogenic lines.
Conclusion: Boron derivatives of antiandrogen drugs show promising antiproliferative effects
on prostate cancer while exhibiting lower toxicity compared to standard drugs. These results
support further research on boron compounds in the field of antiandrogen therapy.
Study program: Doctoral study – Medical Pharmacology | Year of study: 2
ID: 1075