Edible insects: Understanding benzo(a)pyrene toxicokinetics in yellow mealworms for safe and sustainable consumption.

The global interest in edible insects as sustainable protein sources raises concerns about the bioaccumulation of contaminants, including polycyclic aromatic hydrocarbons (PAHs), to problematic levels. Understanding the accumulation dynamics of PAHs in edible insects is highly relevant due to the widespread sources and toxicological profiles; however, the bioaccumulative potential of PAHs in edible insects is unexplored. This study examined the uptake and elimination dynamics of benzo(a)pyrene (B(a)P), a representative and carcinogenic PAH, in yellow mealworm larvae (YMW, Tenebrio molitor). Larvae were exposed to feeding substrate with varying B(a)P concentrations (0.03, 0.3, and 3 mg kg-1), and uptake (21 days in B(a)P-contaminated substrate) and elimination (21 days in B(a)P-free substrate) kinetics were subsequently assessed. The results showed that YMW can eliminate B(a)P, revealing dose-dependent B(a)P bioaccumulation in these insects. Larvae fed on a substrate with 0.03 mg kg-1 accumulated B(a)P over 21 days, presenting values of 0.049 (Standard deviation - 0.011) mg kg-1 and a kinetic-based (BAFkinetic) of 1.93 g substrate g organism-1, exceeding the EU regulatory limits for food. However, with a B(a)P half-life (DT50) of 4.19 days in the larvae, an EU legislation safety criterion was met after a 13-day depuration period in clean substrate. Larvae exposed to substrates with 0.3 and 3 mg kg-1 showed B(a)P accumulation, with BAFkinetic values of 3.27 and 2.09 g substrate g organism-1, respectively, not meeting the current legal standards for food consumption at the end of the exposure to B(a)P. Although the B(a)P half-life values after 35 days were 4.30 and 10.22 days (DT50s), the larvae retained B(a)P levels exceeding permitted food safety limits. These findings highlight a significant oversight in regulating PAHs in animal feed and the need for comprehensive safety evaluations of PAH hazards in edible insects for improved PAH feeding guidelines.

Effects of Metal Promoters (M = Fe, Co, and Cu) in Pt/M x Zr y O z Catalysts and Influence of CO2 and H2O on the CO Oxidation Activity (PROX): Analysis of Surface Properties After Reaction.

In the present paper, the effects of metal promoters (M = Fe, Co, and Cu) in Pt/M x Zr y O z catalysts and the influence of CO2 and H2O on the CO oxidation activity (PROX) were investigated. To do that, characterizations of catalyst structures and surfaces were performed and reported here. The catalyst Pt/Fe x Zr y O z (PFeZ) was the most active at low temperatures among the analyzed ones. The addition of platinum caused strong interaction with the mixed oxide, affecting the structure and the surface composition, blocking basic sites, and thus preventing catalyst deactivation. Particularly, diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) results evidenced the formation of carboxylate and carbonate species. Besides, the addition of CO2 and H2O in the gas feed stream affected the observed CO oxidation results, showing that CO2 competes with O2 on metallic sites. Moreover, DRIFTS and temperature-programmed desorption (TPD) analyses suggested the occurrence of OH- oxidation by CO, leading to the formation of highly reactive compounds that can be easily oxidized.

Clubfoot: Congenital Talipes Equinovarus.

Congenital talipes equinovarus (CTEV), also known as clubfoot, is a common musculoskeletal entity that affects one to two per 1000 live births worldwide. Imaging modalities including radiography, US, and MRI have emerged as valuable tools for the diagnosis, treatment, and monitoring of CTEV. The deformity is characterized by midfoot cavus, forefoot adductus, and hindfoot varus and equinus. The Ponseti method of manipulation and serial casting is the standard treatment of CTEV. Radiography shows the anatomy, position, and relationships of the different bones of the foot. US allows accurate assessment of cartilaginous and bony structures, in addition to its inherent advantages such as absence of ionizing radiation exposure. One of the indications for US is to monitor the response to Ponseti method treatment. MRI enables visualization of bones, cartilage, and soft tissues and allows multiplanar evaluation of deformities, providing a comprehensive imaging analysis of CTEV. An integrated approach that combines clinical examination and imaging findings is essential for effective management of CTEV. The authors provide a comprehensive overview of CTEV with a review of imaging modalities to help evaluate CTEV, focusing on radiography, US, and MRI. Using this article as a guide, radiologists involved in the assessment and treatment of CTEV can contribute to the management of the condition. ©RSNA, 2024 Supplemental material is available for this article.

