Secondary metabolites (SMs) found in medicinal plants are one of main sources of drugs, cosmetics, and health products. With the increase in demand for these bioactive compounds, improving the content and yield of SMs in medicinal plants has become increasingly important. The content and distribution of SMs in medicinal plants are closely related to environmental factors, especially light. In recent years, artificial light sources have been used in controlled environments for the production and conservation of medicinal germplasm. Therefore, it is essential to elucidate how light affects the accumulation of SMs in different plant species. Here, we systematically summarize recent advances in our understanding of the regulatory roles of light quality, light intensity, and photoperiod in the biosynthesis of three main types of SMs , and the underlying mechanisms. This article provides a detailed overview of the role of light signaling pathways in SM biosynthesis, which will further promote the application of artificial light sources in medicinal plant production. Artemisia annua, is widely used for treating malaria  are a class of lactones structurally constructed by a benzene ring fused to α-pyrone ring, such as aesculin, cnidium lactone, and alpha-Angelica lactone. Lignans [(C6-C3)2] are phenolic dimers with a 2,3-dibenzylbutane skeleton, such as phyllanthin, arctiin, and podophyllotoxin. Phenolic acids present compounds containing a carboxylic group among the substituents on the benzene ring, including benzoic acid derivates (C6-C1) and hydroxycinnamic derivates (C6-C3). Flavonoids (C6-C3-C6) refer to compounds consisting of two benzene rings linked by a short three carbon chain, such as chalcones, flavones, flavonols, dihydroflavones, dihydroflavonols, isoflavones, and dihydroisoflavones. Tannins are high molecular polyphenols polymerized by flavonoids units (condensed tannins) or gallic acid esterified with monosaccharide , terpenoids, and alkaloids . Polyphedistinct .A, and paclitaxel), and two triterpenes (cucurbitacin I and oleanolic acid). Terpenoids are synthesized via two parallel upstream pathways: the mevalonate pathway (MVA) and methylerythritol-4-phosphate pathway , and then to various MIAs, such as camptothecin, serpentine, and vinblastine  phytochromes (phyA–phyE), which mainly perceive red light and far-red light , visible light or photosynthetically active radiation , and far-red light (700–800nm). Photoreceptors are indispensable for light sensing and light signal transduction in plants. To date, at least five kinds of photoreceptors have been identified in the model plant species ed light ; (2) cryed light ; (4) zeied light ; and (5)ed light . Among tColeus forskohlii plants treated with supplementary UV-B (ambient +0.042 Wm−2) were 1.34-, 1.67-, and 1.96-fold higher than that in control at 40, 70, and 100 days after transplantation (DAT), respectively. Prunella vulgaris and showed that UV-B radiation significantly increased the contents of total flavonoids, rosmarinic acid, caffeic acid, and hyperoside. In Rosmarinus officinalis plants treated with UV-B (0.359 Wm−2), the contents of rosmarinic acid and carnosic acid were 2.34- and 1.78-fold higher, respectively, than that of the control  significantly promoted the production of caffeic acid and chlorogenic acid  significantly decreased the content of podophyllotoxin as well as transcript levels of 12 related genes, including cinnamic acid 3-hydroxylase (C3H), caffeoyl-CoA O-methyltransferase (CCoAMT), cinnamoyl-CoA reductase (CCR), CAD, dirigent protein oxidase (DPO), pinoresinol-lariciresinol reductase (PLR), secoisolariciresinol dehydrogenase (SDH), cytochrome P450 719A23 (CPY719A23), O-methyltransferase3 (OMT3), cytochrome P450 71CU1 (CYP71CU1), OMT1, and 2-oxoglutarate/Fe(II)-dependent dioxygenase , PAL, CHS, CHI, and FLS. CHS in Betula pendula.The mechanism underlying the regulatory effects of UV-B on the synthesis of polyphenols is not fully understood. However, it has been confirmed that UV-B affects the activities of key polyphenol biosynthetic enzymes and/or transcript levels of the corresponding genes in many medicinal plants . For exaomnifera , 2015. Ie 2-ODD; . In GlycP. vulgaris calli, and found that the calli accumulated most total phenolics (23.9 mg g−1 DW) and total flavonoids (1.65 mg g−1 DW) under blue light. Similarly, Rhodiola imbricata accumulated the highest amounts of salidroside, total phenolics, and total flavonoids under blue light compared with those under red light, green light, RGB (40% red:40% green:20% blue), and white light. Verbena officinalis. Coumarins is an important class of phenols in medicinal plants. The biosynthesis and accumulation of coumarins are significantly affected by blue and red light. For instance, Sarcandra glabra seedlings with different monochromatic lights for 60 days and found that the content of fraxetin and 6-methylcoumarin in red light treated group was 45 and 16% of that in control (under white light), respectively. The content of these two coumarins in blue light treated group was 51 and 11% of that in control, respectively. Eclipta alba. The content of coumarin, wedelolactone, and demethylwedelolactone in the red light treated group  was 3.07-, 1.59-, and 1.59-fold of that in control (white light), respectively. The content of these compounds in the blue light treated group was 2.24-, 1.43-, and 1.29-fold of that in control, respectively. Combined blue and red light are often used to improve the growth and SM content of medicinal plants simultaneously. For example, Ocimum basilicum seedlings under different light conditions, and found that the dry mass, rosmarinic acid, and caffeic acid contents of seedlings were 1.45-, 15-, and 4-fold higher under combined red and blue light (1R:2B), respectively, than under control (white) conditions. Salvia miltiorrhiza seedlings with monochromatic blue light (B), monochromatic red light (R), and combined blue and red light, and showed that seedling growth and phenolic acid production were stimulated under 7R:3B. Cannabis sativa seedlings, and increased the aboveground plant biomass, flower biomass, and flower cannabidiol content by 15.2, 238, and 36.53%, respectively, compared with the control (Seedlings growth under high-pressure sodium light). Pfaffia glomerata and found that equal proportion of red and blue light (1R:1B) was the best light condition for the accumulation of biomass, anthocyanins, and 20-hydroxyecdisone.Blue and red light wavelengths are two important light qualities involved in plant growth and development. Both light qualities are widely recognized as effective elicitors that regulate the accumulation of bioactive compounds in medicinal plants . Fazal ePAL and 4CL, which are required for phlorizin synthesis, are correlated with phlorizin content under red and blue light in Lithocarpus polystachyus. Cyclocarya paliurus leaves, and the flavonoid content was positively correlated with the transcript levels of PAL, 4CL, and CHS. MYB transcription factors and microRNAs are also involved in light-induced polyphenol biosynthesis in some medicinal plants. For example, in Fagopyrum tataricum, FtMYB16 directly binds to the promoter region of the flavanone 3-hydroxylase (F3′H) gene under red and bule light to induce its expression and enhance the flavonoid content , 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR), isopentenyl pyrophosphate isomerase (IPPi), FPS, amorpha-4, 11-diene synthase (ADS) gene, cytochrome P450 71AV1 (CYP71AV1), and dihydroartemisinic aldehyde reductase  increased the content of total tanshinones and the transcript levels of 1-deoxy-D-xylulose-5-phosphate synthase (SmDXS2) and copalyl diphosphate synthase (SmCPS) by 1.5-, 6.2-, and 7.3-fold, respectively, compared with the control  and squalene epoxidase (SE) genes in Gynostemma pentaphyllum. S. miltiorrhiza hairy roots; red light treatment upregulated the expression of SmHMGR, SmDXS2, SmDXR, SmGGPPS, SmCPS, and CYP76AH1 genes and increased the content of tanshinone IIA by 1.4-fold compared with the control, whereas blue light remarkedly suppressed tanshinone IIA biosynthesis and downregulated the expression of key tanshinone IIA biosynthesis genes. Dysphania ambrosioides with different light qualities and showed that blue light inhibited the accumulation of Z-ascaridole in this herb. The effects of red and blue light on terpenoid production are species-specific. For example, both red light and blue light stimulated the biosynthesis of artemisinin and artemisinic acid in A. annua but decreased the production of essential oils in Melissa officinalis  treatment increased the alkaloid content of leaves and roots of C. forskohlii. In Clematis terniflora, UV-B irradiation (1.208 W m−2), followed by dark incubation, increased the content of indole alkaloid (6-hydroxyl-1H-indol-3-yl) carboxylic acid methyl ester by 7-fold (−2), followed by dark incubation, significantly enhanced the contents of protoberberine alkaloids, including berberine, jateorhizine, palmatine, and columbamine, in Mahonia bealei leaves , and found that most of structural genes in the alkaloid biosynthetic pathways were upregulated, among which 10-hydroxygeranioloxidoreductase (10-HGO), tabersonine 16-hydroxylase (T16H), and strictosidine synthase (STR) genes were upregulated by approximately 2-, 4-, and 4-fold, respectively, compared with the control. C. terniflora. The expression of key genes in the tropane alkaloid biosynthetic pathway is also affected by UV-B. In Anisodus luridus hairy roots treated with UV-B (90 W m−2) for 24 h, transcript levels of putrescine N-methyltransferase (PMT), tropinone reductase I (TRI), cytochrome P450 80F1 (CYP80F1), and hyoscyamine 6b-hydroxylase (H6H) genes were increased by 10-, 52-, 16-, and 9-fold, respectively, compared with the control  dramatically increased the production of galanthamine, lycorine, and lycoramine as well as the expression of norbelladine synthase (NBS), OMT, and CYP96T in Lycoris longituba. Camptotheca acuminata seedlings with different light conditions and showed that blue light  promoted the camptothecin content, tryptophan decarboxylase (TDC) and tryptophan synthase (TSB) activities, and TSB, TDC1, and TDC2 transcript levels. Psychotria leiocarp. Red light is considered as another regulator of alkaloid biosynthesis, and its function is dependent on phytochromes as well as secondary messengers , tabersonine-3-reductase (T3R), desacetoxyvindoline-4-hydroxylase (D4H), and DAT. In the dark, PIF1 suppresses the expression of the abovementioned genes, which dramatically decreases the accumulation of vindoline (−2 s−1) significantly enhanced the concentration and yield of vindoline and catharanthine in C. roseus seedlings  stimulated the accumulation of C6C3C6-type phenolics in Eleutherococcus senticosus. Lippia gracilis with different light intensities and