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Comparison of methods for the isolation of RNA from exosome samples of patients with colorectal carcinoma
No full textKolorektalni karcinom (CRC) predstavlja ozbiljan dijagnostički i terapijski problem na globalnoj razini zbog stalno rastuće
prevalencije. Multifaktorska je bolest, specifične etiologije za svakog bolesnika te je potrebna serija genskih mutacija tokom
dužeg razdoblja za razvoj invazivnog karcinoma. Egzosomi koji potječu iz stanica kolorektalnog karcinoma sadrže molekule
miRNA koje se povezuju s lošom prognozom tako što potiču proliferaciju, migraciju i invazivnost tumorskih stanica te
predstavljaju potencijalan biomarker pokazujući značajnu korelaciju s patološkim stanjem i progresijom kolorektalnog
karcinoma. Tekuća biopsija je neinvazivna metoda kojom se mogu izolirati cirkulirajući egzosomi i miRNA iz uzorka krvi što
može voditi velikom napretku u ranoj dijagnostici karcinoma kao i u njegovom praćenju. Cilj diplomskog rada je usporediti
metode Qiagen i ThermoFisher za izolaciju molekula RNA iz egzosoma kod bolesnika s kolorektalnim karcinomom. Početni
je uzorak puna krv iz čije se plazme izolira egzosomalna RNA metodom precipitacije i organske ekstrakcije. Koncentracija
dobivene RNA određuje se spektrofotmetrijski, dok se kvaliteta i integritet određuju metodom kapilarne elektroforeze.
Ekspresija referentnih gena B2M i PPIA određena je metodom Taqman qPCR prema kojoj se određuje relativna ekspresija
ciljnih gena. Također, uspoređuje se ekspresija ciljne miRNA (miR-19a-3p) za čiju se ekspresiju pretpostavlja da bi trebala
biti povišena kod bolesnika s kolorektalnim karcinomom te referentne miRNA miR-103a-3p u odnosu na „spike-in“ miRNA
UniSp6. Iz dobivenih se rezultata može zaključiti kako ne postoji statistički značajna razlika između metoda Qiagen i
ThermoFisher, odnosno metode su jednakovrijedne i može se koristiti bilo koja od dvije metode za određivanje ekspresije
miRNA bez značajne razlike u rezultatima. Unatoč niskoj koncentraciji izolirane RNA, uzorci su dovoljno kvalitetni za
određivanje ekspresije miRNA. Sukladno tome, potvrđuje se prisutnost referentne (miR-103a-3p) i ciljne (miR-19a-3p)
miRNA u svim uzorcima.Colorectal cancer (CRC) represents a serious diagnostic and therapeutic problem at the global level due to its ever-increasing prevalence. It is a multifactorial disease, with a specific etiology for each patient, and a series of gene mutations over a longer period is required for the development of invasive cancer. Exosomes from colorectal cancer cells contain miRNA molecules that are associated with poor prognosis by promoting the proliferation, migration, and invasiveness of tumor cells. Furthermore, they represent a potential biomarker showing a significant correlation with the pathological state and progression of colorectal cancer. Liquid biopsy is a non-invasive method that can isolate circulating exosomes and miRNAs from a blood sample, which could lead to great progress in the early diagnosis of cancer as well as in its monitoring. The aim of this thesis is to compare the Qiagen and ThermoFisher methods for the isolation of RNA molecules from exosomes in patients with colorectal cancer. The initial sample is whole blood where exosomal RNA is isolated by precipitation and organic extraction from plasma exosomes. The concentration of the obtained RNA is determined spectrophotometrically, while the quality and integrity are determined by the capillary electrophoresis method. The expressions of reference genes
B2M and PPIA were determined by the Taqman qPCR method, which determines the relative expression of the target genes.
Additionally, the expression of the target miRNA (miR-19a-3p) is compared, as it is supposed to be elevated in patients with
colorectal cancer and reference miRNA miR-103a-3p relative to “spike-in” miRNA UniSp6. In conclusion, there was no
statistically significant difference between the Qiagen and ThermoFisher methods, therefore, the methods could be
considered equivalent and either of the two can be used to determine miRNA expression without a significant difference in
results. Despite the low concentration of isolated RNA, the samples are of sufficient quality to determine miRNA expression.
Accordingly, the presence of reference (miR-103a-3p) and target (miR-19a-3p) miRNA is confirmed within all samples
Razvoj i terapijski potencijal bispecifičnih protutijela
No full textCilj istraživanja
Cilj ovog specijalističkog rada je opisati svojstva bispecifičnih protutijela odobrenih za kliničku uporabu i onih u kliničkim istraživanjima, analizirati metode njihove proizvodnje te dati uvid u terapijske mogućnosti ovih protutijela.
