The use of drug combinations (DCs) in cancer therapy can prevent the development of drug resistance and decrease the severity and number of side effects. Synthetic lethality (SL), a genetic interaction wherein two nonessential genes cause cell death when knocked out simultaneously, has been suggested as a method of identifying novel DCs. A combination of two drugs that mimic genetic knockout may cause cellular death through a synthetic lethal pathway. Because SL can be context-specific, it may be possible to find DCs that target SL pairs in tumours while leaving healthy cells unscathed. However, elucidating all synthetic lethal pairs in humans would take more than 200 million experiments in a single biological context – an unmanageably large search space. It is thus necessary to develop computational methods to predict human SL. In this thesis, we develop connectivity homology, a novel measure of network similarity that allows for the comparison of interspecies protein-protein interaction networks. We then use this principle to develop Species-INdependent TRAnslation (SINaTRA), an algorithm that allows us to predict SL between species using protein-protein interaction networks. We validate it by predicting SL in S. pombe from S. cerevisiae, then generate over 100 million SINaTRA scores for putative human SL pairs. We use these data to predict new areas of cancer combination therapy, and then test fifteen of these predictions across several cell lines. Finally, in order to better understand synergy, we develop DAVISS (Data-driven Assessment of Variability In Synergy Scores), a novel way to statistically evaluate the significance of a drug interaction.

Leflunomide is widely prescribed for Rheumatoid Arthritis (RA) patients in China. However, a number of RA patients still demonstrated progressive bone erosion (PBE+) after receiving Leflunomide in our clinical data. Moreover, the PBE+ is predicted by high baseline serum CRP level (CRPBH). Further, the changes of serum bone resorption marker (Tartrate-resistant acid phosphatase 5b, TRAP5b) strongly correlated with those of CRP in PBE+ RA patients during Leflunomide treatment. Those were consistently observed in collagen-induced-arthritis (CIA) rats. To precisely address the issue, we screened a series of marketed drugs combined with Leflunomide to inhibit CRP production and CRP-related osteoclastic signaling pathway using bioinformatics analysis. Ligustrazine was postulated as an optimal candidate drug. In vitro studies demonstrated that the combination of Ligustrazine and Leflunomide not only suppressed hepatic CRP production, but also suppressed CRP-related osteoclastic signaling and osteoclast activities. In vivo studies showed that the combination attenuated bone erosion in CIA rats. Further, the randomized parallel controlled clinical trial in 120 CRPBH RA patients showed that the combination therapy reduced serum CRP levels and attenuated bone erosion in those patients (ChiCTR-TRC-10001014). Together, this work presents a precision combination therapy for PBE+ in CRPBH RA patients.

Objetivos: O objetivo deste estudo foi o de identificar compostos seletivamente tóxicos ao carcinoma espinocelular de boca in vitro por meio do reposicionamento de fármacos. Material e Métodos: Por meio de um escaneamento baseado na viabilidade celular de 1.280 fármacos, nós selecionamos três princípios ativos (luteolin, metixene hydrochloride e nitazoxanide) letais às células de câncer de boca SCC-25 e pouco tóxicos às células de queratinócitos cutâneos imortalizados HaCaT. Os fármacos candidatos foram investigados quanto à sua dose- e tempo-resposta bem como comparados e combinados à agentes quimioterápicos padrão por meio do ensaio por colorimetria com brometo de tiazolil azul de tetrazolio (MTT). O impacto dos fármacos na motilidade do SCC-25 e do HaCaT foi verificado pelo ensaio de migração celular e seus mecanismos de ação também foram explorados por meio da verificação dos níveis das proteínas fosforiladas pelo western blotting. Todos os experimentos foram realizados em triplicata e, pelo menos, três vezes independentes. O teste t de student foi utilizado para confrontar as variáveis e nível de significância de 5% foi estabelecido para todos os testes. Resultados: O flavonoide natural luteolin exerceu citotoxicidade potente contra as células de câncer de boca in vitro, apresentando baixa toxicidade ao HaCaT e alta eficiência quando comparado a quimioterápicos como a cisplatina e o AG1478. Do ponto de vista molecular, a luteolin ativou a via de sinalização do dano do DNA e, combinada com