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The impact of omega (ω)-3 fatty acids on prostate cancer is controversial in epidemiological studies but experimental studies suggest a protective effect. However, little is known about the mechanism of action. Here, we studied the effects of purified fatty acid molecules on prostate tumor progression using the TRAMP-C2 syngeneic immunocompetent mouse model. Compared with ω-6 or ω-9–supplemented animals, we observed that late-stage prostate tumor growth was reduced with a monoacylglyceride (MAG)-conjugated form of eicosapentaenoic acid (EPA) supplementation, whereas docosahexanenoic acid (DHA) caused an early reduction. MAG–EPA significantly decreased tumor blood vessel diameter (P < 0.001). RNA sequencing analysis revealed that MAG–EPA downregulated angiogenesis- and vascular-related pathways in tumors. We also observed this tissue vascular phenotype in a clinical trial testing MAG–EPA versus a high oleic sunflower oil placebo. Using anti-CD31 IHC, we observed that MAG–EPA reduced blood vessel diameter in prostate tumor tissue (P = 0.03) but not in normal adjacent tissue. Finally, testing autocrine and paracrine effects in an avascular tumor spheroid growth assay, both exogenous MAG–EPA and endogenous ω3 reduced VEGF secretion and in vitro endothelial cell tube formation and blocked tumor spheroid growth, suggesting that ω3 molecules can directly hinder prostate cancer cell growth. Altogether, our results suggest that fatty acids regulate prostate cancer growth and that a tumor-specific microenvironment is required for the anti-vascular effect of MAG–EPA in patients with prostate cancer.Implications:Increasing the amount of ingested EPA omega-3 subtype for patients with prostate cancer might help to reduce prostate tumor progression by reducing tumor vascularization.

S1. Chronic long-chain omega-3 (LCω3) treatment blocks prostate cancer cell growth. S2. Tables of differentially expressed genes in tumors of MAG-EPA-supplemented mice. S3. Twenty men supplemented for 4-10 weeks with MAG-EPA or placebo before radical prostatectomy. S4. Stable expression of the ω3 fatty acid desaturase 1 (FAT-1) from c. elegans, which convert ω6 into ω3 fatty acids, hinders prostate cancer cell growth. S5. Cytokine profiling of mouse tumors.

PDF file - 657K, Reintroduction of exogenous eIF4E2 restores tumor cell proliferation.

PDF file - 705K, eIF4E2 loss prevents tumor growth and reduces the growth of already-established tumors.

PDF file - 805K, Spheroids and xenografts contain hypoxic cores with inactive mTORC1.

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PDF file - 1255K, Cancer cells require eIF4E2-directed protein synthesis to survive and proliferate under hypoxia.

PDF file - 1124K, Normal cells require eIF4E2-directed protein synthesis to survive and proliferate under hypoxia.

PDF file - 797K, Cancer cells require eIF4E2-directed protein synthesis to evade cell death.

Human tumors display considerable diversity in their genetic makeup but share common physiologic attributes such as a hypoxic microenvironment that contribute to the malignant phenotype. Hypoxic cells switch from eukaryotic initiation factor 4E (eIF4E) to eIF4E2 cap-dependent translation to synthesize a portion of their proteins. Here, we show that genetically distinct human cancer cells exploit eIF4E2-directed protein synthesis to form cellular masses larger than approximately 0.15 mm, the diffusion limit of oxygen. Cancer cells depleted of eIF4E2 are indistinguishable from control cells under normoxic conditions, but are unable to survive and proliferate in low oxygen conditions. Activation of eIF4E2-directed translation is essential for cancer cells to form a hypoxic tumor core in in vitro spheroids and to form detectable tumors in in vivo xenograft assays. In contrast, the eIF4E-directed protein synthesis pathway alone cannot sustain cellular adaptation to hypoxia in vitro or confer tumorigenic potential in xenograft assays. These data demonstrate that the phenotypic expression of the cancer genome requires translation by the eIF4E2-directed hypoxic protein synthesis machinery. Cancer Res; 74(5); 1379–89. ©2014 AACR.

