Transition to invasive breast cancer is associated with progressive changes in the structure and composition of tumor stroma.

Ductal carcinoma in situ (DCIS) is a pre-invasive lesion that is thought to be a precursor to invasive breast cancer (IBC). To understand the changes in the tumor microenvironment (TME) accompanying transition to IBC, we used multiplexed ion beam imaging by time of flight (MIBI-TOF) and a 37-plex antibody staining panel to interrogate 79 clinically annotated surgical resections using machine learning tools for cell segmentation, pixel-based clustering, and object morphometrics. Comparison of normal breast with patient-matched DCIS and IBC revealed coordinated transitions between four TME states that were delineated based on the location and function of myoepithelium, fibroblasts, and immune cells. Surprisingly, myoepithelial disruption was more advanced in DCIS patients that did not develop IBC, suggesting this process could be protective against recurrence. Taken together, this HTAN Breast PreCancer Atlas study offers insight into drivers of IBC relapse and emphasizes the importance of the TME in regulating these processes.

Inter-cellular forces orchestrate contact inhibition of locomotion.

Contact inhibition of locomotion (CIL) is a multifaceted process that causes many cell types to repel each other upon collision. During development, this seemingly uncoordinated reaction is a critical driver of cellular dispersion within embryonic tissues. Here, we show that Drosophila hemocytes require a precisely orchestrated CIL response for their developmental dispersal. Hemocyte collision and subsequent repulsion involves a stereotyped sequence of kinematic stages that are modulated by global changes in cytoskeletal dynamics. Tracking actin retrograde flow within hemocytes in vivo reveals synchronous reorganization of colliding actin networks through engagement of an inter-cellular adhesion. This inter-cellular actin-clutch leads to a subsequent build-up in lamellar tension, triggering the development of a transient stress fiber, which orchestrates cellular repulsion. Our findings reveal that the physical coupling of the flowing actin networks during CIL acts as a mechanotransducer, allowing cells to haptically sense each other and coordinate their behaviors.

Complete biosynthesis of QS-21 in engineered yeast.

QS-21 is a potent vaccine adjuvant and remains the only saponin-based adjuvant that has been clinically approved for use in humans1,2. However, owing to the complex structure of QS-21, its availability is limited. Today, the supply depends on laborious extraction from the Chilean soapbark tree or on low-yielding total chemical synthesis3,4. Here we demonstrate the complete biosynthesis of QS-21 and its precursors, as well as structural derivatives, in engineered yeast strains. The successful biosynthesis in yeast requires fine-tuning of the host's native pathway fluxes, as well as the functional and balanced expression of 38 heterologous enzymes. The required biosynthetic pathway spans seven enzyme families-a terpene synthase, P450s, nucleotide sugar synthases, glycosyltransferases, a coenzyme A ligase, acyl transferases and polyketide synthases-from six organisms, and mimics in yeast the subcellular compartmentalization of plants from the endoplasmic reticulum membrane to the cytosol. Finally, by taking advantage of the promiscuity of certain pathway enzymes, we produced structural analogues of QS-21 using this biosynthetic platform. This microbial production scheme will allow for the future establishment of a structure-activity relationship, and will thus enable the rational design of potent vaccine adjuvants.

Electrically driven organic laser using integrated OLED pumping.

Organic semiconductors are carbon-based materials that combine optoelectronic properties with simple fabrication and the scope for tuning by changing their chemical structure1-3. They have been successfully used to make organic light-emitting diodes2,4,5 (OLEDs, now widely found in mobile phone displays and televisions), solar cells1, transistors6 and sensors7. However, making electrically driven organic semiconductor lasers is very challenging8,9. It is difficult because organic semiconductors typically support only low current densities, suffer substantial absorption from injected charges and triplets, and have additional losses due to contacts10,11. In short, injecting charges into the gain medium leads to intolerable losses. Here we take an alternative approach in which charge injection and lasing are spatially separated, thereby greatly reducing losses. We achieve this by developing an integrated device structure that efficiently couples an OLED, with exceptionally high internal-light generation, with a polymer distributed feedback laser. Under the electrical driving of the integrated structure, we observe a threshold in light output versus drive current, with a narrow emission spectrum and the formation of a beam above the threshold. These observations confirm lasing. Our results provide an organic electronic device that has not been previously demonstrated, and show that indirect electrical pumping by an OLED is a very effective way of realizing an electrically driven organic semiconductor laser. This provides an approach to visible lasers that could see applications in spectroscopy, metrology and sensing.