um inibidor do Chk, apresentou efeitos potencializados. Além disso, nós demonstramos que a nitazoxanide e o metixene hydrochloride são capazes de destruir células SCC-25 porém não as HaCaT de maneira proporcional à dose e ao tempo de tratamento. As combinações entre os três fármacos hit e com a cisplatina ou o AG1478 potencializaram seus efeitos contra as células malignas. Conclusões: O presente estudo traz a luteolin, o metixene hydrochloride e a nitazoxanide como fortes candidatos a agentes terapêuticos contra o câncer de boca uma vez que estes compostos apresentam maior eficácia contra células de câncer de boca e menor toxicidade contra células não tumorais in vitro do que agentes quimioterápicos convencionais. / Objectives: Here we aimed at identifying and reposition approved drugs that could be selectively toxic towards oral squamous cell carcinoma cells. Materials and Methods: Through a cell-based drug screening of 1,280 chemical molecules, we selected 3 compounds (luteolin, metixene hydrochloride, and nitazoxanide) lethal to oral cancer SCC-25 cells, while sparing immortalized keratinocyte HaCaT cells. The drugs were then further challenged for their time- and dose-responses, as well as their comparison and combination to standard chemotherapeutic agents by colorimetric assay 1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan, Thiazolyl blue formazan (MTT). The impact on SCC-25 and HaCaT motility as well as the mode of action of the drugs was then further explored by scratching assay and western blotting, respectively. All the experiments were performed in triplicated and, at least, three independent times. Students t test was performed to verify the differences among the variables and the level of significance was set at 5%. Results: The natural flavonoid luteolin was a potent cytotoxic agent against oral cancer cells in vitro, presenting low toxicity against HaCaT cells and high efficiency as compared to standard-of-care, such as cisplatin and AG1478. From a molecular standpoint, luteolin coopted the DNA-damage pathway and could be efficiently combined with Chk pharmacological inhibitor. Moreover, we demonstrated that nitazoxanide and metixene hydrochloride kill the SCC-25 but not the HaCaT cells in a dose- and time-dependent. The combinations among the three drugs hit and with cisplatin and AG1478 improved their effect against the malignant cells. Conclusions: Luteolin, metixene hydrochloride, and nitazoxanide emerge as strong cytotoxic and/or adjuvant therapy in oral cancer, as these compounds present higher efficiency and lower toxicity against oral cancer cells in vitro than conventional chemotherapeutic agents.

Objetivos: O objetivo deste estudo foi o de identificar compostos seletivamente tóxicos ao carcinoma espinocelular de boca in vitro por meio do reposicionamento de fármacos. Material e Métodos: Por meio de um escaneamento baseado na viabilidade celular de 1.280 fármacos, nós selecionamos três princípios ativos (luteolin, metixene hydrochloride e nitazoxanide) letais às células de câncer de boca SCC-25 e pouco tóxicos às células de queratinócitos cutâneos imortalizados HaCaT. Os fármacos candidatos foram investigados quanto à sua dose- e tempo-resposta bem como comparados e combinados à agentes quimioterápicos padrão por meio do ensaio por colorimetria com brometo de tiazolil azul de tetrazolio (MTT). O impacto dos fármacos na motilidade do SCC-25 e do HaCaT foi verificado pelo ensaio de migração celular e seus mecanismos de ação também foram explorados por meio da verificação dos níveis das proteínas fosforiladas pelo western blotting. Todos os experimentos foram realizados em triplicata e, pelo menos, três vezes independentes. O teste t de student foi utilizado para confrontar as variáveis e nível de significância de 5% foi estabelecido para todos os testes. Resultados: O flavonoide natural luteolin exerceu citotoxicidade potente contra as células de câncer de boca in vitro, apresentando baixa toxicidade ao HaCaT e alta eficiência quando comparado a quimioterápicos como a cisplatina e o AG1478. Do ponto de vista molecular, a luteolin ativou a via de sinalização do dano do DNA e, combinada com um inibidor do Chk, apresentou efeitos potencializados. Além disso, nós demonstramos que a nitazoxanide e o metixene hydrochloride são capazes de destruir células SCC-25 porém não as HaCaT de maneira proporcional à dose e ao tempo de tratamento. As combinações entre os três fármacos hit e com a cisplatina ou o AG1478 potencializaram seus efeitos contra as células malignas. Conclusões: O presente estudo traz a