imagine (Institute of molecular genetics and genetic engineering)
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miRNAs - promissing applications in radiotherapy
No full textUpon 40 years of first microRNA (miRNA) discovery, now it is known that these small non-coding
RNA molecules have regulatory functions in all biological and pathological processes. They play
important roles in regulating gene expression and modulate up to 60% of protein-coding genes in
the human genome. miRNAs are involved in biological processes of several types of cancer where
they can act as both tumor suppressors and oncogenes. Numerous factors, such as the amplification
or deletion of miRNA genes, aberrant transcriptional control of miRNAs, dysregulated epigenetic
modifications, and flaws in the machinery of miRNA biogenesis, contribute to the dysregulation
of miRNA expression in human cancer. Dysregulated miRNAs impact the hallmark of cancer,
such as maintaining proliferative signaling, eluding growth suppressors, preventing cell death,
initiating invasion and metastasis, and triggering angiogenesis. Growing number of studies
revealed miRNAs as potential biomarkers for human cancer diagnosis, prognosis, and therapeutic
targets. miRNAs have remarkable therapeutic potential for radiation treatment, which is of great
interest since it is one of the most often applied treatments in many cancers. In addition to helping
to define the ultimate response to therapy and the risks of recurrence or metastatization, miRNAs
may profile the radioresistance of tumors before to the delivery of treatment and track the response
over the course of the treatment, assisting in the selection of intensification strategies. Promising
applications of miRNAs in radiotherapy, is the therapeutic inhibition or mimicking of miRNAs to
overcome radioresistance during therapy, and the usage of miRNAs as predictive biomarkers for
therapy response and prognosis. Therefore, miRNAs might be seen as promising biomarkers
capable of predicting radiation response prediction and patient-specific tailored therapy planning,
ultimately leading to a new age of personalized medicine for cancer patients
Genetic risk factors in patients with Myasthenia gravis
Get PDFMyasthenia gravis (MG) is a rare autoimmune disease mediated by antibodies against
components of the neuromuscular junction, particularly the acetylcholine receptor
(AChR). The prevalence of MG in Belgrade has been estimated at 189 cases per
1,000,000 inhabitants, which is among the highest prevalence reported to date.
Genetic studies have mainly pointed to specific HLA alleles associated with MG.
However, CTLA-4 and TNFRSF11A, playing a role in the immune response, have
recently been associated with MG in genome-wide association studies. Since CTLA-4
and TNFRSF11A promote other autoimmune diseases, the main objective of this casecontrol
study was to determine the association between these candidate genes and the
risk for developing MG in Serbian population.
Genotyping of rs231735 and rs231770 within the CTLA-4 gene and rs4263037 within
TNFRSF11A in 447 AChR-MG patients and 447 individually sex- and age-matched
controls revealed no association with MG (p=0.344, p=0.923 and p=0.557,
respectively). However, when stratifying patients into those with early-onset (n=183)
and late-onset MG (n=264), we found an association of minor rs231735 allele T with
early-onset MG under the recessive genetic model (OR=0.548, 95% CI=0.339-0.888,
p=0.014, p10e6 permutation=0.014). Haplotype analysis revealed that individuals
with the GC haplotype rs231735-rs231770 had a higher risk for developing earlyonset
MG (OR =1.360, p=0.027, p10e6 permutation =0.027). Considering the
sufficient statistical power of the study (>90%) and the selection criteria for controls,
our results suggest that the CTLA-4 may be associated with early-onset MG in Serbian
population. Analysis of additional variants is needed to understand the association of
CTLA-4 with MGBOOK OF ABSTRACTS: 8th CONGRESS OF SERBIAN NEUROSCIENCE SOCIETY with international participation 31 May – 2 June 2023. Belgrade, Serbi
Transcript CD81-215 may be a long noncoding RNA of stromal origin with tumor-promoting role in colon cancer
No full textThe role of tetraspanin CD81 in malignant transformation is best studied in colorectal cancer, and it appears that other transcripts beside the fully coding mRNA may also be dysregulated in malignant cells. Recent data from a comprehensive pan-cancer transcriptome analysis demonstrated differential activity of two alternative CD81 gene promoters in malignant versus nonmalignant gut mucosa. The promoter active in gut mucosa gives rise to transcripts CD81-203 and CD81-213, while the promoter active in colon and rectal cancer gives rise to transcripts CD81-205 and CD81-215. Our study aimed to explore the biomarker potential of the transcripts from the alternative CD81 gene promoters in colon cancer, as well as to investigate their structure and potential function using in silico tools. The analysis of the transcripts' expression in several colon cell lines cultivated in 2D and 3D and a set of colon cancer and healthy gut mucosa samples by qPCR and RNA sequencing suggested their low expression and stromal origin. Expression patterns in tumor and nontumor tissue along with in silico data suppose that the transcript CD81-215 may be a noncoding RNA of stromal origin with possible involvement in signaling related to malignant transformation
The potential roles of gossypol as anticancer agent: advances and future directions
Get PDFGossypol, a polyphenolic aldehyde derived from cottonseed plants, has seen a transformation in its pharmaceutical application from a male contraceptive to a candidate for cancer therapy. This shift is supported by its recognized antitumor properties, which have prompted its investigation in the treatment of various cancers and related inflammatory conditions. This review synthesizes the current understanding of gossypol as an anticancer agent, focusing on its pharmacological mechanisms, strategies to enhance its clinical efficacy, and the status of ongoing clinical evaluations
Sulforaphane affects morphological changes and cell viability of colon cancer cells: p53 mutation-dependent effect
Get PDFColorectal carcinoma (CRC) is an increasing cause of morbidity and mortality worldwide.