Demographic consequences of phenological asynchrony for North American songbirds.

Changes in phenology in response to ongoing climate change have been observed in numerous taxa around the world. Differing rates of phenological shifts across trophic levels have led to concerns that ecological interactions may become increasingly decoupled in time, with potential negative consequences for populations. Despite widespread evidence of phenological change and a broad body of supporting theory, large-scale multitaxa evidence for demographic consequences of phenological asynchrony remains elusive. Using data from a continental-scale bird-banding program, we assess the impact of phenological dynamics on avian breeding productivity in 41 species of migratory and resident North American birds breeding in and around forested areas. We find strong evidence for a phenological optimum where breeding productivity decreases in years with both particularly early or late phenology and when breeding occurs early or late relative to local vegetation phenology. Moreover, we demonstrate that landbird breeding phenology did not keep pace with shifts in the timing of vegetation green-up over a recent 18-y period, even though avian breeding phenology has tracked green-up with greater sensitivity than arrival for migratory species. Species whose breeding phenology more closely tracked green-up tend to migrate shorter distances (or are resident over the entire year) and breed earlier in the season. These results showcase the broadest-scale evidence yet of the demographic impacts of phenological change. Future climate change-associated phenological shifts will likely result in a decrease in breeding productivity for most species, given that bird breeding phenology is failing to keep pace with climate change.

24-Hour Urinary Sodium and Potassium Excretion and Cardiovascular Risk.

BACKGROUND: The relation between sodium intake and cardiovascular disease remains controversial, owing in part to inaccurate assessment of sodium intake. Assessing 24-hour urinary excretion over a period of multiple days is considered to be an accurate method. METHODS: We included individual-participant data from six prospective cohorts of generally healthy adults; sodium and potassium excretion was assessed with the use of at least two 24-hour urine samples per participant. The primary outcome was a cardiovascular event (coronary revascularization or fatal or nonfatal myocardial infarction or stroke). We analyzed each cohort using consistent methods and combined the results using a random-effects meta-analysis. RESULTS: Among 10,709 participants, who had a mean (±SD) age of 51.5±12.6 years and of whom 54.2% were women, 571 cardiovascular events were ascertained during a median study follow-up of 8.8 years (incidence rate, 5.9 per 1000 person-years). The median 24-hour urinary sodium excretion was 3270 mg (10th to 90th percentile, 2099 to 4899). Higher sodium excretion, lower potassium excretion, and a higher sodium-to-potassium ratio were all associated with a higher cardiovascular risk in analyses that were controlled for confounding factors (P≤0.005 for all comparisons). In analyses that compared quartile 4 of the urinary biomarker (highest) with quartile 1 (lowest), the hazard ratios were 1.60 (95% confidence interval [CI], 1.19 to 2.14) for sodium excretion, 0.69 (95% CI, 0.51 to 0.91) for potassium excretion, and 1.62 (95% CI, 1.25 to 2.10) for the sodium-to-potassium ratio. Each daily increment of 1000 mg in sodium excretion was associated with an 18% increase in cardiovascular risk (hazard ratio, 1.18; 95% CI, 1.08 to 1.29), and each daily increment of 1000 mg in potassium excretion was associated with an 18% decrease in risk (hazard ratio, 0.82; 95% CI, 0.72 to 0.94). CONCLUSIONS: Higher sodium and lower potassium intakes, as measured in multiple 24-hour urine samples, were associated in a dose-response manner with a higher cardiovascular risk. These findings may support reducing sodium intake and increasing potassium intake from current levels. (Funded by the American Heart Association and the National Institutes of Health.).

A Common Variant in the Adaptor Mal Regulates Interferon Gamma Signaling.