Materijali i metode
Rad daje pregled razvoja bispecifičnih protutijela te njihove terapijske primjene. Pregledom znanstvene literature detaljnije su opisane metode za ispravno povezivanje lakih i teških lanaca. Opisana su do sad odobrena bispecifična protutijela te njihova terapijska primjena. Navedena su i protutijela koja se nalaze u različitim fazama razvoja.
Rezultati
Bispecifična protutijela kombiniraju dva ili više elemenata, koji prepoznaju antigene, u jednu cjelinu sa sposobnošću vezanja na dva ili više ciljeva. Djelovanjem na više meta istovremeno povećava se učinkovitost terapije te smanjuje potreba za učestalom primjenom lijeka. Kako bi se povezali različiti teški i laki lanci u jednu molekulu, razvijene su nove metode. Neke od tih metoda su kvadroma tehnologija, metoda "dugme u rupu", zajednički laki lanac, CrossMAb tehnologija, SEED, WuxyBody i dr. Zahvaljujući ovim metodama danas je za kliničku primjenu odobreno sedam bispecifičnih protutijela, a osim njih u pretkliničkim ispitivanjima nalazi se više od 180 bispecifičnih protutijela, dok ih je u različitim fazama kliničkih ispitivanja više od 50.
Zaključak
Razvojem biotehnologije i genetičkog inženjerstva otkrivene su nove metode ispravnog povezivanja lakih i teških lanaca koje su omogućile razvoj funkcionalnih bispecifičnih protutijela koja se koriste u kliničkoj praksi. Bispecifična protutijela pokazala su znatan napredak u učinkovitosti u odnosu na dosadašnju terapiju. Napredak je vidljiv u ishodima liječenja, vremenu potrebnom za postizanje terapijskog odgovora te u vremenu do pogoršanja bolesti. Osim boljih rezultata liječenja, bispecifična protutijela su se pokazala i kao isplativija nego dosadašnja terapija.Objective
The aim of this expert thesis is to describe the properties of bispecific antibodies approved for clinical use and those in clinical trials, to analyse the methods of their production and to provide insight into the therapeutic possibilities of such antibodies.
Materials and methods
The thesis provides an overview of the development of bispecific antibodies and their therapeutic applications. The review of the scientific literature describes in more detail the methods for the correct linking of light and heavy chains. The thesis describes the approved bispecific antibodies and their therapeutic use, as well as those that are in different stages of development.
Results
Bispecific antibodies combine two or more antigen-recognizing elements into a single molecule with the ability to link to two or more targets. By acting on multiple targets simultaneously, the effectiveness of the therapy increases and the need for frequent drug administration is reduced. In order to link different heavy and light chains into one molecule, new methods have been developed. Some of these methods are quadroma technology, buttonhole technique, common light chain, CrossMAb technology, SEED, WuxyBody, etc. Thanks to these methods, today there are seven bispecific antibodies approved for clinical use; in addition to these, there are more than 180 bispecific antibodies approved in preclinical trials, as well as more than 50 in different stages of clinical trials.
Conclusion
The development of biotechnology and genetic engineering has facilitated the discovery of new methods for correct linking of light and heavy chains that have enabled the development of functional bispecific antibodies used in clinical practice. Bispecific antibodies have shown significant improvement in efficacy compared to previous therapy. Progress is evident in treatment outcomes, in the time it takes to achieve a therapeutic response, and in the time to disease worsening. In addition to better treatment results, bispecific antibodies have also been shown to be more cost-effective than the previous therapy
N-glycome and genome in comprehension of type 1 diabetes
No full textN-glikozilacija proteina plazme povezana je s komplikacijama šećerne bolesti tipa 1, dok je uloga ove enzimske modifikacije proteina u nastanku same bolesti neistražena. S razvojem šećerne bolesti tipa 1 do sada je povezano više od 50 gena, među kojima su i glikoziltransferazni geni. Glavni ciljevi ovog istraživanja su identifikacija N-glikana ukupnih proteina plazme i IgG-a karakterističnih za ranu fazu šećerne bolesti tipa 1 te gena kojima su N-glikani ukupnih plazmatskih proteina i IgG-a djece i adolescenata sa šećernom bolesti tipa 1 regulirani. Istraživanje je uključilo 1917 djece i adolescenata (0,6 – 19,1 godina) čiji su uzorci krvne plazme prikupljeni unutar tri mjeseca od dijagnoze šećerne