Among biologically active compounds which have been shown to exert cytotoxic effects on human cancer
cells, including CRC, is sulforaphane (SFN), an isothiocyanate compound extracted from cruciferous
vegetables, especially broccoli. This in vitro study aimed to investigate the effect of SFN on the growth
of human CRC cells and its dependency on the expression of p53. Two human CRC cell lines: HT-29
(a p53 mutated line) and HT-116 (a p53 wild-type line) were treated with SFN at concentrations of 0, 4,
8, 16, and 32 μmol/L for 72 h to test its anticancer effect.
Results indicated that SFN induced cell morphological changes and decreased the total number
of viable cells. Treatment of CRC cells with SFN for 72 h resulted in moderate dose-dependent cytotoxicity.
HCT-116 cells, with a p53-wt, were more sensitive (IC50 = 8.41 μM) than p53-mutated HT-29 cells (IC50
= 24.83 μM). These results indicate that SFN exhibits the anticancer effect against CRC cells in p53 mutation-
dependent manner (Serbia-China project: 451-03-1203/2021-09)
Signifi cance of molecular diagnosti cs in therapy of chronic lymphocytic leukemia
Get PDFChronic lymphocyti c leukemia (CLL) is a malignancy of mature CD5+ B lymphocytes that is characterized by excepti onal
clinical and biological heterogeneity. The Rai and Binet staging systems, developed in the late 1970s to early 1980s,
are used in clinical practi ce to strati fy CLL pati ents into risk categories and to help guide clinical follow-up opti ons: to
treat or to watch and wait. However, in early-stage disease, these systems are unable to predict what pati ents will
face the progression to a more aggressive disease. That means, a number of molecular markers with prognosti c and/
or predicti ve impact exist and their assessment is strongly recommended in all pati ents prior to treatment initi ati on.
One of the fi rst recognized prognosti c genomic aberrati ons in CLL include those detected by fl uorescence in situ
hybridizati on (FISH): del(17p), del(11q), trisomy 12 and del(13q), and the immunoglobulin heavy variable (IGHV) gene
somati c hypermutati on (SHM) status. Moreover, the rapid development of genomics techniques greatly expanded
the understanding of CLL at the molecular level in the past decade. This resulted in the discovery of many newer
prognosti c markers based on chromosomal aberrati ons or gene mutati ons. For instance, next-generati on sequencing
(NGS) studies have led to the discovery of recurrently mutated genes in CLL, such as NOTCH1, SF3B1, BIRC3, XPO1,
POT1, NFKBIE and EGR2, that are associated with poor clinical outcome. Among all of these biomarkers, the disti ncti on
between markers of prognosti c and predicti ve values should be made. Prognosti c markers refer to biomarkers that can
provide informati on regarding the pati ent's outcome regardless of treatment. They are o����� en assessed before treatment
to help guide decisions on to treat or not. Markers associated with overall survival (OS) or ti me to fi rst treatment
(TTFT) represent such examples. On the other hand, predicti ve markers are related to therapeuti c interventi ons
with the ability to predict treatment response to a drug. These markers are normally assessed when pati ents receive
the parti cular therapy. Some markers can be both prognosti c and predicti ve. The Nati onal Comprehensive Cancer
Network guideline recommends testi ng of TP53 geneti c alterati ons, IGHV mutati on status, and several well-established
cytogeneti c markers for CLL prognosti cati on. Of these, TP53 mutati ons, IGHV unmutated status, del(17p), and
del(11q), as well as complex karyotype (the presence of three or more unrelated clonal chromosomal abnormaliti es
in a sample), are associated with poor prognosis. Normal karyotype and trisomy 12 are considered as intermediate
prognosti c factors, whereas del(13q) is associated with a favorable prognosis. The higher frequencies of the
previously menti oned unfavorable markers (except for IGHV) found in the treated populati on usually imply the clonal
evoluti on during disease progression or change in clonal dynamics induced by therapies, especially chemotherapies.
