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Apoptosis is a key cell death pathway in mammalian cells. Understanding this process and its regulation has been a subject of study in the last three decades. Members of the Bcl-2 family of proteins are involved in the regulation of apoptosis through mitochondrial poration with the subsequent initiation of apoptosis. Deregulation of proapoptotic proteins contributes to the progression of many tumor processes. Understanding how these pore-forming Bcl-2 proteins Bak and Bax are activated is key to find new anticancer treatments. As no drug capable of activating Bak has been disclosed yet, the study of the structural features of BH3 peptides-known as Bak activators-relevant for binding along with its binding energy decomposition analysis, becomes essential for designing novel small-molecule mimics of BH3. Interestingly, a BH3 Bim analogue-inactivating Bak has recently been discovered, opening a question on the molecular features that determine the functions of BH3 peptides. Therefore, the present work is aimed at understanding the way BH3 peptides activate or inactivate Bak in order to identify differential structural features that can be used in drug design. For this purpose, complexes of Bak with an activator and an inhibitor have been subjected to a molecular dynamics study. Structural differences were assessed by means of the fluctuations of the corresponding principal components. Moreover, the MMPB/GBSA approach was used to compute the binding free energy of the diverse complexes to identify those residues of the BH3 peptide that exhibit the larger contributions to complex formation. The results obtained in this work show differences between activators and inhibitors, both in structural and energetic terms, which can be used in the design of new molecules that can activate or inactivate proapoptotic Bak

This project was undertaken in attempt to improve the efficacy of the drug delivery while avoiding systemic toxicity, designing a novel implantable drug delivery platform for iontophoretic brain cancer therapy. 3D fabrication is fast becoming key instrument in the medical field. Thus, we explored microfabrication of an implantable drug delivery device, concretely using direct laser writing to electrostatically deliver charged cancer drugs into brain tumours, technology which is called iontophoresis – basically relies on the motion of ions in electric fields. This technology avoids pressure-related problems, such as drug reflux and brain oedema formation, encountered with convection enhanced delivery by solely relying on the motion of ions in electric fields. The reason for the interest in iontophoresis is that this technique is very versatile allowing to deliver substances into tissue which are usually impermeable. Additionally, it opens up the possibility to deliver drugs in a targeted fashion directly into the desired tissue instead of administering it systemically which reduces off-target toxicity and allows higher delivery rates than when relying solely on passive diffusion of the drug. Overall, this study strengthens the idea that very precise microdevices can be fabricated using Nanoscribe while meeting the biocompatibility criteria, which opens the door to future applications in other areas such as neurological diseases and other cancer treatments Aquest projecte es va dur a terme per intentar millorar l'eficàcia del lliurament de fàrmacs, evitant la toxicitat sistèmica, dissenyant una nova plataforma de lliurament de fàrmacs implantables en malalts de càncer cerebral. Concretament es va utilitzar escriptura làser directa per lliurar electrostàticament fàrmacs contra el càncer amb càrrega als tumors cerebrals, tecnologia que s'anomena iontoforesi: bàsicament es basa en el moviment dels ions en camps elèctrics. Aquesta tecnologia evita problemes relacionats amb la pressió, com ara el reflux de fàrmacs i la formació d'edema cerebral. La raó per l'interès de la iontoforesi és que aquesta tècnica és molt versàtil i permet transferir substàncies al teixit que solen ser impermeables. A més, obre la possibilitat de lliurar medicaments de manera específica directament al teixit desitjat en lloc d'administrar-lo d’una forma sistèmica, fet que redueix la toxicitat fora de l'objectiu i permet taxes de lliurament més altes que quan es confia només en la difusió passiva del fàrmac. En general, aquest estudi reforça la idea que es poden fabricar microdispositius molt precisos que compleixen els criteris de biocompatibilitat. Aquest fet obre la porta al futures aplicacions en altres àrees com les malalties neurològiques i altres tractaments contra el càncer

Different EGFR tyrosine kinase inhibitors (TKIs) have been developed for the treatment of non-small cell lung cancer (NSCLC) patients harboring sensitizing mutations in the EGFR gene. Apart from acquired secondary mutations, multiple resistance mechanisms have been reported, such as the overexpression of fatty acid synthase (FASN), a multi-functional enzyme essential for the de novo lipogenesis, or the increase of cancer stem cells, a small subpopulation within the tumor responsible for relapse, metastasis, and resistance to therapies. Hence, the purpose of this work is to evaluate the novel FASN inhibitor AZ12756122, both alone and in combination with gefitinib and osimertinib, in EGFR-mutated (EGFRm) lung adenocarcinoma cell models sensitive and resistant to EGFR-TKIs. The molecular effect of AZ12756122 (alone and in combination with EGFR-TKI) on FASN, EGFR/ STAT3, Akt/mTOR, and MAPK signaling pathways was analyzed using RT-qPCR and Western blot. FASN expression was also evaluated in samples from patients with EGFRm NSCLC through immunohistochemistry. Our findings revealed that AZ12756122 caused cytotoxic effects inducing apoptosis, downregulated FASN expression and activity, decreased the activation of EGFR and Akt/mTOR pathway, and reduced cancer stem-like cells. Furthermore, the combination of AZ12756122 and osimertinib sensitized cells to EGFR-TKI, showing a syner- gistic effect that resulted in a reduction in the activation of EGFR, Akt/mTOR, and MAPK signaling pathways. Our study also showed that FASN+ EGFRm NSCLC patients exhibited a longer mPFS in patients who responded to EGFR-TKI treatment. In conclusion, FASN inhibition should be further studied for the treatment, alone or in combination with EGFR-TKIs, for EGFRm NSCLC patients

Inhibitors of the lipogenic enzyme fatty acid synthase (FASN) have attracted much attention in the last decade as potential targeted cancer therapies. However, little is known about the molecular determinants of cancer cell sensitivity to FASN inhibitors (FASNis), which is a major roadblock to their therapeutic application. Here, we find that pharmacological starvation of endogenously produced FAs is a previously unrecognized metabolic stress that heightens mitochondrial apoptotic priming and favors cell death induction by BH3 mimetic inhibitors. Evaluation of the death decision circuits controlled by the BCL-2 family of proteins revealed that FASN inhibition is accompanied by the upregulation of the pro-death BH3-only proteins BIM, PUMA, and NOXA. Cell death triggered by FASN inhibition, which causally involves a palmitate/NADPH-related redox imbalance, is markedly diminished by concurrent loss of BIM or PUMA, suggesting that FASN activity controls cancer cell survival by fine-tuning the BH3 only proteins-dependent mitochondrial threshold for apoptosis. FASN inhibition results in a heightened mitochondrial apoptosis priming, shifting cells toward a primed-for-death state “addicted” to the anti-apoptotic protein BCL-2. Accordingly, co-administration of a FASNi synergistically augments the apoptosis-inducing activity of the dual BCL-XL/BCL-2 inhibitor ABT-263 (navitoclax) and the BCL-2 specific BH3-mimetic ABT-199 (venetoclax). FASN inhibition, however, fails to sensitize breast cancer cells to MCL-1- and BCL-XL-selective inhibitors such as S63845 and A1331852. A human breast cancer xenograft model evidenced that oral administration of the only clinically available FASNi drastically sensitizes FASN-addicted breast tumors to ineffective single-agents navitoclax and venetoclax in vivo. In summary, a novel FASN-driven facet of the mitochondrial priming mechanistically links the redox-buffering mechanism of FASN activity to the intrinsic apoptotic threshold in breast cancer cells. Combining next-generation FASNis with BCL-2-specific BH3 mimetics that directly activate the apoptotic machinery might generate more potent and longer-lasting antitumor responses in a clinical setting., The authors would like to thank Dr. Kenneth McCreath for editorial support. This work was supported by the NIH National Cancer Institute Grants R01 CA116623 (to Ruth Lupu) and R01 CA166741 (to Scott H. Kaufmann) and by the U.S. Department of Defense (DOD)-Breakthrough 3 Grants BC151072 and BC151072P1 (to Ruth Lupu). Work in the Menendez laboratory is supported by the Spanish Ministry of Science and Innovation (Grants SAF2016-80639-P and PID2019-10455GB-I00, Plan Nacional de l + D + I, founded by the European Regional Development Fund, Spain) and by an unrestricted research grant from the Fundació Oncolliga Girona (Lliga catalana d’ajuda al malalt de càncer, Girona). Joan Montero acknowledges support from the Ramon y Cajal Programme, Ministerio de Economía y Competitividad (RYC-2015-18357) and the Spanish National Plan “Retos Investigación” I + D + I (RTI2018-094533-A-I00) from the Ministerio de Ciencia, Innovación y Universidades. Elisabet Cuyàs holds a research contract “Miguel Servet” (CP20/00003) from the Instituto de Salud Carlos III, Spanish Ministry of Science and Innovation (Spain). All authors have read and agreed to the published version of the manuscript.