bolesti tipa 1 kroz Danski registar dječjeg i adolescentnog dijabetesa, kao i 265 zdravih srodnika. N-glikani ukupnih proteina plazme i IgG-a analizirani su upotrebom tekućinske kromatografije, dok je genotipizacija 1105 djece i adolescenata sa šećernom bolesti tipa 1 provedena pomoću komercijalnog čipa (Immunochip, Illumina Infinium) koji sadrži 183 546 polimorfizama pojedinačnih nukleotida važnih za razvoj bolesti posredovanih imunološkim sustavom. Nakon navedenih analiza provedena je genomska asocijacijska studija. Ovo istraživanje pokazalo je da je šećerna bolest tipa 1 povezana s porastom oligomanoznih N-glikana i N-glikana s račvajućim N-acetilglukozaminom te padom monogalaktoziliranih N-glikana ukupnih plazmatskih proteina i IgG-a, kao i s porastom disijaliniziranih N-glikana IgG-a. Modeli temeljeni na N-glikanima ukupnih plazmatskih proteina i IgG-a su pokazali jako dobru diskriminacijsku moć između djece i adolescenata sa šećernom bolesti tipa 1 i njihovih zdravih srodnika (AUC > 0,9). Tri prethodno identificirana N-glikozilacijska lokusa (MGAT3, MGAT5 i ST6GAL1) povezana su s razinama N-glikana u ovom istraživanju, kao i C3 lokus na kromosomu 19 koji do sada nije bio povezan s N-glikozilacijom. C3 kodira glavni protein sustava komplementa te su dva C3 SNP-a (sinonimni i nesinonimni) povezana s razinama Man9 glikana ukupnih plazmatskih proteina. S obzirom da se takav Man9 glikan nalazi na C3 domeni uključenoj u interakciju patogena i C3 proteina, promjene u razinama Man9 mogle bi potencijalno interferirati s aktivacijom sustava komplementa u šećernoj bolesti tipa 1. Navedena otkrića pružaju temelje za daljnja istraživanja mehanizama koji reguliraju N-glikozilaciju u šećernoj bolesti tipa 1.Alterations of plasma N-glycosylation have mostly been studied in association to diabetes complications, whereas the role of these changes in type 1 diabetes onset is largely unknown. The present study was undertaken to determine plasma N-glycans representative of the type 1 diabetes onset and to gain the knowledge of genes regulating these changes. Plasma and IgG N-glycans were chromatographically analysed in a recent-onset 1917 type 1 diabetes cases (0.6-19.1 years) and their 244 unaffected siblings whose plasma samples were collected through the Danish Registry of Childhood and Adolescent Diabetes. This study identified an increase in the proportion of plasma and IgG high-mannose and bisecting GlcNAc structures, a decrease in monogalactosylation, and an increase in IgG disialylation characteristic of the early phase of type 1 diabetes. A notable discriminative power between children with type 1 diabetes and their healthy siblings was yielded with models including age, gender, and N-glycans with AUCs of 0.915 and 0.869 for addition of plasma and IgG N-glycans, respectively. Increasing the efficacy of models to determine individuals at risk of disease development would be a considerable asset for type 1 diabetes prevention trials. 183,546 single nucleotide polymorphisms (SNPs) on the commercially available Immunochip that covers all major autoimmune diseases were determined for 1105 study participants and genetic association study on plasma and IgG N-glycome data in type 1 diabetes was conducted. Three of the previously established N-glycosylation loci (MGAT3, MGAT5, and ST6GAL1) were significantly associated with N-glycosylation in the discovery cohort, as well as a novel locus on chromosome 19 encoding the C3, the pivotal protein of the complement activation pathway. This study of type 1 diabetes cases revealed a new genetic association of the C3 gene with N-glycosylation, namely of two C3 SNPs (synonymous and non- synonymous) with the Man9 glycan of total plasma proteins. Since such Man9 glycan is located on the domain implicated in pathogen binding on the C3 protein, the identified N-glycan alterations might potentially be implicated in the complement activation in type 1 diabetes. These findings offer starting points for further functional follow-up studies of mechanisms regulating N-glycosylation in type 1 diabetes
Investigation of the presence of sterigmatocystin in beer by thin-layer chromatography
No full textSterigmatocistin (STC) je citotoksični i genotoksični mikotoksin kojeg dominantno proizvode
plijesni roda Aspergillus iz serije Versicolores. Ove su plijesni široko rasprostranjene u prirodi, ali i
unutar zatvorenih prostora, u koje spadaju i oni u kojima se skladišti hrana i sirovine za njenu
proizvodnju, što upućuje na mogućnost kontaminacije te hrane i sirovina ovim mikotoksinom.
