Indian Diabetic Risk Score (IDRS) and Community Based Assessment Checklist (CBAC) are easy, inexpensive, and non-invasive tools that can be used to screen people for Metabolic Syndrome (Met S). The study aimed to explore the prediction abilities of IDRS and CBAC tools for Met S.+ and LR-), Accuracy, and Youden’s index were calculated for different IDRS and CBAC scores cut-offs. Data were analyzed using SPSS v.23 and MedCalc v.20.111.All the people of age ≥30 years attending the selected rural health centers were screened for Met S. We used the International Diabetes Federation (IDF) criteria to diagnose the Met S. ROC curves were plotted by taking Met S as dependent variables, and IDRS and CBAC scores as independent/prediction variables. Sensitivity (SN), specificity (SP), Positive and Negative Predictive Value (PPV and NPV), Likelihood Ratio for positive and negative tests  were found to have Met S. Area Under the Curve (AUC) for IDRS in predicting Met S was 0.73 (95%CI: 0.67–0.79), with 76.3% (64.0%-85.3%) sensitivity and 54.6% (51.2%-57.8%) specificity at the cut-off of ≥60. For the CBAC score, AUC was 0.73 (95%CI: 0.66–0.79), with 84.7% (73.5%-91.7%) sensitivity and 48.8% (45.5%-52.1%) specificity at the cut-off of ≥4 . The AUCs of both parameters (IDRS and CBAC scores) were statistically significant. There was no significant difference (p = 0.833) in the AUCs of IDRS and CBAC [Difference between AUC = 0.00571].The current study provides scientific evidence that both IDRS and CBAC have almost 73% prediction ability for Met S. Though CBAC holds relatively greater sensitivity (84.7%) than IDRS (76.3%), the difference in prediction abilities is not statistically significant. The prediction abilities of IDRS and CBAC found in this study are inadequate to qualify as Met S screening tools. An accumulation of clinical, physiological, biochemical, and metabolic factors which increases the risk of cardiovascular disease, type 2 diabetes mellitus (DM), and all-cause mortality is known as Metabolic Syndrome (Met S) . Rapid uDue to the unique "atherogenic dyslipidemic profile" and "South Asian phenotype," Indians have a significant chance of developing Met S , 5. The Though there are differences in diagnostic criteria of Met S, and the published evidence also varies with respect to the age of study participants and methodology, a significant epidemic of Met S is emerging in the Asia-Pacific region . AccordiIn a country like India, the western world’s influence on dietary patterns, which replace fiber-rich foods with refined carbohydrates and saturated fats, has a significant impact on the expanding obesity epidemic which in turn increases the burden of Met S. A significant association between anthropometric risk factors and Met S among Indian adults has been documented by a systematic review of observational studies which highlighted that people with overweight  or obesity  had higher odds of having Met S than those of normal or low body weight . Due to The Met S is driving the twin global epidemics of type 2 diabetes and CVD. There is an enormous moral, medical, and economic responsibility to identify persons with Met S early so that lifestyle modifications and treatment may avoid the development of diabetes and cardiovascular disease.To prevent and control major NCDs, the Government of India started the National Programme for Prevention & Control of Cancer, Diabetes, Cardiovascular Diseases & Stroke (NPCDCS) in 2010. Grass-root workers use a Community-Based Assessment Checklist (CBAC) under this program. This checklist is based on two non-modifiable (age and family history of NCDs) and four modifiable  known risk factors of NCDs. This checklist helps in screening high-risk individuals for NCDs . The MadThere are no screening standards or procedures in place in India for Met S. These (IDRS and CBAC) easy and inexpensive tools, which have already been scientifically validated and are widely used to screen patients for NCDs, can also be used to screen people for Met S in the country. Before this, it is necessary to assess those tools’ prediction abilities (sensitivity and specificity). So, the current study was conducted to screen persons for Met S who visited primary health centers and to explore the prediction abilities of IDRS and CBAC tools for Met S.This facility-based cross-sectional study was carried out from January to December 2019 in the three Rural Health Training Centres (RHTCs) of the Department of Community Medicine and Family Medicine at a tertiary care hospital in Jodhpur, India. Out of these three centers, one is Community Health Centre (CHC), and the other two are Primary Health Centres (PHCs). The three health centers’ catchment areas serve approximately one lakh sixty thousand population. All the patients of age ≥ 30 years attending these health centers for various ailments were included in the study. Pregnant females and patients already diagnosed with cholesterol gallstones, asthma, Polycystic ovary syndrome (PCOS), sleep apnoea, autoimmune disorders, and cancer of colorectal (colon and rectum), gastric, oesophageal, hepatobiliary , pancreas, lung, bladder, thyroid, renal, leukaemia, malignant melanoma, multiple myeloma, and non-Hodgkin lymphoma were excluded from the study.The detailed methodology of the study is mentioned in another published article .For operational purposes, the subjective assessments about addictions and physical activity were made based on IDRS and CBAC forms. As per the CBAC form, a drinker is a person who consumes alcohol daily, irrespective of quantity. A smoker was categorized as one who used to smoke in the past/sometimes now and another who smokes on a daily basis. According to CBAC, physical activity was defined as any dedicated physical activity for a minimum of 150 minutes in a week. While in IDRS, regular exercise and strenuous work were considered separately. The presence of a family history (any one of the parents or siblings) of high blood pressure, diabetes, and heart disease was considered a positive family history for NCDs, as per CBAC. While in IDRS, different scores were assigned to having a positive family history of only diabetes in either parent or both parents.2, central obesity was assumed irrespective of the waist circumference. The other two criteria used were Blood Pressure (systolic BP ≥ 130 or diastolic BP ≥ 85 mm Hg) and Raised fasting plasma glucose (FPG ≥ 100 mg/dL)  Table 4 provides the sensitivity, specificity, PPV, NPV, LR+, LR-, and accuracy of different cut-offs for IDRS and CBAC scores for predicting Met S. An IDRS value ≥60 and CBAC value of ≥4 had the optimum sensitivity  and specificity  for determining Met S.In this study, facility-based screening of rural people of Western Rajasthan was done to estimate the proportion of newly diagnosed cases of Met S. The prediction capacity of the existing non-invasive IDRS and CBAC scores was calibrated to screen for Met S, and the appropriate cut-offs were determined.Met S was diagnosed among 6.4% of participants in the present study. This is relatively lower than the prevalence (15.6%) reported by a large survey in India among the rural adult population . A studyThough IDRS is used to screen for diabetes, we tried to explore its prediction ability to diagnose Met S in this study. In our study, AUC for IDRS score in predicting Met S was 0.73 (95%CI: 0.67–0.79), with 76.3% sensitivity and 54.6% specificity at the cut-off of ≥60, and with 59.3% sensitivity and 77.9% specificity at the cut-off of ≥70. When the major part of the disease iceberg is hidden in the community, and untreated disease could end up with serious complications, a score that maximizes true positives is preferable. So, the screening cut-off is often set at a lower level, which increases the sensitivity value. Thus, IDRS with a cut-off of ≥60 is more appropriate to predict Met S. The present study’s findings are well supported by a large study conducted in the Southern part of India . The stuThe results of this study also show the prediction ability of the CBAC form, which is extensively used in India by front-line healthcare workers to screen for NCDs. Traditionally, in the screening using the CBAC checklist, a score above 4 indicates that the person may be at risk for NCDs. In our study, AUC for CBAC score in predicting Met S was 0.73 (95%CI: 0.66–0.79), with 84.7% sensitivity and 48.8% specificity at the cut-off of ≥4 . There is a dearth of scientific evidence exploring the prediction capacity of CBAC form for Met S. Based on the findings of this study; the CBAC tool also does not seem promising in identifying people at high risk of developing Met S in primary care settings.The most accurate metric for gauging the effectiveness of a risk score is the total area under the ROC curve. The risk score performs better overall in terms of adequately predicting persons who will get a disease when the area under the curve is higher . The preThis study was non-funded, so the HDL and Triglyceride levels of the study participants could not be assessed. This may underestimate the prevalence of Met S in the study. This is one of the limitations of this study. Besides that, the authors did not collect data related to the diet of study participants, so the effect of diet could not be adjusted for the prediction of Met S using IDRS and CBAC.The current study provides scientific evidence that both IDRS and CBAC have almost 73% prediction ability for Met S. Though CBAC holds relatively greater sensitivity (84.7%) than IDRS (76.3%), the difference in prediction abilities is not statistically significant. The prediction abilities of IDRS and CBAC found in this study are inadequate to qualify as Met S screening tools. Thus, both tools do not seem promising in identifying people at high risk of developing Met S in primary care settings. As these tools are the best available non-invasive tools for screening NCDs and are widely used in India, their prediction abilities need to be explored further with a larger sample size and multicentric basis by considering the limitations mentioned in this study.S1 File(XLSX)Click here for additional data file.


