Fruit and vegetable production across the Mediterranean generates large volumes of by-products that can potentially lessen the livestock feeds deficit in some regions due to climate change effects and other socio-economic factors. However, the utilisation of agro-industrial by-products to their full potential is hindered by several factors such as variability in composition, limited development of preserving technologies and the presence of undesirable substances that may represent a risk for the animal or the final consumer. This work was conducted to study the nutritive value of a range of different alternative by-products from the Spanish agro-industry, assessing the potential to replace conventional ingredients often used in diets for small ruminants and the risk associated with the presence of pesticide residues and minerals. A total of 15 agro-industrial by-products commonly available in different regions of Spain were tested: three different types of apple pomace from three different varieties of apples (Perico, Regona, and Solarina), lemon pulp, avocado seed, broccoli, watermelon silage, olive cake, avocado cake, and six types of silages made with straw and a combination of different by-products (80/20 on fresh matter basis) from the intensive greenhouse horticulture: cucumber, tomato, pepper-cucumber mix (Pep-Cuc silage), aubergine-tomato mix (Aub-Tom silage), tomato-pepper-watermelon mix (Tom-Pep-Wat silage), and watermelon-melon-tomato mix (Wat-Mel-Tom silage). The study involved chemical characterization, including minerals and pesticides residues, and in vitro assessment of the ability to partially replace (10%, 20%, 30%, 50% and 100%) conventional ingredients in a diet for adult sheep (oat hay, barley and sunflower meal), depending on their chemical composition. The inclusion of by-products up to 30% replacement had not effect or improved some of the fermentation parameters and cucumber silage and apple pomace could successfully replace 100% of oat hay and barley, respectively, with only a significant reduction (P < 0.006) of rumen ammonia concentration. The mineral content was highly variable among the by-products with high values of Al in watermelon silage, Cu in olive cake and avocado by-products and Ni and Cr in olive cake. The pesticide residues assessment included 45 different types of compounds commonly used in crop production and three by-products had levels of pesticides residues above the legal limit: watermelon silage had traces of Chlorpyrifos (0.06 mg/kg DM), lemon pulp had Imazalil (0.09 mg/kg DM), and cucumber silage and Wat-Mel-Tom silages had Thiabendazole (0.04 mg/kg DM). Although all by-products showed potential to replace conventional ingredients from a nutritional perspective, the presence of high levels of some minerals and pesticides residues in some by-products requires regular analyses throughout the year or across batches before feeding them to animals.

Most mitigation strategies to reduce enteric methane (CH4) production in the rumen induce an excess of rumen dihydrogen (H2) that is expelled and consequently not redirected to the synthesis of metabolites that can be utilised by the ruminant. We hypothesised that phenolic compounds can be potential H2 acceptors when added to the diet, as they can be degraded to compounds that may be beneficial for the animal, using part of the H2 available when ruminal methanogenesis is inhibited. We performed four in vitro incubation experiments using rumen inoculum from Murciano-Granadina adult goats: Experiment 1 examined the inhibitory potential of Asparagopsis taxiformis (AT) at different concentrations (0, 1, 2, 3, 4 and 5% of the substrate on a DM basis) in 24 h incubations; Experiment 2 investigated the effect of a wide range of phenolic compounds (phenol, catechol, resorcinol, hydroquinone, pyrogallol, phloroglucinol, gallic acid and formic acid) at different doses (0, 2, 4, and 6 mM) on rumen fermentation for 24 h; Experiment 3 evaluated the combined effect of each phenolic compound at 6 mM with AT at 2% DM in sequential batch cultures for 5 days; and Experiment 4 examined the dose-response effect of phloroglucinol at different concentrations (0, 6, 16, 26 and 36 mM) combined with AT in sequential batch cultures for 5 days. Results from Experiment 1 confirmed that AT at 2% DM substantially inhibited CH4 production while significantly increasing H2 accumulation and decreasing the acetate:propionate ratio. Results from Experiment 2 showed that phenolic compounds did not negatively affect rumen fermenta-tion at any dose. In Experiment 3, each phenolic compound at 6 mM combined with AT at 2% DM inhib-ited CH4 production. Phloroglucinol numerically decreased H2 accumulation and significantly increased total gas production (TGP), volatile fatty acid (VFA) production and the acetate:propionate ratio. In Experiment 4, phloroglucinol at increasing doses supplemented with AT at 2% DM significantly decreased H2 accumulation and the abundances of archaea, protozoa and fungi abundances, and increased TGP, total VFA production and the acetate:propionate ratio in a dose-dependent way. In conclusion, combined treatment with AT and phloroglucinol was successful to mitigate CH4 production while preventing the accumulation of H2, leading to an increase in acetate and total VFA production and therefore an improve-ment in rumen fermentation in goats.(c) 2023 Published by Elsevier B.V. on behalf of The Animal Consortium. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