Kontaminacija STC-om do sada je utvrđena u žitaricama, zrnima kave, začinima, orašastim
plodovima, u hrani za životinje te u siru, dok o pojavnosti ovog mikotoksina u pivu nema mnogo
literaturnih podataka. S obzirom na visoku potrošnju ovog popularnog pića u Hrvatskoj i u svijetu
može se pretpostaviti nepovoljan učinak na zdravlje izloženih ljudi uslijed izloženosti ovom
mikotoksinu. Kako bi se detektirala pojavnost STC-a u pivu prikupljeno je 58 uzoraka piva s
hrvatskog tržišta. Ekstrakcija je provedena kombiniranjem ekstrakcije acetonitrilom uz isoljavanje
te pročišćavanje pomoću SPE kolona. Tako pripremljeni ekstrakti analizirani su metodom
tankoslojne kromatografije uz limit detekcije od 1,3 μg/ml. Analizom nije dokazana prisutnost STCa niti u jednom ispitanom uzorku. Međutim u 21/58 uzoraka utvrđeno je prisustvo analita koji po
boji fluorescencije i položaju zadržavanja na TLC ploči upućuju na moguće prisustvo metabolita
STC-a. Analiza istih ekstrakata osjetljivijim kromatografskim tehnikama omogućit će precizniju
interpretaciju ovih opažanja te omogućiti detekciju nižih koncentracija STC-a u uzorcima.Sterigmatocystin (STC) is a cytotoxic and genotoxic mycotoxin predominantly produced by molds
of the genus Aspergillus, section Versicolores. These molds are widespread in nature, but also in
indoor spaces, including those that store food and raw materials for its production, which indicates
the possibility of contamination of food and raw materials with this mycotoxin. STC contamination
has so far been identified in cereals, coffee beans, spices, nuts, animal feed, and cheese, while there
isn't much literature data on the occurrence of this mycotoxin in beer. Given the high consumption
of this popular drink in Croatia and the world, we can expect a negative effect on the health of
exposed people, due to exposure to this mycotoxin. To detect the occurrence of STC in beer, 58
samples of beer were collected from the Croatian market. Extraction was performed by combining
acetonitrile extraction with salting, and purification using SPE columns. Thus prepared extracts were
analyzed by thin-layer chromatography with a limit of detection of 1,3 μg/mL. The STC was not
detected in any of the tested samples. Taking into account resemblence in fluorescence and the spot
on chromatograms 21/58 samples may be related to the metabolates of STC. Further analysis of the
same extracts by application of more sensitive chromatographic techniques will allow a more
accurate interpretation of these observations and allow the detection of lower STC concentrations in
the samples
Kinetics of quercetin and DPPH radical reaction
No full textKvercetin spada u podskupinu flavonola te mu se pripisuje niz bioloških učinaka: antioksidativni, antikarcinogeni,
antialergijski, protuupalni i drugi. Reakcije flavonoida i DPPH• uobičajeno se prate tako što se mjeri smanjenje
apsorbancije na valnoj duljini koja odgovara maksimumu DPPH•. Prema novijoj literaturi (Foti i sur., 2011) smanjenje
apsorbancije u vidljivom području spektra nije povezano izravno s redukcijom DPPH•, te reakcijska otopina, osim
početno, ne sadrži DPPH•. U ovom je diplomskom radu ispitana kinetika reakcije kvercetina i DPPH• radikala u smjesi
otapala dioksan-voda (0,99:0,01 v/v) te u metanolu. Na temelju izmjerene ovisnosti konstante brzine reakcije pseudoprvog reda o koncentraciji kvercetina određena je konstanta brzine reakcije kvercetina i DPPH• u dioksanu, koja iznosi
4,38 mol-1dm3 s -1. Konstante brzine određene na 525 nm i 620 nm su približno iste, iako su odstupanja u izmjerenim
vrijednostima na 525 nm znatno veća u odnosu na 620 nm. Određena je konstanta brzine reakcije kvercetina i DPPH• u
metanolu čija vrijednost iznosi 469 mol-1dm3 s -1. U metanolu nisu utvrđena odstupanja u izmjerenim vrijednostima kao
u dioksanu. Na temelju izmjerene ovisnosti konstante brzine reakcije pseudo-prvog reda o koncentraciji kvercetina u
otapalu dioksan-D2O određena je konstanta brzine reakcije kD koja iznosi 0,568 mol-1dm3 s -1 te kinetički izotopni efekt
koji iznosi 7,71. Ovi podaci nisu prethodno pronađeni u literaturi te su po prvi puta određeni u ovom diplomskom radu.
Za usporedbu su provedena i kinetička i HPLC mjerenja reakcije katehina i DPPH•. Određena je konstanta brzine
reakcije katehina i DPPH• u metanolu koja iznosi 2,70 mol-1dm3 s -1. HPLC analizom je potvrđeno da je tijekom cijelog
perioda u kojemu se prati smanjenje apsorbancije u vidljivom dijelu spektra u reakcijskoj otopini prisutan DPPH• (u
reakciji kvercetina i DPPH• te u reakciji katehina i DPPH•).Quercetin is a flavonol, a subclass of flavonoids that has numerous pharmacological uses due to its antioxidant,
anticarcinogenic, antiallergic and anti-inflammatory properties. The reaction of flavonoids and DPPH• are commonly
monitored by measuring the decrease in absorption at the wavelength corresponding to the maximum DPPH•. According
to new literature (Foti et al., 2011), the decrease in absorption in the visible region of the spectrum is not directly related
to the reduction of DPPH • and the reaction solution, except initially, does not contain the DPPH radical. In this thesis,
the reaction kinetics of quercetin and DPPH• in a solvent mixture of dioxane-water (0,99:0,01 v/v) and methanol were
investigated. Based on the measured dependence of the pseudo-first order reaction rate constant on the quercetin
concentration, the reaction rate constant of quercetin and DPPH • in dioxane was determined to be 4,38 mol-1dm3s-1.
