To evaluate the effect of subgingival administration of various antimicrobials and host-modulating agents in furcation defects as an adjunct to scaling and root planing (SRP) compared to SRP alone or combined with placebo.A systematic review was carried out using MEDLINE-PubMed, Embase, and Scopus for articles up to October 2022 in addition to hand searches. All longitudinal studies that evaluated the effect of subgingival application of antimicrobial and host-modulating agents in furcation defects as adjuncts to SRP compared to SRP alone or SRP + placebo with at least 3 months of follow-up were eligible for inclusion.A total of eight studies were included. Superior clinical treatment outcomes were shown when alendronate, rosuvastatin, boric acid, simvastatin, and tetracycline (only at 3 months) were utilized in furcation defects in conjunction with SRP alone or SRP + placebo. Significant improvement was reported in radiographic bone defect depth and defect depth reduction when SRP was supplemented with alendronate, rosuvastatin, boric acid, and simvastatin.Within the limitations of this review, the adjunctive subgingival administration of medications and host-modulating agents in furcation defects may confer additional clinical and radiographic benefits than non-surgical periodontal treatment alone. Future investigations are needed to confirm their long-term effectiveness.Local host modulators and antimicrobials may be used supplementary to enhance the clinical and radiographic treatment outcomes of conventional periodontal therapy in furcation defects. Periodontitis is a multifactorial inflammatory disease affecting the tissues surrounding the teeth and it is characterized by loss of connective tissue attachment and alveolar bone . UntreatNon-surgical periodontal therapy through scaling and root planing (SRP) is currently considered the gold standard for the treatment of periodontitis and aims at reducing soft tissue inflammation by removing both soft and hard dental biofilm deposits . HoweverWhen teeth with furcation defects are considered, it is well established that they exhibit a higher tooth loss rate than those without such anatomical defects . The anaAdjuncts to non-surgical periodontal treatment have been proposed to enhance the clinical and microbiological outcomes in periodontitis patients, including local delivery of antimicrobials, antibiotics , and host modulation agents , 15. TheThe aim of periodontal treatment is to arrest disease progression by reducing the bacterial load and to reduce the risk of tooth loss, as well as to prevent disease recurrence, even in restricted access areas, such as furcation defects. Local adjunctive periodontal therapy may be utilized in such areas to overcome these limitations. No systematic review has evaluated the available evidence regarding all available agents on the effect of local adjunctive non-surgical periodontal treatments in furcation areas. In this systematic review, both short- and long-term clinical and radiographic findings are assessed following in previously untreated periodontitis patients as well as in patients with recurrent periodontitis during supportive periodontal care. Findings from this systematic review will enable clinicians to exercise evidence-based decision-making for the management of periodontitis patients with furcation defects. Therefore, the aim of this systematic review was to evaluate the effect of subgingival administration of various drugs in furcation defects as an adjunct to SRP compared to SRP alone or combined with placebo.The present systematic review followed a detailed protocol according to the PRISMA  guidelines . The proPopulation: systemically healthy adult patients diagnosed with periodontitis who exhibit at least one furcation defect. Previously untreated periodontitis patients and patients with recurrent periodontitis during supportive periodontal care were included.Intervention: scaling and root planing (SRP) in combination with local application of medications including antimicrobials, antiseptics, host modulators, and biologics in grade II (osseous destruction into the furcation area but not extending to the opposite site) furcation defects.Comparison: scaling and root planing (SRP) alone or SRP + placebo.Outcome: clinical and radiographic changes following periodontal treatment. The primary outcome of the study was the probing pocket depth (PPD) reduction. Secondary outcomes were the following: relative vertical clinical attachment loss (RVAL), relative horizontal clinical attachment loss (RHAL), clinical attachment level (CAL) gain, bleeding on probing reduction, bone defect fill, and furcation closure as well as adverse events.The formulated PICO  question was: “In systemically healthy adult patients diagnosed with periodontitis, does subgingival application of medications in furcation defects as an adjunct to non-surgical periodontal therapy (SRP) improve clinical and radiographic periodontal parameters, when compared to non-surgical periodontal therapy alone or combined with placebo?”