Kniga na apstrakti book of abstrcts

Genetskiot polimorfizam vo GPX1 genot ne e asociran 
so karcinom na prostatata kaj pacienti 
od Republika Makedonija
Arsova-Sarafinoska Z., Matevska. N., Despotovska S., Petrovski D., Xikova S., Banev S.,
Georgiev V., [ikole A., Dimovski A. J.
Institut za kontrola na kvalitet na lekovi, Republi~ki zavod za zdravstvena za{tita, 
Institut za Farmacevtska hemija, Farmacevtski Fakultet, Klinika za uroligija, 
Institut za nefrologija, Institut za patologija, Medicinski Fakultete, Skopje, Makedonija
Poznato e deka oksidativniot stres doveduva do o{tetuvawe na DNA, pri {to mo`e na nastanat
mutacii vo genite koi go reguliraat rastot na tumorite i inicijacijata na kancerogenezata. Glutation
peroksidaza 1 (GPX1) pretstavuva {iroko rasprostranet selen-zavisen enzim. Ovoj enzim igra va`na uloga
vo za{tita na kletkite od oksidativno o{tetuvawe preku redukcija na vodorodniot peroksid i red drugi
organski peroksidi so pomo{ na reduciran glutation. Nukleotidnata supstitucija (C T) vo kodon 198 od
GPX1 genot rezultira so supstitucija na prolin (Pro) so leucin (Leu), mutacija koja spored odredeni istra-
`uvawa e povrzana so zgolemen rizik od karcinomi na belite drobovi, dojkata i mo~niot meur. Ovaa studi-
ja ima za cel da ja ispita povrzanosta na GPX1 genotipot so rizikot od razvoj na kancer na prostata kaj
pacienti vo Republika Makedonija. GPX1 genotipot be{e odreden kaj 60 pacienti so kancer na prostata
i 167 pacienti so benigna hiperplazija na prostata so metodata na real-time PCR i fluorescentno odbele-
`ani TaqMan probi. Kaj pacientite so karinom na prostatata uvrdeni se slednite frekfenciite na Pro/Pro,
Pro/Leu i Leu/Leu genotipovite: 0.57, 0.28 i 0.15, dodeka soodvetnite frekvencii kaj kaj pacientite so benigna
hiperplazija na prostata iznesuvaa 0.56, 0.32 i 0.12. Statisti~kata obrabotka na podatocite poka`a deka
ne postoi statisti~ki zna~ajna razlika vo genotipskata distribucija pome|u pacientite i kontrolite
(OR: 1.2971; 95% CI: 0.5551- 3.0309; p= 0.5530), nezavisno dali pacientite se analizirani vo podgrupi vo odnos
na godini na starotst (OR: 1.1429; 95% CI: 0.274-4.767; p=0.8548) ili vo odnos na stepenot na progresija na
tumorite pri dijagnoza izrazen kako Gleason score (OR: 0.9; 95% CI: 0.2164-3.7427, p= 0.8846). Ovie rezul-
tatite sugeriraat deka GPX1 najverovatno ne e asociran so razvojot na karcinomot na prostatata kaj paci-
entite od Republika Makedonija. 
Macedonian pharmaceutical bulletin 53 (1,2) 164-165 (2007)
PP - 76
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Cyclin D1 G870A Variant is Associated with Increased Risk 
of MSI positive Colorectal Cancer in Young Male Patients 
Matevska N.
1
, Josifovski T.
2
, Hiljadnikova-Bajro M.
1
, Sterjev Z.
1
, Kapedanovska A.
1
,
Serafimoska Z.
1
, Despotovska S.
1
, Petrusevska N.
3
, Panovski M.
2
, Suturkova L.
1
, Dimovski A.J.