showed that the chemical composition of seedlings in the 26 μmol m−2 s−1 group was more complex, with more monoterpene hydrocarbons and less aromatic monoterpenes, than those in other treated groups. In M. breviracema, light intensity had different effects on the production of alkaloids and essential oils; alkaloid content was significantly enhanced in the 30 and 50% sunlight groups, while the accumulation of essential oils increased linearly with the increase in light intensity, reaching maximum levels in the full (100%) sunlight group , 16L/14D, and 14L/16D photoperiods compared with the control (16L/8D). Basella rubra callus cultures under the 16:8 h photoperiod produced the highest amount of phenolics compared with those under continuous light and continuous dark conditions. Echinacea purpurea, and found that the accumulation of caffeic acid derivatives was optimum in cultures grown under 3-h light/21-h dark conditions. There are also many studies which report that continuous light or continuous dark is more efficiency to stimulate the bioactive compounds biosynthesis compared with photoperiod in some medicinal plants. For instance, −1 light was the best light condition for enhancing the growth and essential oil content of Lippia alba seedlings grown in vitro. Linum usitatissimum with different photoperiods, and showed that continuous dark conditions led to the greatest increase in total phenolics (116.85 mg L−1) and total flavonoids (37.05 mg L−1). Photoperiod also affects the chemical composition of medicinal plants. Hypericum perforatum cultured under 16-h light/8-h dark photoperiod showed de novo biosynthesis of two phenolic acids, three flavonol glycosides, and five xanthones compared those cultured under continuous dark. Tanacetum parthenium, while photoperiod showed an opposite effect on the accumulation of these compounds.In many medicinal plants, photoperiod promotes the accumulation of SMs. For example, LONG HYPOCOTYL 5 (HY5) expression. HY5 is a core transcription factor in the light signaling pathway that regulates expression of genes encoding transcription factors or key enzymes involved in the biosynthesis of SMs, to affect the accumulation of SMs under UV-B , and AaORA, to upregulates their expression. These transcription factors then stimulate the expression of related biosynthetic genes and enhance the content of artemisinin  and GATA-type transcription factor gene GATA1, thus repressing the expression of target genes and decreasing the content of vindoline. Under red light, PIF1 is degraded, which stimulates the accumulation of vindoline  reductase (DBR2), and aldehyde dehydrogenase 1 (ALDH1), and dramatically increases the content of artemisinin under light in A. annua , Key project at central government level: The ability establishment of sustainable use for valuable Chinese medicine resources (2060302), the Zhejiang Provincial Natural Science Foundation of China (LR21H280002).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The aim of the study is to evaluate the effectiveness of short whole-body magnetic resonance imaging (WBMRI) protocols for the overall assessment of bone marrow involvement in patients with multiple myeloma (MM), in comparison with standard whole-body MRI protocol. Patients with biopsy-proven MM, who underwent a WBMRI with full-body coverage (from vertex to feet) were retrospectively enrolled. WBMRI images were independently evaluated by two expert radiologists, in terms of infiltration patterns , according to location  and lytic lesions number . The majority of patients showed focal and combined infiltration patterns with bone lesions predominantly distributed in the spine and pelvis. As skull and lower limbs are less frequently involved by focal bone lesions, excluding them from the standard MRI protocol allows to obtain a shorter protocol, maintaining a good diagnostic value. Multiple myeloma (MM) is a plasma cell dyscrasia, characterized by proliferation and accumulation of monoclonal plasma cells . MM is tIn this context, whole-body (WB) imaging techniques, such as computed tomography (CT) and fluorodeoxyglucose positron emission tomography (FDG-PET) acquired increased importance over the past few years. Among them, magnetic resonance imaging (MRI) has a high specificity and sensitivity in the detection of focal bone lesions and bone marrow infiltration, even before the mineralized bone has been destroyed ,4,5,6. AAlthough the clinical use of imaging modalities to diagnose MM is often influenced by the availability and affordability of different techniques, WBMRI remains the best imaging technique for evaluating bone marrow involvement . WBMRI iIn this setting, the present study aims to evaluate the effectiveness of shorter MRI protocols for the overall assessment of bone marrow involvement, according to different infiltration patterns in patients with MM, in comparison with standard whole-body MRI protocol.Local Ethics Committee’s review of the protocol deemed that formal approval was not required owing to the retrospective, observational, and anonymous nature of this study.We retrospectively evaluated all patients who had biopsy-proven MM diagnosed during the period from January 2017 to January 2020, who underwent WBMRI for disease staging.Inclusion criteria were as follows: (1) age > 18 years, (2) MM diagnosis according to the International Myeloma Working Group, (3) WBMRI with full-body coverage (from vertex to feet).Patients with nondiagnostic examinations due to artifacts or premature suspension of the scan were excluded.Whole-body MRI examinations were performed on a 1.5 T magnet . The standard protocol included T1-weighted turbo spin-echo (TSE) and T2-weighted short-tau inversion recovery (STIR) sequences acquired on the coronal plane (from the skull vertex to the feet)  and on tThe patient was positioned supine, headfirst, using two-phased body-array coils, inline for the examination of the thorax, abdomen, pelvic region, and upper and lower limbs, and one head-and-neck coil for the head and neck regions. All WBMRI studies were performed with the stepping-table movement technique.WBMRI images were independently evaluated by two radiologists, with 12 and 7 years of experience in MM and MRI imaging, to identify signs of bone involvement. Images were evaluated in terms of standard infiltration patterns, as normal, focal, diffuse, and combined.The MRI diagnosis of bone marrow involvement was performed using mainly T1-weighted TSE and fat-suppressed sequences (STIR). Bone marrow pathological replacement, either focal or diffuse, leads to a signal intensity modification, namely a decrease on T1-weighted TSE and an increase on T2-weighted STIR images. The diffuse bone marrow involvement could also be seen as speckled and micronodular appearance  with inhomogeneous bone marrow and interposition of fat islands .The DWIBS sequences were used to confirm the type of infiltration pattern found on STIR and T1-TSE, considering that pathologic bone marrow usually exhibits restricted diffusion, with a higher signal on high b-value DWI compared to the very low signal of normal bone marrow .The whole skeleton was divided into six anatomic districts: skull, spine, sternum and ribs, upper limbs, pelvis plus proximal two-thirds of the femur, and remaining parts of lower limbs. Therefore, all the typical focal bone lesions were recorded, according to the location, into two different categories: standard protocol (from vertex to feet) and short protocol (from vertex to thigh bone).For each anatomic district, the two readers recorded the focal bone lesions according to their number as follows: less than 5, between 5–20, or more than 20 focal lesions.The acquisition time of each sequence and the whole protocol was recorded.The detailed MRI protocol is summarized in Categorical variables were presented using median and IQR values, while continuous variables were given as mean ± standard deviation (SD). Intraclass correlation coefficients (ICC) and their 95% CI were calculated based on a mean-rating (k = 2), absolute-agreement, two-way mixed-effects model. Moreover, Kendall’s tau (τ) test was performed between ordinal variables: the tau correlation coefficient returns a value of 0 to 1, where 0 represents no relationship, 1 a perfect positive relationship. The 95% CIs were calculated using bootstrap with 500 iterations and random number seed 978. For both intertest qualitative reliability and inter-reader agreement, ICC values less than 0.5, between 0.5 and 0.75, between 0.75 and 0.9, and greater than 0.90 are indicative of poor, moderate, good, and excellent reliability, respectively.p-value of ≤0.05 was considered statistically significant. All the statistical analyses were performed by using IBM SPSS 26.0 .For each anatomic district involved  a score value was assigned to categorize patients into three different groups: negative , low (1–3), and high (4–6) involvement. Areas under the receiver operating characteristic (AUROC) were calculated, as well as their 95% CIs, and were compared by using the DeLong test. All tests were two-sided, and the By applying the aforementioned inclusion and exclusion criteria, a total of 64 patients were analyzed. The majority was female (M/F = 34/29), with a mean age of 65 years .All MRI examinations were considered diagnostic from both readers, without any significant artifacts.p < 0.001). By grouping lesions’ distribution according to the anatomic district, the two readers showed a very good agreement for spine, skull, sternum and ribs, and pelvis , and perfect or almost perfect agreement for upper and lower limbs . All agreement values with 95% CIs are reported in The overall agreement between readers was very good regarding the pattern (κ = 0.954 (95% CIs: 0.885–1), Considering the good agreement between the two readers, only the results of the most experienced one were used for further analysis.n = 49/64, 76.6%), the majority showed a focal pattern , followed by combined , and diffuse  ones.A total of 15/64 patients (23.4%) showed no lesions in all anatomic districts. Between patients with bone involvement  , sternum and ribs , upper limbs , skull , and lower limbs . Overall, the most common involved district was the spine  , followeTo divide patients into different classes, we assigned one point for each considered district involved by focal lytic lesions, and the final score was computed by summing up each value. In this setting, the final score ranged between 0 and 6 points: “0” stands for patients negative for lytic lesions, while “6” indicates whole-body involvement.n = 17/49 (26.6) and n = 14/49 (21.9), respectively), while only 18/49 (28.1) were categorized in classes 3, 4, and 5 (n = 8 (12.5), n = 7 (11.1), and n = 3 (4.7), respectively). The standard score presented a good sensitivity and specificity (89.8 (95% CIs: 77.8–96-6) and 66.7 (95% CIs: 