Cancer Cells Haploinsufficient for ATM Are Sensitized to PARP Inhibitors by MET Inhibition.

The MET oncogene encodes a tyrosine kinase (TK) receptor. Its activation protects cells from death but also stimulates DNA damage response by triggering excess replicative stress. Transcriptomic classification of cancer cell lines based on MET expression showed that response to the PARP inhibitor (PARPi) olaparib is poorer in MET overexpressing cell lines. Accordingly, a high MET expressing lung carcinoma cell line was sensitized to PARPi by MET TK inhibition. This was not linked solely to MET overexpression: other MET overexpressing cell lines were biochemically but not functionally responsive to combined inhibition. Moreover, exogenously induced MET overexpression was unable to induce resistance to PARPi. The MET overexpressing cell line, responsive to the combined PARP and MET inhibition, carried a heterozygous mutation of the ATM gene and showed an attenuated response of ATM to PARPi. Among the downstream targets of ATM activation, NuMA was phosphorylated only in response to the combined PARP and MET inhibition. Given the role played by NuMA in mitosis, data show that the latter is affected by MET and PARP inhibition in cells with haploinsufficient ATM. This is important as ATM heterozygous mutation is frequently found in human cancer and in lung carcinomas in particular.

Corrigendum to "Drug Delivery Nanoparticles in Skin Cancers".

[This corrects the article DOI: 10.1155/2014/895986.].

OMO-1 reduces progression and enhances cisplatin efficacy in a 4T1-based non-c-MET addicted intraductal mouse model for triple-negative breast cancer.

c-MET is considered a driver of cancer progression, impacting tumor growth and tumor-supporting stroma. Here, we investigated the therapeutic efficacy of OMO-1, a potent and selective c-MET inhibitor, in an immunocompetent intraductal mouse model for triple-negative breast cancer (TNBC). OMO-1 reduced non-c-MET addicted 4T1 tumor progression dose dependently as monotherapeutic and provided additional disease reduction in combination with cisplatin. At the stromal level, OMO-1 significantly reduced neutrophil infiltration in 4T1 tumors, promoted immune activation, and enhanced cisplatin-mediated reduction of tumor-associated macrophages. OMO-1 treatment also reduced 4T1 tumor hypoxia and increased expression of pericyte markers, indicative for vascular maturation. Corroborating this finding, cisplatin delivery to the 4T1 primary tumor was enhanced upon OMO-1 treatment, increasing cisplatin DNA-adduct levels and tumor cell death. Although verification in additional cell lines is warranted, our findings provide initial evidence that TNBC patients may benefit from OMO-1 treatment, even in cases of non-c-MET addicted tumors.

Dual Inhibition of Angiopoietin-TIE2 and MET Alters the Tumor Microenvironment and Prolongs Survival in a Metastatic Model of Renal Cell Carcinoma.

Receptor tyrosine kinase inhibitors have shown clinical benefit in clear cell renal cell carcinoma (ccRCC), but novel therapeutic strategies are needed. The angiopoietin/Tie2 and MET pathways have been implicated in tumor angiogenesis, metastases, and macrophage infiltration. In our study, we used trebananib, an angiopoietin 1/2 inhibitor, and a novel small-molecule MET kinase inhibitor in patient-derived xenograft (PDX) models of ccRCC. Our goal was to assess the ability of these compounds to alter the status of tumor-infiltrating macrophages, inhibit tumor growth and metastases, and prolong survival. Seven-week-old SCID mice were implanted subcutaneously or orthotopically with human ccRCC models. One month postimplantation, mice were treated with angiopoietin 1/2 inhibitor trebananib (AMG 386), MET kinase inhibitor, or combination. In our metastatic ccRCC PDX model, RP-R-02LM, trebananib alone, and in combination with a MET kinase inhibitor, significantly reduced lung metastases and M2 macrophage infiltration (P = 0.0075 and P = 0.0205, respectively). Survival studies revealed that treatment of the orthotopically implanted RP-R-02LM tumors yielded a significant increase in survival in both trebananib and combination groups. In addition, resection of the subcutaneously implanted primary tumor allowed for a significant survival advantage to the combination group compared with vehicle and both single-agent groups. Our results show that the combination of trebananib with a MET kinase inhibitor significantly inhibits the spread of metastases, reduces infiltrating M2-type macrophages, and prolongs survival in our highly metastatic ccRCC PDX model, suggesting a potential use for this combination therapy in treating patients with ccRCC.