IntroductionThis study explores if promoting a complex rumen microbiota represents an advantage or a handicap in the current dairy production systems in which ruminants are artificially reared in absence of contact with adult animals and fed preserved monophyte forage.MethodsIn order to promote a different rumen microbial diversity, a total of 36 newborn goat kids were artificially reared, divided in 4 groups and daily inoculated during 10 weeks with autoclaved rumen fluid (AUT), fresh rumen fluid from adult goats adapted to forage (RFF) or concentrate (RFC) diets, or absence of inoculation (CTL). At 6 months of age all animals were shifted to an oats hay diet to determine their ability to digest a low quality forage.Results and discussionEarly life inoculation with fresh rumen fluid promoted an increase in the rumen overall microbial diversity which was detected later in life. As a result, at 6 months of age RFF and RFC animals had higher bacterial (+50 OTUs) and methanogens diversity (+4 OTUs) and the presence of a complex rumen protozoal community (+32 OTUs), whereas CTL animals remained protozoa-free. This superior rumen diversity and presence of rumen protozoa had beneficial effects on the energy metabolism allowing a faster adaptation to the forage diet, a higher forage digestion (+21% NDF digestibility) and an energetically favourable shift of the rumen fermentation pattern from acetate to butyrate (+92%) and propionate (+19%) production. These effects were associated with the presence of certain rumen bacterial taxa and a diverse protozoal community. On the contrary, the presence of rumen protozoa (mostly Entodinium) had a negative impact on the N metabolism leading to a higher bacterial protein breakdown in the rumen and lower microbial protein flow to the host based on purine derivatives urinary excretion (-17% to -54%). The inoculation with autoclaved rumen fluid, as source of fermentation products but not viable microbes, had smaller effects than using fresh inoculum. These findings suggest that enhancing rumen microbial diversity represents a desirable attribute when ruminants are fed forages in which the N supply does not represent a limiting factor for the rumen microbiota.

Ruminants are able to produce large quantities of saliva which enter into the rumen. Although previous research has indicated that salivary immunoglobulins can partially modulate the rumen microbial activity, the role of the salivary components other than ions on the rumen microbial ecosystem has not been thoroughly investigated in ruminants. A total of 16 semi-continuous in vitro cultures were used to incubate rumen fluid from 4 donor goats inoculated with autoclaved saliva (AUT) as negative control, saliva from the same rumen fluid donor (OWN) as positive control, and either GOAT or SHEEP saliva as experimental interventions. Fermentation was monitored throughout the 7 days of incubation and the prokaryotic communities and metabolome were analysed at day 7 of incubation. Characterization of the salivas used prior to incubation showed a high degree of individual variability in terms of the salivary metabolites and proteins, including immunoglobulins. The prokaryotic community composition in AUT incubators was the most divergent across treatments, suggesting a modulatory effect of active salivary components, which were not affected in the other treatments (OWN, GOAT and SHEEP). The differences across treatments in microbial diversity were mostly caused by a greater abundance of Proteobacteria and Rikenellacea and lower of Prevotellaceae, a key rumen bacterium with greater abundance in GOAT and SHEEP treatments. These results suggest that specific salivary components contribute to host-associated role in selecting the rumen commensal microbiota and its activity.