38.4–88-2), respectively), and an overall good diagnostic accuracy .By using the standard protocol —SP, mostn = 19/49 (29.7) and n = 13/49 (20.3), respectively), while only 17/49 (26.6) were categorized in classes 3, 4, and 5 (n = 9 (14.1), n = 6 (9.4), and n = 2 (3.1), respectively). Overall, this score reported a good sensitivity and specificity (89.8 (95% CIs: 77.8–96.6) and 66.7 (95% CIs: 38.4–88.2), respectively) with a slightly lower diagnostic accuracy .By analyzing the short protocol (shP) removing the values regarding the lower limb involvement —shP1, thn = 25/49, 39.1), followed by 2 , 3 and 4 (n = 7 (10.9) and n = 1 (1.6), respectively). This short protocol showed 81.7% sensitivity (95% CIs: 67.9–91.3) and 73.3% specificity (95% CIs: 44.9–92.2), with a good diagnostic accuracy .By analyzing the short protocol removing the values regarding the skull —shP2, thn = 20/49, 31.3), followed by 2, 3, and 4 (n = 14 (21.9), n = 8 (12.5), and n = 7 (10.9), respectively), showing 89.9% sensitivity (95% CIs: 77.8–96.6) and 66.7% specificity (95% CIs: 38.4–88.2), with a more than acceptable diagnostic accuracy .Finally, excluding both skull and lower limb involvement —shP3, mop = 0.209 and p = 0.141, respectively), while a significant difference was found in the comparison between SP and shP2 (p = 0.031).The pairwise comparison between ROC curves showed no statistically significant differences between standard protocol (SP) and short protocols 1 and 3 (shP1 and shP3) , 32 min and 10 s (±2′10″), 35 min and 40 s (±2′05″), and 28 min and 20 s (±2′15″), respectively.Whole-body MRI (WBMRI) is becoming increasingly relevant for the assessment of patients with MM, due to complete body coverage, excellent sensitivity for bone marrow involvement before or without bone destruction , and availability of advanced techniques such as diffusion-weighted imaging (DWI) and Dixon-based fat-fraction evaluation ,14,15.According to the current literature, WBMRI is considered a valid option for bone marrow imaging by the European Society for Medical Oncology guidelines, and in the UK, it is recommended as first-line imaging for all patients with a suspected new diagnosis of myeloma ,17.Furthermore, the European Myeloma Network guidelines also recommend WBMRI in asymptomatic SMM (smoldering multiple myeloma) patients with no detectable lytic disease on CT , at initIn our series, WBMRI was used to analyze the skeletal and bone marrow involvement in 64 patients with a diagnosis of MM, which was classified into four different infiltration patterns : The majority of patients showed skeletal involvement by bone lytic lesions with both focal and combined patterns, in line with the literature . Only a When analyzing the anatomic distribution of bone marrow involvement in the focal and combined pattern of disease, our results showed that the lytic lesions were predominantly distributed in the spine and the pelvis , in line with previous papers ,22,23. MThe anatomic districts less frequently involved were the skull, the lower limbs , and the upper limbs. Overall, 90.6% of patients did not show any focal bone lesion in the skull or in the lower limbs. Considering the number of lesions recorded, in the skull, no patients showed >20 lesions and only 3% of patients had between 5 and 20 focal lesions.When analyzing the focal lesions in the lower limbs, the total amount of lesions was considerably lower; in fact, no patients with focal involvement of the lower limbs presented >5 lesions in this anatomic district. The obtained results highlighted that most patients with bone lesions in the skull and lower limbs presented >20 focal lesions disseminated in other anatomic districts, being classified in stage III of the Durie and Salmon staging system.Therefore, we may assume that bone lesion detection in these two anatomic districts can have negligible relevance for staging, prognosis, and treatment evaluation.After a careful evaluation of the results, we tried to optimize the standard protocol of WBMRI (from vertex to feet), by shortening it, maintaining adequate sensitivity, specificity, and diagnostic accuracy. Over the past decade, there has been an increased interest in short MRI protocols, aimed to preserve the diagnostic potential while reducing the scanning time and the potential discomfort.In the simulated short protocols, we excluded from the standard protocol the two anatomic districts less involved by bone lesions: skull and lower limbs .The obtained results showed that the short MRI protocols excluding the skull (shP2) or the lower limbs (shP1) presented in both cases a good sensitivity and specificity  and a sufficient diagnostic accuracy  if compared with a standard protocol (SP). Moreover, the shortest MRI protocol, which excludes both skull and lower limbs (shP3), along with good sensitivity and specificity , reached a more than acceptable diagnostic accuracy (88.1%).p > 0.05), highlighting the sufficient efficacy of the shortest protocol (shP3), which might be considered in the clinical practice, in particular for the follow-up.Results in terms of accuracy did not show a statistically significant difference among the MRI short protocols proposed (Excluding both the abovementioned anatomic districts (skull and lower limbs) the MRI protocol allows to save about 15 min in comparison to the standard protocol, thus representing a more acceptable and comfortable diagnostic option for patients with multiple myeloma.Some limitations should be noted, first of all, the retrospective design of the study, and secondly, the small size of the cohort studied, due to the strict inclusion criteria.In conclusion, a short WBMRI protocol, excluding the anatomical regions of the skull and the lower extremities, could represent a reliable diagnostic imaging tool for the evaluation of MM patients, shortening the duration of MRI examination, maintaining an overall good sensitivity, specificity, and diagnostic accuracy and also reducing the patient’s potential discomfort.


Despite global vaccination efforts, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to evolve and spread globally. Relatively high vaccination rates have been achieved in most regions of the United States and several countries worldwide. However, access to vaccines in low- and mid-income countries (LMICs) is still suboptimal. Second generation vaccines that are universally affordable and induce systemic and mucosal immunity are needed. Here we performed an extended safety and immunogenicity analysis of a second-generation SARS-CoV-2 vaccine consisting of a live Newcastle disease virus vector expressing a pre-fusion stabilized version of the spike protein (NDV-HXP-S) administered intranasally (IN), intramuscularly (IM), or IN followed by IM in Sprague Dawley rats. Local reactogenicity, systemic toxicity, and post-mortem histopathology were assessed after the vaccine administration, with no indication of severe local or systemic reactions. Immunogenicity studies showed that the three vaccination regimens tested elicited high antibody titers against the wild type SARS-CoV-2 spike protein and the NDV vector. Moreover, high antibody titers were induced against the spike of B.1.1.7 , B.1.351 (beta) and B.1.617.2 (delta) variants of concern (VOCs). Importantly, robust levels of serum antibodies with neutralizing activity against the authentic SARS-CoV-2 USA‐WA1/2020 isolate were detected after the boost. Overall, our study expands the pre-clinical safety and immunogenicity characterization of NDV-HXP-S and reinforces previous findings in other animal models about its high immunogenicity. Clinical testing of this vaccination approach is ongoing in different countries including Thailand, Vietnam, Brazil and Mexico. The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has afflicted over 250 million people worldwide and has caused the death of at least 5 million people WHO, 2021). To mitigate the burden caused by this virus, diverse countermeasures have been established worldwide, including social distancing, use of face masks, and more recently the rollout of different vaccines . To miti. RecentlWhile there are a handful of vaccines widely available in the United States and many regions worldwide, the number of vaccine doses available and the vaccination rates in places with limited resources is still suboptimal. Particularly, large parts of Africa, Asia, and some countries of Latin-America display poor vaccination rates , 19. MorDifferent COVID-19 vaccines are being currently developed, including: a) additional/alternative mRNA vaccines, b) additional whole virus inactivated vaccines, c) live virus attenuated vaccines, d) recombinant protein (RBD or spike) based vaccines, e) virus like particle (VLP) based vaccines, f) DNA-based vaccines, and g) additional vector-based vaccines. Within this last category, replication competent, replication incompetent and inactivated vector-based vaccines are in development and replication incompetent vectors are authorized already . Here wehttps://www.beiresources.org/). Protein was purified using gravity flow purification with Ni-nitrilotriacetic acid (NTA) agarose (Qiagen) and concentrated and buffer exchanged in Amicon centrifugal units (EMD Millipore). The purified recombinant proteins were analyzed via reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The desired protein folding was confirmed through ELISAs using the Receptor Binding Domain (RBD)-specific monoclonal antibody CR3022 (Live NDV-HXP-S was produced under current Good Manufacturing Practices (cGMP) regulations in the Vaccine and Cell Therapy Laboratory (VCTL) facility (Icahn School of Medicine at Mount Sinai) as previously described . Wild tyy CR3022 . For detad libitum-pelleted rodent food), except during study procedures or activities. The rats were placed on the Certified Rodent Diet® #5002  and fed pellets ad libitum, except during designated procedures.Animal experiments were performed at an animal facility of Charles River Laboratories, Inc . The study was performed in accordance with the U.S. Department of Health and Human Services, Food and Drug Administration, United States Code of Federal Regulations, Title 21, Part 58: Good Laboratory Practice for Nonclinical Laboratory Studies and as accepted by Regulatory Authorities throughout the European Union (OECD Principles of Good Laboratory Practice), Japan (MHLW), and other countries that are signatories to the OECD Mutual Acceptance of Data Agreement. Nine- to ten- week old Sprague Dawley rats were used for all the experiments. Rats were co-housed in solid-bottomed cages by dose group, placing a maximum of 2 rats of the same sex and dose group per cage. Rats from the control group were housed separately, on a different rack, from the rats in the treatment group. The rats were socially housed, provided with Crink-l’ Nest™, a resting platform, and a chewing