Thymineless stress, DNA repair and recombination

描述（由申请人提供）：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。胸苷酸剥夺是通过抑制胸苷酸合成酶（TS）诱导的，并且是几类抗肿瘤药物（如5-氟尿嘧啶和雷替曲塞（Tomudex））的治疗效果。TS的抑制导致复制所必需的TTP的丢失。胸苷酸缺乏导致细胞死亡，不像与其他营养缺乏相关的细胞抑制作用。尽管经过数十年的研究，TS抑制导致死亡的确切机制仍不清楚。对TS抑制的一些细胞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、S期细胞周期停滞和诱导DNA链断裂（可能在复制位点）。一个关键的悬而未决的问题仍然是“在导致细胞死亡的胸苷酸剥夺过程中，损伤的具体性质是什么？“在这个项目中要测试的假设是，在胸苷酸剥夺的条件下，碱基切除修复（BER）的激活和进展导致异常重组，并最终导致细胞凋亡。本项目的具体目的是：目的1：确定胸苷酸剥夺期间BER的启动和进展是否有助于细胞死亡。目的2：确定胸苷酸剥夺过程中BER中间体的命运。目标3：确定胸苷酸剥夺是否诱导染色体重组，并研究BER对胸苷酸剥夺过程中发生的重组的影响。由于BER和重组通常有助于基因组的稳定性，当目前的癌症治疗本身可以诱导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.