Abstract Background The red macroalgae Asparagopsis is an effective methanogenesis inhibitor due to the presence of halogenated methane (CH4) analogues, primarily bromoform (CHBr3). This study aimed to investigate the degradation process of CHBr3 from A. taxiformis in the rumen and whether this process is diet-dependent. An in vitro batch culture system was used according to a 2 × 2 factorial design, assessing two A. taxiformis inclusion rates [0 (CTL) and 2% DM diet (AT)] and two diets [high-concentrate (HC) and high-forage diet (HF)]. Incubations lasted for 72 h and samples of headspace and fermentation liquid were taken at 0, 0.5, 1, 3, 6, 8, 12, 16, 24, 48 and 72 h to assess the pattern of degradation of CHBr3 into dibromomethane (CH2Br2) and fermentation parameters. Additionally, an in vitro experiment with pure cultures of seven methanogens strains (Methanobrevibacter smithii, Methanobrevibacter ruminantium, Methanosphaera stadtmanae, Methanosarcina barkeri, Methanobrevibacter millerae, Methanothermobacter wolfei and Methanobacterium mobile) was conducted to test the effects of increasing concentrations of CHBr3 (0.4, 2, 10 and 50 µmol/L). Results The addition of AT significantly decreased CH4 production (P = 0.002) and the acetate:propionate ratio (P = 0.003) during a 72-h incubation. The concentrations of CHBr3 showed a rapid decrease with nearly 90% degraded within the first 3 h of incubation. On the contrary, CH2Br2 concentration quickly increased during the first 6 h and then gradually decreased towards the end of the incubation. Neither CHBr3 degradation nor CH2Br2 synthesis were affected by the type of diet used as substrate, suggesting that the fermentation rate is not a driving factor involved in CHBr3 degradation. The in vitro culture of methanogens showed a dose-response effect of CHBr3 by inhibiting the growth of M. smithii, M. ruminantium, M. stadtmanae, M. barkeri, M. millerae, M. wolfei, and M. mobile. Conclusions The present work demonstrated that CHBr3 from A. taxiformis is quickly degraded to CH2Br2 in the rumen and that the fermentation rate promoted by different diets is not a driving factor involved in CHBr3 degradation.

Newborn dairy ruminants are usually separated from their dams after birth and fed on milk replacer. This lack of contact with adult animals may hinder the rumen microbiological and physiological development. This study evaluates the effects of rearing newborn goat kids in contact with adult companions on the rumen development. Thirty-two newborn goat kids were randomly allocated to two experimental groups which were reared either in the absence (CTL) or in the presence of non-lactating adult goats (CMP) and weaned at 7 weeks of age. Blood and rumen samples were taken at 5, 7, and 9 weeks of age to evaluate blood metabolites and rumen microbial fermentation. Next-generation sequencing was carried out on rumen samples collected at 7 weeks of age. Results showed that CTL kids lacked rumen protozoa, whereas CMP kids had an abundant and complex protozoal community as well as higher methanogen abundance which positively correlated with the body weight and blood β-hydroxybutyrate as indicators of the physiological development. CMP kids also had a more diverse bacterial community (+132 ASVs) and a different structure of the bacterial and methanogen communities than CTL kids. The core rumen bacterial community in CMP animals had 53 more ASVs than that of CTL animals. Furthermore, the number of ASVs shared with the adult companions was over 4-fold higher in CMP kids than in CTL kids. Greater levels of early rumen colonizers Proteobacteria and Spirochaetes were found in CTL kids, while CMP kids had higher levels of Bacteroidetes and other less abundant taxa (Veillonellaceae, Cyanobacteria, and Selenomonas). These findings suggest that the presence of adult companions facilitated the rumen microbial development prior to weaning. This accelerated microbial development had no effect on the animal growth, but CMP animals presented higher rumen pH and butyrate (+45%) and ammonia concentrations than CTL kids, suggesting higher fibrolytic and proteolytic activities. CMP kids also had higher blood β-hydroxybutyrate (+79%) and lower blood glucose concentrations (-23%) at weaning, indicating an earlier metabolic development which could favor the transition from pre-ruminant to ruminant after the weaning process. Further research is needed to determine the effects of this intervention in more challenging farm conditions.