item  for a total of 100 µL per naris per rat using a pipette with the appropriately sized tip. For intramuscular administration, 100 µL of vaccine or saline (control) were injected into the quadriceps of both hind legs for a total of 200 µL per rat. At the initial dosing, males had body weight ranging from 278 to 400 grams and females 216 to 284 grams. Blood samples (approximately 1 mL) were collected from each rat via the retro-orbital plexus on days 1, 15, and 17 for the ‘main group’ and days 1, 15, and 30 for the ‘recovery group’. Blood samples were centrifuged at 3,000 g for 10 min to separate the serum, then stored and transported under frozen conditions (-60 to -80°C) to the Icahn School of Medicine at Mount Sinai where the immunogenicity analyses took place.After a period of acclimation of at least 6 days, rats were separated to perform two different vaccination experiments. A ‘main’ group, in which assessments were performed on days 1, 15, and 17 after the first vaccine dose administration, and a ‘recovery group’ in which rats were followed for a 12-day period after the administration of the second vaccine dose. In both experiments, 10 males and 10 females in each treatment group were administered 200 µL of live NDV-HXP-S vaccine intranasally (IN), intramuscularly (IM), or The potential toxicity and local site reactogenicity of the NDV-HXP-S vaccine were assessed to examine the vaccine’s impact on the overall health of the rats. To this end, different groups of toxicological parameters, including: a) clinical observations, b) clinical pathology, and c) post-mortem evaluations, were examined.a) Clinical observations consisted of the visual inspection of the injection sites and the area surrounding the intranasal administration, measurements of body temperature, body weight, food intake over two different week periods, ophthalmological examinations, and mortality. For the injection site observations and intranasal evaluations, each rat was evaluated at the administration site on a minimum of four separate occasions, including the day of initial administration (before dosing), and 24 ± 1, 48 ± 1, and 72 ± 1 hours after the vaccine administration. The following score criteria were used (0): no evidence of nasal symptoms (1), nasal rattling or sneezing (2), nasal discharge on external naris, and (3) evidence of mouth breathing. If a non-zero score was recorded at the 72-hour post-dose evaluation, scoring was continued daily (every 24 ± 1 hours) until the observations were resolved or the rat was euthanized. In rats dosed intramuscularly the levels of erythema, edema, atonia, desquamation, and fissuring were scored under the following criteria (0): no symptoms (1), slight symptoms (2), moderate symptoms, or (3) severe symptoms. Scab formation was indicated with an “F” (focus/foci present) or “P” (patches present), while the presence of eschar tissue or necrotic tissue was indicated with a “N”. Ophthalmologic evaluations were performed by board-certified veterinary ophthalmologists at The Animal Eye Center of New Jersey.b) Clinical pathology parameters consisted of hematology, coagulation, clinical chemistry, and urinalysis measurements. Before testing hematology parameters, any blood samples containing clots were discarded from the analysis. The measurements evaluated included red and white blood cell count, red blood cell distribution width, hemoglobin concentration, hematocrit count, mean corpuscular volume, mean corpuscular hemoglobin count and concentration, platelet count, absolute reticulocyte, neutrophil, lymphocyte, monocyte, eosinophil, basophil, and large unstained cell counts. The coagulation parameters were measured using plasma samples produced by transferring blood into tubes containing sodium citrate  and centrifuging the tube within 30 minutes of collection at room temperature for 15 minutes or more. The coagulation of the blood samples was assessed by measuring activated partial thromboplastin and prothrombin times and fibrinogen counts. Clinical chemistry parameters were tested on sera produced by first allowing the blood samples to clot for 20 to 60 minutes in serum separator tubes before being centrifuged at room temperature for at least 15 minutes. These sera were tested for measures of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, gamma-glutamyltransferase, creatine kinase, total bilirubin, urea nitrogen, creatinine, calcium, phosphorus, total protein, albumin, calculated globulin, albumin-to-globulin ratio, glucose, cholesterol, triglycerides, sodium, potassium, and chloride. Finally, urine samples from rats were collected overnight on the day before euthanasia. These urine samples were then evaluated on their color, appearance and clarity, specific gravity, volume, pH, and levels of protein, glucose, bilirubin, ketone, and blood present.c) Post-mortem evaluations consisted of a complete necropsy examination after rats were euthanized by exsanguination. This examination included evaluations of the carcass and musculoskeletal system, external surfaces and orifices, cranial cavity, external surfaces of the brain, and thoracic, abdominal, and pelvic cavities with their associated organs and tissues. In addition, organ weights were measured, and histological examinations of tissue specimens were performed. The tissues utilized for histology were embedded in paraffin, sectioned, mounted on glass slides, and stained with hematoxylin and eosin. Microscopic changes were assessed by microscopy and performed by a board-certified veterinary pathologist.o-phenylenediamine dihydrochloride  for 10 min. The reaction was stopped by adding 50 µL per well of 3M hydrochloric acid . Optical density was measured at 490 nm using a plate reader . Analyses were performed using Microsoft Excel and Prism 9 (GraphPad). Antibody levels were expressed as area under the curve (AUC).Serological testing against SARS-CoV-2 proteins was performed using enzyme-linked immunosorbent assays as previously described . Briefly50) of USA-WA1/2020 for 1 hour at 37°C, 5% CO2 and 120μl of the sera-virus mix were transferred in a mirror-fashion to Vero-E6 plates. Sera-virus mix was removed and additional serum at the same dilution was added to the cells and plates were incubated for 48h at 37°C.Rat sera collected 15 days after administration of the second vaccine dose was used to assess the neutralization of SARS-CoV-2 USA‐WA1/2020 as previously described  with som50), with top and bottom constraints set to 100% and 0% respectively.For staining of the SARS-CoV-2 nucleoprotein (NP) antigen, cells were fixed overnight at 4°C with 200 μl/well of a 10% formaldehyde solution. Plates were washed with PBS (pH 7.4) (Gibco) and permeabilized by adding 150 μl/well of PBS, 0.1% Triton X-100 for 15 min at RT. For blocking, plates were washed with PBS and incubated with PBS, 3% bovine serum albumin (BSA) for 1h at RT. During this time the mAb 1C7C7 (anti-SARS nucleoprotein antibody produced at the Center for Therapeutic Antibody Development at The Icahn School of Medicine at Mount Sinai ISMMS) was biotinylated according to the manufacturer’s instructions (Thermo Scientific EZ-Link NHS-PEG4-Biotin). Blocking solution was discarded and 100 μl/well of biotinylated mAb 1C7C7 at a concentration of 2 µg/ml in PBS, 1% BSA was added for 2h at RT. Plates were washed twice with PBS and 100 μl/well of streptavidin HRP (Thermo Scientific) diluted in PBS, 1% BSA were added at a 1:3000 dilution and plates were incubated for 1h at RT. Plates were washed twice with PBS, and 100 μl/well of OPD  were added for 10 min at RT, followed by addition of 50 μl/well of a 3M HCl solution (Thermo Fisher Scientific). Optical density (OD) was measured (490 nm) using a microplate reader . Analysis was performed using Prism 9 software (GraphPad). After subtraction of background and calculation of the percentage of neutralization with respect to the “virus only” control, a nonlinear regression curve fit analysis was performed to calculate the inhibitory dilution 50% . A regular two-way analysis of variance (ANOVA) with Tukey multiple comparisons test was used. All adjusted P values of <0.05 were considered statistically significant with a confidence interval of 95%.8.87 50% egg infectious dose (EID50) regardless of the administration route  ectodomain from SARS-CoV-2 and the transmembrane domain and cytoplasmic tail (TM/CT) of the NDV fusion protein , 30. TheDifferent parameters were assessed before and after vaccination to determine the safety and immunogenicity of NDV-HXP-S in Sprague Dawley rats. Immunogenicity analyses, comprised of antibody binding and neutralization assessments were performed using sera collected at days 0, 15 and 30. Clinical indicators of overall health, such as weight assessments, were performed on days 0, 1, 2, 8, 15, 16, and 17 post-vaccination, while body temperature (°C) was measured at 0-, 6- and 24-hours post-dosage, either after the prime or the boost vaccine doses. Clinical pathology assessments were performed on days 2, 17, and 30 following the first vaccination. Finally, rats were euthanized on day 17 and day 30 for the main and recovery groups, respectively  of the viral polymerase inhibitor remdesivir, is presented as a control . The study was performed in accordance with the U.S. Department of Health and Human Services, Food and Drug Administration, United States Code of Federal Regulations, Title 21, Part 58: Good Laboratory Practice for Nonclinical Laboratory Studies and as accepted by Regulatory Authorities throughout the European Union (OECD Principles of Good Laboratory Practice), Japan (MHLW), and other countries that are signatories to the OECD Mutual Acceptance of Data Agreement.Conceptualization, FK, PP, AG-S, WS, and JC. Investigation, JC, JT, AR, GS, HK, DB, WS, IG-D, and DS. Data curation, JT, AR, and JC. Formal analysis, JC, JT, and AR. Supervision, JC and FK. Writing - original draft, JC. Writing - review and editing: all authors. All authors contributed to the article and approved the submitted version.This work was mostly funded by institutional support from the Icahn School of Medicine at Mount Sinai with partial support by the Centers of Excellence for Influenza Research and Surveillance  and the Centers of Excellence for Influenza Research and Response , the Collaborative Influenza Vaccine Innovation Centers (CIVIC) contract #75N93019C00051, the JPB Foundation, the Open Philanthropy Project (research grant 2020-215611 (5384)) by anonymous donors.The Icahn School of Medicine at Mount Sinai has filed patent applications relating to NDV-based SARS-CoV-2 vaccines which list WS, FK, AG-S and PP as co-inventors. FK is also listed as inventor on patent applications for SARS-CoV-2 serological assays filed by the Icahn School of Medicine at Mount Sinai. Mount Sinai has spun out a company, Kantaro, to market serological tests for SARS-CoV-2. FK has consulted for Merck and Pfizer (before 2020) as well as Goldman Sachs, and is currently consulting for Pfizer, Seqirus and Avimex. The Krammer laboratory is also collaborating with Pfizer on animal models of SARS-CoV-2. The Adolfo García-Sastre laboratory has received research support from Pfizer, Senhwa Biosciences, Kenall Manufacturing, Avimex, Johnson & Johnson, Dynavax, 7Hills Pharma, Pharmamar, ImmunityBio, Accurius, Nanocomposix, Hexamer, N-fold LLC, Model Medicines and Merck, outside of the reported work. AG-S has consulting agreements for the following companies involving cash and/or stock: Vivaldi Biosciences, Contrafect, 7Hills Pharma, Avimex, Vaxalto, Pagoda, Accurius, Esperovax, Farmak, Applied Biological Laboratories, Pharmamar, and Pfizer.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


