Thymineless stress, DNA repair and recombination

描述(申请人提供)：本申请的长期目标是更好地了解影响正常细胞和癌细胞对化疗反应的DNA损伤机制。胸苷剥夺是由胸苷酸合酶(TS)抑制引起的，是几类抗肿瘤药物如5-氟尿嘧啶和雷替曲塞(Tomudex)的治疗效果。抑制TS会导致复制所必需的TTP的丢失。与其他营养缺乏相关的细胞抑制效应不同，胸苷缺乏会导致细胞死亡。尽管经过了几十年的研究，TS抑制导致死亡的确切机制仍不清楚。细胞对TS抑制的一些反应包括脱氧核苷酸池的改变，包括dUTP水平增加，尿嘧啶掺入DNA，细胞周期停滞在S期，以及诱导DNA链断裂，可能发生在复制部位。一个关键的悬而未决的问题仍然是“在胸苷剥夺过程中导致细胞死亡的损伤的具体性质是什么？”这个项目要检验的假设是，在胸苷剥夺的条件下，碱基切除修复(BER)的激活和进展导致异常重组，最终导致细胞凋亡。该项目的具体目标是：目标1：确定胸腺嘧啶核苷剥夺过程中BER的启动和发展是否有助于细胞死亡。目的2：确定胸腺嘧啶核苷剥夺过程中BER中间体的去向。目的：确定胸腺嘧啶核苷剥夺过程中是否发生染色体重组，并探讨BER对胸腺嘧啶核苷剥夺过程中染色体重组的影响。由于BER和重组通常有助于基因组的稳定，当当前的癌症治疗本身可能导致DNA损伤时，这些问题具有额外的意义。

DESCRIPTION (provided by applicant): The long-term objectives of this application are to better understand the DNA damage mechanisms that influence the response of normal cells and cancer cells to chemotherapy. Thymidylate deprivation is induced by inhibition of thymidylate synthase (TS) and is a therapeutic effect of several classes of antineoplastic drugs, such as 5-fluorouracil and raltitrexed (Tomudex). Inhibition of TS leads to loss of TTP necessary for replication. Thymidylate deprivation leads to cell death, unlike the cytostatic effects associated with other nutritional deficiencies. Despite decades of study, the precise mechanism by which TS inhibition causes death remains unclear. Some cellular responses to TS inhibition include an alteration in deoxynucleotide pools including an increase in dUTP levels, uracil incorporation into DNA, cell cycle arrest during S-phase, and induction of DNA strand breaks, likely at sites of replication. A key unanswered question remains "what is the specific nature of the damage during thymidylate deprivation that results in cell death?" The hypothesis to be tested in this project is that activation and progression of base excision repair (BER) under conditions of thymidylate deprivation lead to aberrant recombination and, eventually, apoptosis. The Specific Aims of this project are: Aim 1: To determine whether the initiation and progression of BER during thymidylate deprivation contributes to cell death. Aim 2: To determine the fate of BER intermediates during thymidylate deprivation. Aim 3: To determine whether chromosomal recombination is induced during thymidylate deprivation and to investigate the influence of BER on recombination occurring during thymidylate deprivation. Because BER and recombination normally contribute to genome stability, these questions have an added significance when current cancer therapies can themselves induce DNA damage.