AbstractBACKGROUNDMeeting the energy and nitrogen (N) requirements of high‐performing ruminants at the same time as avoiding digestive disturbances (i.e. rumen acidosis) is a key priority in ruminant nutrition. The present study evaluated the effect of a cereal ammoniation treatment, in which barley grains are combined with urea and enzymes that catalyze the conversion of urea to ammonia to optimize rumen function. Twelve rumen cannulated sheep were randomly divided into two groups and fed a diet containing 60% of ammoniated barley (AMM) or untreated barley supplemented with urea (CTL) to investigate the impact on rumen fermentation and feed utilization.RESULTSAMM had higher total N content and effective rumen degradable N than untreated barely. AMM sheep had a consistently higher rumen pH throughout the day (6.31 versus 6.03) and tended to have a lower post‐prandial ammonia peak and higher acetate molar proportion (+5.1%) than CTL sheep. The rumen environment in AMM sheep favored the colonization and utilization of agro‐industrial by‐products (i.e. orange pulp) by the rumen microbes leading to a higher feed degradability. AMM sheep also had higher total tract apparent N digestibility (+21.7%) and urinary excretion of purine derivatives (+34%), suggesting a higher N uptake and microbial protein synthesis than CTL sheep.CONCLUSIONThe inclusion of AMM in the diet of ruminants represents a valid strategy for maintaining rumen pH within a physiological range and improving N utilization by the rumen microbes, which could have positive effects on the health and productivity of animals in intensive production systems. These findings warrant further studies under conventional farm conditions. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Background Newborn ruminants possess an underdeveloped rumen which is colonized by microorganisms acquired from adult animals and the surrounding environment. This microbial transfer can be limited in dairy systems in which newborns are separated from their dams at birth. This study explores whether the direct inoculation of fresh or autoclaved rumen fluid from adult goats to newborn kids has a beneficial effect on rumen microbial development and function. Results Repetitive inoculation of young kids with fresh rumen fluid from adult goats adapted to forage (RFF) or concentrate diets (RFC) accelerated microbial colonization of the rumen during the pre-weaning period leading to high protozoal numbers, a greater diversity of bacterial (+ 234 OTUs), methanogens (+ 6 OTUs) and protozoal communities (+ 25 OTUs) than observed in control kids (CTL) without inoculation. This inoculation also increased the size of the core bacterial and methanogens community and the abundance of key rumen bacteria ( Ruminococcaceae, Fibrobacteres, Veillonellaceae, Rikenellaceae, Tenericutes ), methanogens ( Methanobrevibacter ruminantium , Methanomicrobium mobile and Group 9 ), anaerobic fungi ( Piromyces and Orpinomyces ) and protozoal taxa ( Enoploplastron, Diplodinium, Polyplastron, Ophryoscolex, Isotricha and Dasytricha ) before weaning whereas CTL kids remained protozoa-free through the study. Most of these taxa were positively correlated with indicators of the rumen microbiological and physiological development (higher forage and concentrate intakes and animal growth during the post-weaning period) favoring the weaning process in RFF and RFC kids in comparison to CTL kids. Some of these microbiological differences tended to decrease during the post-weaning period, although RFF and RFC kids retained a more complex and matured rumen microbial ecosystem than CTL kids. Inoculation with autoclaved rumen fluid promoted lower development of the bacterial and protozoal communities during the pre-weaning period than using fresh inocula, but it favored a more rapid microbial development during the post-weaning than observed for CTL kids. Conclusions This study demonstrated that inoculation of young ruminants with fresh rumen fluid from adult animals accelerated the rumen microbial colonization which was associated with an earlier rumen functional development. This strategy facilitated a smoother transition from milk to solid feed favoring the animal performance during post-weaning and minimizing stress.