Geopolymer is an environment friendly construction material that could be synthesized using either the natural source or the industrial byproducts such as flyash and GGBS. The characteristics of the Geopolymer rely on the proportion of the flyash and GGBS and the concentration of the activator solution used. In this research work, the effect of partial replacement of flyash with GGBS in proportions such as 0, 10, 20, 30 and 40% is investigated. Also Molarity of NaOH are tested from 8 to 14 M and both the parameters are optimized. In this optimized Geopolymer concrete, the utilization of iron slag as a partial substitute for river sand in various proportions such as 10, 15, 20, 25, 30 35, 40 and 45% are investigated. The optimized Geopolymer concrete with iron slag is investigated for its performance as a paver block with incorporation of banana fiber in proportions such as 0, 0.5, 1 and 1.5 and is compared with conventional cement concrete paver block. The results show that there is a significant enhancement in the properties of Geopolymer concrete with the different levels of optimization and the utilization of natural banana fiber. The developed sustainable paver block was found to with stand medium traffic conditions as per IS 15658:2006. Further this study employed random forest (RF) algorithm for the prediction of compressive strength of geopolymer concrete specimens for the variable parameters such as molarity of alkaline solution, Flyash/GGBS ratio and partial replacement of river sand with iron slag. The performance evaluation parameters represented high accuracy of developed RF model. This research work unleashes a heft potential of Geopolymer concrete to develop economical eco-friendly sustainable paver blocks to the society through mitigation of environmental strain on the ecosystem. The base materials are available as a natural source such as kaolinite, metakaolin or industrial effluents such as Ground Granulated Blast Furnace Slag (GGBS), fly ash, high calcium wood ash, rise husk ash, waste glass powder10. Duxson et al.11 reported early strength attainment with the slag-based geopolymer matrix and enhanced durability with the fly ash-based geopolymer concrete (FGC). FGC concrete requires heat curing for the effective gaining of strength. Yildirim and Prezzi performed various tests and analyzed the suitability of unused slag for the use as construction material and reported superior performance than the conventional materials in the field. Sithole et al., stated the ability of GGBS to produce a sustainable concrete with the inclusion of foundary sand as the fine aggregate instead of natural sand.In the recent decade, significant research works have been concentrated in the vicinity of Geopolymer as a construction material due to its highest versatility. Owing to its outstanding property of being able to synthesise concrete without cement content, geopolymer concrete (GC) gains importance as an eco-friendly and sustainable building material as it addresses the acute environmental issues caused by the manufacture of cement. Geopolymer is inorganic and is synthesized by the activation of alumina silicate base compounds using alkaline solution12 utilized GGBS as a partial substitute for FGC and stated the possibility of synthesis of Geopolymer concrete under ambient curing. Investigational works performed by Hamidi et al.13 and Hardjito et al.14 report that the molarity of the alkaline solution and curing condition hold a major part in the strength-gaining process of GC. The binding potency of GC is primarily affected due to the concentration of the alkaline solution and the category of base materials involved in the process. Hence effort can be concentrated towards the synthesis of Geopolymer concrete with optimum fly ash–slag blend ratio and optimum concentration of an alkaline solution16.Deb et al.26. At high temperatures, geopolymer concrete performs better than cement concrete with increased retained compressive strength. This is due to the sturdy bond that exists within the steel and matrix than the cement concrete28. In addition, geopolymer concrete proves to be cost-beneficial when compared to conventional concrete with a reduced economic index29. The infirmity of Geopolymer is its brittle character. The fracture of Geopolymer was observed to be smoother than the conventional concrete30. Pan et al.31 stated that the brittleness is because of the morphology of the binder material that alters the binder inside the matrix. Contemporary advancement in the concrete industry is fibre-reinforced concrete. The existence of fibres improves the structural integrity and reduces brittleness33. Many research work report that the integration of fibre augments the hardened properties of the concrete37. Additions of synthetic fibres reduce the brittleness and improve the impact energy40. Considering the economic importance, natural fibres can be employed to reduce the brittleness of the GC. Out of many available natural fibres, owing to the high tensile strength and young’s modulus, Banana fibres are preferred to coconut or sugarcane bagasse fibre44. Hence by incorporating banana fibres, an endeavour can be focused towards reducing this brittleness of geopolymer concrete.Many research findings document high compressive properties and increased resistance against acid and fire attacks with low creep in Geopolymer concrete46. Geopolymer paver blocks synthesized in various research works were found to perform well only under low traffic conditions as prescribed by IS 15658: 200647. Analysis of failed paver block claims the low impact energy and brittleness of the Geopolymer concrete as the reason for failure. Hence an approach can be made in the production of a Geopolymer paver block to reduce its brittleness and match the medium traffic conditions as prescribed by IS 15658: 200647.Geopolymer is used in the production of bricks, boat ramps, panels for walls, water tanks and paver blocks48 reported a global imbalance in the supply and demand of the sand market with the major share being held by the construction industry. Another problem faced by the construction sector is the scarcity of river sand. Adding to this point, river sand mining leads to serious environmental problems. Padmalal et al.49 conducted a field survey and claimed that river sand mining accounts for bank erosion, affected surface water and aquatic flora and fauna, the social life of the locality and lowered groundwater. Sand mining leads to the undermining of spillways, abutments and piers. Anthony et al.50 claimed sand mining as an important reason for the reduced supply of sediments to the coast and bank erosion. The intrusion of seawater was observed due to sand mining which affected the cultivation of the locality. Many research works recommended identifying a new material to replace the river sand as fine aggregate52. On the other hand, there is a problem with the disposal of iron slag, a waste residue that comes out of the iron industry. Noufal and Singh53 and Singh and Siddique54 reported the enhancement of mechanical characteristics of conventional cement concrete by replacing river sand with iron slag. Hence an endeavour can be made to replace the river sand with iron slag inside the optimized geopolymer concrete.Gavriletea et al.Many research works concentrate on the development of GC. But only a few focus towards the optimization of different factors that alter the geopolymerization and proceeding with the product development. This research work focuses the limelight on the production of sustainable eco-friendly paver blocks using various industrial wastes like fly ash, GGBS, iron slag and banana fibres. The novelty of this research study is the optimization of important factors for geopolymerization, utilization of iron slag in optimized GC and further optimization of incorporation of natural fibres in optimized Geopolymer concrete. Further, an analytical study using the random forest algorithm is carried out to develop a relationship between the output compressive strength and input variables such as concentration of activator solution, fly ash/GGBS ratio and fractional substitute of river sand with iron slag. The developed model for the prediction of compressive strength is validated using the evaluation parameters. The research hypothesis is to develop a sustainable eco-friendly economical paver block using industrial waste materials such as fly ash, GGBS and iron slag.2SiO3 solution. Low calcium Class F type, collected from Tuticorin thermal power station and High calcium GGBS procured from Salem steel plant, is utilized in this study. SEM view of fly ash and GGBS is depicted in Fig. The preliminary resources used to synthesise geopolymer binder in this research work are binder base materials, fly ash, GGBS, and alkaline solution such as NaOH solution and NaSpecific gravity (SG) of fly ash and GGBS is found as 2.3 and 2.92. The chemical compositions of fly ash and GGBS utilized are listed in Table 2SiO3 solution is used as the alkaline solution. Sodium silicate (Na2SiO3) is a commonly used precursor in the synthesis of geopolymer materials. While it is true that some studies have shown that NaOH alone can be used for geopolymerization, Sithole and Mashifana compared the activation of GGBS with KOH and NaOH and claimed the better performance of Sodium based activator than the potassium based activator. Sithole et al. used different concenterations of NaOH and suggested that NaOH is sufficient for the activation of GGBS based geopolymer concrete. But in this study, a blend of flyash and GGBS is performed, hence combination of NaOH and Na2SiO3 is preferred. The use of sodium silicate as a precursor can have several advantages. sodium silicate is a source of both sodium and silica, which are the two main components required for the formation of geopolymer materials. sodium silicate can help to control the setting and hardening of the geopolymer material. The addition of sodium silicate can lead to the formation of a gel-like precursor, which can be important for controlling the reaction kinetics and ensuring the final properties of the material. NaOH flakes and Na2SiO3 solution are brought as such from chemical industries. SG of NaOH and Na2SiO3 is 1.47 and 1.6. Locally available gravel stones of 10 mm size are used as the coarse aggregate (CA), and locally existing granular river sand is utilized as the fine aggregate. SG and fineness modulus of fine aggregate is 2.62 and 2.3 as per IS: 383-197057. River sand confirms to Zone-2. SG and fineness modulus of CA is 2.97 and 6.95 as per IS: 383-1970.A concoction of NaOH solution and Na58. SG of cement is determined to be 3.14. The initial and final setting times are determined as 30 and 513 min.Conventional cement concrete control specimen is made using Ordinary Portland Cement (OPC) of grade53 that confirms IS: 12269-201359. Application of this plasticizer ensures well-dispersed particle suspension and reduces the risk of bleeding and segregation. Admixture is added during the wet state after the addition of about 60% of the alkaline solution.A Superplasticizer of polycarboxylic ether formulation type, MasterGlenium SKY 8233, is utilized in this research work. This is a high-range water reducer and is applied to boost the workability of the GC significantly. SG of admixture is given as 1.08 by the manufacturer, and the properties are in accordance with ASTM C494Natural fibres are preferred over the utilization of synthetic fibres for economically sustainable development. Owning to the high tensile strength and young’s modulus, Banana fibres are utilized in the present investigation with the objective of increasing the energy absorption capacity of the paver block. Banana fibres used are obtained from the pseudo stem of a banana tree locally available. The properties of banana fibres are tabulated in Table 57. Irons slag confirms Zone I grading. The chemical concentrations of the iron slag are listed in Table Iron slag, a byproduct of the pig iron industry obtained from the Kamachi Iron and Steel industry situated at Gummidipoondi, is utilized in this research work. Iron slag is black–grey in colour and is lighter and more brittle than the river sand. Initially, these iron slags were subjected to screening through the 4.75 mm sieve, and all the oversized particles were removed. SG and FM of Iron Slag Waste are determined as 2.48 and 2.4 as per IS: 383-1970The experimental investigation aims at the development of sustainable high-impact strength paver blocks. The extensive methodology involves the optimization of important parameters that affects geopolymerization, such as the molarity of the alkaline solution and fly ash-GGBS blend ratio. The optimum utilization of iron slag as a fractional substitute of river sand as fine aggregate is investigated over the optimized geopolymer mortar samples. Finally, this optimized geopolymer matrix, along with the iron slag, is subjected to the application-oriented study in the form of a paver block with fibres and is compared with the conventional cement paver blocks for its mechanical properties and water absorption capacity.In the first phase, the optimization of the molarity of the NaOH solution that is utilized as the alkaline solution in various proportions, such as 8, 10, 12, and 14 M, is determined. The geopolymer mortar specimens with the optimized molarity are then investigated for the effective utilization of fly ash and slag as alumina silicate source material in various proportions such as 100/0, 90/10, 80/20, 70/30 and 60/40. These optimized Geopolymer specimens are then investigated for the effectual consumption of iron slag as a fractional substitute for river sand in various fractions such as 0, 10, 15, 20, 25, 30, 35, 40 and 45% to determine the effective utilization of iron slag inside geopolymer matrix. Finally, this optimized geopolymer mix is applied as a paver block of dimensions 200 × 100 × 60 using various proportions of natural banana fibres such as 0.5, 1 and 1.5 percent, and its compressive, tensile, flexural and impact strength and water absorption capacity are contrasted with the conventional cement-based paver blocks. Further SEM analysis is carried out over the optimized specimen to understand the microstructure of the matrix.60 in accordance with IS 10262:200961. To achieve high strength Interfacial