1
1
Institute of Pharmaceutical Chemistry, Faculty of Pharmacy,
2
Clinic for Abdominal Surgery 
and 
3
Institute of Radiotherapy and Oncology, Faculty of Medicine, 
University Ss Cyril and Methodius“, Skopje, Republic of Macedonia
Cyclin D1 (CCND1) is a cell cycle regulatory protein, the overexpression of which is often found in human
tumors and is associated with cell proliferation and poor prognosis. Cyclin D1 plays a key role in cell cycle regula-
tion, particularly in the transition from G1 to S phase, which is regulated by cyclin-dependent kinases. A common
G870A single nucleotide polymorphism at codon 242 in exon 4 of the CCND1 gene has been associated with in an
altered messenger RNA transcript, a longer-life protein and an increased risk of colorectal cancer and adenoma in
some studies. Furthermore, overexpression of CCND1 is reported to modify the effect of mutations in mismatch
repair (MMR) genes and enhance microsatellite instability (MSI), hence influence the age of onset of hereditary non-
polyposis colorectal cancer (HNPCC). This study was designed to evaluate the effect of cyclin D1 gene polymor-
phism on the risk of colorectal cancer in Macedonian population in a case control study of randomly selected 331
colorectal cancer patients and 101 controls without clinical diagnosis of colorectal cancer. Cyclin D1 genotypes (AA,
AG, and GG) were determined using PCR-RFLP analysis and subsequent PAGE electrophoresis. The A allele frequen-
cy was higher (0,533) in our population then in numerous Caucasian populations (0,42-0,43). We did not observe a
significant difference in overall allelic frequencies and genotype distribution of affected and unaffected mutation car-
riers (A allele 0.533 for patients and 0.5 for controls; p = 0.40); (AA 30.21%, AG 46.224%, GG 23.565% for patients
and AA 24.752%, AG 50.495%, GG 24.752% for controls; p = 0.80)}. However, we found a statistically significant
risk in carriers of the CCND1 A allele when patient were stratified in subgroups according to gender, age and MSI
status. A higher risk was observed in patients with MSI tumors (RR 3.0; 95% CI: 1.2926and particularly in male patients with MSI tumors under 60 years of age (RR 5.83; 95% CI: 1.7987p= 0.0006). The consequences of the above observation are reversed in female patients. The data from this study
indicates that the CCND1 A variant might acts by enhancement of CRC progression through a pathway influenced
by estrogens and/or estrogen regulated pathways of cell signalling in colonic epithelia and suggests that this variant
can provide additional information in genetic counselling of families with HNPCC.
Macedonian pharmaceutical bulletin 53 (1,2) 168-169 (2007)
PP - 77
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Ciklin D1 G870A polimorfizmot e asociran so zgolemen rizik 
za razvoj na MSI pozitiven kolorektalen kancer 
kaj mladi pacienti od ma{ki pol 
N. Matevska
1
, T. Josifovski
2
, M. Hiqadnikova-Bajro
1
, Z. Sterjev
1
, A. Kapedanovska
1
, 
Z. Serafimoska
1
, S. Despotovska
1
, N. Petru{evska
3
, M. Panovski
2
, 
L. [uturkova
1
, A. J. Dimovski
1
1
Institut za farmacevtska hemija, Farmacevtski Fakultet, 
2
Klinika za abdominalna hirurgija i 
3
Institut za radioterapija i onkologija, 
Medicinski Fakultet, Univerzitet Sv. Kiril i Metodij, Skopje, R.Makedonija
Proteinot Ciklin D1 (CCND1) igra klu~na uloga vo regulacijata na kleto~niot ciklus, osobeno
pri preodot od G1 vo S fazata. Golem broj na istra`uvawa uka`uvaat na vlijanieto na CCND1 na brzata
proliferacija i lo{ata prognoza kaj humanite tumori. G870A polimorfizamot vo kodon 242 od egzon 4
na CCND1 genot doveduva do promenet mRNA transkript, i konsekventna produkcija na protein so podolg
polu`ivot i najverojatno, zgolemen rizik za razvoj na razli~ni tipovi na kancer. Ovoj efekt e najizrazen
vo slu~aj na prisastvo na dopolnitelni rizik faktori koi se odgovorni za inicijacijata na kancerogeneza-
ta, kako {to se mutaciite vo MMR (mismatch repair) genite kaj familiite so nasleden nepolipozen kolorek-
talen kancer (HNPCC). Ova istra`uvawe ima{e za cel da go ispita vlijanieto na CCND1 G870A polimor-
fizamot na rizikot od kolorektalen kancer kaj pacientite od Republika Makedonija. DNA be{e izolirana
od 331 slu~ajno izbrani pacienti i 101 kontroli bez klini~ka dijagnoza na kolorektalen kancer. CCND1
genotipovite (AA, AG i GG) bea odredeni so PCR-RFLP analiza i poslednovatelna PAGE elektroforeza.