The research about the role of saliva in ruminants has been mainly focused on its buffering capacity together with facilitation of the rumination process. However, the role of salivary bioactive components on modulating the activity of the rumen microbiota has been neglected until recently. This study developed an in vitro approach to assess the impact of different components in saliva on rumen microbial fermentation. Four different salivary fractions were prepared from four goats: (i) non-filtrated saliva (NFS), (ii) filtrated through 0.25 µm to remove microorganisms and large particles (FS1), (iii) centrifuged through a 30 kDa filter to remove large proteins, (FS2), and (iv) autoclaved saliva (AS) to keep only the minerals. Two experiments were conducted in 24 h batch culture incubations with 6 ml of total volume consisting of 2 ml of rumen fluid and 4 ml of saliva/buffer mix. In Experiment 1, the effect of increasing the proportion of saliva (either NFS or FS1) in the solution (0%, 16%, 33% and 50% of the total volume) was evaluated. Treatment FS1 promoted greater total volatile fatty acids (VFA) (+8.4%) and butyrate molar proportion (+2.8%) but lower NH3-N concentrations than NFS fraction. Replacing the bicarbonate buffer solution by increasing proportions of saliva resulted in higher NH3-N, total VFA (+8.0%) and propionate molar proportion (+11%). Experiment 2 addressed the effect of the different fractions of saliva (NFS, FS1, FS2 and AS). Saliva fractions led to higher total VFA and NH3-N concentrations than non-saliva incubations, which suggests that the presence of some salivary elements enhanced rumen microbial activity. Fraction FS1 promoted a higher concentration of total VFA (+7.8%) than the other three fractions, and higher propionate (+26%) than NFS and AS. This agrees with findings from Experiment 1 and supports that 'microbe-free saliva', in which large salivary proteins are maintained, boosts rumen fermentation. Our results show the usefulness of this in vitro approach and suggest that different salivary components can modulate rumen microbial fermentation, although the specific metabolites and effects they cause need further research.

Background The knowledge about blood circulating microbiome and its functional relevance in healthy individuals remains limited. An assessment of changes in the circulating microbiome was performed by sequencing peripheral blood mononuclear cells (PBMC) bacterial DNA from goats supplemented or not in early life with rumen liquid transplantation. Results Most of the bacterial DNA associated to PBMC was identified predominantly as Proteobacteria (55%) followed by Firmicutes (24%), Bacteroidetes (11%) and Actinobacteria (8%). The predominant genera found in PBMC samples were Pseudomonas , Prevotella, Sphingomonas, Acinetobacter, Corynebacterium and Ruminococcus . Other genera such as Butyrivibrivio, Bifidobacterium , Dorea and Coprococcus were also present in lower proportions. Several species known as blood pathogens or others involved in gut homeostasis such as Faecalibacterium prausnitzii were also identified. However, the PBMC microbiome phylum composition differed from that in the colon of goats ( P ≤ 0.001), where Firmicutes was the predominant phylum (83%). Although, rumen liquid administration in early-life altered bacterial community structure and increased Tlr5 expression ( P = 0.020) in colon pointing to higher bacterial translocation, less than 8% of OTUs in colon were also observed in PBMCs. Conclusions Data suggest that in physiological conditions, PBMC microbiome differs from and is not affected by colon gut microbiota in small ruminants. Although, further studies with larger number of animals and covering other animal tissues are required, results point to a common circulating bacterial profile on mammals being phylum Proteobacteria, and genera Pseudomonas and Prevotella the most abundants . All suggest that PBMC microbiome in healthy ruminants could be implicated in homeostatic condition. This study expands our knowledge about PBMC microbiome contribution to health in farm animals.