Advances in Cell-Rich Inks for Biofabricating Living Architectures.

Advancing biofabrication toward manufacturing living constructs with well-defined architectures and increasingly biologically relevant cell densities is highly desired to mimic the biofunctionality of native human tissues. The formulation of tissue-like, cell-dense inks for biofabrication remains, however, challenging at various levels of the bioprinting process. Promising advances have been made toward this goal, achieving relatively high cell densities that surpass those found in conventional platforms, pushing the current boundaries closer to achieving tissue-like cell densities. On this focus, herein the overarching challenges in the bioprocessing of cell-rich living inks into clinically grade engineered tissues are discussed, as well as the most recent advances in cell-rich living ink formulations and their processing technologies are highlighted. Additionally, an overview of the foreseen developments in the field is provided and critically discussed.

Green manure (Crotalaria juncea L.) enhances Origanum vulgare L. biomass accumulation, essential oil yield, and phytochemical properties.

Green manure (GM) may reduce the use of chemical fertilizers, been an ecologically appropriate strategy to cultivation of medicinal plants. Crotalaria juncea, is one of the most used because it adapts to different climatic and high nitrogen content. Origanum vulgare. is widely used in cooking, pharmaceutical, cosmetic industries and food products. The objectives of this study were to evaluate the GM on biomass, essential oil (EO), phenolic and antioxidant. The experiment consisted: control; 150, 300, 450, and 600 g (Sh= leaves+steam) more 200 g roots (R); 600 g aerial part; 200 g roots; and soil with 300 g cattle manure per pot. The highest dry weights were observed in the presence of GM and cattle manure (90 days). The control had an EO production 75% lower in relation to the dose of 450 g GM (Sh+R). Principal component analysis showed that GM and cattle manure positively influenced the dry weight, content, yield, and EO constituents, and total flavonoids. The GM contributed to the accumulation of the major EO compounds (trans-sabinene hydrate, thymol, terpinen-4-ol). The GM management may be beneficial for cultivating, because it can increase the production of biomass and the active components, in addition to being an inexpensive resource.

Post-translational modifications of vertebrate striated muscle myosin heavy chains.

Post-translational modifications (PTMs) play a crucial role in regulating the function of many sarcomeric proteins, including myosin. Myosins comprise a family of motor proteins that play fundamental roles in cell motility in general and muscle contraction in particular. A myosin molecule consists of two myosin heavy chains (MyHCs) and two pairs of myosin light chains (MLCs); two MLCs are associated with the neck region of each MyHC's N-terminal head domain, while the two MyHC C-terminal tails form a coiled-coil that polymerizes with other MyHCs to form the thick filament backbone. Myosin undergoes extensive PTMs, and dysregulation of these PTMs may lead to abnormal muscle function and contribute to the development of myopathies and cardiovascular disorders. Recent studies have uncovered the significance of PTMs in regulating MyHC function and showed how these PTMs may provide additional modulation of contractile processes. Here, we discuss MyHC PTMs that have been biochemically and/or functionally studied in mammals' and rodents' striated muscle. We have identified hotspots or specific regions in three isoforms of myosin (MYH2, MYH6, and MYH7) where the prevalence of PTMs is more frequent and could potentially play a significant role in fine-tuning the activity of these proteins.

Rationale and Design of VOYAGER: Long-term Outcomes of Faricimab and Port Delivery System with Ranibizumab for Neovascular Age-Related Macular Degeneration and Diabetic Macular Edema in Clinical Practice.