Crosstalk between Nrf2 and YAP contributes to maintaining the antioxidant potential and chemoresistance in bladder cancer.

Redox adaptation plays an important role in cancer cells drug resistance. The antioxidant response is principally mediated by the transcription factor Nrf2, that induces the transcriptional activation of several genes involved in GSH synthesis, chemoresistance, and cytoprotection. YAP is emerging as a key mediator of chemoresistance in a variety of cancers, but its role in controlling the antioxidant status of the cells is yet elusive. Here, we show that impairing YAP protein expression reduced GSH content and Nrf2 protein and mRNA expression in bladder cancer cells. Moreover, in YAP knocked down cells the expression of FOXM1, a transcription factor involved in Nrf2 transcription, was down-regulated and the silencing of FOXM1 reduced Nrf2 expression. On the other hand, the silencing of Nrf2, as well as the depletion of GSH by BSO treatment, inhibited YAP expression, suggesting that cross-talk exists between YAP and Nrf2 proteins. Importantly, we found that silencing either YAP or Nrf2 enhanced sensitivity of bladder cancer cells to cytotoxic agents and reduced their migration. Furthermore, the inhibition of both YAP and Nrf2 expressions significantly increased cytotoxic drug sensitivity and synergistically reduced the migration of chemoresistant bladder cancer cells. These findings provide a rationale for targeting these transcriptional regulators in patients with chemoresistant bladder cancer, expressing high YAP and bearing a proficient antioxidant system.

EZH2 Modifies Sunitinib Resistance in Renal Cell Carcinoma by Kinome Reprogramming.

Acquired and intrinsic resistance to receptor tyrosine kinase inhibitors (RTKi) represents a major hurdle in improving the management of clear cell renal cell carcinoma (ccRCC). Recent reports suggest that drug resistance is driven by tumor adaptation via epigenetic mechanisms that activate alternative survival pathways. The histone methyl transferase EZH2 is frequently altered in many cancers, including ccRCC. To evaluate its role in ccRCC resistance to RTKi, we established and characterized a spontaneously metastatic, patient-derived xenograft model that is intrinsically resistant to the RTKi sunitinib, but not to the VEGF therapeutic antibody bevacizumab. Sunitinib maintained its antiangiogenic and antimetastatic activity but lost its direct antitumor effects due to kinome reprogramming, which resulted in suppression of proapoptotic and cell-cycle-regulatory target genes. Modulating EZH2 expression or activity suppressed phosphorylation of certain RTKs, restoring the antitumor effects of sunitinib in models of acquired or intrinsically resistant ccRCC. Overall, our results highlight EZH2 as a rational target for therapeutic intervention in sunitinib-resistant ccRCC as well as a predictive marker for RTKi response in this disease. Cancer Res; 77(23); 6651-66. ©2017 AACR.

Genomic profiling is predictive of response to cisplatin treatment but not to PI3K inhibition in bladder cancer patient-derived xenografts.

PURPOSE: Effective systemic therapeutic options are limited for bladder cancer. In this preclinical study we tested whether bladder cancer gene alterations may be predictive of treatment response. EXPERIMENTAL DESIGN: We performed genomic profiling of two bladder cancer patient derived tumor xenografts (PDX). We optimized the exome sequence analysis method to overcome the mouse genome interference. RESULTS: We identified a number of somatic mutations, mostly shared by the primary tumors and PDX. In particular, BLCAb001, which is less responsive to cisplatin than BLCAb002, carried non-sense mutations in several genes associated with cisplatin resistance, including MLH1, BRCA2, and CASP8. Furthermore, RNA-Seq analysis revealed the overexpression of cisplatin resistance associated genes such as SLC7A11, TLE4, and IL1A in BLCAb001. Two different PIK3CA mutations, E542K and E545K, were identified in BLCAb001 and BLCAb002, respectively. Thus, we tested whether the genomic profiling was predictive of response to a dual PI3K/mTOR targeting agent, LY3023414. Despite harboring similar PIK3CA mutations, BLCAb001 and BLCAb002 exhibited differential response, both in vitro and in vivo. Sustained target modulation was observed in the sensitive model BLCAb002 but not in BLCAb001, as well as decreased autophagy. Interestingly, computational modelling of mutant structures and affinity binding to PI3K revealed that E542K mutation was associated with weaker drug binding than E545K. CONCLUSIONS: Our results suggest that the presence of activating PIK3CA mutations may not necessarily predict in vivo treatment response to PI3K targeted therapies, while specific gene alterations may be predictive for cisplatin response in bladder cancer models and, potentially, in patients as well.