The rate constants determined at 525 nm and 620 nm are approximately the same, although the deviations in the
measured values at 525 nm are significantly larger compared to 620 nm. The reaction rate constant of quercetin and
DPPH• in methanol with a value of 469 mol-1dm3s-1 was determined. No deviations in the measured values were found
in methanol as in dioxane. Based on the measured dependence of the pseudo-first order reaction rate constant on the
quercetin concentration in the solvent dioxane-D2O, a reaction rate constant kD of 0,568 mol-1dm3s-1 and a kinetic
isotopic effect of 7,71 were determined. These data were not previously found in the literature and were determined for
the first time in this thesis. Both kinetic and HPLC measurements of catechin and DPPH • reactions were performed for
comparison. The rate constant of the reaction of catechins and DPPH• in methanol was determined to be 2,70 mol1dm3s-1. HPLC analysis confirmed that DPPH• (in the reaction of quercetin and DPPH • and the reaction catechin and
DPPH •) was present in the reaction solution during the whole period in which the decrease in absorption was monitored
in the visible part of the spectrum
Biological therapy and small molecules in the treatment of moderate to severe plaque psoriasis
No full textPsorijaza je kompleksna, imunološki posredovana upalna bolest koja se javlja u genetski podložnih
pojedinaca, a globalno pogađa približno 2,2% populacije. Tipično se manifestira na koži i
zglobovima no također se povezuje s brojnim komorbiditetnim bolestima ostavljajući negativne
posljedice na kvalitetu života bolesnika, posebice onih s umjerenim do teškim oblikom. S obzirom
na prisustvo komorbiditeta, nuspojave liječenja, različite preferencije te često neadherenciju
pacijenata, liječenje psorijaze još uvijek predstavlja veliki izazov. U većini slučajeva, terapija
uključuje primjenu lokalnih sredstava, fototerapije ili konvencionalnih sistemskih lijekova no
napredak u razumijevanju patofiziologije psorijaze doveo je do razvoja novih, visoko učinkovitih
ciljanih tretmana. Biološki lijekove i male sintetske molekule revolucionirale su liječenje psorijaze
zahvaljujući izvrsnoj djelotvornosti, povoljnom sigurnosnom profilu te poboljšanoj adherenciji
pacijenata. Iako ne pokazuju učinkovitost bioloških lijekova, male molekule posjeduju određene
prednosti poput peroralne i topikalne primjene, manjeg rizika od teških nuspojava te povoljnije
cijene. U biološke lijekove i male molekule odobrene za liječenje plak psorijaze ubrajaju se:
antagonisti TNF-α, inhibitori IL-17, inhibitori IL-12 i IL-23, inhibitori IL-23, inhibitor PDE-4
(apremilast) te ester fumaratne kiseline (dimetil fumarat). Brojni drugi biološki lijekovi i male
molekule koje su trenutno u fazama razvoja pokazuju obećavajuću učinkovitost te potencijal da
postanu sigurnije alternative trenutno dostupnih terapija.Psoriasis is a complex, immune-mediated inflammatory disease that occurs in genetically
susceptible individuals and affects approximately 2.2% of the world's population. It typically
manifests on the skin and joints but is also associated with a number of comorbid diseases leaving
negative consequences on the quality of life of patients, especially those with moderate to severe
form. Given the presence of comorbidities, treatment side effects, different patient preferences and
frequent noncompliance, treating psoriasis remains a major challenge. In most cases, treatment
involves the use of topical agents, phototherapy and conventional systemic medications but advances
in understanding the pathophysiology of psoriasis have led to the development of new highly
effective targeted treatments. Biologics and small synthetic molecules have revolutionized the
treatment of psoriasis thanks to their excellent efficacy, favorable safety profile and improved patient
adherence. Although they do not show the effectiveness of biologics, small molecules have certain
advantages such as oral and topical administration, lower risk of severe side effects and more
favorable price. Biologics and small molecules approved for the treatment of plaque psoriasis
include: TNF-α antagonists, IL-17 inhibitors, IL-12 and IL-23 inhibitors, IL-23 inhibitors, PDE-4
inhibitor (apremilast) and fumaric acid ester (dimethyl fumarate). Numerous other biologics and
small molecules currently under development show promising efficacy and the potential to become
safer alternatives to currently available therapies
Fitoterapijski potencijal crnog papra (Piper nigrum L.)