Purpose: To describe the rationale and design of the VOYAGER (NCT05476926) study, which aims to investigate the safety and effectiveness of faricimab and the Port Delivery System with ranibizumab (PDS) for neovascular age-related macular degeneration (nAMD) or diabetic macular edema (DME) in clinical practice. VOYAGER also aims to understand drivers of clinical practice treatment outcomes by gaining novel insight into the intersection of treatment regimens, decisions, anatomic outcomes, and vision. Design: Primary data collection, noninterventional, prospective, multinational, multicenter clinical practice study. Participants: At least 5000 patients initiating/continuing faricimab or PDS for nAMD/DME (500 sites, 31 countries). Methods: Management will be per usual care, with no mandated scheduled visits/imaging protocol requirements. Using robust methodologies, relevant clinical and ophthalmic data, including visual acuity (VA), and data on treatment clinical setting/regimens/philosophies, presence of anatomic features, and safety events will be collected. Routinely collected fundus images will be uploaded to the proprietary Imaging Platform for analysis. An innovative investigator interface will graphically display the patient treatment journey with the aim of optimizing treatment decisions. Main Outcome Measures: Primary end point: VA change from baseline at 12 months per study cohort (faricimab in nAMD and in DME, PDS in nAMD). Secondary end points: VA change over time and per treatment regimens (fixed, treat-and-extend, pro re nata, and other) and number. Exploratory end points: VA change in relation to presence/location of anatomic features that impact vision (fluid, central subfield thickness, fibrosis, atrophy, subretinal hyperreflective material, diabetic retinopathy severity, and disorganization of retinal inner layers) and per treatment regimen/philosophies. The impact of regional and practice differences on outcomes will be assessed as will safety. Results: Recruitment commenced in November 2022 and will continue until late 2027, allowing for up to 5 years follow-up. Exploratory interim analyses are planned annually. Conclusions: VOYAGER is an innovative study of retinal diseases that will assess the effectiveness and safety of faricimab and PDS in nAMD and DME and identify clinician- and disease-related factors driving treatment outcomes in clinical practices globally to help optimize vision outcomes. Financial Disclosures: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Deep hybrid modeling of a HEK293 process: Combining long short-term memory networks with first principles equations.

The combination of physical equations with deep learning is becoming a promising methodology for bioprocess digitalization. In this paper, we investigate for the first time the combination of long short-term memory (LSTM) networks with first principles equations in a hybrid workflow to describe human embryonic kidney 293 (HEK293) culture dynamics. Experimental data of 27 extracellular state variables in 20 fed-batch HEK293 cultures were collected in a parallel high throughput 250 mL cultivation system in an industrial process development setting. The adaptive moment estimation method with stochastic regularization and cross-validation were employed for deep learning. A total of 784 hybrid models with varying deep neural network architectures, depths, layers sizes and node activation functions were compared. In most scenarios, hybrid LSTM models outperformed classical hybrid Feedforward Neural Network (FFNN) models in terms of training and testing error. Hybrid LSTM models revealed to be less sensitive to data resampling than FFNN hybrid models. As disadvantages, Hybrid LSTM models are in general more complex (higher number of parameters) and have a higher computation cost than FFNN hybrid models. The hybrid model with the highest prediction accuracy consisted in a LSTM network with seven internal states connected in series with dynamic material balance equations. This hybrid model correctly predicted the dynamics of the 27 state variables (R2 = 0.93 in the test data set), including biomass, key substrates, amino acids and metabolic by-products for around 10 cultivation days.

Structure of mavacamten-free human cardiac thick filaments within the sarcomere by cryoelectron tomography.

Heart muscle has the unique property that it can never rest; all cardiomyocytes contract with each heartbeat which requires a complex control mechanism to regulate cardiac output to physiological requirements. Changes in calcium concentration regulate the thin filament activation. A separate but linked mechanism regulates the thick filament activation, which frees sufficient myosin heads to bind the thin filament, thereby producing the required force. Thick filaments contain additional nonmyosin proteins, myosin-binding protein C and titin, the latter being the protein that transmits applied tension to the thick filament. How these three proteins interact to control thick filament activation is poorly understood. Here, we show using 3-D image reconstruction of frozen-hydrated human cardiac muscle myofibrils lacking exogenous drugs that the thick filament is structured to provide three levels of myosin activation corresponding to the three crowns of myosin heads in each 429Å repeat. In one crown, the myosin heads are almost completely activated and disordered. In another crown, many myosin heads are inactive, ordered into a structure called the interacting heads motif. At the third crown, the myosin heads are ordered into the interacting heads motif, but the stability of that motif is affected by myosin-binding protein C. We think that this hierarchy of control explains many of the effects of length-dependent activation as well as stretch activation in cardiac muscle control.

Troponin Structural Dynamics in the Native Cardiac Thin Filament Revealed by Cryo Electron Microscopy.