Humans that are heterozygous for the common S180L polymorphism in the Toll-like receptor (TLR) adaptor Mal (encoded by TIRAP) are protected from a number of infectious diseases, including tuberculosis (TB), whereas those homozygous for the allele are at increased risk. The reason for this difference in susceptibility is not clear. We report that Mal has a TLR-independent role in interferon-gamma (IFN-γ) receptor signaling. Mal-dependent IFN-γ receptor (IFNGR) signaling led to mitogen-activated protein kinase (MAPK) p38 phosphorylation and autophagy. IFN-γ signaling via Mal was required for phagosome maturation and killing of intracellular Mycobacterium tuberculosis (Mtb). The S180L polymorphism, and its murine equivalent S200L, reduced the affinity of Mal for the IFNGR, thereby compromising IFNGR signaling in macrophages and impairing responses to TB. Our findings highlight a role for Mal outside the TLR system and imply that genetic variation in TIRAP may be linked to other IFN-γ-related diseases including autoimmunity and cancer.

Unraveling the Ultrafast Photochemical Dynamics of Nitrobenzene in Aqueous Solution.

Nitroaromatic compounds are major constituents of the brown carbon aerosol particles in the troposphere that absorb near-ultraviolet (UV) and visible solar radiation and have a profound effect on the Earth's climate. The primary sources of brown carbon include biomass burning, forest fires, and residential burning of biofuels, and an important secondary source is photochemistry in aqueous cloud and fog droplets. Nitrobenzene is the smallest nitroaromatic molecule and a model for the photochemical behavior of larger nitroaromatic compounds. Despite the obvious importance of its droplet photochemistry to the atmospheric environment, there have not been any detailed studies of the ultrafast photochemical dynamics of nitrobenzene in aqueous solution. Here, we combine femtosecond transient absorption spectroscopy, time-resolved infrared spectroscopy, and quantum chemistry calculations to investigate the primary steps following the near-UV (λ ≥ 340 nm) photoexcitation of aqueous nitrobenzene. To understand the role of the surrounding water molecules in the photochemical dynamics of nitrobenzene, we compare the results of these investigations with analogous measurements in solutions of methanol, acetonitrile, and cyclohexane. We find that vibrational energy transfer to the aqueous environment quenches internal excitation, and therefore, unlike the gas phase, we do not observe any evidence for formation of photoproducts on timescales up to 500 ns. We also find that hydrogen bonding between nitrobenzene and surrounding water molecules slows the S1/S0 internal conversion process.

How does weight gain since the age of 18 years affect breast cancer risk in later life? A meta-analysis.

Early life factors are important risk factors for breast cancer. The association between weight gain after age 18 and breast cancer risk is inconsistent across previous epidemiologic studies. To evaluate this association, we conducted a meta-analysis according to PRISMA guidelines and the established inclusion criteria. We performed a comprehensive literature search using Medline (Ovid), Embase, Scopus, Cochrane Library, and ClinicalTrials.gov to identify relevant studies published before June 3, 2022. Two reviewers independently reviewed the articles for final inclusion. Seventeen out of 4,725 unique studies met the selection criteria. The quality of studies was assessed using the Newcastle-Ottawa Scale (NOS), and all were of moderate to high quality with NOS scores ranging from 5 to 8. We included 17 studies (11 case-control, 6 cohort) in final analysis. In case-control studies, weight gain after age 18 was associated with an increased risk of breast cancer (odds ratio [OR] = 1.25; 95% CI = 1.07-1.48), when comparing the highest versus the lowest categories of weight gain. Menopausal status was a source of heterogeneity, with weight gain after age 18 associated with an increased risk of breast cancer in postmenopausal women (OR = 1.53; 95% CI = 1.40-1.68), but not in premenopausal women (OR = 1.01; 95% CI = 0.92-1.12). Additionally, a 5 kg increase in weight was positively associated with postmenopausal breast cancer risk (OR = 1.12; 95%CI = 1.05-1.21) in case-control studies. Findings from cohort studies were identical, with a positive association between weight gain after age 18 and breast cancer incidence in postmenopausal women (relative risk [RR] = 1.30; 95% CI = 1.09-1.36), but not in premenopausal women (RR = 1.06; 95% CI = 0.92-1.22). Weight gain after age 18 is a risk factor for postmenopausal breast cancer, highlighting the importance of weight control from early adulthood to reduce the incidence of postmenopausal breast cancer.