.All longitudinal comparative studies with at least 3 months of follow-up were eligible for inclusion—randomized and non-randomized clinical trials that evaluated the effect of subgingival application of drugs in furcation defects as adjuncts to SRP compared to SRP alone or SRP + placebo. Studies performing non-surgical periodontal treatment were only included, while surgical studies were excluded. Included studies reported at least one of the aforementioned outcomes. No language limitations were applied. The exclusion criteria were as follows: (1) abstracts of posters or presentations; (2) studies not including furcation defects; (3) studies including surgical periodontal treatment; (4) studies that did not report treatment outcomes of interest for the present review; (5) studies including less than 3 months of follow-up; (6) case reports; (7) case series.Electronic and manual literature searches were conducted by two independent reviewers  in databases including MEDLINE-PubMed, Embase, and Scopus for articles up to October 2022, without language restriction. In addition, a manual search of the reference lists of the included studies and of relevant journals including the Journal of Dental Research, Journal of Clinical Periodontology, Journal of Periodontal Research, and Journal of Periodontology for publications after 2010 were performed. Grey literature was searched on clinicaltrials.gov to identify potential ongoing or completed RCTs. The two reviewers assessed the eligibility of the studies for inclusion in this systematic review independently based on their title and abstract (1st round) and full text (2nd round). Publications that did not meet the inclusion criteria were excluded. Any disagreement was resolved with discussion. In case of disagreement, a third person (L. T.) was able to advise. The level of agreement between the two reviewers was calculated using kappa statistics.A series of search terms combined with Boolean operators “AND” and “OR” was used. The following search string was developed:  AND (furc* OR molar* OR multirooted* OR multi-rooted* OR radicular).The characteristics of the included publications were extracted by two reviewers (G. S. C and V. P. K.) independently and any discrepancy was resolved with discussion. Among the characteristics extracted were as follows: (1) general information including authors, title, year and journal of publication, country of origin, study setting, study design, length of follow-up, number of clinicians/evaluators; (2) patient characteristics including number of subjects in each group and smoking status; (3) treatment characteristics including type of intervention and controls, number of treated sites, type of teeth included in the study; (4) outcomes including clinical and radiographic parameters, as well as reported adverse events.χ2-based Q-statistic method and I2 measurement. A p value of 0.05 was set as the level of significance. If the necessary data were available, subgroup analyses were to be done to explore differences in treatment outcomes between different medications/agents, smokers/non-smokers, and time of the intervention . Whenever applicable, subgroup analysis would be performed based on the duration of follow-up and type of disease.Meta-analysis of the data reported in the included studies was planned to quantitatively assess the effect of the subgingival application of medications/agents in furcation defects as adjuncts to SRP based on a specific protocol. Baseline clinical and radiographic characteristics were summarized as weighted mean differences and 95% confidence intervals. For each parameter, weighted treatment effects (pre-treatment–post-treatment) were to be calculated and expressed as weighted mean differences and standard deviations. Standard pairwise meta-analyses of direct comparisons (test versus control groups) were to be performed using a random-effect model, and results were to be expressed as mean differences and relative 95% confidence intervals. The results of the meta-analysis were to be summarized with forest plots. Heterogeneity was to be assessed by using the The risk of bias in the included publications was assessed by two reviewers  independently and in duplicate. Any discrepancy was resolved with discussion. The quality of the selected randomized clinical trials (RCTs) was assessed according to the revised RoB 2.0 tool  . In partThe PRISMA flow diagram of the study selection process is shown in Fig. The characteristics of the studies included in the qualitative synthesis are shown in Table Clinical data from the included studies are shown in Table Radiographic data from the included studies and adverse events are shown in Table The studies’ conclusions are demonstrated in Table The risk of bias assessment of the included studies is presented in Fig. The need for additional periodontal treatment, either non-surgical or surgical, following the initial periodontal treatment is often observed in periodontitis cases. This is more frequent especially in molars with furcation defects . AdjunctFindings from this systematic review showed that the majority of the available publications depicted beneficial treatment outcomes, both clinically and radiographically, following the use of local adjuncts to non-surgical periodontal therapy (SRP) in furcation defects. Superiority was primarily noted for PPD reduction and RHAL and RVAL gain as well as bone defect depth reduction. Notably, due to the variability in the agents used in each study and the methodological differences between studies using the same agent, a meta-analysis was not possible. Specifically, alendronate was tested in two RCTs , 27, oneNon-surgical periodontal