Cardiac muscle contraction occurs due to repetitive interactions between myosin thick and actin thin filaments (TF) regulated by Ca2+ levels, active cross-bridges, and cardiac myosin-binding protein C (cMyBP-C). The cardiac TF (cTF) has two nonequivalent strands, each comprised of actin, tropomyosin (Tm), and troponin (Tn). Tn shifts Tm away from myosin-binding sites on actin at elevated Ca2+ levels to allow formation of force-producing actomyosin cross-bridges. The Tn complex is comprised of three distinct polypeptides - Ca2+-binding TnC, inhibitory TnI, and Tm-binding TnT. The molecular mechanism of their collective action is unresolved due to lack of comprehensive structural information on Tn region of cTF. C1 domain of cMyBP-C activates cTF in the absence of Ca2+ to the same extent as rigor myosin. Here we used cryo-EM of native cTFs to show that cTF Tn core adopts multiple structural conformations at high and low Ca2+ levels and that the two strands are structurally distinct. At high Ca2+ levels, cTF is not entirely activated by Ca2+ but exists in either partially or fully activated state. Complete dissociation of TnI C-terminus is required for full activation. In presence of cMyBP-C C1 domain, Tn core adopts a fully activated conformation, even in absence of Ca2+. Our data provide a structural description for the requirement of myosin to fully activate cTFs and explain increased affinity of TnC to Ca2+ in presence of active cross-bridges. We suggest that allosteric coupling between Tn subunits and Tm is required to control actomyosin interactions.

Profiling the neuroproteomics of honeybee brain: A clue for understanding the role of neuropeptides in the modulation of aggressivity.

The aggressivity is modulated in honeybee brain through a series of actions in cascade mode, with the participation of the neuropeptides AmAST A (59-76) and AmTRP (254-262). The aggressivity of honeybees was stimulated by injecting both neuropeptides in the hemocoel of the worker honeybees, which were submitted to behavioral assays of aggression. The brain of stinger individuals were removed by dissection and submitted to proteomic analysis; shotgun proteomic approach of honeybee brain revealed that both neuropeptides activate a series of biochemical processes responsible by production of energy, neuronal plasticity and cell protection. In addition to this, AmTRP (254-262) elicited the expression of proteins related to the processing of the potential of action and lipid metabolism; meanwhile AmAST A (59-76) elicited the metabolism of steroids and Juvenile hormone-related metabolism, amongst others. Apparently, the most complex biochemical process seems to be the regulation of ATP production, which occurs at two levels: i) by a subgroup of proteins common to the three experimental groups, which are over-/under-regulated through glycolysis, pyruvate pathway, Krebbs cycle and oxidative phosphorylation; ii) by a subgroup of proteins unique to the each experimental group, which seems to be regulated through Protein-Protein Interactions, where the protein network regulated by AmTRP (254-262) seems to be more complex than the other two experimental groups. SIGNIFICANCE: Recently we reported the effect of the neuropeptides AmAST A (59-76) and AmTRP (254-262) in the modulation of the aggressive behavior of the worker honeybees. Up to now it is known that the simple presence of the allatostatin and tachykinin-related-peptide in bee brain, is enough for inducing the aggressive behavior. However, nothing was known about how these neuropeptides perform their action, inducing the aggressive behavior. The results of the present study elucidated some of the metabolic pathways that were activated or inhibited to support the complex defensive behavior, which includes the aggressivity. These results certainly will impact the behavioral research of honeybees, since we are paving the way for understanding the molecular base of regulation, of individual /nest defense of honeybees.

Targeted proteomics and metabolomics for biomarker discovery in abdominal aortic aneurysm and post-EVAR sac volume.

BACKGROUND AND AIMS: Abdominal aortic aneurysm (AAA) patients undergo uniform surveillance programs both leading up to, and following surgery. Circulating biomarkers could play a pivotal role in individualizing surveillance. We applied a multi-omics approach to identify relevant biomarkers and gain pathophysiological insights. MATERIALS AND METHODS: In this cross-sectional study, 108 AAA patients and 200 post-endovascular aneurysm repair (post-EVAR) patients were separately investigated. We performed partial least squares regression and ingenuity pathway analysis on circulating concentrations of 96 proteins (92 Olink Cardiovascular-III panel, 4 ELISA-assays) and 199 metabolites (measured by LC-TQMS), and their associations with CT-based AAA/sac volume. RESULTS: The median (25th-75th percentile) maximal diameter was 50.0 mm (46.0, 53.0) in the AAA group, and 55.4 mm (45.0, 64.2) in the post-EVAR group. Correcting for clinical characteristics in AAA patients, the aneurysm volume Z-score differed 0.068 (95 %CI: (0.042, 0.093)), 0.066 (0.047, 0.085) and -0.051 (-0.064, -0.038) per Z-score valine, leucine and uPA, respectively. After correcting for clinical characteristics and orthogonalization in the post-EVAR group, the sac volume Z-score differed 0.049 (0.034, 0.063) per Z-score TIMP-4, -0.050 (-0.064, -0.037) per Z-score LDL-receptor, -0.051 (-0.062, -0.040) per Z-score 1-OG/2-OG and -0.056 (-0.066, -0.045) per Z-score 1-LG/2-LG. CONCLUSIONS: The branched-chain amino acids and uPA were related to AAA volume. For post-EVAR patients, LDL-receptor, monoacylglycerols and TIMP-4 are potential biomarkers for sac volume. Additionally, distinct markers for sac change were identified.