luteolin, o metixene hydrochloride e a nitazoxanide como fortes candidatos a agentes terapêuticos contra o câncer de boca uma vez que estes compostos apresentam maior eficácia contra células de câncer de boca e menor toxicidade contra células não tumorais in vitro do que agentes quimioterápicos convencionais. / Objectives: Here we aimed at identifying and reposition approved drugs that could be selectively toxic towards oral squamous cell carcinoma cells. Materials and Methods: Through a cell-based drug screening of 1,280 chemical molecules, we selected 3 compounds (luteolin, metixene hydrochloride, and nitazoxanide) lethal to oral cancer SCC-25 cells, while sparing immortalized keratinocyte HaCaT cells. The drugs were then further challenged for their time- and dose-responses, as well as their comparison and combination to standard chemotherapeutic agents by colorimetric assay 1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan, Thiazolyl blue formazan (MTT). The impact on SCC-25 and HaCaT motility as well as the mode of action of the drugs was then further explored by scratching assay and western blotting, respectively. All the experiments were performed in triplicated and, at least, three independent times. Students t test was performed to verify the differences among the variables and the level of significance was set at 5%. Results: The natural flavonoid luteolin was a potent cytotoxic agent against oral cancer cells in vitro, presenting low toxicity against HaCaT cells and high efficiency as compared to standard-of-care, such as cisplatin and AG1478. From a molecular standpoint, luteolin coopted the DNA-damage pathway and could be efficiently combined with Chk pharmacological inhibitor. Moreover, we demonstrated that nitazoxanide and metixene hydrochloride kill the SCC-25 but not the HaCaT cells in a dose- and time-dependent. The combinations among the three drugs hit and with cisplatin and AG1478 improved their effect against the malignant cells. Conclusions: Luteolin, metixene hydrochloride, and nitazoxanide emerge as strong cytotoxic and/or adjuvant therapy in oral cancer, as these compounds present higher efficiency and lower toxicity against oral cancer cells in vitro than conventional chemotherapeutic agents.

The area of systems biology evolved in an attempt to introduce mathematical systems theory principles in biology. Although we believe that all biological processes are essentially chemical reactions, describing those using precise mathematical rules is not easy, primarily due to the complexity and enormity of biological systems. Here we introduce a formal approach for modeling biological dynamical relationships and diseases such as cancer. The immediate motivation behind this research is the urgency to find a practicable cure of cancer, the emperor of all maladies. Unlike other deadly endemic diseases such as plague, dengue and AIDS, cancer is characteristically heterogenic and hence requires a closer look into the genesis of the disease. The actual cause of cancer lies within our physiology. The process of cell division holds the clue to unravel the mysteries surrounding this disease. In normal scenario, all control mechanisms work in tandem and cell divides only when the division is required, for instance, to heal a wound platelet derived growth factor triggers cell division. The control mechanism is tightly regulated by several biochemical interactions commonly known as signal transduction pathways. However, from mathematical point of view, these pathways are marginal in nature and unable to cope with the multi-variability of a heterogenic disease like cancer. The present research is possibly one first attempt towards unraveling the mysteries surrounding the dynamics of a proliferating cell. A novel yet simple methodology is developed to bring all the marginal knowledge of the signaling pathways together to form the simplest mathematical abstract known as the Boolean Network. The malfunctioning in the cell by genetic mutations is formally modeled as stuck-at faults in the underlying Network. Finally a mathematical methodology is discovered to optimally find out the possible best combination drug therapy which can drive the cell from an undesirable condition of proliferation to a desirable condition of quiescence or apoptosis. Although, the complete biological validation was beyond the scope of the current research, the process of in-vitro validation has been already initiated by our collaborators. Once validated, this research will lead to a bright future in the field on personalized cancer therapy.