The impact of omega (ω)-3 fatty acids on prostate cancer is controversial in epidemiological studies but experimental studies suggest a protective effect. However, little is known about the mechanism of action. Here, we studied the effects of purified fatty acid molecules on prostate tumor progression using the TRAMP-C2 syngeneic immunocompetent mouse model. Compared with ω-6 or ω-9–supplemented animals, we observed that late-stage prostate tumor growth was reduced with a monoacylglyceride (MAG)-conjugated form of eicosapentaenoic acid (EPA) supplementation, whereas docosahexanenoic acid (DHA) caused an early reduction. MAG–EPA significantly decreased tumor blood vessel diameter (P < 0.001). RNA sequencing analysis revealed that MAG–EPA downregulated angiogenesis- and vascular-related pathways in tumors. We also observed this tissue vascular phenotype in a clinical trial testing MAG–EPA versus a high oleic sunflower oil placebo. Using anti-CD31 IHC, we observed that MAG–EPA reduced blood vessel diameter in prostate tumor tissue (P = 0.03) but not in normal adjacent tissue. Finally, testing autocrine and paracrine effects in an avascular tumor spheroid growth assay, both exogenous MAG–EPA and endogenous ω3 reduced VEGF secretion and in vitro endothelial cell tube formation and blocked tumor spheroid growth, suggesting that ω3 molecules can directly hinder prostate cancer cell growth. Altogether, our results suggest that fatty acids regulate prostate cancer growth and that a tumor-specific microenvironment is required for the anti-vascular effect of MAG–EPA in patients with prostate cancer.Implications:Increasing the amount of ingested EPA omega-3 subtype for patients with prostate cancer might help to reduce prostate tumor progression by reducing tumor vascularization.

Tumor‐associated macrophages (TAMs) support tumor progression within the tumor microenvironment (TME). Many questions remain as to the origin, development, and function of TAMs within the prostate TME. Evaluation of TAMs in prostate cancer (PCa) patients identified the immunosuppressive TAM marker CD163 in adjacent normal epithelium as an independent predictor of metastases or PCa death. Flow cytometry analyses identified prostate TAMs as frequently expressing both proinflammatory M1 (CCR7+) and immunosuppressive M2 (CD163+) markers. In vitro, we demonstrate PCa cells similarly subvert human M1 macrophages toward a mixed M1/M2 macrophage phenotype favoring tumor growth. Further the cytokine milieu‐induced transition between immunosuppressive M2 to proinflammatory M1 (M2→M1) macrophages is abrogated by the presence of PCa cells. RNA sequencing suggests alterations in chemokine expression in prostate TAMs due to the presence of PCa cells. Together, our results suggest that prostate TAMs originate from inflammatory infiltrating macrophages, which are then reprogrammed mainly by PCa cells, but also the cytokine milieu. A better understanding of this subversion of macrophages within the prostate may lead to novel treatment strategies., Our results demonstrate that mixed inflammatory and immunosuppressive macrophages found in the prostate develop because of prostate cancer contact. The presence of these macrophages in adjacent normal epithelium portends worse long‐term clinical outcomes for prostate cancer patients, highlighting the importance to develop effective strategies against their development.

Objective: Inducible nitric oxide (NO) synthase (NOS2) is a well-documented inflammatory mediator of insulin resistance in obesity. NOS2 expression is induced in both adipocytes and macrophages within adipose tissue during high-fat (HF)-induced obesity. Methods: Eight-week-old male mice with adipocyte selective deletion of the Nos2 gene (Nos2AD−KO) and their wildtype littermates (Nos2fl/fl) were subjected to chow or high-fat high-sucrose (HFHS) diet for 10 weeks followed by metabolic phenotyping and determination of brown adipose tissue (BAT) thermogenesis. The direct impact of NO on BAT mitochondrial respiration was also assessed in brown adipocytes. Results: HFHS-fed Nos2AD−KO mice had improved insulin sensitivity as compared to Nos2fl/fl littermates. Nos2AD−KO mice were also protected from HF-induced dyslipidemia and exhibited increased energy expenditure compared with Nos2fl/fl mice. This was linked to the activation of BAT in HFHS-fed Nos2AD−KO mice as shown by increased Ucp1 and Ucp2 gene expression and augmented respiratory capacity of BAT mitochondria. Furthermore, mitochondrial respiration was inhibited by NO, or upon cytokine-induced NOS2 activation, but improved by NOS2 inhibition in brown adipocytes. Conclusions: These results demonstrate the key role of adipocyte NOS2 in the development of obesity-linked insulin resistance and dyslipidemia, partly through NO-dependent inhibition of BAT mitochondrial bioenergetics.

The sensing of neurotransmitters is currently a difficult task for scientists in the field of biomedical research. The design of a new compact and accurate sensor would benefit greatly the research being done on neurotransmitters. In this effect, optoelectronic sensors are ideal candidate as they are non-invasive and non-reactive to the test sample. This work describes the design, fabrication, calibration, and tests of a new type of neurotransmitter sensor using spectral analysis based on a Grism. This new optical sensor was fitted with a self calibrating algorithm and showed superior resolution as well as a smaller footprint than previously observed.