Transition Zone, a high solution-to-binder ratio of 0.61 is fixed. The ratio of Na2SiO3 to NaOH is set as 2.5. The molarity of the NaOH solution and the combined utilization of fly ash-GGBS slag is optimized in the present study.Geopolymer specimens are designed for M35 grade using B.V. Rangan mix design2-W1/W3-W1. The quantity of the compaction factor is evaluated as follows. If the factor is lower than 0.78, 0.85, 0.92, and 0.95, the fresh state of a material are known as very low quality, low quality, medium quality, and high quality, respectively.An alkaline solution is made ready 24 h before the day of casting, and all the proportioned ingredients are assorted using a pan mixer for about 300 s. Geopolymer concrete is then subjected to a compaction factor test to identify the workability of the matrix. The compaction factor test procedure is as follows; (1) the sample of concrete to be tested shall be placed gently in the upper hopper, using the hand scoop, (2) the hopper shall be filled level with its brim, and the trap door shall be opened so that the concrete falls into the lower hopper, (3) certain mixes have a tendency to stick in one or both of the hoppers. If this occurs, the concrete may be helped by pushing the rod gently into the concrete from the top, (4) immediately after the concrete has come to rest, the cylinder shall be uncovered, the trap door of the lower hopper opened, and the concrete allowed to fall into the cylinder, (5) the excess concrete remaining above the level of the top of the cylinder shall then be cut off by holding a trowel, (6) weight the cylinder with concrete to the nearest 10 g. This weight is known as the weight of partially compacted concrete (W1), (7) empty the cylinder and then refill it with the same concrete mix in layers approximately 5 cm deep, each layer being heavily rammed to obtain full compaction, (8) level the top surface, (9) weigh the cylinder fully compacted. This weight is known as the weight of fully compacted concrete (W2), (10) weight of empty cylinder + hand compacted concrete, which is known as W3, (11) Find the weight of the empty cylinder (W). And it was calculated with WConcrete is then cast into moulds for testing. All the geopolymer specimens are immediately subjected to heat curing at 75 degrees Celsius in a hot air oven for 12 h. The specimens are then kept at ambient temperature, and moulds are removed after 1 day. Mix proportions for the different specimens are listed in Table 62. The findings are listed in Table The feasibility of synthesis GC with optimum molarity, fly ash-GGBS slag blend ratio and iron slag content is investigated by studying the workability of the specimens. The compaction factor of the GC specimens is determined using the Compaction factor test as per IS1199-201363.It is pragmatic from Table 14 and Hassan et al.64 also confirm the same.From the compaction factor results, it is observed that the incorporation of GGBS decreases the workability of GC. The reduction of compaction factor is ascribed to the higher specific gravity and specific surface area, texture and irregular shape of GGBS. Also, the CaO content in GGBS gets liquefied immediately in the process of polymer formation resulting in a stiffer matrix. Research works by Deb et al.54 reported the high water-absorbing capacity of iron slag. Hence the decrease in workability at the increased dosage of iron slag could be attributed to the change in its property caused due to its water-absorbing property. According to British Road Note 4 Standards, from Table From Table 65 using a CTM of 2000kN capacity. The outputs are listed in Table The optimization of primary factors of geopolymerization, such as the molarity of the alkaline solution and fly ash-GGBS blend ratio, is investigated through the assessment of the compressive strength of the GC. The influence of utilization of iron slag as a fractional substitute for river sand is also investigated. CS of the various casted GC samples for the different mix constituents, as listed in Table The relationship between the fly ash-GGBS blend and the molarity of the NaOH solution with the CS is depicted in Fig. 66reported enhancement in CS are because of the enhanced leaching of binder base materials by the activator solution in the flash-based GC.G12 specimens with 12 M gained 86% of the 28 days strength in the first seven days itself due to heat curing.Figure 67.From Fig. Figure 63 studied the outcome of utilizing iron fillings in cement concrete and claimed a raise in compressive strength of about 17% with 30 percent utilization of iron slag. Singh and Siddique54 reported increase of about 20 percent in CS with the employment of iron slag in self- compacting concrete.The augment in strength till the optimum percent is due to the presence of Ca and silicates that reacts with the alumina elements in fly ash and GGBS leading to the enhancement of the compressive strength. Silicates present in the iron slag involve actively in the geopolymerization reaction. AlzaedMolarity of the sodium hydroxide solution as 12 M,80/20 fly ash/GGBS ratio as the binder source material,35% of River sand was partially replaced by iron slag.From the above discussions, the following factors are optimized:GC specimens with the optimized molarity, binder source material and iron slag as the filler material are cast as paver blocks of dimension200 × 100 × 60 mm with different proportions of natural banana fibers such as 0.5, 1 and 1.5 percent, and its compressive, tensile, flexural and impact strength and water absorption capacity are determined and contrasted with the cement-based paver blocks.47, and the values are tabulated in Table The performance of the optimized geopolymer paver blocks with and without the banana fiber with regard to compressive strength are assessed and contrasted with the conventional cement concrete paver blocks in accordance with Annex D of IS. 15658:2006Figure 42 reported a rise in CS through the employment of banana fibres in compressed earthen blocks. In the work, an optimized geopolymer paver block to withstand medium traffic conditions as prescribed by IS 15658:2006, which specified a requirement of 40 (MPa) compressive strength for geotechnical road material.From Fig. 47 and are listed in Table The performance of the optimized geopolymer paver blocks with and without the banana fibre with regard to tensile strength are assessed and contrasted with the conventional cement concrete paver blocks as per Annex F of IS. 15658:2006The flexural strength of the geopolymer paver blocks with and without the presence of banana fiber is determined as per Annex G of IS. 15658:2006 and are tabulated in Table 68 reported rise in flexural strength with the integration of banana fibres inside the cement concrete.Figure 22.The impact resistance of paver blocks with and without the presence of banana fibres in the iron ore slag-replaced specimens is assessed by conducting a drop weight impact test in accordance with the standards of ASTM D 2794. The resistance to impact is calculated by counting the quantity of blows needed to commence the first crack to the final crack by visual observation. The ratio between the corresponding absorbed energy at the failure to the first crack is termed residual impact strength ratioThe residual impact strength ratio of the paver blocks fabricated with the help of optimized banana fibre addition is compared with the results of paver without fibres and conventional specimens as calculated in Table WA Capacity of the paver blocks with and without the presence of banana fibre is determined as per Annex C of IS. 15658:2006 (IS:15658-2006) and are listed in Table 42.From Table This investigation is carried out over the optimum geopolymer paver block sample F1OGI35 using Scanning Electron Microscopy (SEM) and Energy Dispersive Spectroscopy (EDS) and is portrayed in Fig. 66 also confirmed that the increase in GGBS content leads to the formation of stable C–A–S–H or C–S–H gel thereby increasing the compressive strength.Figure 72. The ability of RF to respond near accurately for incomplete and noisy data to create indiscriminate results has made RF to be an extremely useful tool. RF algorithm’s ability to train the model from the instances in nature has made it to be one of the most reliable among the other renowned models73. RF is the most effective algorithm, which uses advanced ensemble techniques in predicting the target through multiple decision trees. The diverse applications of RF in various sectors include banking, health care, stock market, E-commerce and data analytics74. In this work key aim of this work is to build a random forest model to predict the compressive strength of GC for different parameters precisely and validate its performance through performance evaluators.From the above experimental discussions, it is explicit that the expected output behavior compressive strength is solely dependent over the variable parameters discussed above, like molarity of the activator solution, % of GGBS utilization and % of partial substitution of river sand with iron slag. Hence developing a tangible relationship between the dependent variables and the output could save a lot of time, cost of materials and labor in this regard. The advent of machine learning (ML) has subsidized the pressure on the civil engineering domain as well. Prediction of the CS of concrete is proven technique by means of statistical models such as artificial neural network (ANN), K-nearest neighbor (KNN), partial least square regression (PLS), principal component regression (PCR) and random forest (RF)The sample representation for the structure of RF algorithm to get a brief idea is represented in Fig. The over-fitting issues faced by decision trees are addressed by RF by improved accuracy of prediction, effective handling of datasets and development of model without hyper-parameter tuning. The key advantage of RF algorithm is that it employs a collective prediction technique which keeps the error low and deep decision trees.2).In this study, the accuracy of the developed model and the predicted values can be ascertained using the performance evaluation parameters such as mean absolute error (MAE), normalized root mean squared error (NRMSE) and coefficient of determination . The value of R2 ranges between 0 and 1, where the value near 1 represents the close association of datasets and the efficiency of the model. The R2 can be used as an indicator to determine the goodness of fit for the developed model.The coefficient of determination RIn the present study, 75% of the data set is used for training the model, and the other 25% is utilized for testing the precision of the developed model. The efficacy of the developed model can be ascertained from the performance evaluation parameters. Table 2 values are very close 0 percent and 1, which claims the accuracy of the predicted model. Figure From Table From the detailed discussions, it is clear that the developed model for the prediction of CS of the GC for input parameters such as molarity of the alkaline solution, % of GGBS utilization and % of partial substitution of river sand with iron slag.Geopolymer concrete exhibited enhancement in the CS with the increase in the molarity of the NaOH solution. 12 M of NaOH solution yielded the maximum rise in the CS.GC exhibited a raise in CS with the little incorporation of GGBS slag in the fly ash-based GC under heat curing. At an increased dosage of GGBS, a slight decrease in strength was observed. 20% partial substitution of fly ash with GGBS slag yielded maximum compressive strength.The incorporation of iron slag as a fine aggregate was determined to be beneficial in the Geopolymer concrete. 35 percent of the partial substitution of river sand with iron slag yielded the maximum compressive strength.Optimized Geopolymer concrete paver block with one percent banana fibre exhibited about a 5 percent rise in CS, 14.8 percent rise in tensile strength and 11.7 percent rise in flexural strength than the GPC paver block without the fibers.Also, it is noticed that the addition of 1% banana fiber in the ore slag optimized paver blocks enhanced the impact resistance by 22% more than the conventional cement concrete specimen and 15% than the pavers without fiber reinforcement.Geopolymer paver block with fiber content exhibited more water absorption capacity than the Geopolymer paver block without fiber. But this water absorption capacity is found to be less than the cement concrete paver blocks and is within permissible limits as prescribed by IS. 15658:2006.Fiber-reinforced geopolymer paver blocks synthesized in this study were determined to withstand medium traffic conditions as prescribed by IS. 15658:2006.The proposed RF model can precisely predict the twenty-eight days CS of GC for different parameters such as molarity of the activator solution, % of GGBS utilization and % of partial substitution of river sand with iron slag.2 to 1 validate the effectiveness of the developed model in predicting the CS.The least values of NRMSE and nearness of RExtensive research work was performed to produce the high-impact strength paver blocks. Important parameters that affect the geopolymerization process, such as the molarity of sodium hydroxide solution and the fly ash-slag blend ratio, were optimized under heat curing. Utilization of iron slag as a partial substitute for river sand was determined. The final optimized geopolymer concrete is then applied as a paver bock with various proportions of the natural banana fibre and tested for its high impact strength that would sustain medium traffic conditions as per IS 15,658:2006. Findings from the test results and discussions could be abridged as follows.The works pave the path for the production of sustainable, eco-friendly paver blocks, and the hypothesis of the research as specified is achieved. This research work would open up the hefty potential of natural fiber-reinforced Geopolymer concrete as paver blocks to replace conventional cement paver blocks and reduce the strain over the environment by utilizing the indusrial btproducts as well. This research contributes an economical and eco-friendly paver block to society. The technique of utilizing a natural fibre to increase the impact strength and reduce brittleness has been proven effective, and further, its scope may be expanded to various applications.