Thymidine kinase expressing human adipose mesenchymal stem cells (TK-hAMSCs) in combination with ganciclovir (GCV) are an effective platform for antitumor bystander therapy in mice models. However, this strategy requires multiple TK-hAMSCs administrations and a substantial number of cells. Therefore, for clinical translation, it is necessary to find a biocompatible scaffold providing TK-hAMSCs retention in the implantation site against their rapid wash-out. We have developed a microtissue (MT) composed by TKhAMSCs and a scaffold made of polylactic acid microparticles and cell-derived extracellular matrix deposited by hAMSCs. The efficacy of these MTs as vehicles for TK-hAMSCs/GCV bystander therapy was evaluated in a rodent model of human prostate cancer. Subcutaneously implanted MTs were integrated in the surrounding tissue, allowing neovascularization and maintenance of TK-hAMSCs viability. Furthermore, MTs implanted beside tumors allowed TK-hAMSCs migration towards tumor cells and, after GCV administration, inhibited tumor growth. These results indicate that TK-hAMSCs-MTs are promising cell reservoirs for clinical use of therapeutic MSCs in bystander therapies., This work was supported by the Severo Ochoa Program for Centers of Excellence in R&D 2016–2019, the European Commission-ERANET (nAngioderm JTC2018-103), the Spanish network of cell therapy (TERCEL), the Spanish Ministry of Science, Innovation and Universities (MAT2015-68906-R), the Spanish State Research Agency (AEI) and the European Regional Development Fund (FEDER) grants. BBF acknowledge finanical support through the BEST Postdoctoral Program, funded by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie (grant agreement no. 712754) and from the Spanish Ministry of Economy and Competitiveness under Severo Ochoa grants (SEV-2014-0425 and CEX2018-000789-S). GRS is thankful to the Spanish Ministry of Economy, Industry and Competitiveness for his fellowship (BES-2016-077182). The authors specially thank Dr. Josep Roca, from Delfos hospital (Dr. Roca I Noguera aesthetic surgery team), for the kind donation of liposuction for hAMSCs preparation; and to the Services of cell culture (IQAC-CISC), animal care (IQAC-CSIC) and cell sorting (CCiT-University of Barcelona) for their technician and specialized support.

Molecular profiling of personal cancer genomes, and the identification of actionable vulnerabilities and drug-response biomarkers, are the basis of precision oncology. Tumors often present several driver alterations that might be connected by cross-talk and feedback mechanisms, making it difficult to mark single oncogenic variations as reliable predictors of therapeutic outcome. In the current work, we uncover and exploit driver alteration co-occurrence patterns from a recently published in vivo screening in patient-derived xenografts (PDXs), including 187 tumors and 53 drugs. For each treatment, we compare the mutational profiles of sensitive and resistant PDXs to statistically define Driver Co-Occurrence (DCO) networks, which capture both genomic structure and putative oncogenic synergy. We then use the DCO networks to train classifiers that can prioritize, among the available options, the best possible treatment for each tumor based on its oncogenomic profile. In a cross-validation setting, our drug-response models are able to correctly predict 66% of sensitive and 77% of resistant drug-tumor pairs, based on tumor growth variation. Perhaps more interesting, our models are applicable to several tumor types and drug classes for which no biomarker has yet been described. Additionally, we experimentally validated the performance of our models on 15 new tumor samples engrafted in mice, achieving an overall accuracy of 75%. Finally, we adapted our strategy to derive drug-response models from continuous clinical outcome measures, such as progression free survival, which better represent the data acquired during routine clinical practice and in clinical trials. We believe that the computational framework presented here could be incorporated into the design of adaptive clinical trials, revealing unexpected connections between oncogenic alterations and increasing the clinical impact of genomic profiling.

Epidermal growth factor receptor (EGFR) tyrosine kinases inhibitors (TKIs) are effective therapies for non-small cell lung cancer (NSCLC) patients whose tumors harbor an EGFR activating mutation. However, this treatment is not curative due to primary and secondary resistance such as T790M mutation in exon 20. Recently, activation of transducer and activator of transcription 3 (STAT3) in NSCLC appeared as an alternative resistance mechanism allowing cancer cells to elude the EGFR signaling. Overexpression of fatty acid synthase (FASN), a multifunctional enzyme essential for endogenous lipogenesis, has been related to resistance and the regulation of the EGFR/Jak2/STAT signaling pathways. Using EGFR mutated (EGFRm) NSCLC sensitive and EGFR TKIs&rsquo, resistant models (Gefitinib Resistant, GR) we studied the role of the natural polyphenolic anti-FASN compound (&minus, )-epigallocatechin-3-gallate (EGCG), and its derivative G28 to overcome EGFR TKIs&rsquo, resistance. We show that G28&rsquo, s cytotoxicity is independent of TKIs&rsquo, resistance mechanisms displaying synergistic effects in combination with gefitinib and osimertinib in the resistant T790M negative (T790M&minus, ) model and showing a reduction of activated EGFR and STAT3 in T790M positive (T790M+) models. Our results provide the bases for further investigation of G28 in combination with TKIs to overcome the EGFR TKI resistance in NSCLC.

Homocisteïna; Càncer de mama; Dieta cetogènica Homocisteína; Cáncer de mama; Dieta cetogénica Homocysteine; Breast cancer; Ketogenic diet Certain dietary interventions might improve the therapeutic index of cancer treatments. An alternative to the “drug plus diet” approach is the pharmacological reproduction of the metabolic traits of such diets. Here we explored the impact of adding metformin to an established therapeutic regimen on the systemic host metabolism of cancer patients. A panel of 11 serum metabolites including markers of mitochondrial function and intermediates/products of folate-dependent one-carbon metabolism were measured in paired baseline and post-treatment sera obtained from HER2-positive breast cancer patients randomized to receive either metformin combined with neoadjuvant chemotherapy and trastuzumab or an equivalent regimen without metformin. Metabolite profiles revealed a significant increase of the ketone body β-hydroxybutyrate and of the TCA intermediate α-ketoglutarate in the metformin-containing arm. A significant relationship was found between the follow-up levels of homocysteine and the ability of treatment arms to achieve a pathological complete response (pCR). In the metformin-containing arm, patients with significant elevations of homocysteine tended to have a higher probability of pCR. The addition of metformin to an established anti-cancer therapeutic regimen causes a fasting-mimicking modification of systemic host metabolism. Circulating homocysteine could be explored as a clinical pharmacodynamic biomarker linking the antifolate-like activity of metformin and biological tumor response. This work was supported by grants from the Ministerio de Sanidad, Servicios Sociales e Igualdad (EC10-125, Ayudas para el Fomento de la Investigación Clínica Independiente to Begoña Martin-Castillo). Work in the Menendez laboratory is supported by the Ministerio de Ciencia e Innovación (Grant SAF2016-80639-P, Plan Nacional de l+D+I, founded by the European Regional Development Fund [EU FEDER], Spain) and by an unrestricted research grant from the Fundació Oncolliga Girona (Lliga catalana d’ajuda al malalt de càncer, Girona).

