Synthesis and characterization of chemically modified oligonucleotides as probes of DNA repair processes.

链间交链(ICL)可以作为脂质过氧化等内源性过程的结果在DNA中形成，是细胞遇到的一些毒性最大的损伤，因为它们阻碍了对复制、转录和重组过程至关重要的两条DNA的解离。ICL对细胞内这些事件的干扰可以在使用双功能烷化剂的癌症化疗方案中加以利用。然而，这些药物的效力被癌细胞修复这些药物诱导的相同损伤的能力所削弱，这导致了耐药性的发生。细胞中存在多种DNA修复途径，包括直接修复、碱基切除修复、核苷酸切除修复、同源重组、非同源末端连接和DNA错配修复。虽然同源重组和核苷酸切除修复与细菌和哺乳动物细胞的ICL修复有关，但其涉及的机制仍然缺乏阐明。对识别和修复DNA损伤的分子过程的详细了解将导致开发更有效的烷化剂和治疗方案。我们的团队开发了一种结合溶液和固相合成的方法来制备足够数量的含有ICL的寡核苷酸，用于生物物理、结构和修复研究。这些研究将评估这种损伤是否会导致DNA的结构变化，这可能会在修复酶的识别中发挥作用。此外，这些ICL DNA将用于DNA修复蛋白的实验，以观察这些ICL是易感的还是逃避修复过程。结构和修复敏感性之间的相关性将有助于我们理解这些治疗相关损伤的处理过程，并将有助于DNA修复领域。

Interstrand cross-links (ICL) can be formed in DNA as a result of endogenous processes such as lipid peroxidation and represent some of the most toxic lesions encountered by cells because they obstruct unwinding of the two strands of DNA which is critical to the processes of replication, transcription and recombination. Interference with these events in the cell by ICLs can be exploited in cancer chemotherapy regimens that employ bifunctional alkylating agents. However, the potency of these agents is diminished by the ability of cancer cells to repair the same lesions these agents induce and this causes the onset of resistance. A number of DNA repair pathways exist in cells including direct repair, base-excision repair, nucleotide-excision repair, homologous recombination, nonhomologous end joining and DNA-mismatch repair. Although homologous recombination and nucleotide excision repair have been implicated in ICL repair in bacteria and mammalian cells, elucidation of the mechanism involved is still lacking. A detailed understanding of the molecular processes by which DNA lesions are recognized and repaired will lead to the development of more effective alkylating agents and treatment regimens. Our group has developed methods using a combination of solution and solid-phase synthesis to prepare sufficient quantities of ICL containing oligonucleotides for biophysical, structural and repair studies. These studies will assess whether such lesions induce a structural change in DNA that may play a role in recognition by repair enzymes. In addition, these ICL DNA will be used in experiments with DNA repair proteins to observe whether these ICL are either susceptible or evade repair processes. Correlations made between structure and repair susceptibility will aid our understanding of the processing of these therapeutically relevant lesions and will contribute to the field of DNA repair.