Analysis of ductal carcinoma in situ by self-reported race reveals molecular differences related to outcome.

BACKGROUND: Ductal carcinoma in situ (DCIS) is a non-obligate precursor to invasive breast cancer (IBC). Studies have indicated differences in DCIS outcome based on race or ethnicity, but molecular differences have not been investigated. METHODS: We examined the molecular profile of DCIS by self-reported race (SRR) and outcome groups in Black (n = 99) and White (n = 191) women in a large DCIS case-control cohort study with longitudinal follow up. RESULTS: Gene expression and pathway analyses suggested that different genes and pathways are involved in diagnosis and ipsilateral breast outcome (DCIS or IBC) after DCIS treatment in White versus Black women. We identified differences in ER and HER2 expression, tumor microenvironment composition, and copy number variations by SRR and outcome groups. CONCLUSIONS: Our results suggest that different molecular mechanisms drive initiation and subsequent ipsilateral breast events in Black versus White women.

Milestone review: GABA, from chemistry, conformations, ionotropic receptors, modulators, epilepsy, flavonoids, and stress to neuro-nutraceuticals.

Arising out of a PhD project more than 50 years ago to synthesise analogues of the neurotransmitter GABA, a series of new chemical entities were found to have selective actions on ionotropic GABA receptors. Several of these neurochemicals are now commercially available. A new subtype of these receptors was discovered that could be a target for the treatment of myopia, the facilitation of learning and memory, and the improvement of post-stroke motor recovery. The development of these new chemical entities over many years demonstrates the importance of neurochemicals with which to investigate selective aspects of GABA receptors and illustrates the significance of collaboration between chemists and biologists in neurochemistry. Vital were the improvements in synthetic organic chemistry and the use of functional human receptors expressed in oocytes. Current interest in ionotropic GABA receptors includes the clinical development of subtype-specific agents and the role of gain-of-function receptor variants in epilepsy. Dietary flavonoids were found to cross the blood-brain barrier to influence brain function. Natural and synthetic flavonoids had a range of effects on GABA receptors, ranging from positive, silent, and negative allosteric modulators, to even second-order modulation of first-order modulators. Flavonoids have been called "a new family of benzodiazepines." Like benzodiazepines, flavonoids reduce stress. Stress produces changes in GABA receptors in the brain that may be because of changes in endogenous modulators, such as neurosteroids and corticosteroids. GABA also occurs naturally in the diet leading to studies of the effects of oral GABA on brain function. This finding has resulted in studies of GABA and related neurochemicals as neuro-nutraceuticals. GABA systems in the gut microbiome are essential to such studies. The actions of oral GABA and of GABA-enriched beverages and foodstuffs are now an area of considerable scientific and commercial interest. GABA is a deceptively simple chemical that can take up many shapes, which may underlie its complex functions. The need for new chemical entities with selective actions for further studies highlights the need for continuing collaboration between chemists and biologists.

Multi-ancestry genome-wide association study of gestational diabetes mellitus highlights genetic links with type 2 diabetes.

Gestational diabetes mellitus (GDM) is associated with increased risk of pregnancy complications and adverse perinatal outcomes. GDM often reoccurs and is associated with increased risk of subsequent diagnosis of type 2 diabetes (T2D). To improve our understanding of the aetiological factors and molecular processes driving the occurrence of GDM, including the extent to which these overlap with T2D pathophysiology, the GENetics of Diabetes In Pregnancy Consortium assembled genome-wide association studies of diverse ancestry in a total of 5485 women with GDM and 347 856 without GDM. Through multi-ancestry meta-analysis, we identified five loci with genome-wide significant association (P < 5 × 10-8) with GDM, mapping to/near MTNR1B (P = 4.3 × 10-54), TCF7L2 (P = 4.0 × 10-16), CDKAL1 (P = 1.6 × 10-14), CDKN2A-CDKN2B (P = 4.1 × 10-9) and HKDC1 (P = 2.9 × 10-8). Multiple lines of evidence pointed to the shared pathophysiology of GDM and T2D: (i) four of the five GDM loci (not HKDC1) have been previously reported at genome-wide significance for T2D; (ii) significant enrichment for associations with GDM at previously reported T2D loci; (iii) strong genetic correlation between GDM and T2D and (iv) enrichment of GDM associations mapping to genomic annotations in diabetes-relevant tissues and transcription factor binding sites. Mendelian randomization analyses demonstrated significant causal association (5% false discovery rate) of higher body mass index on increased GDM risk. Our results provide support for the hypothesis that GDM and T2D are part of the same underlying pathology but that, as exemplified by the HKDC1 locus, there are genetic determinants of GDM that are specific to glucose regulation in pregnancy.