Neurotransmitter sensing is currently a challenge in biomedical research, and optoelectronic sensing may play an important role in this field. In particular, miniature visible-light photo-spectrometers can be very useful for molecular detection. In this respect, this work evaluates a new type of spectrometer geometry based on the grism, or grating-prism, which combines an optical grating and an optical prism. The system was first designed in either transmission or reflection geometry, and then adapted to fit commercially available components so that a prototype could be fabricated in the laboratory. The comparison with a more conventional approach shows that the grism-based spectrometer offers superior resolution as well as a smaller footprint.

Neurotransmitter sensing is an importance component of biomedical research, and optoelectronic sensors play an important role in it. In particular, visible-light photo-spectrometers can be very useful for molecular detection and measurement. This work evaluates two optical design modes for the conception of a Czerny- Turner spectrometer. The system was designed at first using the non-sequential mode and then remade using the sequential mode. By comparing the same design made using both modes, the sequential mode had shown superior optimization potential.

Neurotransmitter sensing is essential in biomedical research to understand brain behavior, with Optical sensing useful for passive measurements. For instance, visible-light spectrophotometry can be used to detect and measure neurotransmitter concentrations in liquid. An important part of the process is the prediction of molecule concentration from the measured data. This work describes an FPGA-based system to measure neurotransmitter concentration in a multi-sensor platform, using a visible-light optical spectrometer and a simple regression predictor. The design required very few hardware resources and it was tested on dopamine data with excellent results.

Introduction and Objective: Recently, gut microbiota emerged as an important factor for success of immunity-based cancer treatments. However, its steady-state interaction and contribution to developing tumors is largely unexplored in non-intestinal cancers. Our objective was to investigate the connection between prostate tumor and the gut microbiota independently of cancer therapies. Methods: Human fecal samples were obtained from men participating into a phase IIb double-blind randomized controlled trial testing 3g/day of monoglyceride-eicosapentaenoic acid (MAG-EPA) versus placebo for a 4-10 week period before their radical prostatectomy (NCT02333435). A second set of samples were from men taking the same intervention of MAG-EPA or placebo after a PSA increase following their radical prostatectomy (NCT03753334). Short chain fatty acids (SCFA) analysis of patient stool samples between baseline and surgery was performed by gas chromatography coupled with flame ionization detection. 16srRNA libraries were amplified by targeting a fragment of the V3-V4 hypervariable region of the bacterial 16S rRNA gene. High-throughput sequencing of the bar-coded amplicons was performed on a MiSeq apparatus and the bioinformatics analysis was conducted using Mothur pipeline. In addition to human fecal samples, fully immunocompetent C57BL/6 mice were injected subcutaneously with TRAMP-C2 or PTEN−/− or PTEN−/− RB1−/− mouse prostate cancer cells to measure changes in the gut microbiota during tumor growth. We also recapitulated the MAG-EPA intervention in our TRAMP-C2 mice model and fed by gavage four different fatty acids (omega-9 (high oleic sunflower oil), omega-6 (MAG-arachidonic acid) and two omega-3 (MAG-docosahexaenoic and MAG-EPA). Results: In human fecal samples from prostate cancer patients, we observed a reduced gut microbiota diversity correlating with tumor stage. We also found that tumor growth was sufficient to modulate the microbiota in three independent prostate cancer syngeneic mouse models. We showed that transplanted human gut flora was sufficient to modulate ectopic prostate tumor growth, supporting the causal impact of gut microbiota for prostate cancer. The analysis of SCFA in patient stool samples between baseline and surgery showed that MAG-EPA prebiotic intervention was associated with a decrease of fecal butyric acid levels in prostate cancer patients with downgrade at surgery. We finally investigated this gut-tumor connection using purified polyunsaturated fatty acids prebiotics in patients and mice. We observed a reduction in the levels of Ruminococcaceae following dietary omega-3 supplementation that correlated with prostate cancer downgrade in patients and reduced tumor growth in mice. Conclusion: Overall our findings suggest that diet-actionable components of the gut microbiome can regulate prostate cancer growth. Citation Format: Jalal Laaraj, Gabriel Lachance, Nikunj Gevariya, Thibaut Varin, Andrei Feldiorean, Fanny Gaignier, Isabelle Boudreau Julien, Hui Wen Xu, Tarek Hallal, Jean-François Pelletier, Sidki Bouslama, Nadia Boufaied, Nicolas Derome, Yves Fradet, Leigh Ellis, Ciriaco A. Piccirillo, Frédéric Raymond, David P. Labbé, Alain Bergeron, André Marette, Karine Robitaille, Vincent Fradet. The gut microbiome-prostate tumor crosstalk is modulated by dietary polyunsaturated fatty acids [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2021 Oct 5-6. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(1 Suppl):Abstract nr P001.

