Pharmacogenetics of toxicity of 5-fluorouracil, doxorubicin and cyclophosphamide chemotherapy in breast cancer patients.

The differences in patients' response to the same medication, toxicity included, are one of the major problems in breast cancer treatment. Chemotherapy toxicity makes a significant clinical problem due to decreased quality of life, prolongation of treatment and reinforcement of negative emotions associated with therapy. In this study we evaluated the genetic and clinical risk factors of FAC chemotherapy-related toxicities in the group of 324 breast cancer patients. Selected genes and their polymorphisms were involved in FAC drugs transport (ABCB1, ABCC2, ABCG2,SLC22A16), metabolism (ALDH3A1, CBR1, CYP1B1, CYP2C19, DPYD, GSTM1, GSTP1, GSTT1, MTHFR,TYMS), DNA damage recognition, repair and cell cycle control (ATM, ERCC1, ERCC2, TP53, XRCC1). The multifactorial risk models that combine genetic risk modifiers and clinical characteristics were constructed for 12 toxic symptoms. The majority of toxicities was dependent on the modifications in components of more than one pathway of FAC drugs, while the impact level of clinical factors was comparable to the genetic ones. For the carriers of multiple high risk factors the chance of developing given symptom was significantly elevated which proved the factor-dosage effect. We found the strongest associations between concurrent presence of clinical factors - overall and recurrent anemia, nephrotoxicity and early nausea and genetic polymorphisms in genes responsible for DNA repair, drugs metabolism and transport pathways. These results indicate the possibility of selection of the patients with expected high tolerance to FAC treatment and consequently with high chance of chemotherapy completion without the dose reduction, treatment delays and decline in the quality of life.

Genetic polymorphisms and response to 5-fluorouracil, doxorubicin and cyclophosphamide chemotherapy in breast cancer patients.

Clinical resistance to chemotherapy is one of the major problems in breast cancer treatment. In this study we analyzed possible impact of 22 polymorphic variants on the treatment response in 324 breast cancer patients. Selected genes were involved in FAC chemotherapy drugs transport (ABCB1, ABCC2, ABCG2, SLC22A16), metabolism (CYP1B1, CYP2C19, GSTT1, GSTM1, GSTP1, TYMS, MTHFR, DPYD), drug-induced damage repair (ERCC1, ERCC2, XRCC1) and involved in regulation of DNA damage response and cell cycle control (ATM, TP53).Apart from preexisting metastases three polymorphic variants were independent prognostic high risk factors of lack of response to FAC chemotherapy. Our results showed that the response to treatment depended of the variability in genes engaged in drugs' transport (ABCC2 c.-24C>T, ABCB1 p.Ser893Ala/Thr) and in DNA repair machinery (ERCC2 p.Lys751Gln). Furthermore, the growing number of high-risk genotypes was reflected in gradual increase in risk of the non-responsiveness to treatment- from OR 2.68 for presence of two genotypes to OR 9.93 for carriers of all three negative genotypes in the group of all patients. Similar gene-dosage effect was observed in the subgroup of TNBCs. Also, TFFS significantly shortened with the increasing number of high-risk genotypes, with median of 54.4 months for carriers of one variant, to 51.5 and 34.9 months for the carriers of two and three genotypes, respectively.Our results demonstrate that results of cancer treatment are the effect of many clinical and genetic factors. It seems that multifactorial polymorphic models could be a potentially useful tool in personalization of cancer therapies. The novelty in our model is the over representation of triple negative breast cancer (TNBC) patients among the carriers of all unfavorable polymorphic variants. This finding contributes to the elucidation of the mechanisms of drug resistance in this subgroup of breast cancer patients.

The truncation of Ku86 in human lymphocytes.

The Ku heterodimer, which consists of Ku70 and Ku86 subunits, is a major sensor of DNA breaks. A truncated form of Ku86 lacking its C-terminus, termed Ku86 variant, has been detected in extracts from different human cells. Here we report that in human lymphocytes the Ku86 variant is not present in vivo but is generated in vitro upon cell lysis by a trypsin-like protease. The resulting Ku86 variant exists exclusively in complexes with Ku70, which possess strong affinity to DNA double strand termini. In different blood donors the levels of Ku86 variant correlated with the magnitude of radiation induced DNA breaks.

The role of chromatin proteins in DNA damage recognition and repair.

The structure of chromatin is the major factor determining the rate and efficiency of DNA repair. Chromatin remodeling events such as rearrangement of nucleosomes and higher order chromatin structures are indispensable features of repair processes. During the last decade numerous chromatin proteins have been identified that preferentially bind to different types of DNA damage. The HMGB proteins, which preferentially interact with DNA intrastrand crosslinks induced by cisplatin, are the archetypal example of such proteins. Several hypothetical models have been proposed describing the role of such damage-binding chromatin proteins. The damage shielding model postulates that binding of chromatin proteins to damaged DNA might disturb damage recognition by repair factors and impair its removal. Alternatively, the damage-recognition/signaling model proposes that the binding of specific chromatin proteins to damaged DNA could serve as a hallmark to be recognized by repair proteins. Additionally, the binding of specific chromatin proteins to damaged DNA could induce chromatin remodeling at the damage site and indirectly affect its repair. This paper aims to critically review current experimental data in relation to such possible roles of chromatin proteins.

Subunit structures and stoichiometries of human DNA fragmentation factor proteins before and after induction of apoptosis.

DNA fragmentation factor (DFF) is one of the major endonucleases responsible for internucleosomal DNA cleavage during apoptosis. Understanding the regulatory checkpoints involved in safeguarding non-apoptotic cells against accidental activation of this nuclease is as important as elucidating its activation mechanisms during apoptosis. Here we address these issues by determining DFF native subunit structures and stoichiometries in human cells before and after induction of apoptosis using the technique of native pore-exclusion limit electrophoresis in combination with Western analyses. For comparison, we employed similar techniques with recombinant proteins in conjunction with atomic force microscopy. Before induction of apoptosis, the expression of DFF subunits varied widely among the cell types studied, and the chaperone/inhibitor subunits DFF45 and DFF35 unexpectedly existed primarily as monomers in vast excess of the latent nuclease subunit, DFF40, which was stoichiometrically associated with DFF45 to form heterodimers. DFF35 was exclusively cytoplasmic as a monomer. Nuclease activation upon caspase-3 cleavage of DFF45/DFF35 was accompanied by DFF40 homo-oligomer formation, with a tetramer being the smallest unit. Interestingly, intact DFF45 can inhibit nuclease activity by associating with these homo-oligomers without mediating their disassembly. We conclude that DFF nuclease is regulated by multiple pre- and post-activation fail-safe steps.