HOMER1 mRNA content and synapse marker synaptophysin protein content). INS and MET treatments showed a reduction in the content of the mature neuronal marker NeuN that was prevented by INS/MET. Similarly, increases in cell size/area, neurite length/area observed with INS and MET, were prevented with INS/MET. These findings indicate that IR and MET impair neuronal markers through distinct pathways and suggest that MET is ineffective in treating IR‐driven impairments in neurons.Insulin resistance (IR) is associated with reductions in neuronal proteins often observed with Alzheimer's disease (AD), however, the mechanisms through which IR promotes neurodegeneration/AD pathogenesis are poorly understood. Metformin (MET), a potent activator of the metabolic regulator AMPK is used to treat IR but its effectiveness for AD is unclear. We have previously shown that chronic AMPK activation impairs neurite growth and protein synthesis in SH‐SY5Y neurons, however, AMPK activation in IR was not explored. Therefore, we examined the effects of MET‐driven AMPK activation with and without IR. Retinoic acid‐differentiated SH‐SY5Y neurons were treated with: (1) Ctl: 24 h vehicle followed by 24 h Vehicle; (2) HI: 100 nM insulin (24 h HI followed by 24 h HI); or (3) MET: 24 h vehicle followed by 24 h 2 mM metformin; (4) HI/MET: 24 h 100 nM insulin followed by 24 h 100 nM INS+2 mM MET. INS and INS/MET groups saw impairments in markers of insulin signaling  demonstrating IR was not recovered with MET treatment. All treatment groups showed reductions in neuronal markers (post‐synaptic marker   AD pathology is traditionally characterized by increased amyloid‐β‐plaque development   is the most prescribed T2D drug and effectively treats peripheral metabolic dysregulation ; (2) High‐insulin ; (3) Metformin ; (4) High‐insulin + Metformin (100 nM HI+2 mM Met). To induce IR, cells were incubated with 100 nM insulin (INS) for 24 h following 5 days of differentiation with RA. This method of high‐insulin‐induced IR has previously been demonstrated in SH‐SY5Y cells and other cell types . Use of this cell line was approved by the Research Ethics Board at Brock University (#17–397). Cells were cultured using high glucose Dulbecco's Modified Eagles Media (DMEM Sigma D6429), containing 10%, 1%, 0.1% (v/v) FBS (Sigma F1051) for seeding, differentiation, and treatments respectively, as well as 5% non‐essential amino acid (Sigma M7145), and 1% Penicillin/Streptomycin (Sigma P4333). Unless otherwise stated, cells were seeded at 150,000 cells/mL on 12‐well culture plates (VWR 10062–894) in triplicate. and cultured until ~90%–95% confluency before differentiation. Differentiation was accomplished with retinoic acid  for 5 days. Differentiated cells were then assigned to one of the following 4 treatments: (1) Control media (H2.22O) or 100 nM INS overnight, after which time fresh 0.1% FBS media containing 100 nM INS was added acutely for 30 min. Cells were then collected and analyzed by WB to assess Akt T308 and S473 phosphorylation status, as well as mTOR subunit S2448 phosphorylation status. Impaired insulin signaling was indicated by a decreased ability of the acutely added INS to increase Akt and mTOR phosphorylation.To examine the effectiveness of the high‐insulin‐induced IR, an acute insulin stimulation experiment was performed. This assessment of insulin responsiveness was adapted from and conducted as previously described  supplemented with 34 μL phenylmethylsulfonyl fluoride and 50 μL protease inhibitor cocktail . Cell lysates were then sonicated . A bicinchoninic acid assay was performed to determine the protein concentration of the lysates. Samples were prepared (1 μg/μL) and equal amounts of protein were then electrophoretically separated on 10% SDS‐PAGE gels and transferred to nitrocellulose membranes . Membranes were blocked for 90 min at room temperature in 5% non‐fat dry milk‐TBST . Membranes were then incubated in primary antibody diluted 1:1000 in 5% BSA (Bovine Serum Albumin)‐TBST overnight at 4°C with gentle agitation. The following day, membranes were incubated for 1 h at room temperature with the appropriate secondary antibodies , #711–035‐152, Goat anti‐mouse IgG (H+L), #115–035‐003 Jackson Immunoresearch) in 1% BSA‐TBST. Membranes were rinsed 3× 5 min in TBST and proteins were visualized by western Lightning Plus‐ECL (Perkinelmer NEL103E001EA) using a ChemiDoc Imaging System (Bio‐Rad).α . Total and phosphorylated AMPK levels were measured using specific primary antibodies  and HOMER1 (Thermo Fischer Hs00188676_m1) as well as GAPDH (thermos Fischer Hs02786624_m1), which was used as a reference gene. Relative differences in mRNA expression were determined using the 2−∆∆CT method . Cells were then fixed using 4% paraformaldehyde and permeabilized using 20% triton X‐100 before being incubated in a buffer containing phalloidin actin stain  where one N represents 3 pooled replicates. Pooled replicates were taken from identical treatments from within the same plate.Differences in protein content/phosphorylation status, cell size, neurite length, and fluorescent levels were determined using a one‐way ANOVA followed by a Fischer's LSD 33.1t‐test, p = 0.3415). Acute insulin responsiveness was then assessed by exposure to fresh media with or without 100 nM insulin for 30 min, followed by WB analysis of the insulin signaling markers, Akt T308 and S473 phosphorylation. Impairment of increases in the phosphorylation of both Akt phosphorylation sites are commonly used markers indicative of impaired insulin signaling. Two‐way ANOVAs indicated an interaction between phosphorylation status and pre‐exposure to insulin for both Akt T308  and Akt S473 . Phosphorylation status of Akt T308 and S473 was increased with a 30‐min insulin stimulation in control cells treated with 0.1% media alone, but this increase was prevented following a 24 h pre‐treatment with 100 nM insulin  . This dose has been used previously by others to disrupt insulin signaling  Figure , indicatp = 0.9382) and both doses showed equivalent increases in AMPK T172 phosphorylation status . As such, the 2 mM dose was selected for use in subsequent experiments.To establish the effective dose of MET, we exposed cells to either 2 mM or 5 mM MET, doses used in previous studies  compared to both control and HI.We next investigated whether HI treatment would impact AMPK phosphorylation and whether MET would still be effective at increased AMPK phosphorylation. Cells were incubated with 100 nM insulin for 24 h followed by 24 h 100 nM insulin (HI) or 100 nM insulin for 24 h followed by both 100 nM insulin and MET (HI/MET). AMPK T172 phosphorylation status was higher following HI/MET treatment  and mTOR S2448 (one‐way ANOVA p = 0.0062) phosphorylation status was higher in both HI  and HI/MET  groups compared to Ctl and MET treatments . IRS‐1 S636 phosphorylation status was next investigated, as IRS‐1 S636 is a target of mTORC1 through the activation of p70s6k  following HI treatment and was higher following HI/MET treatment relative to ctl (p = 0.0039) and MET (p = 0.0017) groups , no significant increase in IRS‐1 S636 phosphorylation was seen in the HI or HI/MET groups relative to control ; only HI was seen to increase relative to MET (post hocHI‐METp = 0.0099). Total IRS‐1 content, however, was found to be higher in the MET group relative to all other groups . Further, insulin receptor‐α (IRα) and insulin receptor‐β (IRβ) were examined as both IRα and IRβ constitute the two major subunits of the insulin receptor that bind insulin to induce downstream insulin action, and reductions in their expression have been associated with the development of IR. IRα receptor levels were higher with MET treatment compared to Ctl  ).The ratio of IRS‐1 S636/IRS‐1 approached significance  Figure  in a sim3.3mTORC1, a major marker of cell growth and proliferation, is a target of both AMPK and Akt, which inhibit or induce mTORC1‐driven protein synthesis/cell growth, respectively. As our previous work showed that AMPK activation and downstream signaling through mTORC1 plays a role in synaptic protein expression of retinoic acid‐differentiated SH‐SY5Y cells . This increase was also observed in ULK S555, the direct downstream target of AMPK when compared to both Ctl and HI groups . Despite the HI/MET treatment not showing higher AMPK T172 phosphorylation (post = hocctl‐HI/METp = 0.0924), HI/MET treatment did show higher ULK S555 phosphorylation , suggesting that treatment of INS with MET induces the activation of ULK. This would be despite AMPK T172 phosphorylation being no different to control , with no change in p70s6k phosphorylation  or postsynaptic  protein content  following any treatment. However, HOMER1 mRNA content was significantly altered  and was reduced in all treatment groups relative to control  indicating that MET activation of AMPK alone, seems to lower HOMER1 mRNA content more than HI treatment.As no changes in protein content were seen, we next investigated whether changes in mRNA content occurred. As the expression of both PSD‐95 and Homer‐1 are paramount for post‐synaptic density and synaptic function, we investigated whether the mRNA expression of these markers was affected by our treatments. We found no change in l Figure . Additiop = 0.0069, post hocctl‐HIp = 0.0036, post hocctl‐METp = 0.0267, post hocHI‐HI/METp = 0.0052, post hocMET‐HI/METp = 0.0341), with reductions in NeuN in both HI and MET groups, compared to Ctl and HI/MET treatments  and were lower in all treatment groups compared to control, with HI/MET treatment showing greater reductions in protein content compared to MET alone , changes in protein content of the whole neuron might not fully reflect adjustments in the synapses nor overall changes in cell function. As such, changes in neuron morphology were analyzed to provide additional information about neuronal and/or synaptic function. The focus of this morphological analysis was directed to changes in soma size , as well as neurites , as they represent the two major regions that define neurons  and total cell area  as well as greater neurite area  and neurite length , compared to controls, however, these increases in cell size were absent with the combined HI/MET treatment . Larger neurite and cell size are commonly associated with greater cell growth and protein synthesis in support of synaptic transmission , the downstream activation of PI3K, increasing the phosphorylation and activation of Akt and leading to eventual mTORC1 activation Figure  Left. ImAlongside changes in synaptic protein content, an important aspect of examining synapses involves quantification of changes in neuronal morphology. Reductions in synapse number and neurite length/size have commonly been reported to occur within AD  , such as protein content ablations in total IRS‐1 and IRβ, and NF‐kB, plays a role in the impairments observed in this study, remains to be explored. Similarly, while some previous evidence does suggest a role for increased neurite size and length under conditions of stress in maintaining effective synaptic transmission despite a loss of synaptic number .The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.This study was conducted in accordance with Brock University's ethical standards and Tri‐Council Policy Statement and has been approved by the Health Science Research Ethics Board of Brock University .