Sugar and sugar-sweetened beverages in relation to premature aging in adult survivors of childhood cancer.

BACKGROUND: Premature aging is a significant concern in adult survivors of childhood cancer as they develop aging-related conditions at a younger age than their peers with no history of childhood cancer. Although modifiable lifestyle factors, such as diet, are postulated to affect aging process, supporting evidence is sparse. METHODS: We examined if the consumption of sugar and sugar-sweetened beverages was related to premature aging in 3322 adult survivors of childhood cancer in the St. Jude Lifetime Cohort. Premature aging was assessed using the Deficit Accumulation Index (DAI) that was a ratio of the number of age-related chronic health conditions each survivor had out of 44 conditions total. Multinomial logistic regressions adjusting for confounders were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS: There were 46% of childhood cancer survivors consumed SSBs once or more times per day. High intake of sugar, especially sugars added to foods during preparation or processing, and habitual consumption of sugar-sweetened beverage were associated with an increased risk of premature aging. DISCUSSION: Our findings support a need to include strategies to reduce sugar and sugar-sweetened beverages consumption in lifestyle interventions to promote healthy aging in adult survivors of childhood cancer.

Predicting the onset of breast cancer using mammogram imaging data with irregular boundary.

With mammography being the primary breast cancer screening strategy, it is essential to make full use of the mammogram imaging data to better identify women who are at higher and lower than average risk. Our primary goal in this study is to extract mammogram-based features that augment the well-established breast cancer risk factors to improve prediction accuracy. In this article, we propose a supervised functional principal component analysis (sFPCA) over triangulations method for extracting features that are ordered by the magnitude of association with the failure time outcome. The proposed method accommodates the irregular boundary issue posed by the breast area within the mammogram imaging data with flexible bivariate splines over triangulations. We also provide an eigenvalue decomposition algorithm that is computationally efficient. Compared to the conventional unsupervised FPCA method, the proposed method results in a lower Brier Score and higher area under the ROC curve (AUC) in simulation studies. We apply our method to data from the Joanne Knight Breast Health Cohort at Siteman Cancer Center. Our approach not only obtains the best prediction performance comparing to unsupervised FPCA and benchmark models but also reveals important risk patterns within the mammogram images. This demonstrates the importance of utilizing additional supervised image-based features to clarify breast cancer risk.

Pectoral muscle removal in mammogram images: A novel approach for improved accuracy and efficiency.

PURPOSE: Accurate pectoral muscle removal is critical in mammographic breast density estimation and many other computer-aided algorithms. We propose a novel approach to remove pectoral muscles form mediolateral oblique (MLO) view mammograms and compare accuracy and computational efficiency with existing method (Libra). METHODS: A pectoral muscle identification pipeline was developed. The image is first binarized to enhance contrast and then the Canny algorithm was applied for edge detection. Robust interpolation is used to smooth out the pectoral muscle region. Accuracy and computational speed of pectoral muscle identification was assessed using 951 women (1,902 MLO mammograms) from the Joanne Knight Breast Health Cohort at Washington University School of Medicine. RESULTS: Our proposed algorithm exhibits lower mean error of 12.22% in comparison to Libra's estimated error of 20.44%. This 40% gain in accuracy was statistically significant (p < 0.001). The computational time for the proposed algorithm is 5.4 times faster when compared to Libra (5.1 s for proposed vs. 27.7 s for Libra per mammogram). CONCLUSION: We present a novel approach for pectoral muscle removal in mammogram images that demonstrates significant improvement in accuracy and efficiency compared to existing method. Our findings have important implications for the development of computer-aided systems and other automated tools in this field.