Ruminants are born with an undeveloped physical, metabolic, and microbial rumen. Rumen development is limited under artificial rearing systems when newborn animals are separated from the dam, fed on milk replacer, and weaned at an early age. This study aims to evaluate the effects of early-life inoculation of young ruminants with rumen fluid from adult animals. Eighty newborn goat kids were randomly allocated to 1 of 4 experimental treatments and inoculated daily from d 1 to wk 11 with autoclaved rumen fluid (AUT), fresh rumen fluid obtained from adult goats fed either a forage diet (RFF) or concentrate-rich diet (RFC), or absence of inoculation (CTL). Goat kids were artificially reared with ad libitum access to milk replacer, starter concentrate, and forage hay. Blood was sampled weekly and rumen microbial fermentation was monitored at 5 (preweaning), 7 (weaning), and 9 wk of age (postweaning). Results indicated that inoculation with fresh rumen fluid accelerated the rumen microbial and fermentative development before weaning. As a result, RFC and RFF animals had higher solid feed intake (+73%), rumen concentrations of ammonia-N (+26%), total volatile fatty acids (+46%), butyrate (+50%), and plasma β-hydroxybutyrate (+48%), and lower milk intake (-6%) than CTL and AUT animals at wk 5. Inoculation with fresh inoculum also promoted early rumen colonization by a complex and abundant protozoal community, whereas CTL animals remained protozoa free. Although all kids experienced moderate growth retardation during 1 wk after weaning, inoculation with fresh rumen fluid favored the weaning process, leading to 2.2 times higher weight gain than CTL and AUT animals during wk 8. Some of these advantages were retained during the postweaning period and RFF and RFC animals showed higher forage intake (up to +44%) than CTL and AUT animals with no detrimental effects on feed digestibility or stress levels. The superior microbial load of RFC compared with RFF inoculum tended to provide further improvements in terms of forage intake, plasma β-hydroxybutyrate, and rumen protozoa, whereas AUT inoculation provided minor (if any) advantages with respect to CTL animals. Although no differences were noted on animal growth, this study suggests that early life inoculation of goat kids with rumen microbiota can represent an effective strategy to accelerate the rumen development, facilitating a smooth transition from milk to solid feed and to the potential implementation of early weaning strategies.

BACKGROUND The rumen microbiota has been used as inoculum for in vitro studies and as a probiotic to improve productivity in young animals. However, great variability across studies has been noted depending on the inoculum considered. The present study aims to assess the relevance of different factors (microbial fraction, collection time, donor animal diet, fermentation substrate and inoculum preservation method) to maximize the rumen inoculum activity and set the standards for further in vitro and in vivo applications. RESULTS Rumen inoculum sampled at 3 h after feeding led to greater microbial growth and activity [+12% volatile fatty acid (VFA), +17% ammonia] compared to before feeding. Similar results were noted when rumen liquid or rumen content were used as inocula. Rumen inoculum adapted to concentrate diets increased microbial activity (+19% VFA) independently of the substrate used in vitro. Freezing-thawing the inoculum, in comparison to fresh inoculum, decreased microbial activity (-14% VFA, -96% ammonia), anaerobic fungi and protozoa, with holotrichs protozoa being particularly vulnerable. Inoculum lyophilization had a stronger negative effect on microbial activity (-51% VFA) and delayed re-activation of the microbes, leading to lower levels of methanogens and anaerobic fungi, as well as almost complete wipe out of rumen protozoa. CONCLUSIONS Fresh rumen fluid sampled at 3 h after feeding from donor animals that were fed concentrate diets should be chosen when the aim is to provide the most diverse and active rumen microbial inoculum. © 2018 Society of Chemical Industry.

Lung cancer is the second most common malignant tumor and the leading cause of cancer death. Epidermal growth factor receptor (EGFR) -mutant non-small cell lung cancer (NSCLC) is a distinct subtype of lung cancer comprising approximately 15–40% of non-squamous tumors. The development of first- and second-generation EGFR tyrosine kinase inhibitors (TKIs) has been a significant step forward in the treatment of patients with EGFR -mutant tumors, and over the last few years has been the therapy of choice in the initial management of patients with activating mutations in EGFR , with some differences in efficacy and toxicity profile. Up to 50% of patients treated with first- and second-generation TKIs develop an EGFR exon 20 T790M mutation at the time of progression. In this context, osimertinib has shown a great benefit in terms of progression-free survival (PFS) in the second-line setting, including central nervous system metastasis control. The FLAURA trial, which compared osimertinib to first-generation inhibitors as first-line therapy, showed a clear PFS advantage for osimertinib and a trend towards an increased overall survival (OS) assessed by investigator review. Although T790M mutation is the most common mechanism of resistance to first- and second-generation EGFR TKIs, other EGFR-dependent and -independent mechanisms have been described, such as HER2 and MET amplifications or BRAF and MEK mutations. Some mechanisms of resistance to osimertinib and other third-generation TKIs have also been described. Several fourth-generation TKIs, targeted drug combinations and immunotherapy strategies are under investigation to overcome resistance to EGFR TKIs in order to improve EGFR -mutant NSCLC patient outcomes.