ESR1 mediating 25 interactions. Several genes  regulated by β-estradiol/ESR1 were highly expressed in non-glaucomatous human trabecular meshwork (TM) or Schlemm’s canal (SC) cells and specifically expressed in TM/SC cell clusters defined by single-cell RNA-sequencing. We confirmed ESR1 gene and protein expression in human TM cells and TM/SC tissue with quantitative real-time PCR and immunofluorescence, respectively. 17β-estradiol was identified in bovine, porcine, and human aqueous humor (AH) using ELISA. In conclusion, we have identified estrogen receptor signaling as a key modulator of several IOP-associated genes. The expression of ESR1 and these IOP-associated genes in TM/SC tissue and the presence of 17β-estradiol in AH supports a role for estrogen signaling in IOP regulation.Elevated intraocular pressure (IOP) is the only modifiable risk factor for primary open-angle glaucoma (POAG). Herein we sought to prioritize a set of previously identified IOP-associated genes using novel and previously published datasets. We identified several genes for future study, including several involved in cytoskeletal/extracellular matrix reorganization, cell adhesion, angiogenesis, and TGF-β signaling. Our differential correlation analysis of IOP-associated genes identified 295 pairs of 201 genes with differential correlation. Pathway analysis identified β-estradiol as the top upstream regulator of these genes with  Glaucoma is the leading cause of irreversible blindness in the world, impacting the lives of approximately 75 million individuals ,2,3. PriAlthough its etiology is not completely understood, several risk factors have been identified, including age, African ancestry, positive family history, and elevated intraocular pressure (IOP) ,7,8,9,10In order to identify genetic factors that contribute to IOP regulation and POAG risk, many genome-wide association studies (GWAS) have been conducted in recent years ,24,25,26TMCO1, FNDC3B, AFAP1, GMDS, OXR1, ANGPT1, LMX1B, and GAS7). Alternatively, some SNPs were located between genes; in such cases, genes on either side of the SNP were included. Furthermore, many genes were identified by both studies. For this reason, the final list of IOP-associated genes included only 157 non-redundant genes  . Ad. AdTES, fication , Table 6 factors ,67. Integlaucoma ,69,70. Lubstrate .AFAP1, ANTXR1, COL4A1, COL8A2, EFEMP1, ETS1, LTBP2, PCSK5, PTPN1, and SEMA3E) also play roles in cytoskeletal reorganization, extracellular matrix (ECM) assembly, and/or cell adhesion, especially through focal adhesions [ETS1  is responsive to both fluid shear [AFAP1 is necessary for c-Src-mediated mechanotransduction [Many other prioritized genes  was angiogenesis, blood vessel morphogenesis, and endothelial cell regulation [However, IOP-associated genes may affect IOP regulation through several other processes. For example, another key biological process shared by several prioritized genes  that are both regulated by estrogen and are associated with IOP. Perhaps these genes and the common biological processes they share can illuminate our understanding of estrogen’s impact on IOP dynamics. However, despite the plethora of data evidencing that estrogen signaling plays an important role in IOP homeostasis, the mechanism of this effect has not been fully explored. Here we present 12 genes  are involved in angiogenesis, endothelial cell function, or other aspects of the vasculature [FOXC1 and VEGFC are also involved in lymphangiogenesis [The majority of these estrogen-regulated, IOP-associated genes  include cytoskeletal reorganization, ECM turnover, and cell adhesion [CAV2 has been shown to be essential for remodeling TM ECM [CAV1/2 have been identified as being more strongly associated in women [CAV2.As seen with the prioritized genes discussed previously, other common biological processes shared by estrogen-regulated genes  regulated by estradiol/ESR1 were also regulated by dexamethasone, suggesting that the two molecules may affect similar processes. Dexamethasone is known to elevate IOP by increasing TM cell and ECM stiffness, thereby increasing outflow resistance [To further elucidate how estrogen signaling may contribute to IOP homeostasis, it may be helpful to note that several of the genes  and SC cells (n = 2) and stretched TM cells (n = 5) [n = 4) used for RNA-Seq by Carnes, et al. [ESR1 expression in male human TM cells and male human TM/SC tissue. Fifth, due to the small number of samples, human AH estradiol concentrations were not able to be statistically analyzed on the basis of sex. Furthermore, the biological sex of bovine and porcine AH was unknown although it was assumed to be male due to commonly accepted abattoir practices. Despite these limitations, including multiple datasets with different approaches and both cells and tissue provides rigor for our prioritization study and ensures that genes of interest have several sources of support.Despite the rigor of consulting multiple data sources, a few limitations must be noted for this study. First, genes were selected based on their proximity to the SNPs identified by GWAS. However, the SNPs in question may occur in regulatory elements for genes much further downstream or upstream. Second, a few of the datasets were limited by small sample sizes. Our own RNA-Seq sample sets of non-glaucomatous TM ( (n = 5)  were sma, et al. . In addiIn the future, additional transcriptional studies with larger sample sizes and more standardized approaches may be consulted to further support our findings. In addition, functional assays may be conducted to determine whether associated SNPs impact IOP regulation through our genes of interest and whether these effects on IOP have pathological consequences. Furthermore, significant effort should be directed towards elucidating the mechanism by which estrogen signaling plays a role in IOP regulation.In summary, we have prioritized a set of 157 genes containing/neighboring IOP-associated SNPs. As a result, we have identified several genes of interest for future functional study as well as identified estradiol as a key regulator of several IOP-associated genes. Therefore, we postulate a role for estrogen signaling in the normal maintenance of IOP homeostasis. Although much evidence for estrogenic effects on IOP regulation has accumulated over the years, the mechanism of action has not been well studied. Here we present a group of genes that are both associated with IOP and regulated by estrogen, thereby offering insight into the mechanisms by which estrogen may impact IOP. Given the function of the estrogen-regulated, IOP-associated genes, we hypothesize that estrogen may mediate its effects on IOP through dilation of the SC/distal vessels and/or through modulation of TM ECM and cellular morphology. The confirmed expression of ESR1 in TM/SC tissue and the presence of estradiol in AH further supports this hypothesis.p-value  [http://genome.ucsc.edu/cgi-bin/hgGateway, accessed on 3 December 2018; ©2000–2018 The Regents of The University of California) [https://gtexportal.org/, accessed on 3 December 2018; ©2021 Broad Institute of MIT and Harvard; V6P release with 544 donors, GENCODE version v19, GRCh37/hg19) [A series of additional filtering criteria were then applied to the 24 genes that were differentially expressed in human TM tissue or SC cells. First, expression of the 10 genes differentially expressed in POAG-affected TM tissue  was deteof Iowa)  was deteof Iowa) . In brieifornia)  was empl37/hg19) ,32,33,34p < 0.05 were considered to be significantly differentially expressed  [Lastly, the specific expression of all 157 IOP-associated genes  was examch 2021)  and applP) of differences between gene-gene correlations in glaucomatous and non-glaucomatous TM tissue. In this way, gene-gene correlations identified in non-glaucomatous tissue that changed significantly  in glaucomatous tissue could be identified, and vice versa. R’s Benjamini-Hochberg procedure function (p.adjust with “BH” parameter) was then used to calculate the FDR from the p-value. Duplicate correlations were removed, and gene-gene correlations with FDR ≤ 0.01 were considered significant and retained for further analysis.Microarray expression data  was usedAll significantly correlated genes were input into Cytoscape   in orderTM Diamond Antifade Mountant with DAPI  from Invitrogen . Mouse anti-ESR1  and CyTM3 goat anti-mouse IgG  antibodies were purchased from Invitrogen and Jackson ImmunoResearch Laboratories, Inc. , respectively  were obtained from Thermo Fisher Scientific , High Sensitivity DNA kits  from Agilent Technologies , and both RiboGone Mammalian kits  and SMARTer Stranded Total RNA Low Input Mammalian Sample Prep kits  from TaKaRa Bio USA, Inc. . RNeasy Mini kits  were purchased from Qiagen  and High-Capacity cDNA Reverse Transcription kits with RNase Inhibitor  were purchased from Applied Biosystems . 17β-estradiol ELISA kits  were obtained from Enzo Life Sciences . All primers were obtained from Integrated DNA Technologies  and nonglaucomatous (n = 4) AH samples were obtained from the Eye Clinic at Augusta University Medical Center. The study was approved by the Institutional Review Board at Augusta University. Written consent was collected from all individuals. Samples were collected from male and female patients undergoing cataract or glaucoma surgery (n = 5) and porcine (n = 4) AH were obtained from the local abattoir. After performing a liquid-liquid extraction of 17β-estradiol with 5:1 (v/v) diethyl ether:sample, the presence of 17β-estradiol in these samples was confirmed with enzyme-linked immunosorbent assay  following the manufacturer’s protocol. Optical density was determined at 405nm using an Infinite M200 Pro plate reader .Glaucomatous  for 30 minutes of darkness at room temperature. ProLongTM Diamond Antifade Mountant  was used to fix coverslips before imaging with an Axio Imager D.2 microscope  and Zeiss Zen 2.3 pro software .Discarded cornea rims n = 4;  were obtThe filtering analyses described in