No full textCilj istraživanja
Cilj ovog specijalističkog rada je pregled podataka dosadašnjih znanstvenih istraživanja o crnom papru (Piper nigrum L.).
Materijali i metode
Istraživanja u okviru ovoga specijalističkog rada su teorijskog karaktera, a uključuju pregled dostupne literature o dosadašnjim nekliničkim i kliničkim istraživanjima crnog papra. U pretraživanju su korištene dostupne elektronske bibliografske baze podataka Current Contents, ScienceDirect, Scopus, PubMed i Medline.
Rezultati
Usitnjeni plod crnog papra je širom svijeta poznat kao začin, a ima i višestoljetnu tradiciju medicinske primjene. Fitokemijskim istraživanjima ustanovljeno je da sadrži fitosterole, triterpene, fenolne spojeve, značajne koncentracije flavonoida (katehina, miricetina i kvercetina), karotenoida (luteina i β-karotena) minerala (kalij, kalcij, magnezij i fosfor) i vitamina C. Crni papar je snažan antioksidans te ima dokazano protupalno, imunomodulacijsko, antitumorsko, citotoksično, antimikrobno i analgetsko djelovanje. Farmakološki učinak zabilježen je kod liječenja astme, prehlade i kašlja, hipertenzije, pretilosti, bolesti probavnih organa, živčanog sustava i štitnjače. Dosadašnja klinička istraživanja ploda crnog papra ograničena su na povećanje terapijskog učinka lijekova i hranjivih tvari jer djeluje kao pojačivač njihove bioraspoloživosti.
Zaključci
Fitoterapijski potencijal crnog papra treba potvrditi daljnjim studijama jer dosadašnja klinička istraživanja ne pružaju dovoljnu razinu dokaza o terapijskoj učinkovitosti crnog papra.Objectives
The aim of this specialist paper is to review the data of previous scientific research on black pepper (Piper nigrum L.).
Materials and methods
The research within this specialist paper is of a theoretical nature, and includes a review of the available literature on previous non-clinical and clinical research on black pepper. Available electronic bibliographic databases Current Contents, ScienceDirect, Scopus, PubMed and Medline were used in the search.
Results
Chopped black pepper is known worldwide as a spice, and has a centuries-old tradition of medical use. Phytochemical studies have shown that it contains phytosterols, triterpenes, phenolic compounds, significant concentrations of flavonoids (catechins, myricetin and quercetin), carotenoids (lutein and β-carotene) minerals (potassium, calcium, magnesium and phosphorus) and vitamin C. Black pepper is a strong. antioxidant and has proven anti-inflammatory, immunomodulatory, antitumor, cytotoxic, antimicrobial and analgesic effects. Pharmacological effect has been reported in the treatment of asthma, colds and coughs, hypertension, obesity, diseases of the digestive organs, nervous system and thyroid gland. Previous clinical trials of black pepper fruit have been limited to increasing the therapeutic effect of drugs and nutrients because it acts as an enhancer of their bioavailability.
Conclusions
The phytotherapeutic potential of black pepper should be confirmed by further studies because previous clinical studies do not provide a sufficient level of evidence on the therapeutic efficacy of black pepper
Farmakogenetika i interakcije direktnih oralnih antikoagulantnih lijekova
No full textDirektni oralni antikoagulansi (DOAK) posljednjih su godina pokazali rastući trend propisivanja zbog povoljne farmakokinetike i farmakodinamike bez potrebe za rutinskim nadzorom koagulacije. Svoj antikoagulantni učinak ostvaruju izravnom inhibicijom trombina (dabigatran) ili izravnom inhibicijom aktiviranog čimbenika X (rivaroksaban, apiksaban i edoksaban). Iako DOAK-i stupaju u manje interakcija s drugim lijekovima u usporedbi s varfarinom, kombinacija DOAK-a i mnogih lijekova zahtijeva oprez te u nekim slučajevima i klinički nadzor. Nedavne studije dokumentirale su interindividualnu varijabilnost u razinama DOAK-a u plazmi i odgovoru pacijenta na lijek. Iako klinički i biokemijski čimbenici mogu dovesti do interindividualnih varijabilnosti prisustvo genskih varijanti ili interakcije lijekova mogu doprinijeti tim razlikama. Potrebno je razumjeti ulogu farmakogenomike prvenstveno polimorfizam gena metaboličkih enzima i prijenosnika lijekova u interindividualnoj varijabilnosti četiri najčešće propisivana DOAK-a.