Avoiding lead-time bias by estimating stage-specific proportions of cancer and non-cancer deaths.

PURPOSE: Understanding how stage at cancer diagnosis influences cause of death, an endpoint that is not susceptible to lead-time bias, can inform population-level outcomes of cancer screening. METHODS: Using data from 17 US Surveillance, Epidemiology, and End Results registries for 1,154,515 persons aged 50-84 years at cancer diagnosis in 2006-2010, we evaluated proportional causes of death by cancer type and uniformly classified stage, following or extrapolating all patients until death through 2020. RESULTS: Most cancer patients diagnosed at stages I-II did not go on to die from their index cancer, whereas most patients diagnosed at stage IV did. For patients diagnosed with any cancer at stages I-II, an estimated 26% of deaths were due to the index cancer, 63% due to non-cancer causes, and 12% due to a subsequent primary (non-index) cancer. In contrast, for patients diagnosed with any stage IV cancer, 85% of deaths were attributed to the index cancer, with 13% non-cancer and 2% non-index-cancer deaths. Index cancer mortality from stages I-II cancer was proportionally lowest for thyroid, melanoma, uterus, prostate, and breast, and highest for pancreas, liver, esophagus, lung, and stomach. CONCLUSION: Across all cancer types, the percentage of patients who went on to die from their cancer was over three times greater when the cancer was diagnosed at stage IV than stages I-II. As mortality patterns are not influenced by lead-time bias, these data suggest that earlier detection is likely to improve outcomes across cancer types, including those currently unscreened.

Development of Corynebacterium glutamicum as a monoterpene production platform.

Monoterpenes are commonly known for their role in the flavors and fragrances industry and are also gaining attention for other uses like insect repellant and as potential renewable fuels for aviation. Corynebacterium glutamicum, a Generally Recognized as Safe microbe, has been a choice organism in industry for the annual million ton-scale bioproduction of amino acids for more than 50 years; however, efforts to produce monoterpenes in C. glutamicum have remained relatively limited. In this study, we report a further expansion of the C. glutamicum biosynthetic repertoire through the development and optimization of a mevalonate-based monoterpene platform. In the course of our plasmid design iterations, we increased flux through the mevalonate-based bypass pathway, measuring isoprenol production as a proxy for monoterpene precursor abundance and demonstrating the highest reported titers in C. glutamicum to date at 1504.6 mg/L. Our designs also evaluated the effects of backbone, promoter, and GPP synthase homolog origin on monoterpene product titers. Monoterpene production was further improved by disrupting competing pathways for isoprenoid precursor supply and by implementing a biphasic production system to prevent volatilization. With this platform, we achieved 321.1 mg/L of geranoids, 723.6 mg/L of 1,8-cineole, and 227.8 mg/L of linalool. Furthermore, we determined that C. glutamicum first oxidizes geraniol through an aldehyde intermediate before it is asymmetrically reduced to citronellol. Additionally, we demonstrate that the aldehyde reductase, AdhC, possesses additional substrate promiscuity for acyclic monoterpene aldehydes.

Systematic engineering for production of anti-aging sunscreen compound in Pseudomonas putida.

Sunscreen has been used for thousands of years to protect skin from ultraviolet radiation. However, the use of modern commercial sunscreen containing oxybenzone, ZnO, and TiO2 has raised concerns due to their negative effects on human health and the environment. In this study, we aim to establish an efficient microbial platform for production of shinorine, a UV light absorbing compound with anti-aging properties. First, we methodically selected an appropriate host for shinorine production by analyzing central carbon flux distribution data from prior studies alongside predictions from genome-scale metabolic models (GEMs). We enhanced shinorine productivity through CRISPRi-mediated downregulation and utilized shotgun proteomics to pinpoint potential competing pathways. Simultaneously, we improved the shinorine biosynthetic pathway by refining its design, optimizing promoter usage, and altering the strength of ribosome binding sites. Finally, we conducted amino acid feeding experiments under various conditions to identify the key limiting factors in shinorine production. The study combines meta-analysis of 13C-metabolic flux analysis, GEMs, synthetic biology, CRISPRi-mediated gene downregulation, and omics analysis to improve shinorine production, demonstrating the potential of Pseudomonas putida KT2440 as platform for shinorine production.