Pancreatic ductal adenocarcinoma is a highly lethal malignancy, which has now become the seventh most common cause of cancer death in the world, with the highest mortality rates in Europe and North America. In the past 30 years, there has been some progress in 5-year survival (rates increasing from 2.5 to 10%), but this is still extremely poor compared to all other common cancer types. Targeted therapies for advanced pancreatic cancer based on actionable mutations have been disappointing, with only 3–5% showing even a short clinical benefit. There is, however, a molecular diversity beyond mutations in genes responsible for producing classical canonical signaling pathways. Pancreatic cancer is almost unique in promoting an excess production of other components of the stroma, resulting in a complex tumor microenvironment that contributes to tumor development, progression, and response to treatment. Various transcriptional subtypes have also been described. Most notably, there is a strong alignment between the Classical/Pancreatic progenitor and Quasi-mesenchymal/Basal-like/Squamous subtype signatures of Moffit, Collinson, Bailey, Puleo, and Chan-Seng-Yue, which have potential clinical impact. Sequencing of epithelial cell populations enriched by laser capture microscopy combined with single-cell RNA sequencing has revealed the potential genomic evolution of pancreatic cancer as being a consequence of a gene expression continuum from mixed Basal-like and Classical cell populations within the same tumor, linked to allelic imbalances in mutant KRAS, with metastatic tumors being more copy number-unstable compared to primary tumors. The Basal-like subtype appears more chemoresistant with reduced survival compared to the Classical subtype. Chemotherapy and/or chemoradiation will also enrich the Basal-like subtype. Squamous/Basal-like programs facilitate immune infiltration compared with the Classical-like programs. The immune infiltrates associated with Basal and Classical type cells are distinct, potentially opening the door to differential strategies. Single-cell and spatial transcriptomics will now allow single cell profiling of tumor and resident immune cell populations that may further advance subtyping. Multiple clinical trials have been launched based on transcriptomic response signatures and molecular subtyping including COMPASS, Precision Promise, ESPAC6/7, PREDICT-PACA, and PASS1. We review several approaches to explore the clinical relevance of molecular profiling to provide optimal bench-to-beside translation with clinical impact. Globallper year . The numectively . In Euroectively . In Nortectively . For Wes cancer) . In 2017 cancer) . In the  cancer) .Most patients present with metastatic disease, with only 10–20% being diagnosed with localized pancreatic cancer that can be surgically removed, while the remaining 20–30% have non-metastatic locally advanced disease that cannot be removed by standard surgical techniques . SystemiThe evolution of molecular targeted therapies, aimed at advancing tumor control and cell killing of pancreatic cancer, has so far met with only very limited progress . While sMutations in genes responsible for producing classical canonical signaling pathways including driver oncogenes and dysfunction of tumor suppressor genes include KRAS, TP53, CDKN2A, and SMAD4 in most cases, and ARID1A, KDM6A, MLL3, TGFBR2, RBM10, BCORL1, and ROBO2 in 5–10% of tumors . Genetic•Apoptosis: CASP10, VCP, CAD, HIP1•DNA damage control: ERCC4, ERCC6, EP300, RANBP2, TP53, BRCA1/2, PALB2, ATM, ATR, MLH1, MSH2, MSH6, RPA1, STK11, FANCA, FANCC•Regulation of G1/S phase transition: CDKN2A, FBXW7, CHD1, APC2•Hedgehog signaling: TBX5, SOX3, LRP2, GLI1, GLI3, BMPR2, CREBBP•Homophilic cell adhesion: CDH1, FAT, PCDH15, PCDHB16, PCDHGA1•Integrin signaling: ITGA4, LAMA1, LAMA4, LAMA5, FN1, ILK•c-Jun N-terminal kinase signaling: MAP4K3, TNF, ATF2, NFATC3•KRAS signaling: KRAS, MAP2K4, RASGRP3•Regulation of invasion: ADAM11, DPP6, MEP1A, PCSK6, APG4A•Small GTPase–dependent signaling: AGHGEF7, ARHGEF9, CDC42BPA•TGF-β signaling: TGFBR2, BMPR2, SMAD4, SMAD3•Wnt/Notch signaling: MYC, PPP2R3A, WNT9A, MAP2, TSC2, GATA6.Single genetic alterations occur in <5% of tumors, notably BRCA1/2 mutations, BRAF gene fusions/mutations, ERBB2 amplifications/mutations, RNF43, TGFBR2, MAP2K4, MLL3, PIK3CA, RBM10, SMARCA4, PBRM1, SLIT2, KDM6A, GATA6, BRAF, ATM, and mismatch repair (MMR) gene mutations . MMR genClinical applicability of genetic biomarkers has been classified by the European Society for Medical Oncology (ESMO) Translational Research and Precision Medicine Working Group into the ESMO Scale of Clinical Actionability for molecular Targets (ESCAT) . There amFOLFIRINOX –preferred for known germline BRCA1/2 or PALB2 mutations.Olaparib – a PARP inhibitor as maintenance therapy in patients who have a germline BRCA1 or BRCA2 mutation and with metastatic pancreatic cancer that had not progressed during first-line platinum-based chemotherapy, resulting in improved progression-free survival.Entrectinib – an inhibitor of tropomyosin receptor kinases (TRKs) of tumors with NTRK or ROS-1 gene fusions.Laroctrenib – an inhibitor of tropomyosin receptor kinases (TRKs) of tumors with NTRK gene fusions.Afatinib – an EGFR tyrosine kinase inhibitor in KRAS wild-type tumors with NRG1 gene fusions.Sotorasib – a small molecule that targets the KRAS p.G12C mutation that is present in 1–2% of PDAC patients  used with gemcitabine, produces an improved survival in metastatic pancreatic cancer, but this benefit is only marginal with increased toxicity.Structural variations amongst the 25,000 defined human genomes include deletions, amplifications, duplications, and translocations . The WadStable (20%), with <50 structural variations per genome, with widespread aneuploidy.Locally rearranged (30%), with >200 structural variants clustered on 1–2 chromosomes. Of these, about a 35% had focal amplifications in KRAS, SOX9 and GATA6, as well as ERBB2, MET, CDK6, PIK3CA, and PIK3R3 but were only present in 1–2% of patients. The remaining local rearrangements involved complex genomic events such as breakage–fusion–bridge or chromothripsis .Scattered (36%), with non-random chromosomal damage in 50–200 structural variants per genome.Unstable (14%), with >200 structural variants distributed across the genome indicating defects in DNA maintenance  and a mutational DNA damage repair (DDR) deficiency, with potential sensitivity to DNA-damaging agents. The unstable structural variation subtype is responsive to platinum therapy and BRCA1/2 germline carriers also sensitive to both platinum and PARP inhibitors.It is estimated that 24% of all pancreatic cancers may be sensitive to platinum therapy based on an unstable genomic structural variation subtype, and/or somatic and germline mutations in BRCA genes, and/or a BRCA-type mutational signature . MSI-H oMSI-H tumors express a large number of neoantigens, potentially rendering them more susceptible to immunotherapy in comparison to those tumors with relatively few mutations. DNA replication stress producing single-stranded DNA will induce DDR of which the DNA damage checkpoint kinase ATR [Ataxia-Telangiectasia Mutated (ATM) and Rad3-related protein kinase] is a critical component. Genes encoding subunits of SWI/SNF (BAF) chromatin remodeling complexes, each composed of approximately 15 protein subunits, include ARID1A, ARID1B, ARID2, PBRM1, SMARCA4, and SMARCB1. ARID1A deficiency will impair cells to recruit topoisomerase 2A to chromatin causing cell cycle defects. The consequence is increased reliance on ATR checkpoint activity and thereby increased sensitivity to ATR inhibitor therapy .Various transcriptional pancreatic cancer subtypes have also been described, most notably the Moffit, Collinson, Bailey, Puleo, and Chan-Seng-Yue signatures amongst others, which have potential clinical impact . Each stExocrine-like: characterized by relatively high expression of tumor cell derived digestive enzyme genes.Classical: demonstrating high expression of adhesion-associated and epithelial genes, and epithelial cell terminal differentiation genes, notably GATA6; KRAS mRNA levels elevated relative to the other subtypes; Classical subtype cell lines are more sensitive to erlotinib.Quasi-mesenchymal: has high expression of mesenchyme associated genes; a relatively high proportion of high-grade tumors and poor patient outcomes; low GATA6 expression; QM-PDA subtype cell lines are relatively more sensitive to gemcitabine than those with the Classical subtype.Classical: characterized by overlapping signature with the genes described in the Collisson classification including GATA6, and overall a better prognosis.Basal-like: associated with a worse prognosis than the Classical subtype but may have a better response to adjuvant therapy.Squamous: is characterized by enrichment for TP53 and KDM6A mutations; upregulation of the TP63ΔN transcriptional network; hypermethylation of pancreatic endodermal cell-fate determining genes and is associated with a poor clinical prognosis.Pancreatic progenitor: is defined by preferential expression of genes involved in early pancreatic development notably FOXA2/3, PDX1, and MNX1 and also by gene programs involved in metabolism.Immunogenic: is classed by the enrichment of genes associated with specific immune cell populations, including T-cells and B-cells.Aberrantly differentiated endocrine exocrine (ADEX): is featured by upregulation of genes that regulate networks involved in KRAS activation, and exocrine (NR5A2 and RBPJL) and endocrine differentiation (NEUROD1 and NKX2-2).The Squamous subtype overlaps with the Quasi-mesenchymal subtype of Collisson but has notable pan-squamous features, including a significant association with adenosquamous PDAC histology . There iThe Pancreatic progenitor subtype has four key characteristics.(i) Transcriptional networks containing transcription factors PDX1, MNX1, HNF4G, HNF4A, HNF1B, HNF1A, FOXA2, FOXA3, and HES1, which are pivotal for pancreatic endoderm cell-fate determination toward a pancreatic lineage and are linked to maturity onset diabetes of the young(ii) Gene programs regulating metabolism notably fatty acid oxidation, steroid hormone biosynthesis, and drug metabolism(iii) O-linked glycosylation of mucins, notably apomucins MUC5AC and MUC1, but not MUC2 or MUC6, that define the IPMN pancreatobiliary subtype with PDAC-associated IPMN clustering(iv) TGFBR2 inactivating mutations.The ADEX subtype was defined by both exocrine and endocrine lineage features in later stages of pancreatic development and differentiation , and could be considered a subclass of Pancreatic progenitor tumors. There are two main transcriptional networks.(i) Acinar cell differentiation and pancreatitis/regeneration, transcription factors NR5A2, MIST1 (BHLHA15A) and RBPJL and their downstream targets.