Unlocking the Role of sMyBP-C: A Key Player in Skeletal Muscle Development and Growth.

Skeletal muscle is the largest organ in the body, responsible for gross movement and metabolic regulation. Recently, variants in the MYBPC1 gene have been implicated in a variety of developmental muscle diseases, such as distal arthrogryposis. How MYBPC1 variants cause disease is not well understood. Here, through a collection of novel gene-edited mouse models, we define a critical role for slow myosin binding protein-C (sMyBP-C), encoded by MYBPC1, across muscle development, growth, and maintenance during prenatal, perinatal, postnatal and adult stages. Specifically, Mybpc1 knockout mice exhibited early postnatal lethality and impaired skeletal muscle formation and structure, skeletal deformity, and respiratory failure. Moreover, a conditional knockout of Mybpc1 in perinatal, postnatal and adult stages demonstrates impaired postnatal muscle growth and function secondary to disrupted actomyosin interaction and sarcomere structural integrity. These findings confirm the essential role of sMyBP-C in skeletal muscle and reveal specific functions in both prenatal embryonic musculoskeletal development and postnatal muscle growth and function.

Corrigendum: The paradigm of IL-23-independent production of IL-17F and IL-17A and their role in chronic inflammatory diseases.

[This corrects the article DOI: 10.3389/fimmu.2023.1191782.].

Local protection bubbles: an interpretation of the slowdown in the spread of coronavirus in the city of São Paulo, Brazil, in July 2020.

After four months of fighting the pandemic, the city of São Paulo, Brazil, entered a phase of relaxed social distancing measures in July 2020. Simultaneously, there was a decline in the social distancing rate and a reduction in the number of cases, fatalities, and hospital bed occupancy. To understand the pandemic dynamics in the city of São Paulo, we developed a multi-agent simulation model. Surprisingly, the counter-intuitive results of the model followed the city's reality. We argue that this phenomenon could be attributed to local bubbles of protection that emerged in the absence of contagion networks. These bubbles reduced the transmission rate of the virus, causing short and temporary reductions in the epidemic curve - but manifested as an unstable equilibrium. Our hypothesis aligns with the virus spread dynamics observed thus far, without the need for ad hoc assumptions regarding the natural thresholds of collective immunity or the heterogeneity of the population's transmission rate, which may lead to erroneous predictions. Our model was designed to be user-friendly and does not require any scientific or programming expertise to generate outcomes on virus transmission in a given location. Furthermore, as an input to start our simulation model, we developed the COVID-19 Protection Index as an alternative to the Human Development Index, which measures a given territory vulnerability to the coronavirus and includes characteristics of the health system and socioeconomic development, as well as the infrastructure of the city of São Paulo.

Aneurysm Sac Dynamics and its Prognostic Significance Following Fenestrated and Branched Endovascular Aortic Aneurysm Repair.