HDAC 1 and 6 modulate cell invasion and migration in clear cell renal cell carcinoma.

BACKGROUND: Class I histone deacetylases (HDACs) have been reported to be overexpressed in clear cell renal cell carcinoma (ccRCC), whereas the expression of class II HDACs is unknown. METHODS: Four isogenic cell lines C2/C2VHL and 786-O/786-OVHL with differential VHL expression are used in our studies. Cobalt chloride is used to mimic hypoxia in vitro. HIF-2α knockdowns in C2 and 786-O cells is used to evaluate the effect on HDAC 1 expression and activity. Invasion and migration assays are used to investigate the role of HDAC 1 and HDAC 6 expression in ccRCC cells. Comparisons are made between experimental groups using the paired T-test, the two-sample Student's T-test or one-way ANOVA, as appropriate. ccRCC and the TCGA dataset are used to observe the clinical correlation between HDAC 1 and HDAC 6 overexpression and overall and progression free survival. RESULTS: Our analysis of tumor and matched non-tumor tissues from radical nephrectomies showed overexpression of class I and II HDACs (HDAC6 only in a subset of patients). In vitro, both HDAC1 and HDAC6 over-expression increased cell invasion and motility, respectively, in ccRCC cells. HDAC1 regulated invasiveness by increasing matrix metalloproteinase (MMP) expression. Furthermore, hypoxia stimulation in VHL-reconstituted cell lines increased HIF isoforms and HDAC1 expression. Presence of hypoxia response elements in the HDAC1 promoter along with chromatin immunoprecipitation data suggests that HIF-2α is a transcriptional regulator of HDAC1 gene. Conversely, HDAC6 and estrogen receptor alpha (ERα) were co-localized in cytoplasm of ccRCC cells and HDAC6 enhanced cell motility by decreasing acetylated α-tubulin expression, and this biological effect was attenuated by either biochemical or pharmacological inhibition. Finally, analysis of human ccRCC specimens revealed positive correlation between HIF isoforms and HDAC. HDAC1 mRNA upregulation was associated with worse overall survival in the TCGA dataset. CONCLUSIONS: Taking together, these results suggest that HDAC1 and HDAC6 may play a role in ccRCC biology and could represent rational therapeutic targets.

GSH-targeted nanosponges increase doxorubicin-induced toxicity "in vitro" and "in vivo" in cancer cells with high antioxidant defenses.

Several reports indicate that chemo-resistant cancer cells become highly adapted to intrinsic oxidative stress by up-regulating their antioxidant systems, which causes an increase of intracellular GSH content. Doxorubicin is one of the most widely used drugs for tumor treatment, able to kill cancer cells through several mechanisms. However, doxorubicin use is limited by its toxicity and cancer resistance. Therefore, new therapeutic strategies able to reduce doses and to overcome chemo-resistance are needed. A new class of glutathione-responsive cyclodextrin nanosponges (GSH-NS), is able to release anticancer drugs preferentially in cells having high GSH content. Doxorubicin-loaded GSH-NS, in the cancer cells with high GSH content, inhibited clonogenic growth, cell viability, topoisomerase II activity and induced DNA damage with higher effectiveness than free drug. Moreover, GSH-NS reduced the development of human tumor in xenograft models more than free drug. These characteristics indicate that GSH-NS can be a suitable drug delivery carrier for future applications in cancer therapy.

Improved Anti-Tumoral Therapeutic Efficacy of 4-Hydroxynonenal Incorporated in Novel Lipid Nanocapsules in 2D and 3D Models.