We measured solar light transmitted through ice and snow cover with fiber optic probes, and examined the benefits and trade-offs of using diffusers and optics to reduce signal variations due to non-uniform illumination.

Epigenetic regulation of gene expression by DNA methylation plays a central role in the maintenance of cellular homeostasis. Here we present evidence implicating the DNA methylation program in the regulation of hypoxia-inducible factor (HIF) oxygen-sensing machinery and hypoxic cell metabolism. We show that DNA methyltransferase 3a (DNMT3a) methylates and silences the HIF-2α gene (EPAS1) in differentiated cells. Epigenetic silencing of EPAS1 prevents activation of the HIF-2α gene program associated with hypoxic cell growth, thereby limiting the proliferative capacity of adult cells under low oxygen tension. Naturally occurring defects in DNMT3a, observed in primary tumors and malignant cells, cause the unscheduled activation of EPAS1 in early dysplastic foci. This enables incipient cancer cells to exploit the HIF-2α pathway in the hypoxic tumor microenvironment necessary for the formation of cellular masses larger than the oxygen diffusion limit. Reintroduction of DNMT3a in DNMT3a-defective cells restores EPAS1 epigenetic silencing, prevents hypoxic cell growth, and suppresses tumorigenesis. These data support a tumor-suppressive role for DNMT3a as an epigenetic regulator of the HIF-2α oxygen-sensing pathway and the cellular response to hypoxia.

Hypoxia activates a translation initiation pathway that escapes global inhibition of protein synthesis. When cells are starved of oxygen, they downregulate the machinery that makes proteins by sequestering a translation-initiation factor, eIF4E, that binds the 5× cap of mRNAs. However, some proteins escape this mechanism and continue to be translated. Stephen Lee and colleagues show that in conditions of low oxygen tension, an alternative pathway of translation initiation is activated. A novel complex is formed that recognizes a specific hypoxia-responsive sequence on certain mRNAs. After binding this element, the complex is able to substitute for the cap-binding activity of eIF4E and thereby promote translation. Protein synthesis involves the translation of ribonucleic acid information into proteins, the building blocks of life. The initial step of protein synthesis is the binding of the eukaryotic translation initiation factor 4E (eIF4E) to the 7-methylguanosine (m7-GpppG) 5′ cap of messenger RNAs1,2. Low oxygen tension (hypoxia) represses cap-mediated translation by sequestering eIF4E through mammalian target of rapamycin (mTOR)-dependent mechanisms3,4,5,6. Although the internal ribosome entry site is an alternative translation initiation mechanism, this pathway alone cannot account for the translational capacity of hypoxic cells7,8. This raises a fundamental question in biology as to how proteins are synthesized in periods of oxygen scarcity and eIF4E inhibition9. Here we describe an oxygen-regulated translation initiation complex that mediates selective cap-dependent protein synthesis. We show that hypoxia stimulates the formation of a complex that includes the oxygen-regulated hypoxia-inducible factor 2α (HIF-2α), the RNA-binding protein RBM4 and the cap-binding eIF4E2, an eIF4E homologue. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP)10 analysis identified an RNA hypoxia response element (rHRE) that recruits this complex to a wide array of mRNAs, including that encoding the epidermal growth factor receptor. Once assembled at the rHRE, the HIF-2α–RBM4–eIF4E2 complex captures the 5′ cap and targets mRNAs to polysomes for active translation, thereby evading hypoxia-induced repression of protein synthesis. These findings demonstrate that cells have evolved a program by which oxygen tension switches the basic translation initiation machinery.

Human tumors display considerable diversity in their genetic makeup but share common physiologic attributes such as a hypoxic microenvironment that contribute to the malignant phenotype. Hypoxic cells switch from eukaryotic initiation factor 4E (eIF4E) to eIF4E2 cap-dependent translation to synthesize a portion of their proteins. Here, we show that genetically distinct human cancer cells exploit eIF4E2-directed protein synthesis to form cellular masses larger than approximately 0.15 mm, the diffusion limit of oxygen. Cancer cells depleted of eIF4E2 are indistinguishable from control cells under normoxic conditions, but are unable to survive and proliferate in low oxygen conditions. Activation of eIF4E2-directed translation is essential for cancer cells to form a hypoxic tumor core in in vitro spheroids and to form detectable tumors in in vivo xenograft assays. In contrast, the eIF4E-directed protein synthesis pathway alone cannot sustain cellular adaptation to hypoxia in vitro or confer tumorigenic potential in xenograft assays. These data demonstrate that the phenotypic expression of the cancer genome requires translation by the eIF4E2-directed hypoxic protein synthesis machinery. Cancer Res; 74(5); 1379–89. ©2014 AACR.