Future Perspectives of Personalized Oncology

Based upon an individual’s molecular make-up, personalized molecular medicine provides information regarding the origin of disease, its treatment and progression, while personalized molecular pharmacology advises on drug prescription and patient response to it, thus ensuring drug effectiveness and preventing drug toxicity or lack of response. Interindividual differences in drug responses are mostly due to structural variation in parts of genome, e.g. in genes participating in drug metabolism, transport or targeting. However, a wide variety of diseases and accompanying health conditions, including patient’s therapy or drug response, also have epigenetic or epigenomic etiology. High priority for personalized oncologic research stems from inter/intraindividual tumor heterogeneity provoked by gradual acquisition of multiple random, or programmed mutations and rearrangements as well as epigenetic alterations or by stochastic fluctuations in cell components, all in tight feedback interaction with tumor’s environmental or therapy conditions. Natural selection subsequently shapes inter/intraindividual tumor heterogeneity by promoting clonal expansion of cells that have acquired advantageous mutations for tumor population. Hence, the main rationale of personalized molecular oncology should focus on treating disease by relying on relevant structure and state of patient’s whole molecular network (genome/transcriptome/RNome/proteome/metabolome/metabonome) in interaction with its unique environmental conditions, thus implying right therapy for the right patient at the right dose and time. The future of personalized oncology should therefore rely on the methods of systems biology applied in cytology and pathology in order to develop and utilize the efficient and effective diagnostic, prognostic and predictive biomarkers, consequently providing the molecular information on tumor origin, its potential for metastasis, adequate therapy, tumor specific therapy responsiveness, and the probability of its recurrence

Nove psihoaktivne tvari u uzorcima urina prikupljenima tijekom Ultra Europe festivala u Splitu

We believe that analysing pooled urine samples for recreational drugs used at mass events can provide useful information about trends in drug use. An opportunity arose with the Ultra Europe music festival, which is attended by more than 150,000 people from over 150 countries every year. We analysed 30 pooled urine samples collected from portable chemical toilets located at or close to the Ultra Europe music festival venue in Split, Croatia in 2016–2018 to detect the presence of classic and new psychoactive substances (NPS). Four urine samples collected in 2016 were from a toilet without added chemicals (otherwise used to kill the smell) while the remaining samples were collected from toilets with added chemicals. Samples were qualitatively analysed with gas chromatography-mass spectrometry (GC/MS) using the fullscan mode. Data were compared with the Wiley mass spectral library of designer drugs and our in-house library containing about 1000 compounds and metabolites. We identified forty-six different substances and metabolites, 26 of which were classic substances/metabolites, mostly from the stimulants group, while 20 were NPS. In the NPS group, most of them were phenethylamines and cathinones. The variety of substances was the highest on the first day of the festival regardless of the year, but 2018 showed a significant drop compared to the previous two years. The results of our study revealed a stable trend of classic drug consumption, while NPS trend changed from one year to another.Analiza uzoraka urina prikupljenih iz prijenosnih kemijskih zahoda korisna je u otkrivanju droga za rekreaciju koje su se koristile tijekom Ultra Europe glazbenoga festivala. Zbog prisutnosti više od 150 000 ljudi iz više od 150 svjetskih zemalja, moguće je dobiti više informacija o svjetskom trendu NPS-a. Analiza uzoraka urina koji su se prikupljali od 2016. do 2018. tijekom glazbenoga festivala Ultra Europe u Splitu provedena je radi otkrivanja prisutnosti novih psihoaktivnih tvari (NPS). Analizirano je trideset uzoraka urina prikupljenih iz prijenosnih kemijskih zahoda smještenih na mjestu festivala ili blizu njega. U 2016. godini prikupljeno je osam uzoraka urina iz dvaju zahoda koji su bili smješteni na dvama različitim mjestima, i to jedan s dodanim kemikalijama, a drugi bez njih. U 2017. godini prikupljeno je deset uzoraka urina iz triju zahoda na trima mjestima, svi iz zahoda s dodanim kemikalijama. U 2018. godini prikupljeno je dvanaest uzoraka urina iz devet prijenosnih kemijskih zahoda na četirima mjestima, svi s dodatkom kemikalija. Uzorci su kvalitativno analizirani tehnikom plinske kromatografije-spektrometrije masa (GC/MS) korištenjem načina ukupnog skeniranja. Podatci su uspoređeni s bazom spektara masa Wiley (DD2015), kao i s vlastitom bibliotekom koja sadržava oko 1000 spojeva i metabolita. U analiziranim uzorcima pronađeno je 46 različitih tvari i metabolita, od kojih 26 klasičnih tvari/metabolita, uglavnom iz skupine stimulansa, te 20 tvari iz skupine novih psihoaktivnih tvari (NPS). U skupini NPS-a najviše ih je bilo iz skupina fenetilamina i katinona. U svakoj promatranoj godini, prvoga festivalskog dana otkriveno je više tvari nego u ostalim danima. Suprotno 2016. i 2017. godini, u 2018. godini broj otkrivenih tvari bio je znatno manji. Rezultati našeg istraživanja pokazali su stabilnost konzumacije klasičnih droga, ali su se trendovi NPS-a mijenjali