4-hydroxynonenal (HNE), a lipid peroxidation product, is a promising anti-neoplastic drug due to its remarkable anti-cancer activities. However, this possibility has not been explored, because the delivery of HNE is very challenging as a result of its low solubility and its poor stability. This study intentionally designed a new type of lipid nanocapsules specifically for HNE delivery. They consist of a medium chain triglyceride liquid oil core surrounded by a polymer shell. A ß-cyclodextrin-poly(4-acryloylmorpholine) conjugate was selected as the shell component. HNE-loaded nanocapsules were about 350 nm in size with a negative surface charge. They were stable for two years when stored in suspensions at 4 degrees C. In vitro experiments showed that HNE was released from the nanocapsules at a considerable rate. Nanocapsule uptake into cells was evaluated using a fluorescent formulation that revealed rapid internalisation. Cytotoxicity studies demonstrated the safety of the formulation. Enhanced anti-tumoral activity against various cell lines, depending on increased HNE stability, was obtained by using HNE-loaded nanocapsules. In particular, we have demonstrated an increase in anti-proliferative, pro-apoptotic and differentiative activity in several tumour cell lines from different tissues. Moreover, we evaluated the effects of these new nanocapsules on a three-dimensional human reconstructed model of skin melanoma. Interestingly, the encouraging results obtained with topical administration on the epidermal surface could open new perspectives in melanoma treatments.

Generation of Adducts of 4-Hydroxy-2-nonenal with Heat Shock 60 kDa Protein 1 in Human Promyelocytic HL-60 and Monocytic THP-1 Cell Lines.

Heat shock 60 kDa protein 1 (HSP60) is a chaperone and stress response protein responsible for protein folding and delivery of endogenous peptides to antigen-presenting cells and also a target of autoimmunity implicated in the pathogenesis of atherosclerosis. By two-dimensional electrophoresis and mass spectrometry, we found that exposure of human promyelocytic HL-60 cells to a nontoxic concentration (10 µM) of 4-hydroxy-2-nonenal (HNE) yielded a HSP60 modified with HNE. We also detected adducts of HNE with putative uncharacterized protein CXorf49, the product of an open reading frame identified in various cell and tissue proteomes. Moreover, exposure of human monocytic THP-1 cells differentiated with phorbol 12-myristate 13-acetate to 10 µM HNE, and to light density lipoprotein modified with HNE (HNE-LDL) or by copper-catalyzed oxidation (oxLDL), but not to native LDL, stimulated the formation of HNE adducts with HSP60, as detected by immunoprecipitation and western blot, well over basal levels. The identification of HNE-HSP60 adducts outlines a framework of mutually reinforcing interactions between endothelial cell stressors, like oxLDL and HSP60, whose possible outcomes, such as the amplification of endothelial dysfunction, the spreading of lipoxidative damage to other proteins, such as CXorf49, the activation of antigen-presenting cells, and the breaking of tolerance to HSP60 are discussed.

Role of 4-hydroxynonenal-protein adducts in human diseases.

SIGNIFICANCE: Oxidative stress provokes the peroxidation of polyunsaturated fatty acids in cellular membranes, leading to the formation of aldheydes that, due to their high chemical reactivity, are considered to act as second messengers of oxidative stress. Among the aldehydes formed during lipid peroxidation (LPO), 4-hydroxy-2-nonenal (HNE) is produced at a high level and easily reacts with both low-molecular-weight compounds and macromolecules, such as proteins and DNA. In particular, HNE-protein adducts have been extensively investigated in diseases characterized by the pathogenic contribution of oxidative stress, such as cancer, neurodegenerative, chronic inflammatory, and autoimmune diseases. RECENT ADVANCES: In this review, we describe and discuss recent insights regarding the role played by covalent adducts of HNE with proteins in the development and evolution of those among the earlier mentioned disease conditions in which the functional consequences of their formation have been characterized. CRITICAL ISSUES: Results obtained in recent years have shown that the generation of HNE-protein adducts can play important pathogenic roles in several diseases. However, in some cases, the generation of HNE-protein adducts can represent a contrast to the progression of disease or can promote adaptive cell responses, demonstrating that HNE is not only a toxic product of LPO but also a regulatory molecule that is involved in several biochemical pathways. FUTURE DIRECTIONS: In the next few years, the refinement of proteomical techniques, allowing the individuation of novel cellular targets of HNE, will lead to a better understanding the role of HNE in human diseases.