(ii) Endocrine differentiation and maturity onset diabetes of the young, including INS, NEUROD1, NKX2-2, and MAFA.Pure basal-like tumors are composed of poorly differentiated tumors with predominant Gly12Asp and Gly12Val KRAS mutations; they have a low stromal signal.Stroma Activated tumors are moderately differentiated, specifically enriched in the activated stroma component defined by high a-SMA, SPARC, and FAP.Desmoplastic tumors are also moderately differentiated with a predominant basal association, characterized by a low tumoral component and a large stromal transcriptomic signal, including immune and inflammatory stroma components and, particularly, a high expression of structural and vascularized stroma components.Pure-classical and Immune classical tumors are histologically well differentiated with fewer CDKN2A and TP53 mutations than basal-like tumors, and are also enriched with the Gly12Arg KRAS mutation, and associated with hENT1 expression; predicted to be Moffitt–Classical, and Bailey–Progenitor subtypes.Maurer et al. used laser capture microdissection (LCM) epithelial cell enriched samples for mRNA sequencing to profile the expression of 60 matched pairs of human PDAC malignant epithelial and stroma samples . They de(1)Genes used to define the Collisson–Classical, Moffitt–Classical, Moffitt–Basal-like, and Bailey–Progenitor subtypes predominantly provide information about the malignant compartment regardless of the amount of stromal cell infiltration.(2)Genes used to define the Moffitt–Activated, Moffitt–Normal, and Bailey–Immunogenic subtypes report on stromal expression that is largely independent of the malignant compartment (but see below).(3)Gene sets in the Collisson–Quasi-Mesenchymal and Bailey–Squamous subtypes represent a mixture of epithelial and stromal identity, indicative of a more poorly differentiated state.(4)Most genes that define the Collisson–Exocrine and Bailey–ADEX subtypes are largely derived from bulk tumor tissue samples are arguably mostly absent from LCM samples (but see below).Basal-like A: these tumors were associated with the epithelial mesenchymal transition (EMT) program; TP53 gene and TGF-β signaling enriched; 5% of stage I/II (resectable); and 24% of stage IV (metastatic) tumors.Basal-like B: these tumors were associated with the EMT program; TP53 and TGF-β signaling enriched; 9% of stage I/II (resectable), 7% of stage III , and 12% of stage IV (metastatic) tumors.Hybrid: this subtype was found in 24% of stage I/II (resectable), 43% of stage III , and 18% of stage IV (metastatic) tumors.Classical A: this subtype was found in 44% of stage I/II (resectable), 43% of stage III , and 36% of stage IV (metastatic) tumors.Classical B: this subtype was found in 8% of stage I/II (resectable), 7% of stage III , and 10% of stage IV (metastatic) tumors.Ma) favoring the mutant allele over the wild-type allele occurred in only 4% of primary tumors compared to 29% in metastatic disease  compared to metastatic Classical A/B tumors (14%), and KRASMa tumors were also more chemoresistant  both Basal-like and Classical cell populations, and (b) linked to allelic imbalances in mutant (mt) KRAS, with metastatic tumors being more copy number-unstable compared to primary tumors . In prim+) and regulatory T cells (CD4+CD25+FOXP3+ Tregs) are predominant  infiltrate PDACs to activate tissue-specific tumor immunity, inferring another novel immunoregulatory target . Enhancetivation . Tumor iine IL33 . Pre-cliine IL33 . The PD-ine IL33 . Inhibitine IL33 .Targeted therapies for advance pancreatic cancer based on next generation sequencing has been disappointing, with only 3–5% showing any clinical benefit in terms of actionable mutations, and limited to only a few months of additional survival . In real+/HNF1A–) subtype, which was associated with the shortest survival; the Exocrine-like (KRT81–/HNF1A+) subtype which was associated with the longest survival; and the Classical (KRT81–/HNF1A–) subtype, which was associated with intermediate survival -1A and KRT81 that enabled stratification of tumors into different molecular subtypes by using immunohistochemistry . The twosurvival . Exocrinsurvival . CYP3A5 subtypes .+ T-cell, CD8+ T-cell, and Toll-like receptor signaling pathways  and HAVCR2 (TIM3); and PDL-2 (PDCD1LG2) was expressed in all but the Pure classical subtypes, which overall do not up-regulate any of the immune checkpoints  study based in Canada aims to sequence metastatic pancreatic tumors of 400 patients through two clinical trials (COMPASS and PanGen), both of which are generating molecular and phenotypic signatures of individual tumors in a clinically relevant timeframe and related to chemotherapy responses1 .Precision Promise was created by the Pancreatic Cancer Action Network and 15 USA clinical academic sites, in cooperation with the FDA and pharmaceutical partners. Precision Promise is an active adaptive Phase II/III clinical trial platform (NCT04229004) that allows rapid evaluation of novel therapeutic options in patients with metastatic pancreatic cancer. The protocol utilizes an adaptive randomization design and includes several trial designs and statistical innovations. All patients undergo pre- and on-treatment biopsies with state-of-the-art genomic, transcriptomic, and immune analysis, along with collection of blood research samples throughout the study. Focused on both first- and second-line treatment of metastatic PDAC, 30% of patients are randomized to one of the two common control arms (gemcitabine plus nab-paclitaxel and FOLFIRINOX), while 70% of patients are randomized to an experimental treatment arm. The platform currently has one experimental arm open , with two additional experimental arms to be added in 2021. Compared to traditional trial designs, Precision Promise has several advantages: multiple investigational treatments can be evaluated simultaneously using common controls; only 175 patients per experimental arm are required to initiate a regulatory registration, and it is expected that this platform will significantly accelerate the time to evaluate a new therapy, with an anticipated cost savings of 30–50%. With its unique design and novel method of data sharing, Precision Promise serves as a new clinical trial ecosystem to accelerate drug development for PDAC.The ESPAC-6 adjuvant trial in resectable patients and the ESPAC-7 neoadjuvant trial in locally advanced patients are evaluating oxaliplatin- or gemcitabine-based chemotherapy response of PDAC patients that will be randomized according to standard clinical criteria (control arms) or by transcriptomic stratification signatures .The PREDICT-PACA co-clinical trial funded by the German Cancer Aid is being conducted as a biopsy-based approach to predict response to chemotherapy for patients with metastasized pancreatic cancer. The consortium has established robust and highly predictive transcriptomic signatures linked to specific chemotherapy response profiles. Using microfluidic card-based qRT-PCR marker panels, clinical utility of the signatures is validated in a prospective collection of core biopsies from metastases of stage IV PDAC patients that receive one of the current chemotherapies upon clinical decision. In parallel, potential alternative therapies for third-line treatment are identified by high-throughput drug screening using patient-derived organoids and by next-generation sequencing-based detection of actionable mutations.A major area of discussion is whether to discard the tumor microenvironment for molecular classification and replace this with a tumor-cell intrinsic classification system, but this may seem short-sighted as the stroma is a uniquely powerful biological phenomenon in pancreatic cancer. The relationship between molecular subtypes, most notably the immunogenic subtype as described by XZ, KH, PB, CS, SR, TG, MB, and JN prepared the initial drafts of the manuscript which were then critically reviewed by RK, BB, JA, KW, TP, MWB, and TH. All of the authors approved the final version of the manuscript.XZ is in receipt of funding from the Chinese Scholarship Council; PB declares grants from Horizon Europe 2020 (Marie Skłodowska Curie Innovative Training Network); SR declares grants from German Cancer Aid grants (70112720 and 70113167); JN declares grants from the Dietmar Hopp Stiftung GmbH, the Stiftung Deutsche Krebshilfe, and the Heidelberger Stiftung Chirurgie. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


XRD results showed the presence of dopant ions and a crystalline structure. FESEM images showed that the surface morphology of the synthesized nanocomposites formed a rod-like structure. The highlight of this study is the in-situ synthesis of the novel Cu/La/CeO2/GO nanocomposites, which manifest higher photodegradation of harmful organic dyes (Rhodamine B (RhB), Sunset Yellow (SY), and Cibacron Red (CR)). In Cu/La/CeO2/GO nanocomposites, the dopant materials restrict the rapid recombination of photoinduced electron–hole pairs and enhance the photocatalytic activity. The degradation percentages of RhB, SY, and CR dye solution are 80%, 60%, and 95%, respectively. In summary, the synthesized nanocomposites degrade toxic organic dyes with the help of visible light and are suitable for future industrial applications.Environmental pollution is a major threat that increases day by day due to various activities. A wide variety of organic pollutants enter the environment due to petrochemical activities. Organic contamination can be unsafe, oncogenic, and lethal. Due to environmental issues worldwide, scientists and research communities are focusing their research efforts on this area. For the removal of toxic organic pollutants from the environment, photocatalysis-assisted degradation processes have gained more attention than other advanced oxidation processes (AOPs). In this manuscript, we report a novel photocatalysis of copper and lanthanum incorporating cerium oxide (CeO Every living organism in the world depends on water. Moreover, only a small percentage of the freshwater in the world is available for drinking and agriculture . However3) oxidation processes [•, H2O2, O3, O2, and O2•−) [2) photocatalysis for the oxidation of organic compounds has intrinsic advantages over other types of photocatalysis because of its properties [To develop suitable techniques for the degradation and mineralization of dye-polluted wastewater, many techniques are applied to degrade dye effluent, as follows: (1) physical methods, such as membrane filtration, flotation, and sedimentation; (2) biological methods; and (3) chemical methods, such as chemical coagulation, photocatalysis, Fenton or Fenton-like oxidation, and ozone  ,18. In Aoperties . Ceria i2 [4+ ionic state to Ce3+ with the incorporation of oxygen vacancies, makes it a strong contender for use as a catalyst or photocatalyst. Copper oxide (CuO) is a narrow-bandgap (1.2 eV) p-type semiconductor material