Polimorfizmi gena ABCB1, CYP2B6 i CYP3A4 ne utječu na metadonsku terapiju održavanja u bolesnika s HCV-om

The aim of this study was to determine the influence of ABCB1, CYP2B6, and CYP3A4 genetic polymorphisms on methadone metabolism in patients with hepatitis C virus (HCV) undergoing methadone maintenance treatment (MMT).The study included 35 participants undergoing MMT, who were divided in three groups: HCV-positive (N=12), HCVnegative (N=16), and HCV clinical remission (CR) (N=7). The concentrations of methadone and its main metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) were determined with gas chromatography-mass spectrometry. The patients were genotyped for ABCB1 rs1045642, CYP2B6 rs3745274, CYP3A4 rs2242480, and CYP3A4 rs2740574 polymorphisms. Differences between single nucleotide polymorphism (SNP) genotypes and methadone-to-EDDP ratio were analysed with one-way ANOVA, which showed no significant difference between the genes (p=0.3772 for ABCB1 rs1045642, p=0.6909 for CYP2B6 rs3745274, and p=0.6533 for CYP3A4 rs2242480). None of the four analysed SNP genotypes correlated with methadone-to-EDDP concentration ratio. A major influence on it in hepatitis C-positive patients turned out to be the stage of liver damage.Cilj ovoga istraživanja bio je utvrditi utjecaj polimorfizama gena ABCB1, CYP2B6 i CYP3A4 na metabolizam metadona u bolesnika/ovisnika s pozitivnim nalazom virusa hepatitisa C (HCV) na metadonskoj terapiji održavanja. Istraživanje je provedeno na uzorku od 35 sudionika na metadonskoj terapiji održavanja, podijeljenih u sljedeće skupine: HCV pozitivni (N=12), HCV negativni (N=16) i oni s kliničkom remisijom HCV-a (CR) (N=7). Koncentracije metadona i njegova glavnog metabolita 2-etiliden-1,5-dimetil-3,3-difenilpirolidina (EDDP) utvrđene su plinskom kromatografijom – masenom spektrometrijom. U sudionika su analizirani genski polimorfizmi ABCB1 rs1045642, CYP2B6 rs3745274, CYP3A4 rs2242480 i CYP3A4 rs2740574. Jednosmjerna analiza varijance (engl. one-way ANOVA) nije pokazala statistički značajne razlike između genotipova jednonukleotidnih polimorfizama (engl. single-nucleotide polymorphism, krat. SNP) u omjeru koncentracije metadona i EDDP-a (p=0,3772 za ABCB1 rs1045642; p=0,6909 za CYP2B6 rs3745274 F=0,374 i p=0,6533 za CYP3A4 rs2242480). Nijedan od četiriju analiziranih SNP genotipova nije korelirao s omjerom koncentracije metadona i EDDP-a. U bolesnika/ovisnika koji su bili pozitivni na HCV na taj je omjer ponajviše utjecao stupanj oštećenja jetre