Combination strategy targeting VEGF and HGF/c-met in human renal cell carcinoma models.

Alternative pathways to the VEGF, such as hepatocyte growth factor or HGF/c-met, are emerging as key players in tumor angiogenesis and resistance to anti-VEGF therapies. The aim of this study was to assess the effects of a combination strategy targeting the VEGF and c-met pathways in clear cell renal cell carcinoma (ccRCC) models. Male SCID mice (8/group) were implanted with 786-O tumor pieces and treated with either a selective VEGF receptor tyrosine kinase inhibitor, axitinib (36 mg/kg, 2×/day); a c-met inhibitor, crizotinib (25 mg/kg, 1×/day); or combination. We further tested this drug combination in a human ccRCC patient-derived xenograft, RP-R-01, in both VEGF-targeted therapy-sensitive and -resistant models. To evaluate the resistant phenotype, we established an RP-R-01 sunitinib-resistant model by continuous sunitinib treatment (60 mg/kg, 1×/day) of RP-R-01-bearing mice. Treatment with single-agent crizotinib reduced tumor vascularization but failed to inhibit tumor growth in either model, despite also a significant increase of c-met expression and phosphorylation in the sunitinib-resistant tumors. In contrast, axitinib treatment was effective in inhibiting angiogenesis and tumor growth in both models, with its antitumor effect significantly increased by the combined treatment with crizotinib, independently from c-met expression. Combination treatment also induced prolonged survival and significant tumor growth inhibition in the 786-O human RCC model. Overall, our results support the rationale for the clinical testing of combined VEGF and HGF/c-met pathway blockade in the treatment of ccRCC, both in first- and second-line setting.

Tasquinimod modulates suppressive myeloid cells and enhances cancer immunotherapies in murine models.

A major barrier for cancer immunotherapy is the presence of suppressive cell populations in patients with cancer, such as myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM), which contribute to the immunosuppressive microenvironment that promotes tumor growth and metastasis. Tasquinimod is a novel antitumor agent that is currently at an advanced stage of clinical development for treatment of castration-resistant prostate cancer. A target of tasquinimod is the inflammatory protein S100A9, which has been demonstrated to affect the accumulation and function of tumor-suppressive myeloid cells. Here, we report that tasquinimod provided a significant enhancement to the antitumor effects of two different immunotherapeutics in mouse models of cancer: a tumor vaccine (SurVaxM) for prostate cancer and a tumor-targeted superantigen (TTS) for melanoma. In the combination strategies, tasquinimod inhibited distinct MDSC populations and TAMs of the M2-polarized phenotype (CD206(+)). CD11b(+) myeloid cells isolated from tumors of treated mice expressed lower levels of arginase-1 and higher levels of inducible nitric oxide synthase (iNOS), and were less immunosuppressive ex vivo, which translated into a significantly reduced tumor-promoting capacity in vivo when these cells were coinjected with tumor cells. Tumor-specific CD8(+) T cells were increased markedly in the circulation and in tumors. Furthermore, T-cell effector functions, including cell-mediated cytotoxicity and IFNγ production, were potentiated. Taken together, these data suggest that pharmacologic targeting of suppressive myeloid cells by tasquinimod induces therapeutic benefit and provide the rationale for clinical testing of tasquinimod in combination with cancer immunotherapies.

Sunitinib dose escalation overcomes transient resistance in clear cell renal cell carcinoma and is associated with epigenetic modifications.

Sunitinib is considered a first-line therapeutic option for patients with advanced clear cell renal cell carcinoma (ccRCC). Despite sunitinib's clinical efficacy, patients eventually develop drug resistance and disease progression. Herein, we tested the hypothesis whether initial sunitinib resistance may be transient and could be overcome by dose increase. In selected patients initially treated with 50 mg sunitinib and presenting with minimal toxicities, sunitinib dose was escalated to 62.5 mg and/or 75 mg at the time of tumor progression. Mice bearing two different patient-derived ccRCC xenografts (PDX) were treated 5 days per week with a dose-escalation schema (40-60-80 mg/kg sunitinib). Tumor tissues were collected before dose increments for immunohistochemistry analyses and drug levels. Selected intrapatient sunitinib dose escalation was safe and several patients had added progression-free survival. In parallel, our preclinical results showed that PDXs, although initially responsive to sunitinib at 40 mg/kg, eventually developed resistance. When the dose was incrementally increased, again we observed tumor response to sunitinib. A resistant phenotype was associated with transient increase of tumor vasculature despite intratumor sunitinib accumulation at higher dose. In addition, we observed associated changes in the expression of the methyltransferase EZH2 and histone marks at the time of resistance. Furthermore, specific EZH2 inhibition resulted in increased in vitro antitumor effect of sunitinib. Overall, our results suggest that initial sunitinib-induced resistance may be overcome, in part, by increasing the dose, and highlight the potential role of epigenetic changes associated with sunitinib resistance that can represent new targets for therapeutic intervention.