Acquired resistance to trastuzumab is a major clinical problem in the treatment of HER2-positive (HER2+) breast cancer patients. The selection of trastuzumab-resistant patients is a great challenge of precision oncology. The aim of this study was to identify novel epigenetic biomarkers associated to trastuzumab resistance in HER2+ BC patients. Methods: We performed a genome-wide DNA methylation (450K array) and a transcriptomic analysis (RNA-Seq) comparing trastuzumab-sensitive (SK) and trastuzumab-resistant (SKTR) HER2+ human breast cancer cell models. The methylation and expression levels of candidate genes were validated by bisulfite pyrosequencing and qRT-PCR, respectively. Functional assays were conducted in the SK and SKTR models by gene silencing and overexpression. Methylation analysis in 24 HER2+ human BC samples with complete response or non-response to trastuzumab-based treatment was conducted by bisulfite pyrosequencing. Results: Epigenomic and transcriptomic analysis revealed the consistent hypermethylation and downregulation of TGFBI, CXCL2, and SLC38A1 genes in association with trastuzumab resistance. The DNA methylation and expression levels of these genes were validated in both sensitive and resistant models analyzed. Of the genes, TGFBI presented the highest hypermethylation-associated silencing both at the transcriptional and protein level. Ectopic expression of TGFBI in the SKTR model suggest an increased sensitivity to trastuzumab treatment. In primary tumors, TGFBI hypermethylation was significantly associated with trastuzumab resistance in HER2+ breast cancer patients. Conclusions: Our results suggest for the first time an association between the epigenetic silencing of TGFBI by DNA methylation and trastuzumab resistance in HER2+ cell models. These results provide the basis for further clinical studies to validate the hypermethylation of TGFBI promoter as a biomarker of trastuzumab resistance in HER2+ breast cancer patients This work was supported in part by the Spanish Instituto de Salud Carlos III (ISCIII; FIS PI11/00692 and PI14/00329; to’ T. Puig), Fundacion Ramon Areces (to T. Puig), the support of Catalonian Government (2017SGR00385), and La Marató de TV3 (20131530, TPuig), financial support was from the University of Girona (MPCUdG2016/036), and the University of Girona and La Caixa Foundation awarded S. Palomeras with a predoctoral grant. A.D.-L. is funded by a “Juan Rodés” contract (JR17/00016) from ISCIII. Ana B Crujeiras is funded by a research contract “Miguel Servet” (CP17/00088) from the ISCIII

Cancer is a worldwide known disease that affects a big part of the population, reason why it’s being studied nowadays in biomedical research. The tyrosine kinase receptors family ErbB is highly studied in cancer therapy, due to its implication in cell proliferation and differentiation pathways. The epidermal growth factor (EGF) is a ligand of this family that binds to its receptor EGFR causing a dimerization and consequently autophosphorylation, driving to the activation of the signaling path. Targeted cancer therapy it’s about guiding a drug to the tumoral cells through determined proteins (usually receptors) involved in the tumoral process. The idea of this project is to address the attack to the epidermal growth factor receptor (EGFR), which is involved in the uncontrolled massive cell proliferation characteristic of this disease, and lock it with a derivative of its own EGF ligand. To convert this ligand into a blocker agent is necessary to do some modifications to the residues that composes it, with the intention of maintain the natural affinity of this peptide to its receptor but avoiding that this causes dimerization, eluding the activation of the cell growth pathway. This final degree work is focused in the study of the natural interaction between the EGF ligand and its EGFR receptor, comparing it with the truncated model (EGFt) done by Panosa et al. in 2013. By molecular dynamics the trajectories of both models are generated and after analyzed by Amber16. Finally, docking of proteins is done by ClusPro 2.0 to foresee the interaction that would be between the modified ligands (EGFm) and the receptor

Gramicidin (GA), a very hydrophobic pentadecapeptide with important biological activities (i.e. in addition to its well-known antimicrobial and antibiotic activities, GA has been recently identified as a potent therapeutic agent against different carcinomas), has been loaded by electrospraying in poly(tetramethylene succinate) (PE44), a biodegradable and biocompatible aliphatic polyester. Microspheres (average diameter: 5.0 ± 0.7 μm) were successfully obtained from the mixture of GA and PE44 solutions in ethanol and chloroform, respectively. The loading of the peptide, which has been proved by FTIR and X-ray photoelectron spectroscopies, essentially occurred at the surface of the microspheres, as was reflected by scanning electron microscopy micrographs and atomic force microscopy phase images. In spite of this, the thermal stability of the polyester matrix remained essentially unaltered, even though the wettability decreased. The release of GA in phosphate buffer saline (PBS) was limited by the very low solubility of the peptide in aqueous solution, a fast burst effect followed by the establishment of equilibrium after 5 days of being observed in this hydrophilic environment. The release behaviour was very different when the hydrophilicity of the medium was reduced by adding ethanol. In this case, a very fast but sustained release was identified during the first few hours. On the other hand, biological tests have demonstrated that GA retains its antimicrobial activity after loading and does not alter the biocompatibility of PE44. Our results prove that, despite its hydrophobicity and relatively large number of residues, the loading of GA in a polymeric matrix represents an alternative strategy for the release of this versatile peptide in cancer therapy., Authors thank supports from MINECO and FEDER (MAT2015-69367-R, MAT2015-69547-R and CTQ2013-40855-R) and Gobierno de Aragón – Fondo Social Europeo (research group E40). Support for the research of C.A. was received through the prize “ICREA Academia” for excellence in research funded by the Generalitat de Catalunya.

Atmospheric pressure plasma jets (APPJ) have great potential in wound healing, bacterial disinfection and in cancer therapy. Recent studies pointed out that hydrogels can be used as screens during APPJ treatment, or even be used as reservoirs for reactive oxygen and nitrogen species generated by APPJ in liquids. Thus, novel applications are emerging for hydrogels which deserve fundamental exploration of the possible modifications undergone by the polymers in solution due to the reactivity with plasmas. Here we investigate the possible modifications occurred by APPJ treatment of an amphiphilic poly(ethylene oxide)-based triblock copolymer (tPEO) photo-crosslinkable hydrogel. While APPJ treatments lead to a certain degradation of the self-assembly of the polymeric chains at low concentrations (2 g/L), the polymeric chains are unaffected by APPJ and the hydrogel forming ability is kept. APPJ treatments induced a pre-crosslinking of the network with an increase of the mechanical properties of the hydrogel. Overall, the small modifications induced allow thinking of polymer solutions with hydrogel forming ability a new platform for several applications related to plasma medicine, and thus, with potential in different therapies.

This work was supported by the Agencia Estatal de Investigación (AEI)[MICINN/Fondo Europeo de Desarrollo Regional (FEDER) (RTC2016-4603-1 to EntreChem SL and SAF-2016-75286-R toR.R.), ISC III/FEDER [Miguel Servet Program] (CPII16/00049 to R.R.), Sara Borrell Program (CD16/00103 toS-T.M.) and Consorcio CIBERONC (CB16/12/00390)] and the Plan de Ciencia Tecnología e Innovación del Principado de Asturias (GRUPIN14-003). We want to particularly acknowledge for its collaboration, the Principado de Asturias BioBank (PT17/0015/0023),financed jointly by Servicio de Salud del Principado de Asturias, Instituto de Salud Carlos III and Fundación Bancaria Cajastur and integrated in the Spanish National Biobanks Network., Estupiñan, O., Santos, L., Rodriguez, A., Fernandez-Nevado, L., Costales, P., Perez-Escuredo, J., Hermosilla, M.A., Oro, P., Rey, V., Tornin, J., Allonca, E., Fernandez-Garcia, M.T., Alvarez-Fernandez, C., Braña, A., Astudillo, A., Menendez, S.T., Moris, F., Rodriguez, R.