Nasljedni karcinom dojke i jajnika - iskustva Kliničkog bolničkog centra Split

Aim: To investigate the clinical and pathohistological tumor characteristics, treatment, and treatment outcomes in patients with hereditary breast and ovarian cancer who were diagnosed, treated, and monitored at the University Hospital of Split from October 1999 to April 2021. Methods: The data were collected retrospectively from the medical history of 15 patients. They included the patient’s age at diagnosis, family history of malignancies, histological subtype, grade, breast cancer immunophenotype, stage of disease, status and types of BRCA mutations, type of surgical and oncological treatment, the specifics of metachronous bilateral breast cancers, the specifics of synchronous breast and ovarian cancers, and the outcome of treatment through overall survival (OS). Results: The median age of patients at the time of diagnosis of breast cancer was 53 years, and for ovarian cancer it was 56 years. A positive family history was confirmed in 13 patients (87%). All ovarian cancer patients had a high-grade serous histologic type, most often diagnosed in FIGO stages III and IV. Breast cancers were most commonly diagnosed in stages IA and IIA, with equally represented triple-negative and luminal immunophenotypes. The most common mutation was BRCA1 c.5266dup. The median OS of our patients was not reached. Conclusion: The clinical features of patients, pathohistological characteristics of tumors, and treatment outcomes in our study population are comparable with reports in the literature, respecting the specifics of different nations and races.Cilj: Istražiti kliničke osobitosti, patohistološke karakteristike tumora, način i ishode liječenja bolesnica s nasljednim karcinomom dojke i jajnika koje su dijagnosticirane, liječene i praćene u Kliničkom bolničkom centru Split od listopada 1999. do travnja 2021. godine. Metode: Podatci su prikupljeni retrospektivno iz povijesti bolesti 15 bolesnica. Uključivali su dob bolesnica kod dijagnoze bolesti, obiteljsku anamnezu za zloćudne bolesti, histološki podtip, gradus, imunofenotip karcinoma dojke, stadij bolesti, status i tip BRCA mutacija, osobitosti kirurškog i onkološkog liječenja, specifičnosti metakrono nastalih bilateralnih karcinoma dojke, specifičnosti sinkrono nastalih karcinoma dojke i jajnika te ishod liječenja kroz ukupno preživljenje. Rezultati: Medijan dobi bolesnica u trenutku dijagnoze raka dojke bio je 53 godine, a za karcinom jajnika 56 godina. Pozitivna obiteljska anamneza potvrđena je u 13 (87%) bolesnica. Karcinom jajnika je kod svih bolesnica bio seroznog papilarnog histološkog podtipa visokog gradusa i najčešće dijagnosticiran u FIGO stadiju III i IV. Karcinom dojke je najčešće dijagnosticiran u stadiju IA i IIA, jednake zastupljenosti trostruko negativnog i luminalnog imunofenotipa. Najčešća mutacija je bila BRCA1 c.5266dup. Medijan ukupnog preživljenja naših bolesnica nije dosegnut. Zaključak: Kliničke osobitosti bolesnica, patohistološke karakteristike tumora kao i ishodi liječenja u našoj studijskoj populaciji su usporedivi s izvješćima iz literature, respektirajući specifičnosti različitih naroda i rasa

Hereditary breast and ovarian cancer – University Hospital of Split experiences

Aim: To investigate the clinical and pathohistological tumor characteristics, treatment, and treatment outcomes in patients with hereditary breast and ovarian cancer who were diagnosed, treated, and monitored at the University Hospital of Split from October 1999 to April 2021. Methods: The data were collected retrospectively from the medical history of 15 patients. They included the patient’s age at diagnosis, family history of malignancies, histological subtype, grade, breast cancer immunophenotype, stage of disease, status and types of BRCA mutations, type of surgical and oncological treatment, the specifics of metachronous bilateral breast cancers, the specifics of synchronous breast and ovarian cancers, and the outcome of treatment through overall survival (OS). Results: The median age of patients at the time of diagnosis of breast cancer was 53 years, and for ovarian cancer it was 56 years. A positive family history was confirmed in 13 patients (87%). All ovarian cancer patients had a high-grade serous histologic type, most often diagnosed in FIGO stages III and IV. Breast cancers were most commonly diagnosed in stages IA and IIA, with equally represented triple-negative and luminal immunophenotypes. The most common mutation was BRCA1 c.5266dup. The median OS of our patients was not reached. Conclusion: The clinical features of patients, pathohistological characteristics of tumors, and treatment outcomes in our study population are comparable with reports in the literature, respecting the specifics of different nations and races

Clinicopathological Characteristics of BRAF V600E Mutated Melanomas in the Dalmatian Region of Croatia

A high proportion of cutaneous melanomas harbor activating mutations of the BRAF or NRAS genes, which are components of mitogen-activated protein kinase (MAPK) signal transduction pathway. The importance of BRAF V600E mutation in melanoma is not only related to the possibility of the administration of the targeted therapy, but also to the fact that BRAF V600E mutated melanomas have distinct clinicopathological features. We investigated the clini-copathological features of 80 primary skin melanomas with known BRAF V600E mutation status excised in the Dalmatian region of Croatia, with comparison of these features between the mutated and wild-type group. The frequency of BRAF V600E mutation was 47.5%. In comparison with wild-type melanomas, BRAF V600E mutated melanomas were significantly associated with younger age and female sex (P=0.014 and P=0.011, respectively). The mutated melanomas were more often located on the extremities, of a nodular type, ulcerated, and with higher median of mitotic index but without significant difference in comparison with wild-type tumors. There were no differences in the depth of invasion and the presence of lymphovascular invasion, tumor infiltrating lymphocytes, and regression between the investigated groups. The frequency of BRAF V600E mutation in our cohort of primary skin melanomas and the clinicopathological features of mutated tumors were similar to those reported in the literature, except for the higher proportion of women observed in our group with mutation