OBJECTIVE: This study aimed to assess aneurysm sac dynamics and its prognostic significance following fenestrated and branched endovascular aneurysm repair (F/BEVAR). METHODS: Patients undergoing F/BEVAR for degenerative complex aortic aneurysm from 2008 to 2020 at two large vascular centres with two imaging examinations (30 day and one year) were included. Patients were categorised as regression and non-regression, determined by the proportional volume change (> 5%) at one year compared with 30 days. All cause mortality and freedom from graft related events were assessed using Kaplan-Meier methods. Factors associated with non-regression at one year and aneurysm sac volume over time were examined for FEVAR and BEVAR independently using multivariable logistic regression and linear mixed effects modelling. RESULTS: One hundred and sixty-five patients were included: 122 FEVAR, of whom 34% did not regress at one year imaging (20% stable, 14% expansion); and 43 BEVAR, of whom 53% failed to regress (26% stable, 28% expansion). Following F/BEVAR, after risk adjusted analysis, non-regression was associated with higher risk of all cause mortality within five years (hazard ratio [HR] 2.56, 95% confidence interval [CI] 1.09 - 5.37; p = .032) and higher risk of graft related events within five years (HR 2.44, 95% CI 1.10 - 5.26; p = .029). Following multivariable logistic regression, previous aortic repair (odds ratio [OR] 2.56, 95% CI 1.11 - 5.96; p = .029) and larger baseline aneurysm diameter (OR/mm 1.04, 95% CI 1.00 - 1.09; p = .037) were associated with non-regression at one year, whereas smoking history was inversely associated with non-regression (OR 0.21, 95% CI 0.04 - 0.96; p = .045). Overall following FEVAR, aneurysm sac volume decreased significantly up to two years (baseline vs. two year, 267 [95% CI 250 - 285] cm3vs. 223 [95% CI 197 - 248] cm3), remaining unchanged thereafter. Overall following BEVAR, aneurysm sac volume remained stable over time. CONCLUSION: Like infrarenal EVAR, non-regression at one year imaging is associated with higher five year all cause mortality and graft related events risks after F/BEVAR. Following FEVAR for juxtarenal aortic aneurysm, aneurysm sacs generally displayed regression (66% at one year), whereas after BEVAR for thoraco-abdominal aortic aneurysm, aneurysm sacs displayed a concerning proportion of growth at one year (28%), potentially suggesting a persistent risk of rupture and consequently requiring intensified surveillance following BEVAR. Future studies will have to elucidate how to improve sac regression following complex EVAR, and whether the high expansion risk after BEVAR is due to advanced disease extent.

Microencapsulation of Bacillus thuringiensis strains for the control of Aedes aegypti.

In this study, we investigated the microencapsulation of two strains of the entomopathogenic bacteria Bacillus thuringiensis (B. thuringiensis) (BtMA-750 and BtMA-1114), which are biopesticides of high toxicity for the mosquito vector Aedes aegypti. The encapsulation of different concentrations of microorganisms in starch microparticles was evaluated, and the inverse suspension polymerization technique was explored. It was possible to observe that the higher amounts of the biopesticide caused a slight decrease in the diameter of the particles; however, even when encapsulated, the biopesticide still presents an average diameter that is able to be consumed by the larvae of Aedes aegypti. Furthermore, it was noticed that the presence of both of the B. thuringiensis strains did not affect the thermal stability of the particles. The microencapsulated bacterial strains presented a high number of viable spores and preserved the expression of proteins with molecular masses corresponding to the insecticidal toxins Cry and Cyt, indicating that the encapsulation process was conducted satisfactorily. Finally, the encapsulated strains were tested against Ae. aegypti larvae and maintained 100% larval mortality even after 35 days. Therefore, microencapsulation of B. thuringiensis not only guarantees the bacterial activity, but also prolongs the action of the biopesticide. Collectively, such findings highlight the great potential of the new biopesticides, which may help to reduce the population indices of the mosquito vector Ae. aegypti via a sustainable and environment-friendly route.

A Strategy Utilizing Protein-Protein Interaction Hubs for the Treatment of Cancer Diseases.

We describe a strategy for the development of a rational approach of neoplastic disease therapy based on the demonstration that scale-free networks are susceptible to specific attacks directed against its connective hubs. This strategy involves the (i) selection of up-regulated hubs of connectivity in the tumors interactome, (ii) drug repurposing of these hubs, (iii) RNA silencing of non-druggable hubs, (iv) in vitro hub validation, (v) tumor-on-a-chip, (vi) in vivo validation, and (vii) clinical trial. Hubs are protein targets that are assessed as targets for rational therapy of cancer in the context of personalized oncology. We confirmed the existence of a negative correlation between malignant cell aggressivity and the target number needed for specific drugs or RNA interference (RNAi) to maximize the benefit to the patient's overall survival. Interestingly, we found that some additional proteins not generally targeted by drug treatments might justify the addition of inhibitors designed against them in order to improve therapeutic outcomes. However, many proteins are not druggable, or the available pharmacopeia for these targets is limited, which justifies a therapy based on encapsulated RNAi.