The Epidermal Growth Factor Receptor (EGFR) is a glycoprotein that belongs to the family of tyrosine kinase receptors and that it is usually altered in tumor cells. It has been reported that its inhibition decreases tumor proliferation and thus, it is an attractive target for antitumor therapies. There are currently two main strategies of EGFR inhibition: monoclonal antibodies and tyrosine kinase inhibitors (TKI). Unfortunately, the appearance of resistance and other factors limit often the effectiveness of these treatments. In order to study how certain changes in EGFR can affect target therapies against this receptor, we must understand its conformational dynamics. EGFR activation requires the binding of two monomers that, induced by the presence of a ligand, allow then the interaction of the kinase domains and initiate the downstream signaling. However, the deregulation of this receptor may be modulated by several factors, such as the glycosylation of its ectodomine. Tumor progression has been related with an alteration of EGFR glycosylation and it has been observed that aberrant glycosylations may affect both the binding of the ligand and its activation. Accordingly, to study the overall effect of glycosylation on the response to EGFR directed treatments requires a more profound investigation. This is the objective of this Degree Final Project. Based on the existing bibliographic information, it is widely accepted that the site and type of glycosylation influence EGFR conformation and activity. In spite of the controversy in some studies, it seems that terminal fucosylation and sialylation decrease the activity of this receptor. In contrast, other glycosylations such as core fucosylation and Lewis epitopes, increase the activity of EGFR. Moreover, it seems that the alteration of glycosylation could have an effect on EGFR response to treatments. According to current information, EGFR glycosylation does not seem to alter the binding of antibodies to the extracellular domain. However, studies on another member of the EGFR family, HER2, show the opposite. The most relevant result obtained in this project is that extracellular glycosylation of EGFR indirectly affects intracellular sensitivity to TKI. Finally, the need to find alternative treatments to fight emerging resistance and the demand for higher treatment specificity opens an avenue for glycosylation itself as a potential novel target against cancer

Supplemental Digital Content is Available in the Text., Basal cell carcinoma (BCC) seems to originate from ultraviolet light-induced mutations involving the bulge or the outer sheath of the hair follicle cells. However, the etiopathogenic mechanisms involved in the development of these tumors in nonphotoexposed and in hairless areas remain unclear. The cytokeratin (CK) profile (including CK5/6, CK7, CK14, CK15, CK17, and CK19) from a series of different BCC subtypes developing in sun-exposed and non–sun-exposed areas, including hairless regions, was evaluated. The authors have observed that CK7 expression in BCC is associated with the anatomical localization of the tumor and its sun-exposition, but not with other factors such as histological subtype. The expression of this CK is higher in BCCs located in non–sun-exposed and nonhairy areas, such as the vulvar semimucosa and the nipple. Because CK7 is a marker of simple glandular epithelia, the authors suggest a glandular origin for BCCs located in hairless and nonphotoexposed areas.

The undecapeptide KKLFKKILKKL-NH2 (BP16) is a non-toxic cell-penetrating peptide (CPP) that is mainly internalized into cancer cells through a clathrin dependent endocytic mechanism and localizes in late endosomes. Moreover, this CPP is able to enhance the cellular uptake of chlorambucil (CLB) improving its cytotoxicity. In this work, we further explored the cell-penetrating properties of BP16 and those of its arginine analogue BP308. We investigated the influence on the cytotoxicity and on the cellular uptake of conjugating CLB at the N- or the C-terminal end of these undecapeptides. The effect of incorporating the cathepsin B-cleavable sequence Gly-Phe-Leu-Gly in CLB-BP16 and CLB-BP308 conjugates was also evaluated. The activity of CLB was significantly improved when conjugated at the N- or the C-terminus of BP16, or at the N-terminus of BP308. While CLB alone was not active (IC50 of 73.7 to >100 μM), the resulting conjugates displayed cytotoxic activity against CAPAN-1, MCF-7, PC-3, 1BR3G and SKMEL-28 cell lines with IC50 values ranging from 8.7 to 25.5 μM. These results were consistent with the internalization properties observed for the corresponding 5(6)-carboxyfluorescein-labeled conjugates. The presence of the tetrapeptide Gly-Phe-Leu-Gly at either the N- or the C-terminus of CLB-BP16 conjugates further increased the efficacy of CLB (IC50 of 3.6 to 16.2 μM), which could be attributed to its selective release in the lysosomal compartment. Enzymatic assays with cathepsin B showed the release of CLB-Gly-OH from these sequences within a short time. Therefore, the combination of BP16 with an enzymatic cleavable sequence can be used as a drug delivery system for the effective uptake and release of drugs in cancer cells This work was supported by Consolider Ingenio CSD/CSD2010-00065 from MICINN of Spain. We also thank the Catalan DIUE of the Generalitat de Catalunya (2009SGR637). X. R. acknowledges financial support from INNPLANTA project INP-2011-0059-PCT-420000-ACT1

Tumor progression, limited efficacy of current standard treatments, and the rise in patient mortality are associated with gene expression caused by the synergistic action of intratumoral hypoxia and HIF-1α activation. For this reason, recent investigations have focused on HIF-targeting therapeutic agents, with encouraging preclinical and clinical results in solid tumors. Here we describe the efficacy of a HIF-1α inhibitor, Acriflavine, and demonstrate its potency against brain cancer. This safe antibacterial dye induces cell death and apoptosis in several glioma cell lines, targets HIF-1α–mediated pathways, and decreases the level of PGK1, VEGF and HIF-1α in vitro and in vivo. Administered locally via biodegradable polymers, Acriflavine provides significant benefits in survival resulting in nearly 100% long term survival, confirmed by MRI and histological analyses. This study reports preclinical evidence that this safe, small molecule can contribute to brain tumor therapy and highlights the significance of HIF-1α-targeting molecules.

Cancer is one of the most important health problems in the world, affecting a large part of the population. Although there are different treatments such as surgery, radiotherapy or chemotherapy, the cancer currently has no cure, in the strict sense of the word. The numbers of those affected continue to increase, and the future forecasts are not very comforting. Cancer occurs due to alterations in the genes involved in cell growth, survival and propagation, resulting in a set of common characteristics. These characteristics are known as the hallmarks of cancer, and allow to organize and rationalize this complex disease. ErbB (erythroblastic) receptors play a very important role, they’re essential mediators of cell proliferation and differentiation in the development of the embryo and adult tissues, and their inappropriate activation is associated with the development and severity of the cancer. Members of this tyrosine kinase receptor family are: EGFR (ErbB1 or HER1), HER2 (Neu or ErbB2), HER3 (ErbB3) and HER4 (ErbB4). EGFR and HER2 are associated with cancer, since its alteration is found in most cancers. Recently, it has been seen that HER3 receptor also has an important role, in addition, it can act as a new target to treat this disease. Each of these receptors have several ligands with which they bind to be activated, with the exception of HER2 that isn’t known any ligand. Once activated, the receptors dimerize: homodimerization (among them) or heterodimerization (between family members). The heterodimers formed by HER2 and HER3 form the most robust signalling complexes of the ErbB family. One of the characteristics of HER3 is its unique and powerful ability to activate the downstream signalling path PI3K and Akt. It’s key to the control of many biological processes critical for tumorigenesis. In addition, HER3 is also associated with cancer resistance, which is the main problem of current therapies. With these evidences emphasizing the importance of HRE3 signalling in human cancers and their resistance, today there are more than 40 different therapeutic agents that are being tested targeting HER3, in addition to combinatorial therapies that act against different combinations of receptors