Frekfencijata na CCND1 A alelot vo na{ata populacija e povisoka (0.533) vo sporedba so drugi popu-
lacii od belata rasa (0.42-0.43). Nema statisti~ki zna~ajna razlika vo alelnite frekvencii i genotip-
skata distribucija pome|u pacientite i kontrolite {(A alel 0.533 kaj pacientite i 0.5 kaj kontrolite; p
= 0.40); (AA 30.21%, AG 46.224%, GG 23.565% kaj pacientite i AA 24.752%, AG 50.495%, GG 24.752% kaj
kontrolite; p= 0.80)}. Za razlika od ova, postoi statisti~ki zna~aen rizik kaj pacientite dokolku rezul-
tatite se razgleduvaat vo odnos na polot, vozrasta pri dijagnoza i MSI statusot na tumorite. Najgolem
rizik e zabele`an kaj pacienti so MSI tumori (RR 3.0; 95% CI: 1.2926pacienti so MSI tumori od ma{ki pol pomladi od 60 godini (RR 5.83; 95% CI: 1.7987Interesno e toa {to rizikot kaj ma{kata polulacija se namaluva so zgolemuvawe na godinite na starost,
dodeka pak kaj `enskata populacija e zabele`an sprotiven efekt. Ovie podatoci uka`uvaat na toa deka
CCND1 A alelot najverojatno doveduva do zabrzuvawe na proliferacijata kaj kolorektalniot kancer preku
pati{ta vkrsteni so estrogenite i/ili estrogenskata kleto~na signalizacija vo epitelot na kolonot i
sugeriraat deka ovoj marker mo`e da bide od korist pri genetskoto sovetuvawe kaj familii so HNPCC.
Macedonian pharmaceutical bulletin 53 (1,2) 168-169 (2007)
PP - 77
169
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170

FARMACEVTSKI ANALIZI / 
OBEZBEDUVAWE KVALITET / REGULATIVA
Sekciski vovedni predavawa  SPL 9-11
Kratki usmeni soop{tenija  SOP 4-5
Posterski prezentcii  PP 78-123
PHARMACEUTICAL ANALISIS / 
QUALITY ASSURANCE / REGULATORY AFFAIRS
Section plenary lectures  SPL 9-11
Short oral presentation  SOP 4-5
Poster presentation  PP 78-123

Integral development of high quality Pharmaceutical products
Ales ROTAR
KRKA d.d., Novo Mesto, Slovenia
Development of a new pharmaceutical product, it's specifications as well as production technologies and
analytical methodology is a process which starts long before the product is launched and is essentially never con-
cluded. Today scientific, economic and social development in the world requires continuous contribution of all the
subjects in the market: regulators, industry and academia and last but not least the users to strive for the optimum
quality of pharmaceutical care. Quality, efficacy and safety of pharmaceuticals are fundamentals of the mission of
our profession. Therefore specification and methodology together with general quality assurance mechanisms rep-
resent one of the core elements of regulatory environment of pharmaceutical market.
During the last decade it has been confirmed that an integral model of product development is essential if the
company wants to bring a safe, efficacious and quality product to the market. In order to carry out a successful proj-
ect a team of pharmacists, chemists, physicians and several other professions has to be able to combine different sci-
entific disciplines in order to solve problems of ever increasing complexity.
Product and process specifications play an important role in the process of development, regulatory and
launch of the products. Different aspects of specification development with regards to chemical and physicochemi-
cal as well as pharmaceutical parameters are going to be discussed. Active substance related issues require different
approach than the issues related to pharmaceutical product. Nevertheless, in order to find a scientifically and regu-
latory sound solution a combination of solutions is required. On top of the scientific questions intellectual property
issues as patents pose additional challenges to the development teams. 
Successful product development projects, where Krka's R&D team has been able to implement latest scien-
tific achievements are going to be presented. Products like Vasilip (simvastatin) tablets, Lanzul (lansoprazole)
pellets/capsules and Yasnal (donepezil) tablets represent significant contribution to portfolio of high quality gener-
ic products.
The topic of  the presentation is also going to be continuous development process throughout the lifecycle
of a product. In the recent period several cases of development and revision of the monographs of European
Pharmacopoeia have confirmed this approach. Krka is one of the companies contributing to quality of European
pharmaceutical markets through everlasting product monitoring and upgrade of quality specifications for the benfit
of the patients. At the end the role of specifications, methods, monographs and link to marketing authorisations with-
in a general regulatory network as well as global quality systems is going to be discussed.