Diff erent molecular and genomic techniques are employed for detecti ng molecular biomarkers in CLL. For IGHV
mutati on status, the preferred method is Sanger sequencing to detect mutati ons in genomic DNA or cDNA following
PCR, and align the resulti ng sequences to the germline IGHV using the IMGT/V-QUEST analyti c tool, where ≥ 98%
homology to the germ line is interpreted as unmutated, >2% nonhomology as mutated, and 97.0% to 97.9% is
interpreted as borderline. Prognosti cally signifi cant chromosomal abnormaliti es are frequently detected using
fl uorescence in situ hybridizati on, array comparati ve genomic hybridizati on or conventi onal karyotyping. Fluorescence
in situ hybridizati on, although off ers a high sensiti vity and specifi city, requires prior knowledge of chromosomal
lesions for the probe designs and are limited to the chosen panel genes. The technique has limitati ons in detecti ng
possible complex cytogeneti c abnormaliti es, as well. On the other hand, karyotyping and array comparati ve genomic
hybridizati on provide genome-wide coverage. Despite the fact that array comparati ve genomic hybridizati on does not
eff ecti vely detect balanced chromosomal rearrangements, it uncovers more genomic abnormaliti es than karyotyping
as the probe-based technology examines the chromosomal structure at a much higher resoluti on. Development in
NGS technology in the past two decades, made the technique, especially targeted sequencing of gene panels, much
less costly and accessible. Currently, in Serbia, geneti c techniques such as FISH, conventi onal karyotyping, Sanger
sequencing and NGS are available for detecti on of CLL biomarkers. Advances in the understanding of CLL pathogenesis
have consequently led to the development of several highly eff ecti ve targeted therapies, including Bruton tyrosine
kinase (BTK), phosphati dylinositol 3-kinase, and BCL2 apoptosis regulator (BCL2) directed inhibitors. B-cell survival
and proliferati on is regulated by the BCR signaling pathway. In normal B cells, BCR is triggered by anti gen ligati on,
leading to acti vati on of a cascade of tyrosine kinases, including BTK. BCR signaling is aberrantly acti vated in many B-cell
malignancies, including CLL. Ibruti nib has demonstrated high clinical effi cacy acti ng as an irreversible potent inhibitor of
Bruton's tyrosine kinase and targets several key components of the BCR pathway. However, despite having 80% to 90%
response rate, 10% to 15% of CLL pati ents, who respond initi ally, develop ibruti nib resistance and disease relapse in 2 to
3 years on ibruti nib treatment, mainly because of the acquisiti on of a BTK C481S mutati on. The mutati on prevents the
drug from forming a covalent bond with the C481 residue that weakened the drug-BTK binding by 500-fold. As a result,
BCR signaling and cell proliferati on were restored in the tumor cells. BTK mutati ons may be found in approximately 70%
of CLL pati ents who progressed on ibruti nib treatment. Another resistance mechanism is through acquired acti vati ng
mutati ons in PLCG2, which is found in approximately 10% of the cases. Given these evidences, the current Nati onal
Comprehensive Cancer Network guideline recommends testi ng for BTK and PLCG2 mutati ons for CLL pati ents receiving
ibruti nib who are suspected of having disease progression. NGS has become the opti mal method for detecti ng BTK
or PLCG2 mutati ons in the se����� ng of ibruti nib treatment, as multi ple mutati ons in both genes may occur in the same
specimen. Currently, approximately 20% of CLL pati ents who progressed on ibruti nib do not have either BTK or PLCG2
mutati ons; thus, with NGS, it is possible to uncover other less common but yet undefi ned drug-resistance mutati ons.