We aimed to assess changes in the total effective xenoestrogen burden (TEXB) -a biomarker of combined effect of mixtures of xenoestrogens- in breast cancer patients at surgery (breast adipose tissue) and at different time points during an 18-month follow-up (abdominal adipose tissue), and to analyze the potential influence of socio-demographic, reproductive, tumor, and treatment characteristics on TEXB levels. TEXB-alpha (due to persistent organohalogenated chemicals) and TEXB-beta (due mainly to endogenous estrogens) were quantified in 44 women. TEXB values significantly increased over follow-up (p

Silvia Cufi 1,2 , Bruna Corominas-Faja 1,2 , Alejandro Vazquez-Martin 1,2 , Cristina Oliveras-Ferraros 1,2 , Joan Dorca 2,3 , Joaquim Bosch-Barrera 2,3 , Begona Martin-Castillo 2,4 , Javier A. Menendez 1,2 1 Translational Research Laboratory, Catalan Institute of Oncology (ICO), Girona, Spain 2 Girona Biomedical Research Institute (IDIBGi), Girona, Spain 3 Medical Oncology, Catalan Institute of Oncology (ICO), Girona, Spain 4 Clinical Research Unit, Catalan Institute of Oncology (ICO), Girona, Spain Received: April 27, 2012, Accepted: April 28, 2012, Published: May 4, 2012, Keywords: Metformin, trastuzumab, HER2, cancer stem cells, breast cancer Correspondence: Javier A. Menendez, email: // // Abstract Trastuzumab-refractory breast cancer stem cells (CSCs) could also explain the high rate of primary resistance to single-agent trastuzumab in HER2 gene-amplified breast cancer patients. The identification of agents with strong selective toxicity for trastuzumab-resistant breast CSCs may have tremendous relevance for how HER2+ breast cancer patients should be treated. Using the human breast cancer cell line JIMT-1, which was established from the pleural metastasis of a patient who was clinically resistant to trastuzumab ab initio , we examined whether preferential killing of the putative CD44 + CD24 -/low breast CSC population might be sufficient to overcome primary resistance to trastuzumab in vivo . Because recent studies have shown that the anti-diabetic biguanide metformin can exert antitumor effects by targeted killing of CSC-like cells, we explored whether metformin’s ability to preferentially kill breast cancer initiating CD44 + CD24 -/low cells may have the potential to sensitize JIMT-1 xenograft mouse models to trastuzumab. Upon isolation for breast cancer initiating CD44 + CD24 -/low cells by employing magnetic activated cell sorting, we observed the kinetics of metformin-induced killing drastically varied among CSC and non-CSC subpopulations. Metformin’s cell killing effect increased dramatically by more than 10-fold in CD44 + CD24 -/low breast CSC cells compared to non-CD44 + CD24 -/low immunophenotypes. While seven-weeks treatment length with trastuzumab likewise failed to reduce tumor growth of JIMT-1 xenografts, systemic treatment with metformin as single agent resulted in a significant two-fold reduction in tumor volume. When trastuzumab was combined with concurrent metformin, tumor volume decreased sharply by more than four-fold. Given that metformin-induced preferential killing of breast cancer initiating CD44 + CD24 -/low subpopulations is sufficient to overcome in vivo primary resistance to trastuzumab, the incorporation of metformin into trastuzumab-based regimens may provide a valuable strategy for treatment of HER2+ breast cancer patients.

Ribonucleases are proteins whose use is promising in anticancer therapy. We have previously constructed different human pancreatic ribonuclease variants that are selectively cytotoxic for tumor cells by introducing a nuclear localization signal into their sequence. However, these modifications produced an important decrease in their stability compromising their behavior in vivo. Here, we show that we can significantly increase the thermal stability of these cytotoxic proteins by introducing additional disulfide bonds by site-directed mutagenesis. One of these variants increases its thermal stability by around 17 &deg, C, without affecting its catalytic activity while maintaining the cytotoxic activity against tumor cells. We also show that the most stable variant is significantly more resistant to proteolysis when incubated with proteinase K or with human sera, suggesting that its half-live could be increased in vivo once administered.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License.-- et al., [Introduction]: Endocrine therapies targeting cell proliferation and survival mediated by estrogen receptor α (ERα) are among the most effective systemic treatments for ERα-positive breast cancer. However, most tumors initially responsive to these therapies acquire resistance through mechanisms that involve ERα transcriptional regulatory plasticity. Herein we identify VAV3 as a critical component in this process. [Methods]: A cell-based chemical compound screen was carried out to identify therapeutic strategies against resistance to endocrine therapy. Binding to ERα was evaluated by molecular docking analyses, an agonist fluoligand assay and short hairpin (sh)RNA-mediated protein depletion. Microarray analyses were performed to identify altered gene expression. Western blot analysis of signaling and proliferation markers, and shRNA-mediated protein depletion in viability and clonogenic assays, were performed to delineate the role of VAV3. Genetic variation in VAV3 was assessed for association with the response to tamoxifen. Immunohistochemical analyses of VAV3 were carried out to determine its association with therapeutic response and different tumor markers. An analysis of gene expression association with drug sensitivity was carried out to identify a potential therapeutic approach based on differential VAV3 expression. [Results]: The compound YC-1 was found to comparatively reduce the viability of cell models of acquired resistance. This effect was probably not due to activation of its canonical target (soluble guanylyl cyclase), but instead was likely a result of binding to ERα. VAV3 was selectively reduced upon exposure to YC-1 or ERα depletion, and, accordingly, VAV3 depletion comparatively reduced the viability of cell models of acquired resistance. In the clinical scenario, germline variation in VAV3 was associated with the response to tamoxifen in Japanese breast cancer patients (rs10494071 combined P value = 8.4 × 10-4). The allele association combined with gene expression analyses indicated that low VAV3 expression predicts better clinical outcome. Conversely, high nuclear VAV3 expression in tumor cells was associated with poorer endocrine therapy response. Based on VAV3 expression levels and the response to erlotinib in cancer cell lines, targeting EGFR signaling may be a promising therapeutic strategy. [Conclusions]: This study proposes VAV3 as a biomarker and a rationale for its use as a signaling target to prevent and/or overcome resistance to endocrine therapy in breast cancer., This work was supported by grants from the Eugenio Rodríguez Pascual Foundation (2012, to MAP), the Government of Catalonia (2009-SGR283, to AV and MAP), the National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health (R01 DK015556, to JAK), the Red Cooperative Research Thematic Network on Cancer (RTICC) (12/0036/0002 to XRB and 12/0036/0008 to XRB and MAP) and the Spanish Ministry of Health, Fund for Health Research–Institute of Health Carlos III (11/00951 to AU and 12/01528 to MAP).

Content from this work may be used under the terms of the Creative Commons Attribution 3.0 licence., Glioblastomas are highly diffuse, malignant tumors that have so far evaded clinical treatment. The strongly invasive behavior of cells in these tumors makes them very resistant to treatment, and for this reason both experimental and theoretical efforts have been directed toward understanding the spatiotemporal pattern of tumor spreading. Although usual models assume a standard diffusion behavior, recent experiments with cell cultures indicate that cells tend to move in directions close to that of glioblastoma invasion, thus indicating that a biased random walk model may be much more appropriate. Here we show analytically that, for realistic parameter values, the speeds predicted by biased dispersal are consistent with experimentally measured data. We also find that models beyond reaction-diffusion-advection equations are necessary to capture this substantial effect of biased dispersal on glioblastoma spread. © IOP Publishing and Deutsche Physikalische Gesellschaft., This work was supported by MEC (FIS-2012-31307 grant, JF), the Generalitat de Catalunya (2009-SGR-374 grant, JF), the Marcelino Bot́ín Foundation (RVS) and the Santa Fe Institute (RVS).