For acute and chronic soft tissue infections, radical surgical debridement is required and is considered the gold standard, along with its immediate systemic antibiotic therapy. Treatment with local antibiotics and/or antibiotic-containing materials is commonly used as an additional tool in clinical practice. Spraying with fibrin and antibiotics is a newer technique that has been studied for some antibiotics. However, for gentamicin, data are not yet available on absorption, optimal application, antibiotic fate at the site and transfer of antibiotic into the blood. In an animal study involving 29 Sprague Dawley rats, 116 back wounds were sprayed with gentamicin using either gentamicin alone or one of two possible spray combinations of gentamicin and fibrin. Simultaneous application of gentamicin and fibrin via a spray system to soft tissue wounds resulted in significant antibiotic concentration over a long period of time. The technique is easy and cost-effective. The systemic crossover was significantly minimized in our study, which may have led to fewer side effects in patients. These results could lead to an improvement in local antibiotic therapy. Staphylococcus aureus. Infections with Streptococcus pyogenes or Pseudomonas—aeruginosa also occur frequently. Other pathogens for typical wound infections are Escherichia coli, Enterococcus, Proteus and Klebsiella [stapylococci, Pseudomonas—aeruginosa and E. coli has been described [The current gold standard treatment for acute and chronic soft tissue infections involves radical surgical wound debridement , accompaebsiella ,15. Gentescribed ,17. It wescribed . The spoescribed ,19.The question of stable retention at the application site, duration of retention and tissue concentration of antibiotics remains in most cases unanswered ,21. ManyIn 2016, our group presented a new approach to fixate antibiotics in a wound ,23,24. IFibrin spraying is a common method of hemostasizing surfaces, such as the liver and maxillary sinuses ,26. HoweSeveral clinical cases at our institution have documented the good clinical effects of fibrin-fixed antibiotics , but in In order to answer the questions of resorption, the best possible type of application, retention of antibiotic on site and transfer of antibiotic into blood, an animal model was used in this study. We measured the concentration of the antibiotic gentamicin in the soft tissue and blood of the animals to investigate the best type of antibiotic application and its fixation.n = 12 per group. Two animals died during the experiment, so 29 animals were used. Animals that were 8 weeks old were used in the experiment. The cages were housed in constantly ventilated, temperature—(15–21 °C) and light-controlled (14 h day: 10 h night) rooms of the Central Research Institute (ZFE) of the Goethe University Frankfurt am Main (Germany).All animal experiments were performed in strict compliance with the applicable standards for animal experiments in Germany, and according to the official approval by the Darmstadt Regional Council (project number FK/1133). A total of 29 male rats of the strain Sprague Dawley , weighing 335–385 g, were kept in cages with three to four animals each and received food and water ad libitum. For the experiment in 9 groups, 27 animals were required to arrive at Gentamicin sulfate 40 mg/mL  was dissolved in 9 mL of 0.9% isotonic NaCl solution , creating a concentration of 4 mg/mL. A total of 10 mg was applied per wound.2 each were placed under aseptic conditions with a scalpel and scissors to the muscle/fascia level. The fascia was roughened with the scalpel. Then, the wounds were sprayed according to the different groups  and Xylavet 20 mg/mL  at weight-adjusted doses with an injection volume of 1 to 2 mL. The back of the anesthetized rat was shaved and disinfected with Octeniderm . After sterile covering of the surface with a sterile perforated cloth, four wounds of 100 mmt groups .After surgery, animals were placed in single cages and killed depending on the group after 1, 2, and 4 h by an overdose of pentobarbital 500 mg/kg i.p. and the creation of bilateral pneumothoraces. Tissue samples were collected from the wound surfaces. Blood was transferred to EDTA tubes and tissue samples were transferred to 1 mL Eppendorf cups and stored on dry ice at −80 °C in the laboratory until further preparation and measurement.Gentamicin was sprayed without fixation (G), sequentially with gentamicin and fibrin (GF−), and simultaneously with gentamicin and fibrin (GF+) . Prefillg), 100 µL of supernatant was used for LC-MS/MS analysis. For the analysis of gentamicin, 20 µL of an internal standard  and 300 µL acetonitrile with 1% formic acid were added to 100 µL of homogenate. After vortexing and centrifugation  and the corresponding standard .® HILIC column  was utilized with a flow of 0.35 mL/min and a column temperature of 40 °C. As solvents, 1% formic acid (solvent A) and acetonitrile with 1% formic acid (solvent B) were used with the following gradients : 0.0 min, 90%; 1.0 min, 90%; 2.0 min, 75%; 2.5 min, 20%; 7.0 min, 20%; 7.3 min, 90%; 11.0 min, 90%. A total of 0.5 µL of each sample was injected.The LC-MS/MS analysis was carried out using an Agilent 1290 Infinity LC-system with a binary HPLC pump, column oven and autosampler , coupled with a triple quadrupole mass spectrometer QTRAP 5500 . A LunaData acquisition was performed using Analyst Software 1.7.1, and quantification was performed using MultiQuant Software 3.0.3 . Curves were calculated by using linear regression with a weighting factor of 1/x and calibration.p < 0.05 (Bonferroni-Holm corrected) was considered significant. BiAS for Windows version 11.12 dated 01/2021 was used .The primary study variable was antibiotic concentration after 1, 2, and 4 h. Each group consisted of 12 samples. For post hoc analysis, a Kruskal–Wallis test with multiple Conover–Iman comparisons was performed to identify the significant features of each group’s results. An effect size of Considering the results of the tissue concentration of gentamicin in ng per mg of tissue in the different groups after one hour, uptake of the antibiotic into the tissue was evident on all sides. Of the 10 mg of gentamicin applied, a mean of 60.688 ng/mg (standard deviation 5.733) was detected after 1 h in the group treated with gentamicin (G) alone. The group with sequentially applied gentamicin and fibrin (GF−) 55.680 ng/mg (standard deviation 15.776) and the group with simultaneously applied gentamicin and fibrin (GF+) 67.629 ng/mg (standard deviation 15.061) showed similar values. No significant differences were found within the 1 h group A. p < 0.05 vs. GF− and GF+). The group that received consecutive spraying of gentamicin and fibrin (GF−)  and simultaneous spraying of gentamicin and fibrin (GF+)  showed significantly higher values   showed the highest values of the antibiotic that still remained in the tissue . Lower values were found in the group that received consecutive application of gentamicin and fibrin (GF−) , but these were not statistically significantly lower than those in the GF+ group. However, the tissue levels were significantly lower in the group in which only gentamicin was administered (G)  C.p < 0.05 vs. G, GF−, GF+ 4 h). In the 2 h group, tissue concentration was significantly lower than in the 1 h group only in the gentamicin group without fixation (G) . The decrease in tissue concentration over time is clearly visible in A comparison of the three groups over the three time points is shown in p < 0.05) . However, significant differences were observed after 4 h. In the simultaneously sprayed group (GF+) , significantly lower levels of gentamicin or its metabolites were found in the blood of the animals than in groups G  and GF−  ( < 0.05) .With the in vivo experiments shown here, we were able to demonstrate a new technique for antibiotic fixation in wounds for gentamicin, as we did previously for vancomycin. Its application as a spray and fixation with fibrin benefits both the widespread distribution of the antibiotic substance and its ease of application. The antibiotic can be sprayed specifically on a particular area or even on the entire wound site. Even irregular surfaces and soft tissue-bone interfaces are no longer a problem due to the finely misted substances. This concept has already been clinically tested due to its very simple application. In the case of gentamicin, however, the type of application that results in the longest retention of the antibiotic in tissues has never been clearly investigated and established.S. aureus to stabilized fibrin matrices, as used in our trial, promoted a local, macrophage-mediated antimicrobial response [We examined the efficacy of spraying gentamicin alone, spraying gentamicin and fibrin sequentially, and simultaneously spraying gentamicin and fibrin to determine which form of application leads to the highest tissue concentration. The data revealed that there are significant differences depending on the mode of application. Spraying with an antibiotic alone or clinical irrigation with an antibiotic solution without any additional fixation had an insufficient effect. Furthermore, it was shown that the binding of response . It was response ,24, thatresponse .The use of pellets, foams and gels as antibiotic carriers is widespread in clinical practice ,29. The Antibiotic spraying allows a large area of infected tissue to be covered, and the antibiotic to be applied in a targeted and uniform manner. A clinically proven nebulizer for spraying fibrin glue is used to spray the antibiotic ,33. The With all the application techniques tested in this study, gentamicin sprayed into a wound showed a significant concentration in the wound area after one hour. As a result of the well-perfused tissue in the wound area, the concentration of antibiotics in the wound decreases rapidly B–D. WhenConsidering the detection of gentamicin and its metabolic components in the blood of the animals, no metabolites or gentamicin could be systematically detected in the 1 h and 2 h groups. In the 4 h group only, animals receiving gentamicin without fibrin fixation had systemic gentamicin levels twice as high as those receiving gentamicin and fibrin simultaneously.Therefore, when gentamicin and fibrin are applied simultaneously, the antibiotic substance will remain in the tissue for a longer period and a smaller systemic effect will be produced.An antibiotic drug fixation technique, such as the one shown here, should not be the only method of treating a local infection. The treatment concept involves comprehensive surgical debridement, nebulizing an antibiotic simultaneously with fibrin glue and next-look surgery, if necessary. It is important to note that intravenous antibiotics are administered as usual ,35. In addition to treating soft tissue infections, this spray technique can also be used to treat early postoperative soft tissue and bone infections ,36,37. TIn this study, we demonstrated the possibility of antibiotic fibrin spraying in an animal wound model. In an infected wound animal model, however, this procedure has not yet been tested, and comparative studies on other delivery methods are still lacking, but the clinical cases already performed showed good clinical results .By spraying gentamicin and fibrin onto soft tissue wounds simultaneously, a high level of antibiotic concentration is achieved over a long period of time. A wide range of indications in surgery and wound therapy can be treated with this simple and inexpensive technique. Systemic crossover is minimized, which can lead to fewer side effects in patients. However, further studies are needed, especially for clinical applications in vivo, to demonstrate the therapeutic potential of simultaneously sprayed gentamicin and fibrin.