The impact of IL-6 and IL-28B gene polymorphisms on treatment outcome of chronic hepatitis C infection among intravenous drug users in Croatia

Background Several genes and their single nucleotide polymorphisms (SNPs) are associated with either spontaneous resolution of hepatitis C infection or better treatment-induced viral clearance. We tested a cohort of intravenous drug users (IVDU) diagnosed with chronic hepatitis C virus (HCV) for treatment response and its association with the SNPs in the interleukin-6 (rs1800795-IL6) and the interleukin-28B (rs12979860-IL28B) genes. Methods The study included 110 Croatian IVDU positive for anti-HCV antibody. Genotyping was performed by polymerase chain reaction (PCR) based approach. Patients were treated by standard pegylated-interferon/ribavirin and followed throughout a period of four years, during which sustained virological response (SVR) was determined. All data were analysed with statistical package SPSS 19.0 (IBM Corp, Armonk, NY, USA) and PLINK v1.07 software. Results Patients showed a significantly better response to treatment according to the number of copies of the C allele carried at rs1800795-IL6 (P = 0.034). All but one of the patients with CC genotype achieved SVR (93%), whereas the response rate of patients with GG genotype was 64%. The association of rs1800795-IL6 with SVR status remained significant after further adjustment for patients’ age, fibrosis staging, and viral genotype (OR 2.15, 95% CI 1.16–4.68, P = 0.019). Distributions of allele frequencies at the locus rs12979860-IL28B among the study cohort and the underlying general population were suggestive of a protective effect of CC genotype in acquiring chronic hepatitis C in the Croatian IVDU population. Discussion The rs1800795-IL6 polymorphism is associated with positive response to treatment in IVDU patients positive for HCV infection. A protective role of rs12979860-IL28B CC genotype in acquiring chronic hepatitis C is suggested for Croatian IVDU population

Real-Time PCR Method as Diagnostic Tool for Detection of Periodontal Pathogens in Patients with Periodontitis

The most common type of periodontal disease is chronic periodontitis, an inflammatory condition caused by pathogenic bacteria in subgingival plaque. The aim of our study was the development of a real-time PCR test as a diagnostic tool for the detection and differentiation of five periodontopathogenic bacteria, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, and Treponema denticola, in patients with periodontitis. We compared the results of our in-house method with the micro-IDent® semiquantitative commercially available test based on the PCR hybridization method. DNA was isolated from subgingival plaque samples taken from 50 patients and then analyzed by both methods. Comparing the results of the two methods, they show a specificity of 100% for all bacteria. The sensitivity for A. actinomycetemcomitans was 97.5%, for P. gingivalis 96.88%, and for P. intermedia 95.24%. The sensitivity for Tannerella forsythia and T. denticola was 100%. The Spearman correlation factor of two different measurements was 0.976 for A. actinomycetemcomitans, 0.967 for P. gingivalis, 0.949 for P. intermedia, 0.966 for Tannerella forsythia, and 0.917 for T. denticola. In conclusion, the in-house real-time PCR method developed in our laboratory can provide information about relative amount of five bacterial species present in subgingival plaque in patients with periodontitis. It is likely that such a test could be used in dental diagnostics in assessing the efficacy of any treatment to reduce the bacterial burden

Differences in Plasma Cannabidiol Concentrations in Women and Men: A Randomized, Placebo-Controlled, Crossover Study

The potential therapeutic benefits of cannabidiol (CBD) require further study. Here, we report a triple-blind (participant, investigator, and outcome assessor) placebo-controlled crossover study in which 62 hypertensive volunteers were randomly assigned to receive the recently developed DehydraTECH2.0 CBD formulation or a placebo. This is the first study to have been conducted using the DehydraTECH2.0 CBD formulation over a 12-week study duration. The new formulation’s long-term effects on CBD concentrations in plasma and urine, as well as its metabolites 7-hydroxy-CBD and 7-carboxy-CBD, were analyzed. The results of the plasma concentration ratio for CBD/7-OH-CBD in the third timepoint (after 5 weeks of use) were significantly higher than in the second timepoint (after 2.5 weeks of use; p = 0.043). In the same timepoints in the urine, a significantly higher concentration of 7-COOH-CBD was observed p < 0.001. Differences in CBD concentration were found between men and women. Plasma levels of CBD were still detectable 50 days after the last consumption of the CBD preparations. Significantly higher plasma CBD concentrations occurred in females compared to males, which was potentially related to greater adipose tissue. More research is needed to optimize CBD doses to consider the differential therapeutic benefits in men and women