Macedonian pharmaceutical bulletin 53 (1,2) 172 (2007)
SPL - 9
172
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Electroanalytical methods in drug analysis 
– validation and biovalidation
V. Kapetanovic
1,
M. Aleksic
2
1
Institute of Analytical Chemistry, Faculty of Pharmacy, University of Belgrade, 11000, Serbia
2
Institute of Physical Chemistry, Faculty of Pharmacy, University of Belgrade, 11000, Serbia
Modern voltammetric techniques, such as differential pulse voltammetry (DPV), linear scan voltammetry
(LSV), square – wave voltammetry (SQW) and the corresponding stripping techniques are the most accurate, as well
as most sensitive and very rapid electrolytic instrumental methods. The potential range over which voltammetric
techniques can be used will depend on the electrode material, the solvent, the supporting electrolyte, and the acidi-
ty of the solution. If a platinum electrode is used in aqueous medium, the limiting positive potential would be oxi-
dation of water, unless the supporting electrolyte contains a more easily oxidizable ion (e.g.,Cl-). The negative lim-
iting potential will be from the reduction of hydrogen ions, and since the platinum has a low hydrogen over-voltage
at low current densities, this will occur at about -0.1V. Carbon electrodes are frequently used for voltammetry. Their
positive potential limit is the same as with platinum ones, but more negative potentials can be reached because hydro-
gen has a rather high over-voltage on carbon. Potentials of about -1V versus SCE or more can be used, depending
on the pH. While carbon electrodes can be used at fairly negative potentials, a dropping mercury electrode (DME)
is often preferred ( potentials of -2V vs. SCE ) and better reproducibility can be achieved. This is because the elec-
trode surface is constantly renewed (small mercury drops fall from a capillary).
The modern voltammetric techniques mentioned above are performing with hanging mercury electrode
(HMDE) – as a working , Ag/AgCl – as a reference and Pt-as a auxiliary electrode.
This presentation deals with the analytical application of the modern voltammetric techniques for determi-
nation of drugs, especially antibiotics - cephalosporins.
Taking into consideration the general knowledge about the cephalosporins
1
and its electrochemical behav-
ior
2
, its acid-base equilibrium, recently established
3
and its adsorptive properties
4
, the idea was born to develop the
modern voltammetric methods for determination of these drugs in water solution and in biological matrix as well.
For this purpose the basic analytical demands were established for the corresponding methods, the validation param-
eters were recognized for determination in buffer-water solution and the corresponding real humane sample, such as
urine is. The selectivity of the method proposed was checked over the main degradation product-as impurity or as
its main metabolite, in the case of biological sample. Two model compounds were presented, cefotaxime (CFX) and
its active metabolite - desacetylcefotaxime (DCFX)
5
.
The methods applied were adsorptive stripping voltammetry (AdSV) in the case of cefetamet
6
(CEF) and
square-wave voltammetry (SQW) in the case of CFX and DCFX.
5
The validated methods enabled determination of the cited cephalosporins with high selectivity related to the
main impurities and metabolites of the corresponding drugs. The methods proposed are highly sensitive, accurate
and rapid for performance.
Macedonian pharmaceutical bulletin 53 (1,2) 173 (2007)
SPL - 10
173
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Regulatory affairs affected by new achievements 
in pharmaceutical science and professional practice
Vesna Koblar
Agency for Medicinal Products and Medical Devices, Ljubljana, Einspielerjeva 6, Slovenia, www.jazmp.si
Regulatory Affairs encompass scientific knowledge, legal form and health or drug policy. 
Common principals, reflected in the whole EU pharmaceutical legislative body are:
• ensuring high level of public health and
• ensuring competitiveness of pharmaceutical industry.
Area of pharmaceuticals is very dynamic. New methods, new molecules, new processes, new sources, new
forms and changes have become an every-day matter, a way of life within the area. Even more, it gains in momen-
tum and becomes more widespread.
Achievements of pharmaceutical science and professional practice are always resulting from strive for:
• high level of health protection, and
• high level of competitiveness of pharmaceutical industry. 
Legislation must follow state-of-the-art science and technology. Regulatory affairs must take into consider-
ation best practices and guidelines in order to keep flexibility, necessary to enable further progress. This is why the
regulatory environment for medicines is changing on daily basis. Pharmaceutical acquis is the most regulated area
of the acquis communautaire. Eighty thousand pages of EU legislation have been produced in order to follow the
science and technology, to protect health, to set rules of the game, to set updated standards for quality, safety and
efficacy of medicinal products as well as standards for knowledge, competency and ethics.