Cilj istraživanja: Pregledno prikazati kako farmakogenetička predispozicija, prvenstveno polimorfizmi gena CYP3A4/5 i CES1, koji kodiraju metaboličke enzime, te ABCB1 i ABCG2, koji kodiraju transportne proteine, uz interakcije lijekova u politerapiji mogu doprinijeti interindividualnoj varijabilnost razina DOAK-a u plazmi, te učinkovitosti i sigurnosti njihove primjene. Materijali i metode: Prilikom izrade ovog rada provedena je opsežna pretraga nedavno objavljenih izvornih i preglednih znanstvenih radova, kliničkih ispitivanja, in vitro studija, terapijskih smjernica i publikacija stručnih društava. Naglasak je na polimorfizmima farmakogena bitnih za farmakokinetiku DOAK-a: gena CYP3A4/5 i CES1, za enzime uključene u biotransformaciju, te gena ABCB1 i ABCG2, za transportere lijekova i njihovom doprinosu u interindividualnoj varijabilnosti i bioraspoloživosti DOAK-a, te rizicima razvoja neželjenih učinaka prvenstveno krvarenja. Pretražena je baza podataka PubMed, Web of Science, Cochrane, Google Schoolar koristeći ključne riječi: apiksaban, CES1, CYP3A4, CYP3A5, dabigatran, direktni oralni antikoagulansi, edoksaban, farmakogenetika, farmakogenomika, genske varijante, interakcije gen-lijek, interakcije lijek-lijek,
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krvarenje, novi oralni antikoagulansi, polimorfizam gena transportera ABCB1, ABCG2, rivaroksaban, SNP, tromboembolijski događaji u razdoblju od zadnjih 10-ak godina. Prikazani su interesantni slučajevi pacijenata iz kliničke prakse koji su zbog farmakogenetičke predispozicije i interakcije lijekova razvili nuspojave DOAK-a u vidu krvarenja. Raspravljene su prisutne dileme, te predočeno trenutno stanje u Hrvatskoj uspoređeno sa situacijom u Europi i drugdje u svijetu.
Rezultati: Nekoliko varijanti gena CES1 i ABCB1 se smatra potencijalno odgovornim za nastanak interindividualnih varijabilnosti DOAK-a. Utvrđeno je da su varijante gena CES1 rs2244613 i rs8192935 povezane s nižim trough koncentracijama dabigatrana i nižim rizikom od krvarenja. S druge strane varijanta gena ABCB1 rs1045642 povezuje s povećanim vršnim koncentracijama dabigatrana i rivaroksabana, te povećanom učestalošću krvarenja. Farmakogenetička analiza varijanti gena ABCB1 rs2032582 (C.2677G> T) i rs104562 (C3435C> T) mogla bi biti opravdana i od koristi kod pacijenata na terapiji rivaroksabanom. Da bi se spriječili neželjeni učinci tijekom terapije apiksabanom kod nositelja nefunkcionalnih alela gena CYP3A5 potrebano je pažljivo odabrati dozu i pratiti nuspojave. Postoji veliki rizik od razvoja neželjenih učinaka lijeka kad se edoksaban kombinira s lijekovima koji inhibiraju CYP izoenzime u homozigotnih nositelja nefunkcionalnih alela za gen CYP3A5.
Zaključak: Polimorfizam gena zajedno s istovremenom primjenom inhibitora/induktora transportnih proteina ili enzima odgovornih za metabolizam lijeka može povećati rizik od nastanka štetnih događaja vezanih uz promjenu DOAK-a.Due to good pharmacokinetics and pharmacodynamics without the requirement for routine coagulation monitoring, direct oral anticoagulants (DOAK) have seen an increase in prescribing in recent years. DOAC achieves its anticoagulant effect by direct inhibition of thrombin (dabigatran) or direct inhibition of activated factor X (rivaroxaban, apixaban and edoxaban). Although DOACs interact less with other drugs compared to warfarin, the combination of DOAC and many drugs requires caution and clinical supervision in some cases. Recent studies have documented interindividual variability in DOAC plasma levels and patient response to the drug. Although clinical and biochemical factors may lead to interindividual variability, the presence of genetic variants or drug interactions may contribute to these differences. The role of pharmacogenomics, namely polymorphism of metabolic enzyme genes and drug transporters, in the interindividual variability of the four most often prescribed DOACs, must be understood.
Objectives: Demonstrate how pharmacogenetic predisposition, primarily polymorphisms of CYP3A4 / 5 metabolic enzyme genes, and CES1 and ABCB1 and ABCG2 transport proteins with drug interactions in polytherapy, may contribute to interindividual variability in DOAC plasma levels, and their efficiency and safety.