Dll4 blockade potentiates the anti-tumor effects of VEGF inhibition in renal cell carcinoma patient-derived xenografts.

BACKGROUND: The Notch ligand Delta-like 4 (Dll4) is highly expressed in vascular endothelium and has been shown to play a pivotal role in regulating tumor angiogenesis. Blockade of the Dll4-Notch pathway in preclinical cancer models has been associated with non-productive angiogenesis and reduced tumor growth. Given the cross-talk between the vascular endothelial growth factor (VEGF) and Delta-Notch pathways in tumor angiogenesis, we examined the activity of a function-blocking Dll4 antibody, REGN1035, alone and in combination with anti-VEGF therapy in renal cell carcinoma (RCC). METHODS AND RESULTS: Severe combined immunodeficiency (SCID) mice bearing patient-derived clear cell RCC xenografts were treated with REGN1035 and in combination with the multi-targeted tyrosine kinase inhibitor sunitinib or the VEGF blocker ziv-aflibercept. Immunohistochemical and immunofluorescent analyses were carried out, as well as magnetic resonance imaging (MRI) examinations pre and 24 hours and 2 weeks post treatment. Single agent treatment with REGN1035 resulted in significant tumor growth inhibition (36-62%) that was equivalent to or exceeded the single agent anti-tumor activity of the VEGF pathway inhibitors sunitinib (38-54%) and ziv-aflibercept (46%). Importantly, combination treatments with REGN1035 plus VEGF inhibitors resulted in enhanced anti-tumor effects (72-80% growth inhibition), including some tumor regression. Magnetic resonance imaging showed a marked decrease in tumor perfusion in all treatment groups. Interestingly, anti-tumor efficacy of the combination of REGN1035 and ziv-aflibercept was also observed in a sunitinib resistant ccRCC model. CONCLUSIONS: Overall, these findings demonstrate the potent anti-tumor activity of Dll4 blockade in RCC patient-derived tumors and a combination benefit for the simultaneous targeting of the Dll4 and VEGF signaling pathways, highlighting the therapeutic potential of this treatment modality in RCC.

Drug delivery nanoparticles in skin cancers.

Nanotechnology involves the engineering of functional systems at nanoscale, thus being attractive for disciplines ranging from materials science to biomedicine. One of the most active research areas of the nanotechnology is nanomedicine, which applies nanotechnology to highly specific medical interventions for prevention, diagnosis, and treatment of diseases, including cancer disease. Over the past two decades, the rapid developments in nanotechnology have allowed the incorporation of multiple therapeutic, sensing, and targeting agents into nanoparticles, for detection, prevention, and treatment of cancer diseases. Nanoparticles offer many advantages as drug carrier systems since they can improve the solubility of poorly water-soluble drugs, modify pharmacokinetics, increase drug half-life by reducing immunogenicity, improve bioavailability, and diminish drug metabolism. They can also enable a tunable release of therapeutic compounds and the simultaneous delivery of two or more drugs for combination therapy. In this review, we discuss the recent advances in the use of different types of nanoparticles for systemic and topical drug delivery in the treatment of skin cancer. In particular, the progress in the treatment with nanocarriers of basal cell carcinoma, squamous cell carcinoma, and melanoma has been reported.

Dietary protein restriction inhibits tumor growth in human xenograft models.