Brain tumor segmentation consists of separating the different tumor tissues (solid or active tumor, edema,and necrosis) from normal brain tissues: gray matter (GM), white matter (WM), and cerebrospinal fluid (CSF). In brain tumor studies, the existence of abnormal tissues may be easily detectable most of the time. However, accurate and reproducible segmentation and characterization of abnormalities are not straightforward. In the past, many researchers in the field of medical imaging and soft computing have made significant survey in the field of brain tumor segmentation. Both semiautomatic and fully automatic methods have been proposed. Clinical acceptance of segmentation techniques has depended on the simplicity of the segmentation, and the degree of user supervision. Interactive or semiautomatic methods are likely to remain dominant in practice for some time, especially in these applications where erroneous interpretations are unacceptable. This article presents an overview of the most relevant brain tumor segmentation methods, conducted after the acquisition of the image. Given the advantages of magnetic resonance imaging over other diagnostic imaging, this survey is focused on MRI brain tumor segmentation. Semiautomatic and fully automatic techniques are emphasized.

The recent identification of HER2 gene amplification as a novel predictor of resistance to the EGFR (HER2)-targeted antibody cetuximab and of response to combination therapies against EGFR and HER2 in wild-type KRAS tumor settings may represent a further step toward personalized medicine for patients with colorectal cancer. Herein, we show that transcriptional upregulation of HER2 expression in the absence of HER2 gene amplification is a molecular phenomenon that takes place in EGFR-dependent, wild-type KRAS squamous cell carcinoma (SCC) cells that acquire resistance to cetuximab. Since cetuximab activity against cetuximab-refractory SCC cells can be fully restored in the presence of the anti-HER2 monoclonal antibody trastuzumab, our findings suggest that, beyond HER2 gene amplification, we might need to redefine the threshold values for HER2 positivity to improve the accuracy of the selection of cetuximab-refractory patients with wild-type KRAS that may benefit from receiving a cetuximab/trastuzumab combination.

Ribonucleases are promising agents for use in anticancer therapy. Engineering a nuclear localization signal into the sequence of the human pancreatic ribonuclease has been revealed as a new strategy to endow this enzyme with cytotoxic activity against tumor cells. We previously described a cytotoxic human pancreatic ribonuclease variant, named PE5, which is able to cleave nuclear RNA, inducing the apoptosis of cancer cells and reducing the amount of P-glycoprotein in different multidrug-resistant cell lines. These results open the opportunity to use this ribonuclease in combination with other chemotherapeutics. In this work, we have investigated how to improve the properties of PE5 as an antitumor drug candidate. When attempting to develop a recombinant protein as a drug, two of the main desirable attributes are minimum immunogenicity and maximum potency. The improvements of PE5 have been designed in both senses. First, in order to reduce the potential immunogenicity of the protein, we have studied which residues mutated on PE5 can be reverted to those of the wild-type human pancreatic ribonuclease sequence without affecting its cytotoxicity. Second, we have investigated the effect of introducing an additional nuclear localization signal at different sites of PE5 in an effort to obtain a more cytotoxic enzyme. We show that the nuclear localization signal location is critical for the cytotoxicity. One of these variants, named NLSPE5, presents about a 10-fold increase in cytotoxicity respective to PE5. This variant induces apoptosis and kills the cells using the same mechanism as PE5 This work has been supported by Grant BFU2009-06935 from MICINN (Spain) and by Grant GRCT04 from the University of Girona. We are very grateful to Dr. Milica Pesic and Dr. Sabera Ruzdijićfor providing us with the NCI-H460/R cell line and to Dr. Ramon Colomer for providing us with the NCI/ ADR-RES cell line. A.V., J.C., and P.T. acknowledge their fellowship from MINEDU, Universitat de Girona, and MEC (Spain), respectively

Ribonucleases are small basic proteins that have shown remarkable antitumor activity linked to their ability to destroy RNA. Therefore, they are a second line of cancer chemotherapeutics as they are not genotoxic. This chapter summarizes the main biochemical characteristics of these enzymes and the key factors responsible for their cytotoxic mechanism. Some of them are shared by most cytotoxins, but each RNase has particular cancer cell killing abilities. The effects on the cell cycle and the induced apoptosis mechanism are cell dependent. The knowledge obtained from the cytotoxic mechanism of natural cytotoxic RNases has been used to artificially engineer more potent and selective RNA-degrading enzymes. These approaches are also described. The chapter ends with a brief description of the results of the clinical trials performed with RNases This work has been supported by grants BFU2009-06935 from MICINN (Spain) and GRCT04 from the University of Girona

Introduction. Inhibiting the enzyme Fatty Acid Synthase (FASN) leads to apoptosis of breast carcinoma cells, and this is linked to human epidermal growth factor receptor 2 (HER2) signaling pathways in models of simultaneous expression of FASN and HER2. Methods. In a xenograft model of breast carcinoma cells that are FASN+ and HER2+, we have characterized the anticancer activity and the toxicity profile of G28UCM, the lead compound of a novel family of synthetic FASN inhibitors. In vitro, we have analyzed the cellular and molecular interactions of combining G28UCM with anti-HER drugs. Finally, we have tested the cytotoxic ability of G28UCM on breast cancer cells resistant to trastuzumab or lapatinib, that we developed in our laboratory. Results. In vivo, G28UCM reduced the size of 5/14 established xenografts. In the responding tumours, we observed inhibition of FASN activity, cleavage of poly- ADPribose polymerase (PARP) and decrease of p-HER2, p- protein kinase B (AKT) and p-ERK1/2, which were not observed in the nonresponding tumours. In the G28UCM-treated animals, no significant toxicities occurred, and weight loss was not observed. In vitro, G28UCM showed marked synergistic interactions with trastuzumab, lapatinib, erlotinib or gefitinib (but not with cetuximab), which correlated with increases in apoptosis and with decreases in the activation of HER2, extracellular signal-regulated kinase (ERK)1/2 and AKT. In trastuzumab-resistant and in lapatinib-resistant breast cancer cells, in which trastuzumab and lapatinib were not effective, G28UCM retained the anticancer activity observed in the parental cells. Conclusions. G28UCM inhibits fatty acid synthase (FASN) activity and the growth of breast carcinoma xenografts in vivo, and is active in cells with acquire resistance to anti-HER2 drugs, which make it a candidate for further pre-clinical development

Includes supplementary materials for the online appendix. Includes supplementary materials for the online appendix. Anaplastic lymphoma kinase (ALK) genomic alterations have emerged as a potent predictor of benefit from treatment with ALK inhibitors in several cancers. Currently, there is no information about ALK gene alterations in urothelial carcinoma (UC) and its correlation with clinical or pathologic features and outcome. Samples from patients with advanced UC and correlative clinical data were collected. Genomic imbalances were investigated by array comparative genomic hybridization (aCGH). ALK gene status was evaluated by fluorescence in situ hybridization (FISH). ALK expression was assessed by immunohistochemistry (IHC) and high-throughput mutation analysis with Oncomap 3 platform. Next generation sequencing was performed using Illumina Genome Analyzer IIx, and Illumina HiSeq 2000 in the FISH positive case. 70 of 96 patients had tissue available for all the tests performed. Arm level copy number gains at chromosome 2 were identified in 17 (24%) patients. Minor copy number alterations (CNAs) in the proximity of ALK locus were found in 3 patients by aCGH. By FISH analysis, one of these samples had a deletion of the 5′ALK. Whole genome next generation sequencing was inconclusive to confirm the deletion at the level of the ALK gene at the coverage level used. We did not observe an association between ALK CNA and overall survival, ECOG PS, or development of visceral disease. ALK genomic alterations are rare and probably without prognostic implications in UC. The potential for testing ALK inhibitors in UC merits further investigation but might be restricted to the identification of an enriched population.