BackgroundAcute abdomen is often complicated by intra-abdominal infection requiring antibiotic therapy. Danish regional antibiotic guidelines emphasize the restricted use of broad-spectrum antibiotics such as cephalosporins. In this study, we aimed to evaluate antibiotic practices in relation to hospitalized patients with acute abdomen.MethodologyThis retrospective quality assurance study was conducted among patients admitted to the surgical emergency department at the North Denmark Regional Hospital during a four-month observation period. Data were extracted from electronic patient journals and entered in the Research Electronic Data Capture data management system for further analytical work.ResultsOf 331 patients, 174 (53%) were treated with antibiotics, of whom 98 (56%) had been treated with cephalosporin, 47 (27%) with benzylpenicillin and gentamicin, 22 (13%) with piperacillin/tazobactam, and seven (4%) with ciprofloxacin. Use of a cephalosporin-based antibiotic regimen was significantly more common in patients with acute appendicitis (75%) compared to other diagnostic groups, such as acute cholecystitis (57%), incarcerated hernia with strangulation (56%), acute pancreatitis (50%), and acute diverticulitis (30%). However, patients with uncomplicated diverticulitis (53%) were significantly more often treated with benzylpenicillin and gentamicin, whereas patients with complicated diverticulitis Hinchey stage 3-4 were significantly more often treated with piperacillin/tazobactam. In addition, as the severity of acute cholecystitis increased, it was more frequently treated with piperacillin/tazobactam.ConclusionsThe study revealed that cephalosporins are frequently used in patients hospitalized with acute abdomen. This finding conflicts with current regional antibiotic guidelines. Reinforcement of the guidelines is required as an essential measure against the development of antibiotic resistance associated with the use of cephalosporins. Acute abdomen is frequently associated with intra-abdominal infections (IAIs), such as acute appendicitis, diverticulitis, and cholecystitis, but may also be caused by other inflammatory conditions and non-specific abdominal pain in general. IAI is a broad term used for conditions in which inflammation of the peritoneum is observed due to local contamination with microorganisms, often followed by exudation in the abdominal cavity and a systemic inflammatory response .Clostridium difficile infections related to the use of broad-spectrum agents, in particular cephalosporins [Timely diagnosis and management of IAI by use of antibiotics and surgery as the key elements of intervention is an essential strategy ,3. Indicosporins ,5, are tosporins ,7.However, the use of antibiotics in the hospital sector in Denmark has increased by 25% in the last decade , which rTherefore, this study aimed to assess antibiotic practice with reference to the existing guidelines for patients with acute abdomen admitted to the surgical emergency department in a Danish regional hospital.Study design and populationThis quality assurance study was conducted at the North Denmark Regional Hospital to evaluate antibiotic therapy practice in the surgical emergency department. The target population comprised patients with acute abdomen admitted to the department during a four-month observation period. Patients were excluded from the study if they had received antibiotics within the previous four weeks before admission. In addition, patients diagnosed with gastrointestinal hemorrhage, trauma, small abscesses, and other medical diseases were also excluded.Data collection and managementThe Research Electronic Data Capture data management system was used in this study . DemograDiagnostic categories of intra-abdominal infectionsGroups for discharge diagnoses, including acute appendicitis, gallstones (with and without cholecystitis), diverticulitis, pancreatitis, incarcerated hernia, and non-specific abdominal pain, were created according to surgical findings together with radiological and pathological results. Moreover, subgroups within these groups were determined based on existing classification criteria. Acute appendicitis cases were categorized as either phlegmonous, gangrenous, or perforation/abscess based on the pathological examinations of resected specimens . DivertiAntibioticsData on the antibiotic used in the perioperative periods were collected. Three principal antibiotic regimens were used according to the North Denmark regional antibiotic guidelines for patients admitted to the surgical emergency department. These included benzylpenicillin and gentamicin, cephalosporin , and piperacillin/tazobactam. Moreover, quinolones  were occasionally used. The listed antibiotics were often used in combination with metronidazole. Overall, antibiotic treatments were given only during their hospitalization.Study registrationThe study was registered in the North Denmark Region clinical research registry. Approval by the North Denmark Region Committee on Health Research Ethics was not deemed necessary as it is a clinical quality assurance study.Statistical analysisNominal and ordinal parameters were described with frequency analysis. Means and standard deviations were used to describe scale parameters. Pearson’s chi-square and Fisher’s exact tests were used for differences between nominal and ordinal parameters. The Kolmogorov-Smirnov test was used for normality tests of scale parameters. The Mann-Whitney U and Kruskal-Wallis tests were used for differences between non-normally distributed scale parameters, whereas the independent-samples t-test was used for normally distributed scale parameters. Spearman’s rho correlation analysis was used for correlations between non-parametric variables. SPSS for Windows version 25.0  was used with a 95% confidence interval and 0.05 significance level.In total 331 patients with acute abdomen were included in this study. The mean age was 51 (SD = ±21) years and within the range of 8 to 95 years. Overall, 43% of the patients were males. The mean BMI of all patients was 27.4 (SD = ±6.1). The discharge diagnoses were as follows: non-specific abdominal pain , acute appendicitis , acute diverticulitis , acute cholecystitis , incarcerated hernia , pancreatitis , and other diagnoses , including malign colon, sigmoid volvulus, small bowel ileus, perforated ulcer, and ischemic bowel (Table Surgery was performed on 121 (37%) patients. Antibiotic treatment was given to 174 (53%) patients. The mean age of the antibiotic recipients was 54 (SD = ±20), which was significantly higher than the 46 years (SD = ±21) of the non-antibiotic recipients (p < 0.05) , cephalosporins , piperacillin/tazobactam , and quinolone   Table . In the Of the 23 patients with an incarcerated hernia, 30% did not receive antibiotic treatment, although it is recommended to initiate antibiotics in mesh repair hernia Table . MoreoveIn this study, cephalosporin was found to be the most prescribed antibiotic with an overall rate of 56%, as opposed to 27% for benzylpenicillin and gentamicin, although the latter regimen is the first-line antibiotic therapy in accordance with the regional antibiotic guidelines, especially in patients with acute appendicitis, acute diverticulitis, and acute cholecystitis. Preferably, cephalosporin should mainly be used in patients with penicillin allergy. In our study population, two patients only treated with cephalosporin had been registered with penicillin allergy.Antibiotic treatment is generally recommended for the prevention of postoperative infectious complications . In aligAccording to the Tokyo Guidelines 2018, in patients with moderate-to-severe acute cholecystitis, antibiotic therapy should be initiated in conjunction with cholecystectomy, whereas mild acute cholecystitis should not be treated with antibiotics . In our It is suggested that patients with incarcerated hernia operated on with a mesh repair should receive antibiotic treatment . Among pAntibiotic treatment is not recommended in patients with acute mild pancreatitis . About 4The use of cephalosporins is associated with an increased risk of antibiotic resistance development, which is a growing public health concern worldwide. Unjustified use of antibiotics and ineffective infection control policies are major causes of the spread of antibiotic resistance . In accoC. difficile infection is a common hospital-acquired infection and is an increasingly frequent cause of morbidity and mortality among the elderly and fragile hospitalized patients receiving broad-spectrum antibiotics, such as cephalosporins [C. difficile infection has been reported in 2.6% of surgical patients treated with cephalosporins. C. difficile emerges in the intestinal tract when the normal gut flora has been disrupted in conjunction with antibiotic therapy, in particular when using broad-spectrum agents, and is the causative organism of antibiotic-associated colitis including pseudomembranous colitis [osporins . The dev colitis ,25.The study has revealed a considerable lack of compliance with the regional antibiotic treatment guidelines among clinicians prescribing antibiotics to patients. There seems to exist a preference toward second or third-generation cephalosporins over the benzylpenicillin and gentamicin antibiotic regimen, which is recommended as the first choice in the regional antibiotic guidelines. A plausible reason for this lack of compliance could be that clinicians are skeptical or cautious about the use of the benzylpenicillin and gentamicin antibiotic regimen due to concern about the risk of nephrotoxicity and ototoxicity associated with gentamicin administration and the risk of allergic reactions to penicillin. Another reason for the preference for cephalosporins could be that international antibiotic recommendations generally opt for this antibiotic because penicillin is not an option in many parts of the world due to widespread resistance. However, penicillin is still a solid option in Denmark as a result of many years of restrictive antibiotic treatment policy, thereby having prevented penicillin resistance in a successful manner. Moreover, the potential risk of nephrotoxicity and ototoxicity associated with gentamicin administration is carefully addressed in the regional antibiotic guidelines, e.g., by routine control of serum creatinine and plasma gentamicin concentrations.There are some major limitations of this study. First, this was a single-center quality assurance study, which challenges the extent to which the findings can be generalized to other hospitals in Denmark. Second, the fact that it was a retrospective study limited the measurement and inclusion of other diagnoses that could be associated with the decisions for or against antibiotic therapy, or the use of different antibiotics. Finally, low patient numbers in certain diagnostic subgroups limited the potential outcomes of the statistical analyses.C. difficile infection, in conjunction with the uncritical use of cephalosporins.Our findings suggest a critically common use of cephalosporin in patients admitted to the hospital with acute abdomen. Hence, there is an urgent need for reinforcement of continuing medical training programs with a focus on an antibiotic stewardship strategy to increase adherence to the regional antibiotic guidelines. Such a strategy in Denmark aims to reduce the risk of antibiotic resistance development and adverse outcomes, such as