In additi on to BTK and PLCG2 mutati ons known to confer ibruti nib resistance, other molecular markers have
been associated with an upfront high risk of relapse on ibruti nib treatment. It has been reported that complex
karyotype, del(17p)/TP53 mutati on, and del(18p) at baseline before ibruti nib treatment are strongly associated
with disease relapse. Other approved targeted agents for CLL treatment include the phosphati dylinositol 3-kinase
inhibitors idelalisib and duvelisib and BCL2 inhibitor venetoclax. For venetoclax, a novel BCL2-G101V mutati on was
identi fi ed to prevent drug acti vity through drug-protein interacti on. Each pati ent with CLL may have several clinical
and molecular markers of confl icti ng prognosti c signifi cance simultaneously, making the precise prognosti cati on
challenging. Today is of the greatest importance to apply ultrasensiti ve techniques to reveal molecular relapses
a����� er therapy initi ati on and to detect minimal residual disease a����� er pati ents achieve complete responses
Prothrombin influences proliferation and migration of colon cancer in vitro
Get PDFIntroduction: Thrombin, crucial member of the coagulation cascade, can influence growth and development of different types of cancer. Prothrombin, thrombin precursor, although predominantly secreted
from the liver into the bloodstream, can also be expressed in the cancer cells. According to latest data prothrombin can bind in vitro to transmembrane receptors, which have previously been shown to be up-regulated in cancers and activate migration and invasion. Despite the significant amount of data on the
effects of thrombin in cancer progression, there are little data of prothrombin´s effect. The aim of this
study was to further examine the effects of prothrombin and thrombin in cancer cell lines.
Methods: Colon cancer cell lines (Caco2, SW480, SW620, HT29 and HCT116) were treated with prothrombin, thrombin and direct thrombin inhibitor, dabigatran, for 24h and 48h. To assess the effects of
treatment on cell viability and proliferation MTT test was used, and wound healing assay was used for cell
migration potential.
Results: Detected effects of treatment with prothrombin, thrombin and dabigatran varied between cell
lines. Trend of lower cell viability, proliferation and migration was observed in cells treated with prothrombin in comparison to untreated controls.
Conclusion: Our resultsindicate that prothrombin, although considered an inactive zymogen, can exert
an effect on colon cancer cells proliferation and migration in vitro
Plant growth promoting Halomonas from roots of halophytes as a strategy to improve crop resistance to soil salinity
Get PDFsoil salinity has numerous negative effects on plant growth and poses
a significant threat to agricultural productivity worldwide. The diversity of
endophytic bacterial communities found in various indigenous halophytes
such as Salicornia europaea, Suaeda maritima, and Camphorosma annua, may
reveal beneficial bacteria with various plant growthpromoting (PGP) properties.
Members of Halomonas, the largest genus of the family Halomonadaceae,
are rod-shaped, gram-negative, moderately halophilic bacteria that have been
increasingly isolated from various saline environments, including halophyte
tissues. As part of the halophytic microbiota, halotolerant plant growth-promoting
endophytic bacteria can contribute to plant host growth, productivity, and
fitness under abiotic and biotic stresses. The use of halotolerant PGP bacteria
represents a good alternative strategy to mitigate the effects of salt stress in
crops and to recover saline soils. The aim of this study was to isolate halotolerant
bacteria from halophyte roots, and evaluate their PGP ability and their effect on
germination parameters of selected crops. The endophytic bacterial communities
in the roots of halophytes from natural saline soil in Slano Kopovo (Serbia)
were analyzed using culture-dependent techniques. The genus Halomonas was
the predominant among the halotolerant root-associated bacteria of analyzed
halophytes. We have isolated seven Halomonas sp. strains, six of which were
closely related (>99,86% similarity) to Halomonas songnenensis, a moderately
halophilic bacterium from saline and alkaline soils. Obtained isolates were tested
at different sodium chloride concentrations for a range of PGP traits at
different concentrations of sodium chloride. The selected strains were used to
inoculate seeds iand monitor of the germination process of barley, sunflower,
and wheat under conditions of elevated salinity. Most of the isolates tolerate
the NaCl concentration up to 18% and showed multiple PGP features with some
of them retained even in the presence of additional salt concentrations. The
germination test results showed that Halomonas seed inoculation positively
affected barley germination parameters in conditions with and without added
salt. The observed traits can be further exploited to improve plant growth and
augment the salt tolerance of agricultural production on saline soils.Book of abstract: ICGEB WORKSHOP; Trends in microbial solutions for sustainable agriculture, 13 – 15 September 2023. Belgrade, Serbi
EXOPOLYSACCHARIDE-PRODUCING GUT BACTERIA MODULATE HOST AGEING
Get PDFIntroduction: New theories of ageing recognized gut microbiota as one of twelve hallmarks of ageing (1).