<p>  </p> <p>Metastatic breast cancer (MBC) is an extremely recalcitrant disease capable of overcoming targeted therapies and evading immune surveillance via the engagement of complicated signaling networks. Resistance to targeted therapies and therapeutic failure of immune checkpoint blockade (ICB) are two major challenges in treating MBC. To survive in the dynamic tumor microenvironment (TME) during metastatic progression, shared signaling nodes are required for MBC cells to regulate the signaling networks efficiently, which are potential multifunctional therapeutic targets. SH2 containing protein tyrosine phosphatase-2 (SHP2) is a druggable oncogenic phosphatase that is a key shared node in both tumor cells and immune cells. How tumor-cell autonomous SHP2 manages its signaling inputs and outputs to facilitate the growth of tumor cells, drug resistance, immunosuppression, and the limited response of ICB in MBC is not fully understood. Herein, we used inducible genetic depletion and two distinct types of pharmacological inhibitors to investigate anti-tumor effects with immune reprogramming during SHP2 targeting. </p> <p>We first focus on the signaling inputs and outputs of SHP2. We find that phosphorylation of SHP2 at Y542 predicts the survival rates of breast cancer patients and their immune profiles. Phosphorylation of SHP2 at Y542 is elevated with differential activation mechanisms under a growth-factor-induced and extracellular matrix (ECM)-rich culture environment. Phosphorylation of SHP2 at Y542 is also elevated in HER2 positive MBC cells upon acquired resistance to the HER2 kinase inhibitor, neratinib. The resistant cells can be targeted by SHP2 inhibitors. SHP2 inhibitors block ERK1/2 and AKT signaling and readily prevented MBC cell growth induced by multiple growth factors. Inhibition of SHP2 also blocks these signaling events generated from the ECM signaling. In fact, the inhibitory effects of SHP2 blockade are actually enhanced in the ECM-rich culture environment. We utilize the <em>in vitro</em> T-cell killing assays and demonstrate that pretreatment of tumor cells with FGF2 and PDGF reduces the cytotoxicity of CD8+ T cells in a SHP2-dependent manner. Both growth factors and ECM-rich culture environment transcriptionally induce PD-L1 via SHP2. SHP2 inhibition balances MAPK signaling and STAT1 signaling, which prevents growth factor-mediated suppression of INF-γ-induced expression of MHC class I. </p> <p>Next, we evaluate the efficacy of SHP2 inhibitors. Blockade of SHP2 in the adjuvant setting decreased pulmonary metastasis <em>in vivo</em> and extended the survival of systemic tumor-bearing mice. Tumor-cell autonomous depletion of SHP2 reduces pulmonary metastasis and relieves exhaustion markers on CD8+ and CD4+ cells. Meanwhile, both systemic SHP2 inhibition and tumor-cell autonomous SHP2 depletion reduce tumor-infiltrated CD4+ T cells and M2-polarized tumor associated macrophages. </p> <p>Finally, we investigate potential combination therapies with SHP2 inhibitors. The combination of SHP2 inhibitors and FGFR-targeted kinase inhibitors synergistically blocks the growth of MBC cells. Pharmacological inhibition SHP2 sensitizes MBC cells growing in the lung to α-PD-L1 antibody treatment via relieving T cell exhaustion induced by ICB. </p> <p>Overall, our findings support the conclusion that MBC cells are capable of simultaneously engaging several survival pathways and immune-suppressive mechanisms via SHP2 in response to multiple growth factors and ECM signaling. Inhibition of SHP2, potentially in combination with other targeted agents and ICB, holds promise for the therapeutic management of MBC.</p>