Ribonucleases represent a new class of antitumor RNA-damaging drugs. However, many wild-type members of the vertebrate secreted ribonuclease family are not cytotoxic because they are not able to evade the cytosolic ribonuclease inhibitor. We previously engineered the human pancreatic ribonuclease to direct it to the cell nucleus where the inhibitor is not present. The best characterized variant is PE5 that kills cancer cells through apoptosis mediated by the p21WAF1/CIP1 induction and the inactivation of JNK. Here, we have used microarray-derived transcriptional profiling to identify PE5 regulated genes on the NCI/ADR-RES ovarian cancer cell line. RT-qPCR analyses have confirmed the expression microarray findings. The results show that PE5 cause pleiotropic effects. Among them, it is remarkable the down-regulation of multiple genes that code for enzymes involved in deregulated metabolic pathways in cancer cells This work has been supported by grants BFU2009-06935 and BIO2013-43517 from MINECO (Spain) and SING12/0 from UdG (Spain).

Cell 166(3), 740-754 (2016). doi:10.1016/j.cell.2016.06.017, Published by Elsevier, New York, NY

// Elisabet Cuyas 1,2 , Salvador Fernandez-Arroyo 3 , Bruna Corominas-Faja 1,2 , Esther Rodriguez-Gallego 3 , Joaquim Bosch-Barrera 2,4 , Begona Martin-Castillo 2,5 , Rafael De Llorens 6 , Jorge Joven 3 and Javier A. Menendez 1,2 1 Metabolism and Cancer Group, Translational Research Laboratory, Catalan Institute of Oncology (ICO), Girona, Catalonia, Spain 2 Molecular Oncology Group, Girona Biomedical Research Institute (IDIBGI), Girona, Catalonia, Spain 3 Unitat de Recerca Biomedica (URB-CRB), Institut d’Investigacio Sanitaria Pere i Virgili (IISPV), Universitat Rovira i Virgili, Reus, Catalonia. Spain 4 Medical Oncology, Catalan Institute of Oncology (ICO), Girona, Catalonia, Spain 5 Clinical Research Unit, Catalan Institute of Oncology (ICO), Girona, Catalonia, Spain 6 Biochemistry and Molecular Biology Unit, Department of Biology, University of Girona, Girona, Catalonia, Spain Correspondence to: Javier A. Menendez, email: // Keywords : IDH1, oncometabolites, metformin, anaplerosis, cancer Received : January 17, 2015 Accepted : March 11, 2015 Published : March 30, 2015 Abstract Metabolic flexibility might be particularly constrained in tumors bearing mutations in isocitrate dehydrogenase 1 (IDH1) leading to the production of the oncometabolite 2-hydroxygluratate (2HG). To test the hypothesis that IDH1 mutations could generate metabolic vulnerabilities for therapeutic intervention, we utilized an MCF10A cell line engineered with an arginine-to-histidine conversion at position 132 (R132H) in the catalytic site of IDH1, which equips the enzyme with a neomorphic α-ketoglutarate to 2HG reducing activity in an otherwise isogenic background. IDH1 R132H/+ and isogenic IDH1 +/+ parental cells were screened for their ability to generate energy-rich NADH when cultured in a standardized high-throughput Phenotype MicroArrayplatform comprising >300 nutrients. A radical remodeling of the metabotype occurred in cells carrying the R132H mutation since they presented a markedly altered ability to utilize numerous carbon catabolic fuels. A mitochondria toxicity-screening modality confirmed a severe inability of IDH1-mutated cells to use various carbon substrates that are fed into the electron transport chain at different points. The mitochondrial biguanide poisons, metformin and phenformin, further impaired the intrinsic weakness of IDH1-mutant cells to use certain carbon-energy sources. Additionally, metabolic reprogramming of IDH1-mutant cells increased their sensitivity to metformin in assays of cell proliferation, clonogenic potential, and mammosphere formation. Targeted metabolomics studies revealed that the ability of metformin to interfere with the anaplerotic entry of glutamine into the tricarboxylic acid cycle could explain the hypersensitivity of IDH1-mutant cells to biguanides. Moreover, synergistic interactions occurred when metformin treatment was combined with the selective R132H-IDH1 inhibitor AGI-5198. Together, these results suggest that therapy involving the simultaneous targeting of metabolic vulnerabilities with metformin, and 2HG overproduction with mutant-selective inhibitors (AGI-5198-related AG-120 [Agios]), might represent a worthwhile avenue of exploration in the treatment of IDH1-mutated tumors.

Pregunta d’investigació: És més efectiva la combinació d’exercici terapèutic i Mindfulness envers únicament activitat física per a l’abordatge simptomàtic en pacients amb càncer de mama tractats amb quimioteràpia?. Objectius: Com a objectiu general, valorar la efectivitat d’un tractament basat en la combinació d’exercici terapèutic combinat amb Mindfulness per l’abordatge simptomàtic en pacients amb càncer de mama tractats amb quimioteràpia. I com a objectiu específic valorar la fatiga, qualitat del son, el dolor i la depressió en aquests pacients. Metodologia: es realitzarà un estudi controlat aleatoritzat amb una mostra de 64 pacients en període de reminiscència de càncer de mama els quals dividirem en dos grups de 32 persones. La intervenció té una durada de 8 setmanes. El grup control realitzarà activitat terapèutica ( 3 dies per setmana 1h cada sessió) i el grup experimental realitzarà activitat terapèutica combinada amb treball formal Mindfulness (90 minuts setmanalment, un total de 12 hores) i treball informal de 45 minuts diaris. Es valorarà la fatiga mitjançant el qüestionari de Piper de la Fatiga, la qualitat del son mitjançant l’índex de la qualitat del son de Pittsburgh, el dolor mitjançant l’escala EVA i la depressió mitjançant l’escala de Hamilton per a la Depressió HAM-D. Es realitzaran 3 valoracions: una pre-tractament, una altra post-tractament a les 8 setmanes d’intervenció i una de llarg termini a l’any des de la primera intervenció. Research question: Is the combination of therapeutic exercise and Mindfulness more effective only for physical activity for the symptomatic approach in breast cancer patients treated with chemotherapy? Objectives: As a general objective, assess the effectiveness of a treatment based on the combination of therapeutic exercise combined with Mindfulness for the symptomatic approach in patients with breast cancer treated with chemotherapy. And as a specific objective, assess fatigue, sleep quality, pain and depression in these patients. Methodology: A randomized controlled study will be carried out with a sample of 64 patients reminiscent of breast cancer, which will be divided into two groups of 32 people. The intervention lasts for 8 weeks. The control group will perform therapeutic activity (3 days per week 1h each session) and the experimental group will perform therapeutic activity combined with formal Mindfulness work (90 minutes per week, a total of 12 hours) and informal work 45 minutes daily. Fatigue will be assessed through the Piper Fatigue questionnaire, sleep quality through the Pittsburgh sleep quality index, pain through the EVA scale and depression through the Hamilton scale for depression HAM-D. Three assessments will be made: one pre-treatment, another post-treatment at 8 weeks of intervention and one long-term per year from the first intervention.

Conjugates of a dicarba analogue of octreotide, a potent somatostatin agonist whose receptors are overexpressed on tumor cells, with [PtCl 2(dap)] (dap = 1-(carboxylic acid)-1,2-diaminoethane) (3), [(η 6-bip)Os(4-CO 2-pico)Cl] (bip = biphenyl, pico = picolinate) (4), [(η 6-p-cym)RuCl(dap)] + (p-cym = p-cymene) (5), and [(η 6-p-cym)RuCl(imidazole-CO 2H)(PPh 3)] + (6), were synthesized by using a solid-phase approach. Conjugates 3-5 readily underwent hydrolysis and DNA binding, whereas conjugate 6 was inert to ligand substitution. NMR spectroscopy and molecular dynamics calculations showed that conjugate formation does not perturb the overall peptide structure. Only 6 exhibited antiproliferative activity in human tumor cells (IC 50 = 63 ± 2 μ in MCF-7 cells and IC 50 = 26 ± 3 μ in DU-145 cells) with active participation of somatostatin receptors in cellular uptake. Similar cytotoxic activity was found in a normal cell line (IC 50 = 45 ± 2.6 μ in CHO cells), which can be attributed to a similar level of expression of somatostatin subtype-2 receptor. These studies provide new insights into the effect of receptor-binding peptide conjugation on the activity of metal-based anticancer drugs, and demonstrate the potential of such hybrid compounds to target tumor cells specifically.