There is minimal data regarding optimal duration of therapy in gram-negative bloodstream infections (GN-BSI) with obstructing nephrolithiasis/urolithiasis. Prescribing practices often include standard antibiotic duration (fixed duration with an antibiotic free period prior to stone management) or extended duration (continuous antibiotics until definitive stone management). This study compares outcomes between standard versus extended antibiotic durations in patients with GN-BSI with obstructing nephrolithiasis/urolithiasis.This IRB approved retrospective cohort study screened patients treated between 1/1/2016 and 1/1/2022. The standard duration group had an antibiotic free period of ≥ 7 days before definitive stone management and the extended duration group had continuous antibiotics without an antibiotic free period ≥ 7 days before definitive stone management. The primary outcome was infection recurrence before definitive stone management. Select secondary outcomes included revisit to care, antimicrobial resistance, and mortality. Continuous data utilized a t-test or Mann-Whitney U test and categorical data utilized either Chi-square or Fisher’s exact test.After screening 405 encounters, there were 39 standard and 29 extended group patients. Median antibiotic duration was 16  vs. 19  days, p = 0.06 and time to definitive stone management was 52  vs. 22  days, p< 0.01 in the standard and extended group respectively. Infection recurrence was no different in the two groups, 5% (n=2) vs. 0%, p=0.5. Revisit to care was significantly higher in the standard group 28% (n=11) vs. 3% (n=1), p< 0.01, with the most common patient complaint of “concern for infection.”This study found no difference in infection recurrence between standard and extended antibiotic treatment strategies but was underpowered. A higher rate of revisit to care in patients who received standard courses of antibiotics is concerning and may also be influenced by a prolonged time to definitive stone management. Further data is necessary to determine the optimal antibiotic treatment strategy and optimal time of definitive stone management after GN-BSI with obstructing stones.All Authors: No reported disclosures


MET exon 14 skipping is one of the rare mutations in non-small cell lung cancer (NSCLC), involving its pathogenesis and progression. The performances of several MET inhibitors in clinical trials have been validated based on NGS, immunohistochemistry (IHC), and gene copy number assessments. Thus, a detailed understanding of the relationship between these markers and prognosis is required.This study has recruited patients (n = 17) with MET exon 14 skipping mutation and initially screened genes (n = 10) by polymerase chain reaction (PCR) from 257 specimens of NSCLC, including small biopsies and surgical resection. Further, the IHC analysis detected MET overexpression and recorded the score using the MetMAb trial  with MET exstainings). Finally, the fluorescence in situ hybridization (FISH) resulted in the MET amplification with a MET copy number initially screened genes (n = 10) by p.PCR results indicated strong MET staining ( 3+) in more than 50% of tumor cells. Among the recruited 17 cases of MET exon 14 skipping, 9 cases presented MET amplification, and 10 cases with MET overexpression. These attributes were not correlated to the clinicopathological characteristics and overall survival. In addition, 4 cases showed gene amplification, and 3 cases presented polyploidy condition. The correlation analysis showed a significant relationship between MET amplification and MET overexpression .Together, these findings indicated a significant correlation between MET overexpression and MET amplification in NSCLC patients but no correlation to prognosis.The online version contains supplementary material available at 10.1186/s12890-023-02482-9. Indeed, the MET proto-oncogene located in the 7q31 locus of chromosome 7 encodes the receptor tyrosine kinase (RTK) enzyme. The binding to a ligand, i.e., hepatocyte growth factor (HGF), often induces its downstream signaling. The regulated expression of the MET proto-oncogene is often related to embryogenesis, wound healing, liver regeneration, angiogenesis, and immunomodulation in diverse physiological processes. Nevertheless, the aberrant MET signaling drives the cells toward tumor development through uncontrolled cell proliferation, survival, invasion, and metastasis, as well as angiogenesis . Due to Over the past decade, several reports indicated the development of various therapeutic strategies addressing the MET exon 14 skipping. In a case, it was reported approximately 5% of NSCLC cases upon progression to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) and approximately 10% of patients treated with osimertinib (a mutant-selective EGFR TKI) for EGFR T790M-positive NSCLC. Nonetheless, some patients resulted in genetic alteration in the MET receptor, leading to amplified levels of MET proteins and/or the MET gene and resulting in the catastrophe of osimertinib actions. In another case, MET amplification exhibited resistance to ALK inhibitors in the ALK-rearranged NSCLC. In addition, some studies based on MET expression failed to show their potential in HGF/MET targeted therapies. Despite the detection of overexpression, the predominant reason could be the lack of standardization of methodologies, specifically in the development of the scoring systems applied in immunohistochemistry (IHC) determinations or the absence of oncogenic addiction of cancer cells to the MET pathway. Nevertheless, determining the amplification and mutation aberrations often suffer from these pitfalls . SavannaInspired by these aspects, in this study, we have recruited NSCLC patients (n = 17) with MET exon 14 skipping mutation in our hospital to conduct relevant clinicopathological analysis. In addition, gene amplification and MET protein overexpression in cases with MET exon 14 skipping mutation were tested to substantially explore the relationship and provide ideas for clinical diagnosis and treatment.In this study, the patient recruitment was performed following the criteria, i.e., the patient with non-small cell lung carcinoma (NSCLC) histology and availability of sufficient cancerous tissues for the extensive studies. Following the criteria, 17 patients with MET exon 14 skipping mutation tumor specimens were included from the total of 257 patients diagnosed with NSCLC at our hospital between 2019 and 2022. Further, IHC and fluorescence in situ hybridization (FISH) validations of C-MET were performed to detect the correlation of MET exon 14 skipping mutations in the recruited cases. Then, the collected tissues, either from surgical resections or core-needle biopsies from the recruited subjects, were used for further analyses. To establish a correlation, the clinical data were extracted from the medical records, including age, sex, tumor disease stage, and clinical follow-up information, among others.Initially, the IHC evaluations were performed using anti-total c-MET  rabbit monoclonal antibody as a primary antibody and revealed using an ultraView Universal DAB Detection Kit . The IHC staining procedure was carried out according to the manufacturer’s instructions using the Discovery ULTRA platform (Ventana BenchMark Ultra). Briefly, the primary antibody was first incubated for 16 min, and the antigen treatment was performed using the repair solution , which was presented in more than 50% of tumor cells. In contrast, some cases were categorized as low MET as they showed no such criteria. The MET staining pattern, i.e., predominantly membranous vs. cytoplasmic, was assessed as described elsewhere. This systematic pattern was applied to evaluate both proportion and staining intensity: 0 referred to as no staining or < 50% tumor cells with any intensity; 1 + denoted as ≥ 50% of tumor cells staining with weak or higher staining but < 50% with moderate or higher strong intensity; 2 + indicated as ≥ 50% of tumor cells with moderate or higher staining but < 50% with strong intensity; and 3 + referred to as ≥ 50% of tumor cells staining with strong intensity. Accordingly, the samples that exhibited 2 + or 3 + immunostaining were considered positive for MET overexpression.The FISH evaluation of MET was performed using the unstained, formalin-fixed, and paraffin-embedded (FFPE) tumor tissue sections, as previously described. The FISH analysis was performed using a MET/CEP7 7q31) probe cocktail  using the FISH method. The mean cMET/CEN7 ratio was calculated for each probe by scoring the number of c-MET and CEN7 each nucleus independently. The attributes of MET copy number ≥ 5 and/or MET:CEP7 signal ratio ≥ 2 were denoted as amplification. In contrast, the MET:CEP7 ratio of < 2 was classified as MET non-amplification. While patients with MET copy numbers of ≥ 5 but a ratio of < 2 were classified as MET gain, and both MET and CEP7 copy numbers of ≥ 3 but < 5 were classified as MET polysomy.Statistical analyses were performed using GraphPad Prism 9.0. Comparing categorical variables was done using the Fisher’s exact test. Using either the Mann-Whitney U test, continuous variables expressing mean ± SD were compared. The correlation of MET amplification and MET overexpression was analyzed using Pearson’s correlation coefficient. Survival curves were generated using Prism. Survival time was regarded as the period from the date of surgery to the date of death of any cause or the date of last follow-up. P < 0.05 was considered significant.The retrospective analysis was performed on the selected cases of NSCLC detected using the ARMS polymerase chain reaction (PCR) from January 2019 to July 2022 in our hospital. Notably, these selected cases of NSCLC included patients with MET exon 14 skipping mutation, 1 case with MET exon 14 skipping mutation combined with KRAS mutation, a case with ALK fusion, and a case with EGFR L858R mutation. Among the total recruited 17 subjects, there were 10 males and 7 females, with a median age of 69 years. The carcinoma status included a case of well-differentiated adenocarcinoma, a case of moderately differentiated adenocarcinoma, and 15 cases of poorly differentiated adenocarcinoma and sarcomatoid carcinoma. Among them, 3 of the recruited cases were in clinical stage I, and 14 were in stage IV (Table ) or have a more jagged outline with central luminal spaces surrounded by the tumor cells , moderately positive staining in 3 cases , weakly positive in a case , polyploid in 3 cases, and negative cases in 7 cases (Fig. The FISH analysis of 16 samples indicated that the MET cluster amplification in 6 cases Fig. K and L, ses Fig. . No statses Fig. B.Further, the correlation between the C-MET IHC and MET FISH of the selected 16 cases was demonstrated. The