Materials and methods: During the preparation of this paper, an extensive search of recently published original and reviewed scientific papers, clinical trials, in vitro studies, therapeutic guidelines and publications of professional societies was conducted. The emphasis is on pharmacogen polymorphisms that are important for DOAC pharmacokinetics: CYP3A4 / 5 and CES1 genes, for enzymes involved in biotransformation, and ABCB1 and ABCG2 genes, for drug transporters and their contribution to interindividual variability and bioavailability of DOACs, effects primarily of bleeding. The database PubMed, Web of Science, Cochrane, Google Schoolar were searched using keywords: apixaban, CES1, CYP3A4, CYP3A5, dabigatran, direct oral anticoagulants, edoxaban, pharmacogenetics, pharmacogenomics, gene variants, drug-drug interactions, gene-drug interactions, bleeding, new oral anticoagulants, polymorphism of transporter genes ABCB1, ABCG2, rivaroxaban, SNP, thromboembolic events in the period of the last 10 years.
Interesting cases of patients from clinical practice who have developed DOAC side effects in the form of bleeding due to pharmacogenetic predisposition and drug interactions are presented. The present dilemmas were discussed, and the current situation in Croatia was compared to the situation in Europe and elsewhere in the world.
Results: Several variants of the CES1 and ABCB1 genes are thought to be potentially responsible for the occurrence of interindividual variability in DOAC. Variants of the CES1 rs2244613 and rs8192935 genes were found to be associated with lower trough out dabigatran concentrations and a lower risk of bleeding. On the other hand, the ABCB1 gene variant rs1045642 is associated with increased peak concentrations of dabigatran and rivaroxaban, and increased bleeding frequency. Pharmacogenetic analysis of the ABCB1 gene variants rs2032582 (C.2677G>T) and rs104562 (C3435C>T) could be justified and useful in patients on rivaroxaban therapy. To prevent side effects during apixaban therapy in carriers of non-functional alleles of the CYP3A5 gene, the dose should be carefully selected and adverse reactions monitored. There is a high risk of developing adverse drug reactions when edoxaban is combined with drugs that inhibit CYP isoenzymes in homozygous carriers of non-functional alleles for the CYP3A5 gene.
Conclusion: Gene polymorphism in combination with the concomitant use of inhibitors / inducers of transport proteins or enzymes responsible for drug metabolism may increase the risk of adverse events associated with alterations in DOACs
Digitalni lijekovi suvremeni oblici za unapređenje adherencije u liječenju mentalnih bolesti
No full textMental disorders are the leading global cause of mortality and
morbidity. Pharmacological therapy represents the basis for the
treatment of mental disorders and medication adherence is extremely important
due to long-term treatment and serious consequences if the treatment is stopped
prematurely. Intentional and unintentional non-adherence to antipsychotics is
the main problem in the treatment of schizophrenia, which occurs due to the
serious side effects of antipsychotics and because the patient is unaware of the
illness. The first approved digital medication Abilify MyCite contains the antipsychotic
aripiprazole as the active substance. lt represents the digital medicine
system which contains a tablet with an ingestible sensor and active substance, a
wearable patch, an application on a smartphone and a web portal. It works on
the principle of transmitting the digital signal from the sensor to the wearable
patch when the sensor comes into contact with gastric fluid. Data on drug ingestion,
as well as other data on health and life habits, are stored on the web portal
and become available to the doctor and other individuals, to whom the patient
grants access. Another digital system in research is the ID-Cap system, with a
sensor built in the capsule. lt works on the principle of transmitting a low-intensity
radio signal to a reading device when the capsule comes into contact with
gastric fluid. AiCure is a visual recognition system that records video and uses
facial image analysis to verify that the patient has taken the prescribed medication.
Digital medicine systems have been developed to improve adherence, thereby
providing better therapy outcomes and reduced healthcare costs. However,
there is no clinical evidence confirming that digital medications can consistently
monitor medication ingestion in real-time or improve adherence. Furthermore,
digital medications are accompanied by several ethical issues such as the negative
impact on patient autonomy and privacy, the subjective feeling of coercion
and control, the disturbed trust in the relationship between doctor and patient
and the impact of pharmaceutical companies on the doctor. Further clinical
research should be conducted with an emphasis on assessing the adherence and
quality of life of patients taking digital medications. Also, doctors and patients
should be educated about digital medications
Uvod u anatomiju - znanstvena pozadina odabranih slučajeva
No full textIn this paper few clinical cases from the popular series Grey's
Anatomy are presented. These cases are related to biotransformation
of drugs and xsenobiotics: the patient who kept waking up from anesthesia because she was a rapid metabolizer of an anesthetic, the changed color of urine
that is the result of hydroxylation and glucuronidation of the drug amitriptyline,
neurotoxicity of cobalt is a consequence of its influence on the hemostasis of
complex biochemical reactions, the interaction of a herbal drug and chemotherapeutic
agent that led to the formation of a neurotoxin. The inspiration for
these cases was found in biomedical journals from real case reports. Case reports
are used as a means of introducing scientists with rare and unusual occurrences
which can lead to form new hypothesis. Television series such as Grey's Anatomy
popularize science and are likely to kindle a spark of interest in students to study
in fields such as medicine and pharmacy