The members of the epidermal growth factor (EGF)/ErbB family are prime targets for cancer therapy. However, the therapeutic efficiency of the existing anti-ErbB agents is limited. Thus, identifying new molecules that inactivate the ErbB receptors through novel strategies is an important goal on cancer research. In this study we have developed a shorter form of human EGF (EGFt) with a truncated C-terminal as a novel EGFR inhibitor. EGFt was designed based on the superimposition of the three-dimensional structures of EGF and the Potato Carboxypeptidase Inhibitor (PCI), an EGFR blocker previously described by our group. The peptide was produced in E. coli with a high yield of the correctly folded peptide. EGFt showed specificity and high affinity for EGFR but induced poor EGFR homodimerization and phosphorylation. Interestingly, EGFt promoted EGFR internalization and translocation to the cell nucleus although it did not stimulate the cell growth. In addition, EGFt competed with EGFR native ligands, inhibiting the proliferation of cancer cells. These data indicate that EGFt may be a potential EGFR blocker for cancer therapy. In addition, the lack of EGFR-mediated growth-stimulatory activity makes EGFt an excellent delivery agent to target toxins to tumours over-expressing EGFR.

Projecte realitzat en col.laboració amb el centre ICFO-Institut de Ciències Fotòniques Emodin (6-methyl-1,3,8 trihydroxyanthraquinone) is considered as a potential candidate for photodynamic therapy for the treatment of cancer. A success in using of the molecule depends among other factors on the knowledge how fast it penetrates through the cell membrane and then diffuses inside the cell. A spatial distribution of emodin inside cancerous cells is also an important factor. The aim of this study is to investigate the diffusion of emodin in cancerous cells by means of a non invasive and sensitive technique: Surface Enhanced Raman scattering which permits us to study the evolution of the small amount of emodin inside single living cells. The experimental results were compared with a theoretical model that simulated the Brownian diffusion of particles. From this comparison and comparing with previous results in fluorescence spectroscopy we have found that: for a long time, emodin is accumulated in the cytoplasm in a region close to the membrane. The result is discussed in terms of emodin being stored in cell organelles located in the cytoplasm.

Purpose: Triple-negative breast cancer (TNBC) lacks an approved targeted therapy. Despite initial good response to chemotherapy, 30% of the patients relapse within 5 years after treatment. EGFR overexpression is a common marker in TNBC, and its expression has been correlated with poor outcome. Inhibition of fatty acid synthase (FASN) activity leads to apoptosis of human carcinoma cells overexpressing FASN. We tested the hypothesis that blocking FASN in combination with anti-EGFR signaling agents would be an effective antitumor strategy in sensitive and chemoresistant TNBC. Experimental Design: Several TNBC cell lines and 29 primary tumors were included to determine whether FASN is a potential target in TNBC. Doxorubicin-resistant TNBC cell lines (231DXR and HCCDXR) have been developed and characterized in our laboratory. Cellular and molecular interactions of anti-FASN compounds (EGCG and C75) with cetuximab were analyzed. In vivo tumor growth inhibition was evaluated after cetuximab, EGCG, or the combination in TNBC orthoxenograft models. Results: TNBC cell lines showed overexpression of FASN enzyme and its inhibition correlated to FASN levels. FASN staining was observed in all of the 29 TNBC tumor samples. In vitro, EGCG and C75 plus cetuximab showed strong synergism in sensitive and chemoresistant cells. In vivo, the combination of EGCG with cetuximab displayed strong antitumor activity against the sensitive and chemoresistant TNBC orthoxenografts, without signs of toxicity. Conclusions: Our results show that the simultaneous blockade of FASN and EGFR is effective in preclinical models of sensitive and chemoresistant TNBC. Clin Cancer Res; 22(18); 4687–97. ©2016 AACR.

Curs 2014-2015, Els avenços en l’estudi de les bases moleculars del càncer han permès el desenvolupament de nous fàrmacs personalitzats a les necessitats de cada pacient, per tal de maximitzar el seu benefici. Estudis recents suggereixen que els exosomes, vesícules que presenten una mida compresa entre els 30 i els 100 nm, secretats per les cèl·lules tumorals i que contenen diferents biomolècules (mARN, miARN, proteïnes oncogèniques i angiogèniques...), poden ser utilitzades per la caracterització biomolecular del càncer i així, poder dissenyar una teràpia apropiada. No obstant això, la manca de sistemes de detecció de petites concentracions d’exosomes en mostres reals és un obstacle a l’hora de desenvolupar un test de diagnòstic, ja que no és possible normalitzar la quantitat de biomarcador respecte al nombre d’exosomes. L’objectiu del projecte en el qual s’engloba aquest Treball de Fi de Grau és el desenvolupament d’un biosensor ràpid, precís i de simple maneig basat en les guies bimodals per a la detecció d’exosomes, mitjançant un immunoassaig dirigit contra proteïnes de membrana, i la posterior detecció de mARN, corresponent a la variant 1 i 3a/b del gen de fusió EML4-ALK en càncer de pulmó, per mitjà d’un assaig genòmic. El biosensor de guies bimodals possibilita la detecció sense cap tipus de marcatge i la seva integració en dispositius point-of-care, on l’anàlisi es pot realitzar sense cap instrumentació addicional. A més a més, la seva elevada sensibilitat podria permetre detectar ARN directament en mostres reals sense cap amplificació. En conclusió, s’ha desenvolupat un biosensor específic per a la detecció d’exosomes amb la capacitat de detectar-los en mostres de medis tumorals amb un límit de detecció de exosomes/mL. A més a més, en aquest treball s’ha aconseguit dissenyar els bioreceptors que possibilitaran la detecció del mARN contingut, fent els primers passos cap al desenvolupament d’un dispositiu integrat que permetrà la normalització del biomarcador respecte al nombre d’exosomes a temps real., Advances in the study of the molecular basis in cancer have enabled the development of new personalized drugs customized for each patient in order to maximize their benefits. Recent studies suggest that exosomes, vesicles which have a size between 30 and 100 nm, secreted by tumor cells and which contain different biomolecules (mRNA, miRNA, oncogenic and angiogenic proteins…),can be used for biomolecular cancer characterization and thus, designing an appropriate therapy. However, the lack in the detection systems of small concentrations of exosomes on real samples is an obstacle in developing a diagnostic test, because it is not possible to standardize the amount of biomarker compared the number of exosomes. The aim of the project included in this Final Year Project is the development of a fast, specific and simple handling biosensor, based on bimodal waveguides biosensors for the detection of exosomes using an immunoassay against membrane proteins, and subsequent detection of mRNA corresponding to variant 1 and 3a/b of the fusion gene EML4-ALK in lung cancer, using a genomic test. Bimodal waveguide biosensor makes possible the detection without labelling and their integration in point-of-care devices, where the analysis can be performed without extra instruments. In addition, its high sensitivity could allow directly detection of RNA in real samples without amplification. In conclusion, we have developed a specific biosensor for exosomes detection with the ability to detect them in tumor cell culture with a limit detection of exosomes/mL. Moreover, in this project we have achieved to design bioreceptors that will enable the detection of mRNA content, giving the first steps for the development of an integrated device that will allow the normalization of the biomarker compared to the number of exosomes at real time.