Regulatory affairs are a safeguard for patients and industry and tool for governments in achieving necessary
balance between the two. Combination of scientific knowledge, legal and business issues enable that product develop-
ment, manufacturing, marketing and use of medicinal products meet or exceed the government requirements.
Rules should be simple, clear, understandable, up-to date to technological progress, user friendly, used only
when necessary and of a high quality. Burdens that they impose should be proportionate to their aim. This is why
European Commission is now focusing on improving the existing body of legislation and on producing new pieces
of legislation that reflect the need to fill the existing gaps and to follow progress of science and technology.
Recent new documents confirm those efforts. 
Recently adopted Pediatric regulation is aimed at ensuring proper use of medicines for children. Over 50%
of medicines used in children may not been studied on this age group. Children represent 20% of EU population. It
is a vulnerable group physiologically and psychologically different from adults. Children are not „small people“, so
the absence of suitable authorized products to treat conditions in children causes „off label“ use of authorized prod-
ucts and use of unauthorized products or giving medicines to population in which they have not been tested. Pediatric
regulation requires studies on pediatric population if medicinal product is to be used for children. The Regulation
introduces incentives for industry to perform the trials. Centralized expertise in the EMEA enables proper assess-
ment of all the submitted studies on heterogeneous pediatric population.
Recently reached agreement on Regulation on advanced therapy medicinal products (ATP) defines and reg-
ulates those products in the light of the existing legislation on medicinal products. ATPs are gene therapy products,
cell therapy products and tissue engineering products. 
Recently proposed changes of provisions on pharmacovigilance are aimed at improving safety of products
and particularly at addressing to a new generation of product.
Pharmaceutical regulatory affairs, affected by new achievements in pharmaceutical science and profession-
al practice are a tool for achieving a high level of health protection and high level of competitiveness of pharmaceu-
tical industry.
Macedonian pharmaceutical bulletin 53 (1,2) 174 (2007)
SPL - 11
174
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Practical aspects of analytical method transfer
Aneta Dimitrovska, Suzana Trajkovic-Jolevska
Faculty of Pharmacy, Center for drug quality control, 1000 Skopje, R. of Macedonia
In a regulated environment of drug quality control, once developed and validated methods in the originator
laboratory („sending“ laboratory) are commonly transferred to another laboratory („receiving“ laboratory). Analytical
method transfer (AMT) is the verification that the analytical method works as well in the receiving laboratory as in
the originator's laboratory and meets all acceptance criteria.
There are several different options for AMT. These include comparative testing complete or partial method
validation or revalidation, covalidation between the two laboratories and the omission of formal transfer, sometimes
called transfer „waiver“. The choice of which option to use depends upon the stage of development of the method
(early or late stage), the type of methods (simple or complex, compendia or noncompendia) and the experience and
capabilities of the laboratory personnel. In order to achieve a successful AMT it is necessary to fulfill a lot of require-
ments, starting with preapproved test protocol, description of method and test procedures, description and rationale
of test requirements, acceptance criteria, documentation of results and ending with transfer report. As in any valida-
tion process, the heart of the technical transfer process is documentation, both for the process and the results. The
AMT report certifies that the acceptance criteria have been met, and that the receiving laboratories are fully trained
and qualified to run the method. By anticipating that instruments, experience and training, and procedure interpre-
tations can differ from laboratory to laboratory, many of the common pitfalls encountered during AMT can be pre-
vented with a little up-front work.
The rapid increase of high-performance liquid chromatography (HPLC) as a premier technique in the phar-
maceutical industry provokes development of fool-proof procedures that can be transferred from one laboratory to
another. This review will discuss sources of failures to reproduce HPLC procedures, running from sample handling
and preparation, through equipment, instrument consideration (injector, pump, column, and detector), mobile phase,
staff training and data manipulation problems. There are many precautions that should be considered when initially
developing a HPLC method to minimize future problems and how to handle problem assay. As conclusion, due to
the fact that HPLC methods' omnipresence, attention should be paid to development of robust and reproducible pro-
cedures liable to easy transfer from one laboratory to another. 
Macedonian pharmaceutical bulletin 53 (1,2) 175-176 (2007)
SOP - 4
175
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