C-MET cytoplasm and membrane staining demonstrated strong positive staining in 10 cases and cluster amplification in 3 cases. Further, the FISH analysis indicated polyploid in 3 cases, moderately positive in 3 cases, a case of polyploid, a case of cluster amplification, and a case of negative (Fig. According to the statistical data, MET exon 14 skipping mutation is evident in about 3% of NSCLC patients, indicating its prognosis and progression . DespiteNSCLC patients with MET exon 14 skipping mutation are often affected in people of an average age of over 70 years or older, dissimilar in cancerous patients with other oncogene-driven forms of NSCLC . PreviouNotably, the MET expression is heterogeneous. Accordingly, we have observed different levels of molecular heterogeneity, including intra-tumor variability Fig. . These dIn general, the MET exon 14 skipping mutation may not result in MET overexpression . HoweverIHC staining analysis is often used in clinics due to its feasibility and facile operation. In addition, protein expression levels reflected in the IHC results could be correlated to the amount of gene amplification . HencefoIn a study of 933 patients with non-squamous NSCLC, the findings displayed that none of the patients with MET exon 14 skipping mutation showed activating mutations in KRAS, EGFR or ERBB2, or rearrangements involving ALK, ROS1, or RET . ContrarFor the treatment of cancers where MET is amplified/overexpressed and activated, MET inhibitors hold tremendous potential. This is crucial since MET amplification, overexpression, and activation in tumor tissue from cancer patients have been linked to poor overall survival. Previously,NSCLC patients with MET exon 14 mutations and MET amplification had unfavcorable clinical outcome , 22. OurThe present study does, however, have certain drawbacks, including the low quality of the statistics because of the small sample size. Multivariate analysis could not be applied due to the small number of cases and that analysis with a larger sample size will be necessary in the future. Additionally, the limitation seemed to be due to the primer’s incapacity to detect all known exon 14 skipping events. The probe used in DNA-based methods using amplicons may not sufficiently cover the region of interest.In conclusion our study analyzed the between MET gene copy number and MET protein expression in 17 NSCLC patients. Among the recruited 17 cases of MET exon 14 skipping, 9 cases presented MET amplification, and 10 cases with MET overexpression. 4 cases showed gene amplification, and 3 cases presented polyploidy condition. A significant relationship between MET amplification and MET overexpression were observed. And yet, no associations of MET copy amplification or overexpression were showed with clinical characteristics and overall survival. Our findings provide baseline biomarker data for MET inhibition-based clinical trials against NSCLC.Below is the link to the electronic supplementary material.Supplementary Material 1


MET) proto-oncogene encodes the MET receptor tyrosine kinase. MET aberrations drive tumorigenesis in several cancer types through a variety of molecular mechanisms, including MET mutations, gene amplification, rearrangement, and overexpression. Therefore, MET is a therapeutic target and the selective type Ib MET inhibitor, tepotinib, was designed to potently inhibit MET kinase activity. In vitro, tepotinib inhibits MET in a concentration-dependent manner irrespective of the mode of MET activation, and in vivo, tepotinib exhibits marked, dose-dependent antitumor activity in MET-dependent tumor models of various cancer indications. Tepotinib penetrates the blood–brain barrier and demonstrates strong antitumor activity in subcutaneous and orthotopic brain metastasis models, in-line with clinical activity observed in patients. MET amplification is an established mechanism of resistance to EGFR tyrosine kinase inhibitors (TKI), and preclinical studies show that tepotinib in combination with EGFR TKIs can overcome this resistance. Tepotinib is currently approved for the treatment of adult patients with advanced or metastatic non–small cell lung cancer harboring MET exon 14 skipping alterations. This review focuses on the pharmacology of tepotinib in preclinical cancer models harboring MET alterations and demonstrates that strong adherence to the principles of the Pharmacological Audit Trail may result in a successful discovery and development of a precision medicine.The mesenchymal–epithelial transition factor ( MET) proto-oncogene, which is located on chromosome 7q21–31, encodes the MET receptor tyrosine kinase  skipping  phosphorylation site Ser985; Tyr1003, which is phosphorylated to activate recruitment of the E3 ubiquitin ligase Casitas B-lineage lymphoma (CBL); and a caspase cleavage site involved in apoptosis  tyrosine kinase inhibitors ,  in patients with target or non-target brain lesions (n = 43) and an intracranial objective response rate (ORR) of 66.7%  in patients with target brain lesions (n = 15), according to Response Assessment in Neuro-Oncology Brain Metastases criteria , gefitinib and afatinib had no inhibitory effect on tumor growth whereas tepotinib alone delayed tumor growth significantly, and tepotinib combined with EGFR-TKIs caused complete tumor regression (EGFR Del19 and METamp), tepotinib, alone or combined with EGFR-TKIs , induced complete tumor regression , monotherapy with tepotinib or rociletinib only moderately affected tumor growth, whereas afatinib and erlotinib had no effect. Tepotinib in combination with afatinib or erlotinib also moderately inhibited tumor growth but tepotinib combined with the third-generation EGFR-TKI rociletinib induced complete regression  may help overcome HER2 inhibitor resistance among patients with cooperating pan-HER and MET dysregulation . A maximMETamp and MET overexpressions are associated with anti-EGFR resistance in NSCLC, it was hypothesized that MET contributes to anti-EGFR resistance in TNBC  cell lines . In TNBC in TNBC . In TNBC in TNBC .in vivo preclinical models, to optimize assessment and selection of the clinical dose and schedule. Moreover, the characteristics of in vivo models and their predictive value in humans should be established and carefully considered.Establishing a dose that is pharmacodynamically active and safe is a mainstay in drug development, especially for targeted treatments that are less prone to toxicities. It is therefore essential to develop a thorough understanding of the PK/PD relationship on the basis of METamp, METex14 skipping, and MET fusions, as described above  phospho-MET inhibition for ≥24 hours was required to achieve tumor regression in the KP-4 model . After aMET alterations, including gastric cancer (NCT05439993) and brain tumors (NCT05120960).As of December 2022, the tepotinib clinical development program includes 17 phase I and II clinical studies  sponsored by the healthcare business of Merck KGaA, Darmstadt, Germany see . Of thesMETamp, consistent with the previously described preclinical pharmacology data  and RP2D of tepotinib  or high-level METamp  from liquid biopsy (LBx) samples or with an RNA-based approach using fresh or archival tissue biopsy (TBx) samples. Independently assessed ORR was consistent between the LBx  and TBx groups , underscoring the value of non-invasive LBx testing  NSCLC harboring MET overexpression or METamp, who had failed prior EGFR-TKI therapy. Although no significant difference was seen in the overall population, the tepotinib and gefitinib combination greatly improved outcomes versus chemotherapy in the subgroup with METamp, with a median progression-free survival (mPFS) of 16.6 months  versus 4.2 months  and median overall survival (mOS) of 37.3 months  versus 13.1 months , respectively, in the final analysis  testing and ≥9 months’ follow-up  have previously been reported in detail (METex14 skipping NSCLC (N = 255), tepotinib had a manageable safety profile with a low frequency of treatment discontinuation due to AEs, the most frequent all-cause AEs were mostly mild/moderate and included : edema , nausea (26.7%/0.8%), diarrhea (26.3%/0.4%), blood creatinine increase (25.9%/0.4%), hypoalbuminemia , pleural effusions (13.3%/5.1%), vomiting (12.9%/1.2%), and alanine transaminase (ALT) and/or aspartate transaminase (AST) increase , seven patients had MET codon D1228 or Y1230 mutations at the time of progression (METamp) showed emergence of MET kinase domain mutations in 2/9 patients (22.2%) with available end-of-treatment biomarker profiles (D1228H/N/Y, Y1230C/H, and D1231N in one patient, and D1213N, D1228N/H, and Y1230H in the other; ref. Clinical evidence for these variants as potential resistance mutations was provided by biomarker analyses from VISION. In Cohort A (gression . More reRAS mutations may activate the MAPK pathway and could thereby impair the downstream effect of MET inhibition (RAS mutations (in vitro (in vitro study suggests that MET inhibitor–induced autophagy may mediate resistance to MET inhibitors, including tepotinib, specifically in gastric cancer models (hibition . Preclinutations . In EBC-in vitro . Anotherr models . In thisr models .Thus, a MET inhibitor in combination with another targeted therapy to block alternative signaling pathways may be a strategy for further investigation to overcome MET inhibitor resistance . OngoingMETex14 skipping NSCLC. In this respect, the development of tepotinib is fully consistent with the subsequently published PhAT, which describes key questions to be addressed during discovery and development of a molecularly targeted anticancer drug (The discovery and development of tepotinib followed a stepwise approach, including the early identification of a robust PD biomarker and development of a deep understanding of the PK/PD relationship with respect to target/pathway modulation, which supported successful clinical dose and schedule selection. Alongside stringent use of biomarker-selected and predictive preclinical models to demonstrate preclinical proof of concept, these studies undergirded a successful clinical development program that enabled approval of tepotinib for advanced/metastatic cer drug . ImportaMETex14 skipping in LBx samples in VISION significantly accelerated recruitment and generated important data to understand tepotinib resistance (LBx is a powerful diagnostic tool for precision oncology that has recently been integrated into routine practice and is highlighted as an emerging technology in the PhAT, both to identify appropriate patients for biomarker-driven therapy, and to enable exploratory analysis of resistance mechanisms and alternative response measures . Implemesistance . CarefulMET alterations (METex14 skipping alterations, and was followed by approvals in multiple other countries/regions (METex14 skipping metastatic NSCLC (METamp metastatic NSCLC (By selectively targeting MET, the precision medicine tepotinib has shown durable activity in patients with hard-to-treat aggressive NSCLC tumors harboring specific oncogenic erations . The app/regions . Tepotinic NSCLC , and as ic NSCLC .