that shows excellent catalytic properties [+/Cu2+ and Ce3+/Ce4+ results in higher interfacial redox activity, and at the same time, the catalytic ability of Cu-doped CeO2 is changed or enhanced [3+, Eu3+, Sc3+, Gd3+, and La3+, among others. La3+ has important applications in various fields, particularly in electronic devices, photocatalysis, and optical and solid oxide fuel cells. When trivalent La ions are doped into the ceria crystal lattice, the La3+ ions replace Ce4+ ions and create a charge imbalance, which introduces oxygen vacancies into the surface of the material. These surface defects are mainly responsible for the improvement of the photocatalytic properties of the photocatalyst. Moreover, an efficient strategy is to deposit the photocatalysts on conducting substrates to enhance the conductivity and dispersibility [For these applications, the research community constantly seeks to enhance the physicochemical properties of ceria to attain better performance. Ceria is an emerging photocatalyst for dye wastewater treatment, and it has a bandgap of 3.2–3.4 eV, which is similar to the bandgap of TiO2 ,21. Hencoperties ,23,24. Tenhanced . Moreovesibility ,27,28,292 catalysts on graphene is an important solution for engineering the photocatalytic activity of Cu/La/CeO2. To the best of our knowledge, no Cu/La/CeO2/GO-related photocatalytic application has been achieved under visible light to date. In this study, we synthesized Cu/La/CeO2/GO nanocomposites through hydrothermal techniques. The photocatalytic ability of the synthesized nanocomposites was studied using three different organic dyes  under irradiation with visible light. The role of reactive radicals was studied with the help of free radical scavenging techniques. A possible degradation reaction mechanism for RhB, SY, and CR dyes was explored to illustrate the reaction involved in photocatalysis. The results revealed that Cu/La/CeO2/GO nanocomposites have a high potential for degradation of organic compounds under visible-light irradiation.Graphene, as a two-dimensional, hexagonal, single-layered carbon, has attracted attention to non-metal catalysts. Anchoring Cu/La/CeO3)3.6H2O), copper (II) nitrate trihydrate (CuH6N2O9), lanthanum (III) nitrate hexahydrate (La(NO3)3.6H2O), and sodium hydroxide (NaOH) were purchased from Sigma-Aldrich, Taiwan. Any other modifications and purifications were allowed for these materials. Moreover, double-distilled (DI) water was used in all experiments. Cerium (III) nitrate hexahydrate (Ce(NOModified Hummers’ preparation techniques were used to prepare the graphene oxide (GO) . In this2/GO composites. Equal (0.1) molar ratios of copper, cerium, and lanthanum precursor were placed into separate beakers containing 20 mL of DI water. Those solutions were simultaneously stirred magnetically for 30 min to obtain homogenous mixtures. After that, the copper and lanthanum solutions were added dropwise into the cerium solution under magnetic stirring, and 0.2 M NaOH solution was added as a reducing agent. The mixed solution was steadily stirred for 1 h at ambient temperature. After stirring, 100 mg of reduced graphene oxide was added to the above solution, which was further stirred for 1 h The resulting composite solution was transferred to a statin-less, steel-lined autoclave and then heated in a hot-air oven at 120 °C for 12 h. After the hydrothermal process, the autoclave was cooled naturally at room temperature. The Cu/La/CeO2/GO composites were centrifuged and washed with DI water and ethanol several times, and then the obtained composites were dried in an oven at 90 °C for 24 h. The obtained powder composites were calcinated at 600 °C for 3 h. The crystalline structure, surface morphology, functional groups, and elemental surface chemistry of the synthesized composites were characterized by methods such as X-ray diffractometry (XRD) and field-emission scanning electron microscopy (FE–SEM). Hydrothermal techniques were used to synthesize Cu/La/CeOThe hydrothermally prepared nanocomposites were characterized with various techniques. The diffraction patterns of the nanocomposite were observed using X-ray diffraction ). The surface structure and topography were obtained with a field-emission scanning electron microscope equipped with an energy-dispersive X-ray spectroscope , and ), and by transmission electron microscopy . In addition, the degradation of wastewater polluted with organic compounds was analyzed with a UV–visible spectrophotometer .−4 moles. A 50 W tungsten-halogen lamp  was used as the visible-light source. For the degradation process, 100 mL of dye solution was irradiated under visible light. For this experiment, 20 mg of nanocomposites was added to the dye solution, which was constantly stirred. Before the experiment, the dye solution with a catalyst added was stirred continuously at 30 min under the dark condition to acquire an adsorption–desorption equilibrium condition. The solution mixture was irradiated with visible light for 90 min under magnetic stirring, and every 10 min, 5 mL of the solution was extracted for the study of the degradation/decolorization of dye with a UV–visible spectrometer. Before the absorbance analysis, the extracted samples were filtered using Whatman filter paper to remove the photocatalyst.The photocatalytic performance of the hydrothermally synthesized nanocomposites in the degradation of three different organic dyes, namely, RhB, SY, and CY,  under vi2/GO nanocomposites are depicted in 2 also indicated the (111), (200), and (220) planes, which were attributed to the cubic fluorite-structured CeO2 crystals (JCPDS: 03-065-0859) [2O3-related peaks were attributed to the (002), (011), and (003) planes, which were indexed to the hexagonal structure of La2O3 crystals (JCPDS: 01-083-1349) [2+ (77 pm), La3+ (103 pm), and Ce4+ (102 pm) ions, respectively. The XRD patterns of the hydrothermally synthesized Cu/La/CeO65-0859) ,20,21,2283-1349) ,24,25,26The nanocomposite crystalline size was also calculated using Scherrer’s equation:2/GO morphology and structures were studied by FE–SEM analysis  of ceria nanoparticles [3/2 and Cu 2p1/2 peaks at 935.5 and 954.9 eV, respectively, indicated the formation of the Cu+ (Cu2O) oxidation state in the nanocomposites. Concurrently, the Cu2+ oxidation peaks also formed at 942.1, 946.9, 960.5, and 966.2 eV, indicating the presence of the Cu2+ (CuO) oxidation state. The results revealed that the copper was present in two forms (CuO and Cu2O) of oxidation states in the nanocomposites. 5/2 and La 3d3/2 states of La3+ oxidation. 2/GO nanocomposites, which had four oxygen-related functional groups at 285.8, 287.02, 288.78, and 289.5 eV. These oxygen-related functional groups were attributed to the C–O (hydroxyl), C=O (carbonyl), O–C=O (carboxylic acid), and carbonate bonds, respectively [mposites a. The hiarticles ,32,33. Fectively ,37,38. M2 < CeO2/GO < Cu/La/CeO2 . Additionally, the recombination rate of the photogenerated charge carrier in the CeO2 lattice was effectively restricted by the ions of the dopant materials. Therefore, the visible light on the surface of the photocatalyst absorbed more energy, resulting in electrons transferring from the valence band to the conduction band. Photogenerated electron–hole pairs were generated, as were electrons in the conduction band and positive holes in the valence band. The positive holes in the valence band reacted with H2O to generate more active hydroxyl radicals (OH●), while the excited electrons reacted with adsorbed oxygen to give superoxide radicals (O2●−). These reactive species were the major reason for the degradation of the targeted organic pollutants. Hence, these reactive species directly interacted with organic molecules, causing oxidation, and excitation occurred. Then the organic molecules transformed into non-hazardous organic molecules, water, and carbon dioxide. The photocatalytic degradation of organic effluents under visible light is expressed in the following equations [The catalyst-assisted photocatalytic degradation reaction is an advanced oxidation process organized by a photocatalyst under visible-light irradiation. Pure CeOe region ,21,22,26quations ,28,29:hν0 are the concentrations of dyes before and after the treatment. The rate constant and treatment time are denoted as k and t, respectively. −1, and the correlation coefficients (R2) were 0.9876, 0.94062, and 0.93102, respectively. Therefore, the rate constant mainly depended on the treatment time because the treatment time increased with increases in the rate constant. As a result, the degradation of RhB, SY, and CR was influenced by the generation of higher concentrations of various reactive species [The photocatalytic degradation of three different dyes was further evaluated with the help of a pseudo-first-order kinetic rate constant, which can be expressed by the following equation : ln  and isopropanol (IPA) were used as superoxide radicals and hydroxyl radical scavengers, respectively [2EDTA and IPA, respectively. The results strongly suggested that OH● was generated as the major active species for the degradation process, though the holes also contributed to the degradation process in the photocatalytic process. Photocatalyst-assisted degradation mainly depends on the generation of various reactive species, electrons, and holes. During the degradation reaction, the photocatalyst produces electron–hole pairs under visible-light irradiation, which generates reactive species such as OHectively ,40. Figu2/GO. The synthesized nanocomposite showed excellent efficiency (86%) of RhB dye degradation even in the third cycle, demonstrating the stability of the nanocomposites.To inspect the stability of the synthesized nanocomposites, a recycling experiment was conducted, and the results are depicted in 2) is a highly efficient photocatalyst, as it absorbs more visible light than TiO2 and ZnO. The crystalline structure, surface morphology, and topography of the hydrothermally synthesized nanocomposites were studied. Cerium oxide was engineered by doping with Cu and La metal ions to increase visible-light absorption. Moreover, the efficiencies of the photocatalytic degradation of RhB, SY, and CR organic dyes were 80% (RhB), 60% (SY), and 95% (CR) with treatment times of 90, 90, and 50 min, respectively. The photocatalytic degradation was improved by doping the metal ions, as well as the graphene oxide, due to the higher electron mobility, high light absorption capability, and restrictions of electron–hole pair recombination. In this regard, higher amounts of various reactive radicals were generated, which facilitated the effective degradation of the organic compounds. Accordingly, the hydrothermally synthesized Cu/La/CeO2/GO nanocomposites appear to be promising photocatalysts for the degradation of toxic organic dyes in aqueous solution.Cerium oxide (CeO