SYNTHESIS, CHARACTERIZATION, ANTICANCER ACTIVITY AND REACTIVITY TOWARD BIOLOGIC

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。这项建议的主要目的是寻找与目前使用的基于金属的抗癌药物如顺铂、卡铂等相比具有更好性能的新的铂类药物，并通过研究它们与一些有希望发现的药物的生物重要分子(DNA、寡核苷酸)的反应性来发现其细胞毒性的分子基础。为了实现这些目标，我们计划确定合成和表征两个系列铂(II)配合物的较好条件。1.-铂(II)的阴离子三卤络合物，其中苯基苯并杂唑或重要生物活性羧酸的酰胺衍生物将被用作配体。这些化合物是根据已报道的苯并杂唑三溴阴离子配合物的抗癌活性设计的，与顺铂相比，它显示了不同的交联型式。虽然还没有对这些配合物进行毒性研究，但我们希望通过在其他系列化合物上用与铂(II)中心结合的氯化物取代溴配体来减少任何可能的有害毒性。为了研究配体对这些配合物的影响，将通过在苯并唑和/或苯并基上放置其他取代基(给电子和/或吸电子)以及通过改变配体上苯并唑部分的杂原子(NH，S，O)来对新的苯并苯并杂唑配体进行修饰。其他用于阴离子三卤络合物的配体包括从已报道的生物活性羧酸衍生的酰胺，如2-苯乙酸、苯丁酸、支链和不支链饱和脂肪酸以及不饱和和多不饱和脂肪酸。2.中性铂(II)络合物，其中母体药物中的一个或两个氯离开基团被上述生物活性羧酸取代。母体药物涉及使用顺铂、顺铂(DACH)Cl2和顺铂(En)Cl2，其中DACH代表1，2-二氨基环己烷，En代表乙二胺。这些配体的使用有望增加新配合物的脂亲性和膜通透性，并可能诱导与主要生物活性靶点的新的相互作用方式。此外，羧酸基的存在有望以一种已知的方式产生一种多功能化合物，即离开基团在它与DNA结合之前由铂配合物实现。因此，游离的羧酸盐应该在细胞上发挥自己的生物活性。所有这些化合物都将通过元素分析、UV-Vis光谱、1H、3C、195PT和二维核磁共振实验进行纯化和表征。它们还将被结晶，以获得足够好的晶体，用于X射线衍射研究。针对癌细胞株的生物测试将在里约热内卢皮德拉斯校区UPR的生物测试实验室进行。拟议项目的长期目标是研究有前景的配合物与重要生物分子的相互作用，例如DNA，这是已知铂配合物的主要目标。在这些相互作用之后，将采用标准程序，并通过核磁共振波谱和高效液相色谱对其产品进行表征。为了获得有关新络合物的首选结合部位的信息，将对铂化的DNA进行酶消化，并对其加合物进行层析分离，并通过1H核磁共振波谱进行鉴定。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The major objective of this proposal is to search for new platinum drugs with improved properties compared to currently used metal based anticancer agents such as cisplatin, carboplatin, etc..., and to discover the molecular basis of their cytotoxicity by studying their reactivity toward biological important molecules (DNA, oligonucleotides) of some of the promising discovered drugs. In order to achieve these goals we plan to determine the better conditions for the synthesis and characterization of two series of platinum(II) complexes. 1.- anionic trihalo complexes of platinum(II) in which either a styrylbenzoheterazole or amide derivatives of important biological active carboxylic acids will be used as ligands. These complexes were designed based on the reported anticancer activity of previously synthesized tribromo anionic complexes of styrylbenzoheterazole which demonstrated a different pattern of cross-linking compared to cisplatin. Although no toxicity studies have been performed on these complexes, we expect to decrease any possible unwanted toxicity by substitution of the bromide ligands by chloride bound to the platinum(II) center on other series of compounds. To study the effect of the ligand on these complexes, the new styrylbenzoheterazole ligands to be used will be modified by placing other substituents (electrondonating and/or electronwithdrawing) on either the benzazolo and/or the styryl moieties, and also by changing the heteroatom (NH, S, O) on the benzazolo part of the ligands. Other ligands to be used for the anionic trihalo complexes include amides derived from reported bioactive carboxylic acids, such as 2-phenylacetic acid, phenylbutyric acid, branched and unbranched saturated fatty acids and unsaturated and polyunsaturated fatty acids. 2.- Neutral platinum(II) complexes in which the one or both of the chloride leaving groups in the parent drugs are substituted by the biological active carboxylic acids mentioned above. The parent drugs involves the use of cisplatin, cis-Pt(DACH)CI2, and cis-Pt(en)CI2, where DACH stands for 1,2-diaminocyclohexane and en for ethylenediamine. The use of these ligands are expected to increase the lipophyllicity and membrane permeability of the new complexes and probably to induce new ways of interactions with the main targets for bioactivity. Also the presence of the carboxylate groups are expected to produce a multifunctional compound in such a way that is known that the leaving groups are realized by the platinum complexes before it is bound to DNA. So, the freed carboxylates should exert their own bioactivity on the cells. All of these compounds will be purified and characterized by elemental analysis, UV-Vis spectroscopy, 1H, 3C, 195Pt and two dimension NMR experiments. They will be also crystallized to obtain crystal good enough for X-Ray Diffraction studies. The biological testing against cancer cell lines will be performed at the Biotesting Laboratory at UPR, Rio Piedras Campus. The long term objectives of the proposed projects is to study the interaction of promising complexes with important biological molecules, such as DNA, which is the main target of known platinum complexes. These interactions will be followed by standard procedures andby NMR spectroscopy and HPLC, and their products characterized. In order to obtain information on the preferred sites of binding of the new complexes, the platinated DNA will be enzymatically digested and their adducts separated chromatographically and identified by 1H NMR spectroscopy.