Verazine biosynthesis from simple sugars in engineered Saccharomyces cerevisiae.

Steroidal alkaloids are FDA-approved drugs (e.g., Zytiga) and promising drug candidates/leads (e.g., cyclopamine); yet many of the ≥697 known steroidal alkaloid natural products remain underutilized as drugs because it can be challenging to scale their biosynthesis in their producing organisms. Cyclopamine is a steroidal alkaloid produced by corn lily (Veratrum spp.) plants, and it is an inhibitor of the Hedgehog (Hh) signaling pathway. Therefore, cyclopamine is an important drug candidate/lead to treat human diseases that are associated with dysregulated Hh signaling, such as basal cell carcinoma and acute myeloid leukemia. Cyclopamine and its semi-synthetic derivatives have been studied in (pre)clinical trials as Hh inhibitor-based drugs. However, challenges in scaling the production of cyclopamine have slowed efforts to improve its efficacy and safety profile through (bio)synthetic derivatization, often limiting drug development to synthetic analogs of cyclopamine such as the FDA-approved drugs Odomzo, Daurismo, and Erivedge. If a platform for the scalable and sustainable production of cyclopamine were established, then its (bio)synthetic derivatization, clinical development, and, ultimately, widespread distribution could be accelerated. Ongoing efforts to achieve this goal include the biosynthesis of cyclopamine in Veratrum plant cell culture and the semi-/total chemical synthesis of cyclopamine. Herein, this work advances efforts towards a promising future approach: the biosynthesis of cyclopamine in engineered microorganisms. We completed the heterologous microbial production of verazine (biosynthetic precursor to cyclopamine) from simple sugars (i.e., glucose and galactose) in engineered Saccharomyces cerevisiae (S. cerevisiae) through the inducible upregulation of the native yeast mevalonate and lanosterol biosynthetic pathways, diversion of biosynthetic flux from ergosterol (i.e., native sterol in S. cerevisiae) to cholesterol (i.e., biosynthetic precursor to verazine), and expression of a refactored five-step verazine biosynthetic pathway. The engineered S. cerevisiae strain that produced verazine contains eight heterologous enzymes sourced from seven different species. Importantly, S. cerevisiae-produced verazine was indistinguishable via liquid chromatography-mass spectrometry from both a commercial standard (Veratrum spp. plant-produced) and Nicotiana benthamiana-produced verazine. To the best of our knowledge, this is the first report describing the heterologous production of a steroidal alkaloid in an engineered yeast. Verazine production was ultimately increased through design-build-test-learn cycles to a final titer of 83 ± 3 µg/L (4.1 ± 0.1 µg/g DCW). Together, this research lays the groundwork for future microbial biosynthesis of cyclopamine, (bio)synthetic derivatives of cyclopamine, and other steroidal alkaloid natural products.

Outcomes of sugar reduction policies, United Kingdom of Great Britain and Northern Ireland.

Poor diets are the major cause of death and disease globally, driving high levels of obesity and noncommunicable diseases. Cheap, heavily marketed, ultra-processed, energy-dense and nutrient-poor food and drinks that are high in fat, sugar and salt play a major role. The high-sugar content of these products leads to consumption levels much higher than recommended. The World Health Organization recommends that sugar intake should be reduced to just 5% of energy intake by using fiscal policies and food and drink reformulation strategies. Over the previous decade, the government of the United Kingdom of Great Britain and Northern Ireland has implemented several policies aimed at reducing sugar intake. We compare the soft drinks industry levy and the sugar reduction programme, examining how differences in policy design and process may have influenced the outcomes. Success has been mixed: the mandatory levy achieved a reduction in total sugar sales of 34.3%, and the voluntary reduction programme only achieved a 3.5% reduction in sugar levels of key contributors to sugar intake (despite a target of 20%). Both policies can be improved to enhance their impact, for example, by increasing the levy and reducing the sugar content threshold in the soft drinks industry levy, and by setting more stringent subcategory specific targets in the sugar reduction programme. We also recommend that policy-makers should consider applying a similar levy to other discretionary products that are key contributors to sugar intake. Both approaches provide valuable learnings for future policy in the United Kingdom and globally.

