Spatially defined peptidomimetics for pain relief and for targeted cancer therapy and diagnostics

Cancer is a disease that has plagued scientists for decades, and how to treat cancer and its complications are inevitable topics in current scientific research. Cancer pain is a major factor that reduces the quality of life of patients. Therefore, the development of analgesic agents with minimal adverse side effects, especially with low addiction, has attracted more and more attention. Among them, opioid analgesics are widely used to alleviate cancer pain and improve the quality of life of patients with advanced cancer, such as in the palliative therapy. Although peptide drugs are efficient, selective and safe, they have several unignorable disadvantages such as poor biological stability, rapid excretion, difficulty in penetrate blood brain barrier. In order to solve these problems, peptidomimetics were developed by introducing unnatural/modified amino acids, decorated peptide backbone, conformational restrictions and secondary structure mimics in peptide sequence. Compared with peptides, peptidomimetics have improved biological stability, increased bioavailability, high affinity and selectivity for receptor binding, and decreased adverse side effects. As the second part of this thesis, I explored the opportunity to design peptide-functionalized responsive biomaterials for the detection of cancer cell and the selective delivery of cytotoxic drugs. The conjugation of peptides with biomaterials enhanced the stability of the loaded drugs, improved targeted delivery, decreased side effects, and increased bioavailability. The precise and controllable drug delivery platform has profound application prospects in cancer treatment. Grafting specific peptides sequence on the surface of biomaterials can satisfy different drug delivery demands according to the characteristics of both peptides and biomaterials. For example, the introduction of tumor-targeting peptides can guide biomaterials into tumor lesions, and blood-brain barrier (BBB) shuttle peptides can lead biomaterials to penetrate the BBB, etc.

The role of Proteasome in the development, maintenance and management of Chronic Pain

In chronic pain, opioids represent the gold standard analgesics, but their use is hampered by the development of several side effects, as development of analgesic tolerance and opioid-induced hyperalgesia. Evidence showed that many molecular mechanisms (changes in opioid receptors, neurotransmitter release, and glia/microglia activation) are involved in their appearance, as well as in chronic pain. Recently, a crucial role has been proposed for oxidative stress and proteasome in chronic pain and in treatment-related side effects. To better elucidate these aspects, the aim of this PhD thesis was to investigate the effects of opioids on cell oxidative stress, antioxidant enzymatic machinery and proteasome expression and activity in vitro. Also, the involvement of proteasome in the development of chronic pain conditions was investigated utilizing an experimental model of oxaliplatin-induced neuropathy (OXAIN), in vivo. Data showed that morphine, fentanyl, buprenorphine and tapentadol alter differently ROS production. The ROS increasing effect of morphine is not shared by the other opioids, suggesting that the different pharmacological profile could influence this parameter. Moreover, these drugs produced different alterations of β2trypsin-like and β5chymotrypsin-like activities. In fact, while morphine and fentanyl increased the proteolytic activity after prolonged exposure, a different picture was observed for buprenorphine and tapentadol, suggesting that the level of MOR agonism could be strongly related with proteasome activation. In vivo studies revealed that rats treated with oxaliplatin showed a significant increase in β5, in the thalamus (TH) and somatosensory cortex (SSCx). Moreover, a selective up-regulation of β5 and LMP7 subunit gene expression was assessed in the SSCx. Furthermore, our study revealed that oprozomib, a selective β5 inhibitor normalized the spinal prodynorphin gene expression upregulation induced by oxaliplatin, and reverted mechanical/thermal allodynia and mechanical hyperalgesia in oxaliplatin-treated rats. These results underline the role of proteasome in the OXAIN and suggest new pharmacological targets to counteract it.

New senthesis and derivatizations of peptidomimetics and peptide-conjugates for theranostic applications

In the contest of a modern green chemistry approach, we firstly tried to substituent the classic peptide synthesis approach with the use of N-carboxyanhydrides in the presence of Hydroxyapatite, a high biocompatible inorganic base. Despite the great results, further developments are necessary for a daily use in laboratory and for our research, we decided to proceed with solid phase or liquid phase synthesis. In the first chapter, the treatment of pain with the use of opioids is introduced. The abuse and misuse of these kind of potent analgesics, led to the necessity of developing new drugs with less side effects. Starting from a previous study, where the introduction of a lactam-like structure in the place of the proline of Endomorphine1, switched the selectivity from MOR to KOR, we designed and synthetized three different libraries by placing a different trans inducer element to gain the desired selectivity and activity forcing the structure to adopt a linear rather than folded position. In the second chapter, we focused on lactate dehydrogenase, an enzyme overexpressed when the cells in hypoxia conditions, like in a tumour mass, need to produce energy through the transformation of pyruvate into lactate. We synthetized different cyclic peptidomimetics, designed to be inhibitors, as powerful tool to contrast cancer cells growing. Biological assays produced satisfactory preliminary results, but further studies are necessary for a definitive output. Finally in the last chapter, the cancer treatment problem is also approached through the design of nanoparticles, able to deliver drugs with efficacy and selectivity. We firstly synthetized silica core nanoparticles, built with toxic peptide sequences conjugated through click chemistry with Pluronic acid and then, in collaboration with Miriam Royo’s research group, we synthetized multivalent platforms for used drugs for the treatment of advanced colorectal cancer.