A broad clinical applicability of some next-generation sequencing (NGS) assays might be limited by analytic difficulties and tissue amount requirements. We successfully applied an amplicon-based NGS panel in advanced non-small-cell lung cancers (NSCLCs; n = 109). In nonsquamous tumors, immunohistochemistry tests for ALK and ROS1 with DNA NGS were combined. Forty NSCLCs had actionable mutations and 10 patients received tailored treatments. Introduction: A substantial fraction of non-small-cell lung cancers (NSCLCs) harbor targetable genetic alterations. In this study, we analyzed the feasibility and clinical utility of integrating a next-generation sequencing (NGS) panel into our routine lung cancer molecular subtyping algorithm.Patients and Methods: After routine pathologic and molecular subtyping, we implemented an amplicon-based gene panel for DNA analysis covering mutational hot spots in 22 cancer genes in consecutive advanced-stage NSCLCs. Results: We analyzed 109 tumors using NGS between December 2014 and January 2016. Fifty-six patients (51%) were treatment-naive and 82 (75%) had lung adenocarcinomas. In 89 cases (82%), we used samples derived from lung cancer diagnostic procedures. We obtained successful sequencing results in 95 cases (87%). As part of our routine lung cancer molecular subtyping protocol, single-gene testing for EGFR, ALK, and ROS1 was attempted in nonsquamous and 3 squamous-cell cancers (n = 92). Sixty-nine of 92 samples (75%) had sufficient tissue to complete ALK and ROS1 immunohistochemistry (IHC) and NGS. With the integration of the gene panel, 40 NSCLCs (37%) in the entire cohort and 30 NSCLCs (40%) fully tested for ALK and ROS1 IHC and NGS had actionable mutations. KRAS (24%) and EGFR (10%) were the most frequently mutated actionable genes. Ten patients (9%) received matched targeted therapies, 6 (5%) in clinical trials. Conclusion: The combination of IHC tests for ALK and ROS1 and amplicon-based NGS is applicable in routine clinical practice, enabling patient selection for genotype-tailored treatments. Copyright (C) 2017 Elsevier Inc. All rights reserved.

Although studies in rodents and humans have evidenced a weaker effect of fat in comparison to carbohydrates on the suppression of food intake, very few studies have been carried out in this field in dogs. This study investigates the effects of a high-carbohydrate (HC) and a high-fat (HF) diets on subsequent food intake and blood satiety-related hormones in dogs. Diets differed mainly in their starch (442 vs. 271 g/kg dry matter) and fat (99.3 vs. 214 g/kg dry matter) contents. Twelve Beagle dogs received the experimental diets at maintenance energy requirements in two experimental periods, following a cross-over arrangement. In week 7 of each period, blood concentrations of active ghrelin, glucagon-like peptide (GLP-1), peptide YY, insulin, and glucose were determined before and at 30, 60, 120, 180, and 360 min post-feeding. The following week, intake of a challenge food offered 180 min after the HC and HF diets was recorded over two days. In comparison to the dogs on the HC diet, those on the HF diet had a higher basal concentration of GLP-1 (p = .010) and a higher total area under the curve over 180 min post-prandial (tAUC0-180 ) (p = .031). Dogs on the HC diet showed a higher elevation of ghrelin at 180 min (p = .033) and of insulin at 360 min (p = .041), although ghrelin and insulin tAUC0-180 did not differ between the two diets (p ˃ .10). Diet had no effect on challenge food intake (p ˃ .10), which correlated with the tAUC0-180 of ghrelin (r = .514, p = .010), insulin (r = -.595, p = .002), and glucose (r = -.516, p = .010). Feeding a diet high in carbohydrate or fat at these inclusion levels does not affect the feeding response at 180 min post-prandial, suggesting a similar short-term satiating capacity.