La malnutrition est l'une des principales causes de décès et de pathologies dans le monde, entraînant des taux élevés d'obésité et un grand nombre de maladies non transmissibles. Massivement commercialisés, les aliments et boissons bon marché, ultra-transformés, riches en énergie et pauvres en nutriments, à forte teneur en graisse, en sucre et en sel jouent un rôle majeur. La quantité de sucre contenue dans ces produits engendre une consommation qui dépasse largement les recommandations en la matière. L'Organisation mondiale de la Santé conseille de réduire la proportion de sucre afin que ce dernier ne représente plus que 5% de l'apport énergétique grâce à des politiques fiscales et des stratégies de révision de la composition des aliments et des boissons. Au cours des dix dernières années, le gouvernement du Royaume-Uni de Grande-Bretagne et d'Irlande du Nord a adopté plusieurs politiques visant à réduire la consommation de sucre. Dans le présent document, nous comparons la taxe sur l'industrie des sodas avec le programme de réduction du sucre, en examinant comment les différences de conception et de mise en œuvre des politiques pourraient avoir influencé les résultats. Le succès s'est révélé mitigé: la taxe obligatoire a permis de faire chuter le total des ventes de sucre de 34,3%, alors que le programme de baisse volontaire n'a pas permis de faire diminuer ce taux de plus de 3,5% chez les acteurs clés de l'apport en sucre (bien loin des 20% ciblés). Les deux politiques peuvent être améliorées pour renforcer leur impact, par exemple en augmentant la taxe et en réduisant la teneur en sucre maximale applicable à l'industrie des sodas, mais aussi en définissant des objectifs spécifiques plus stricts dans les sous-catégories du programme de réduction du sucre. Nous encourageons en outre les responsables politiques à instaurer une taxe similaire sur d'autres produits non essentiels qui contribuent eux aussi à la consommation de sucre. Les deux approches fournissent des renseignements précieux pour de futures mesures au Royaume-Uni et partout dans le monde.

Las dietas inadecuadas son la principal causa de muerte y enfermedad en todo el mundo. Además, impulsan altos niveles de obesidad y enfermedades no transmisibles. Los alimentos y las bebidas baratos, muy comercializados, ultraprocesados, hipercalóricos y pobres en nutrientes, con un alto contenido en grasas, azúcar y sal, desempeñan una función importante. El alto contenido en azúcar de estos productos conduce a niveles de consumo muy superiores a los recomendados. La Organización Mundial de la Salud recomienda reducir el consumo de azúcar a solo el 5% de la ingesta energética mediante políticas fiscales y estrategias de reformulación de alimentos y bebidas. En la última década, el gobierno del Reino Unido de Gran Bretaña e Irlanda del Norte ha aplicado varias políticas encaminadas a reducir la ingesta de azúcar. Comparamos el impuesto del sector de las bebidas no alcohólicas y el programa de reducción del azúcar, examinando cómo las diferencias en el diseño y el proceso de las políticas pueden haber influido en los resultados. El éxito ha sido desigual: el impuesto obligatorio logró una reducción de las ventas totales de azúcar del 34,3%, y el programa de reducción voluntaria solo consiguió una reducción del 3,5% en los niveles de azúcar de los principales contribuyentes a la ingesta de azúcar (a pesar de un objetivo del 20%). Se pueden mejorar ambas políticas para aumentar su impacto, por ejemplo, aumentando el impuesto y reduciendo el umbral de contenido de azúcar en el impuesto del sector de las bebidas no alcohólicas, y estableciendo objetivos específicos por subcategorías más estrictos en el programa de reducción de azúcar. También recomendamos a los responsables de formular las políticas que estudien la posibilidad de aplicar un impuesto similar a otros productos discrecionales que contribuyen decisivamente a la ingesta de azúcar. Ambos enfoques aportan valiosas enseñanzas para las futuras políticas del Reino Unido y del resto del mundo.