Recent data conducted on Caenorhabditis elegans imply a potential role of Lactobacillus species and other
commensal bacteria
in regulation of ageing highlighting TFEB/HLH-30-dependent autophagy, p38
MAPK/PMK-1 signalling and mitochondrial function as activated longevity-associated mechanisms (2,3).
Here, we explore the potential of bacterial polysaccharides
loosely attached to bacterial cell wall
(exopolysaccharides), considering it is still unknown which bacterial molecules could activate longevity
signalling pathways.
Materials & Methods: Caenorhabditis elegans was used as ageing model. Evaluation of worm’s lifespan
and locomotion rate were performed by feeding worms with six exopolysaccharide-producing lactobacilli.
Worms fed with two selected strains were subjected to RNAseq analysis. Identified upregulated genes were
confirmed by qPCR and expression of their mammalian orthologs checked in human HepG2 cell.
Results: Two strains of lactobacilli showed the most pronounced effect on worms’ lifespan. RNAseq analysis
identified core gene signature associate with exopolysaccharide-induced longevity. The expression of
identified fmo-2, gsto-1, nlp-29, and clec-47 genes were confirmed by qPCR, while upregulation of FMO-5
was confirmed in HepG2 cells.
Conclusion: Overall, our results imply that bacteria-derived exopolysaccharides could stimulate longevity-
promoting flavin-containing monooxygenase 2 to regulate lifespan in Caenorhabditis elegansInternational Society of Microbiota 10th ISM World Congress on Targeting Microbiota October 17-19, 2023 – Venice, Ital
Brza progresija hronične limfocitne leukemije u Rihterov sindrom kod bolesnika sa kariotipom blizu triploidnog broja hromozom
Get PDFIntroduction. The presence of aneuploidy in patients diagnosed with chronic lymphocytic leukemia (CLL), except trisomy 12, is considered quite uncommon. Hyperdiploidy or near-tetraploidy (occurring in 1–3% of all CLL patients) usually confer a poor prognosis. Case report. We report a patient in a progressive phase of CLL with near–triploid karyotype. The prognosis of the disease was more precisely determined by applying the cytogenetic analysis of the karyotype and was complemented with molecular methods and pathohistological examination. The complex karyotype was accompanied by the TP53, C-MYC, and IGH gene disruptions, the most probable cause of rapid evolution into Richter’s syndrome. Conclusion. The use of comprehensive contemporary diagnostic techniques is highly recommended in patients who are in the progressive phase of CLL, primarily for the adequate choice of management strategy. The presented case confirms that aneuploidy in CLL patients indicates poor prognosis, which is in accordance with previous publications reporting on cases of CLL patients with aneuploidy.Uvod. Prisustvo aneuploidije kod bolesnika sa dijagnozom hronične limfocitne leukemije (HLL), sa izuzetkom trizomije 12, smatra se retkom pojavom. Pojava hiperdiploidnog ili kariotipa blizu tetraploidnog broja hromozoma (koji se javlja kod 1–3% svih bolesnika sa HLL) smatra se lošim prognostičkim parametrom. Prikaz bolesnika. Prikazan je bolesnik u uznapredovaloj fazi HLL sa kariotipom blizu triploidnog broja hromozoma. Prognoza bolesti je preciznije određena citogenetičkom analizom kariotipa bolesnika, i dopunjena molekularnim metodama i patohistološkom analizom. Otkriveno je prisustvo kompleksnog kariotipa udruženog sa poremećajima u genima TP53, C-MYC i IGH, što je najverovatnije bio uzrok brze progresije u Rihterov sindrom. Zaključak. Primena savremenih dijagnostičkih metoda veoma je značajna kod bolesnika u uznapredovaloj fazi HLL, prvenstveno zbog adekvatnog terapijskog pristupa. Prikazani slučaj ukazuje da je prisustvo aneuploidije kod bolesnika sa HLL loš prognostički znak, što je u saglasnosti sa prethodno publikovanim prikazima bolesnika sa HLL i sa aneuploidijom u kariotipu