Dacomitinib is a second-generation, irreversible, covalent pan-HER tyrosine-kinase inhibitor (TKI). It showed potent EGFR signaling inhibition in experimental models, including first-generation TKI-resistant non-small cell lung cancer (NSCLC) cell lines. This preclinical efficacy did not translate into clinically meaningful treatment benefits for advanced, pretreated, molecularly unselected NSCLC patients enrolled in two parallel phase III trials. Dacomitinib and erlotinib showed overlapping efficacy data in chemotherapy-pretreated EGFR wild-type (WT) patients in the ARCHER 1009 trial. Similarly, it failed to demonstrate any survival benefits as compared to placebo in EGFR WT subsets progressing on chemotherapy and at least one previous first-generation TKI (erlotinib or gefitinib) in the BR. 26 trial. In the case of EGFR-mutant NSCLCs, a pooled analysis of the ARCHER 1009 and ARCHER 1028 trials comparing the efficacy of dacomitinib vs. erlotinib in chemotherapy-pretreated, EGFR TKI-naive patients showed a trend to a longer progression-free survival (PFS) and overall survival in favor of dacomitinib that did not reach statistical significance, with a higher rate of treatment related adverse events (mainly skin rash, paronychia, and gastrointestinal toxicities). On the other hand, the clinical activity in patients with EGFR-mutant NSCLCs with acquired TKI resistance that were included in phase II/III trials was equally poor (response rate < 10%; PFS 3-4 months). Therefore, with the results of the ARCHER 1050 trial (NCT01774721) still pending, the current clinical development of dacomitinib is largely focused on EGFR-mutant, TKI-naive patients. Here, we review the most relevant clinical data of dacomitinib in advanced NSCLC. We discuss the potential role of dacomitinib in pretreated EGFR WT and EGFR-mutant (TKI-naive and TKI-resistant) patients. Finally, we briefly comment the available clinical data of dacomitinib in HER2-mutant NSCLC patients.

Dacomitinib is a second-generation, irreversible, covalent pan-HER tyrosine-kinase inhibitor (TKI). It showed potent EGFR signaling inhibition in experimental models, including first-generation TKI-resistant non-small cell lung cancer (NSCLC) cell lines. This preclinical efficacy did not translate into clinically meaningful treatment benefits for advanced, pretreated, molecularly unselected NSCLC patients enrolled in two parallel phase III trials. Dacomitinib and erlotinib showed overlapping efficacy data in chemotherapy-pretreated EGFR wild-type (WT) patients in the ARCHER 1009 trial. Similarly, it failed to demonstrate any survival benefits as compared to placebo in EGFR WT subsets progressing on chemotherapy and at least one previous first-generation TKI (erlotinib or gefitinib) in the BR.26 trial. In the case of EGFR-mutant NSCLCs, a pooled analysis of the ARCHER 1009 and ARCHER 1028 trials comparing the efficacy of dacomitinib vs. erlotinib in chemotherapy-pretreated, EGFR TKI-naïve patients showed a trend to a longer progression-free survival (PFS) and overall survival in favor of dacomitinib that did not reach statistical significance, with a higher rate of treatment related adverse events (mainly skin rash, paronychia, and gastrointestinal toxicities). On the other hand, the clinical activity in patients with EGFR-mutant NSCLCs with acquired TKI resistance that were included in phase II/III trials was equally poor (response rate <10%; PFS 3–4 months). Therefore, with the results of the ARCHER 1050 trial (NCT01774721) still pending, the current clinical development of dacomitinib is largely focused on EGFR-mutant, TKI-naïve patients. Here, we review the most relevant clinical data of dacomitinib in advanced NSCLC. We discuss the potential role of dacomitinib in pretreated EGFR WT and EGFR-mutant (TKI-naïve and TKI-resistant) patients. Finally, we briefly comment the available clinical data of dacomitinib in HER2-mutant NSCLC patients.