PURPOSE: Data from epidemiological and experimental studies suggest that dietary protein intake may play a role in inhibiting prostate and breast cancer by modulating the IGF/AKT/mTOR pathway. In this study we investigated the effects of diets with different protein content or quality on prostate and breast cancer. EXPERIMENTAL DESIGN: To test our hypothesis we assessed the inhibitory effect of protein diet restriction on prostate and breast cancer growth, serum PSA and IGF-1 concentrations, mTOR activity and epigenetic markers, by using human xenograft cancer models. RESULTS: Our results showed a 70% inhibition of tumor growth in the castrate-resistant LuCaP23.1 prostate cancer model and a 56% inhibition in the WHIM16 breast cancer model fed with a 7% protein diet when compared to an isocaloric 21% protein diet. Inhibition of tumor growth correlated, in the LuCaP23.1 model, with decreased serum PSA and IGF-1 levels, down-regulation of mTORC1 activity, decreased cell proliferation as indicated by Ki67 staining, and reduction in epigenetic markers of prostate cancer progression, including the histone methyltransferase EZH2 and the associated histone mark H3K27me3. In addition, we observed that modifications of dietary protein quality, independently of protein quantity, decreased tumor growth. A diet containing 20% plant protein inhibited tumor weight by 37% as compared to a 20% animal dairy protein diet. CONCLUSIONS: Our findings suggest that a reduction in dietary protein intake is highly effective in inhibiting tumor growth in human xenograft prostate and breast cancer models, possibly through the inhibition of the IGF/AKT/mTOR pathway and epigenetic modifications.

Interaction of aldehydes derived from lipid peroxidation and membrane proteins.

A great variety of compounds are formed during lipid peroxidation of polyunsaturated fatty acids of membrane phospholipids. Among them, bioactive aldehydes, such as 4-hydroxyalkenals, malondialdehyde (MDA) and acrolein, have received particular attention since they have been considered as toxic messengers that can propagate and amplify oxidative injury. In the 4-hydroxyalkenal class, 4-hydroxy-2-nonenal (HNE) is the most intensively studied aldehyde, in relation not only to its toxic function, but also to its physiological role. Indeed, HNE can be found at low concentrations in human tissues and plasma and participates in the control of biological processes, such as signal transduction, cell proliferation, and differentiation. Moreover, at low doses, HNE exerts an anti-cancer effect, by inhibiting cell proliferation, angiogenesis, cell adhesion and by inducing differentiation and/or apoptosis in various tumor cell lines. It is very likely that a substantial fraction of the effects observed in cellular responses, induced by HNE and related aldehydes, be mediated by their interaction with proteins, resulting in the formation of covalent adducts or in the modulation of their expression and/or activity. In this review we focus on membrane proteins affected by lipid peroxidation-derived aldehydes, under physiological and pathological conditions.

The inclusion complex of 4-hydroxynonenal with a polymeric derivative of ß-cyclodextrin enhances the antitumoral efficacy of the aldehyde in several tumor cell lines and in a three-dimensional human melanoma model.

4-Hydroxynonenal (HNE) is the most studied end product of the lipoperoxidation process, by virtue of its relevant biological activity. The antiproliferative and proapoptotic effects of HNE have been widely demonstrated in a great variety of tumor cell types in vitro. Thus, it might represent a promising new molecule in anticancer therapy strategies. However, the extreme reactivity of this aldehyde, as well as its insolubility in water, a limiting factor for drug bioavailability, and its rapid degradation by specific enzymes represent major obstacles to its possible in vivo application. Various strategies can used to overcome these problems. One of the most attractive strategies is the use of nanovehicles, because loading drugs into nanosized structures enhances their stability and solubility, thus improving their bioavailability and their antitumoral effectiveness. Several natural or synthetic polymers have been used to synthesize nanosized structures and, among them, ß-cyclodextrin (ßCD) polymers are playing a very important role in drug formulation by virtue of the ability of ßCD to form inclusion compounds with a wide range of solid and liquid molecules by molecular complexation. Moreover, several ßCD derivatives have been designed to improve their physicochemical properties and inclusion capacities. Here we report that the inclusion complex of HNE with a derivative of ßCD, the ßCD-poly(4-acryloylmorpholine) conjugate (PACM-ßCD), enhances the aldehyde stability. Moreover, the inclusion of HNE in PACM-ßCD potentiates its antitumor effects in several tumor cell lines and in a more complex system, such as a human reconstructed skin carrying melanoma tumor cells.