treatment is considered the gold standard in periodontal therapy, but subgingival instrumentation may still lead to residual pockets . ResiduaWith regards to the host-modulating agents, this review identified studies reporting on the use of alendronate and statins, such as rosuvastatin, atorvastatin, and simvastatin. Specifically, the local application of alendronate in mandibular grade II furcation defects demonstrated significantly enhanced clinical and radiographic treatment outcomes, when used in conjunction with SRP, compared to SRP with placebo in step II and step III periodontal treatment , 27. AleMoreover, statins including rosuvastatin, atorvastatin, and simvastatin have been tested for their additional effect as adjunctive treatment to SRP for the treatment of mandibular grade II furcation defects and have shown significantly improved clinical and radiographic treatment outcomes, compared to SRP + placebo , 28. TheWith regard to antiseptics, this review identified a study reporting on the adjunct use of boric acid. Boric acid has demonstrated antimicrobial and antiinflammatory properties by inhibiting the lipopolysaccharide-induced tumor necrosis factor-α (TNF-α) , 45. In Finally, with regard to antibiotics, this review identified studies reporting on doxycycline and tetracycline fibers. Doxycycline is a third-generation tetracycline leading to better absorption, protein binding, and diffusion into the tissues and exhibiting prolonged action, when compared to tetracycline . FurtherTo conclude, according to the present systematic review, host-modulating agents including alendronate, statins, and antiseptic such as boric acid may lead to notable improvement after non-surgical periodontal treatment of grade II furcation defects. However, it is important to consider the small number of included studies and the different therapeutic products and protocols assessed. Specifically, two studies utilized these agents during the initial periodontal treatment , 25, wheIn conclusion, based on the currently available evidence derived from RCTs, local antimicrobial and host-modulating agents including alendronate, rosuvastatin, boric acid, simvastatin, and tetracycline may enhance the clinical treatment outcomes when used as adjuncts to non-surgical periodontal treatment in furcation defects. Host modulators including alendronate, rosuvastatin, and simvastatin as well as boric acid demonstrated more favorable radiographic treatment outcomes than conventional periodontal therapy in molars with furcation defects. All agents appeared to be overall safe and well-tolerated and were not associated with adverse reactions. Future and adequately powered studies are needed to confirm their long-term effectiveness including different populations and further explore the effects of these agents on challenging clinical scenarios.


Colorectal tumorigenesis is driven by alterations in genes and proteins responsible for cancer initiation, progression, and invasion. This multistage process is based on a dense network of protein–protein interactions (PPIs) that become dysregulated as a result of changes in various cell signaling effectors. PPIs in signaling and regulatory networks are known to be mediated by short linear motifs (SLiMs), which are conserved contiguous regions of 3–10 amino acids within interacting protein domains. SLiMs are the minimum sequences required for modulating cellular PPI networks. Thus, several in silico approaches have been developed to predict and analyze SLiM-mediated PPIs. In this review, we focus on emerging evidence supporting a crucial role for SLiMs in driver pathways that are disrupted in colorectal cancer (CRC) tumorigenesis and related PPI network alterations. As a result, SLiMs, along with short peptides, are attracting the interest of researchers to devise small molecules amenable to be used as novel anti-CRC targeted therapies. Overall, the characterization of SLiMs mediating crucial PPIs in CRC may foster the development of more specific combined pharmacological approaches. Moreover, it is predicted that the global incidence of CRC will more than quadruple by 2035, with the greatest increase occurring in less developed countries due to the growing number of cases detected in the older population [Colorectal cancer (CRC) is mainly caused by abnormal proliferation of glandular epithelial cells in the colon. CRC is classified into three types: sporadic, hereditary, and colitis-associated . About 6From a molecular perspective, CRC initiation and progression are complex multistage processes involving a dense network of protein–protein interactions (PPIs). These PPIs mediate cell-signaling cascades that are deregulated as a result of CRC-related alterations .It has long been believed that PPIs were mediated by large, structured domains ,8. NonetPPIs involved in signaling and regulatory networks are now known to be mediated by an important subclass of disordered interfaces termed short linear motifs (SLiMs). SLiMs are typically conserved, contiguous sequences consisting of about 3–10 amino acids within interacting protein regions ,12. ThesSLiMs are the minimum sequence determinants that can finely modulate cellular PPI networks independently of the wider sequence/structure context where they function . FrequenFrom a molecular point of view, cancer-causing mutations are typically thought to affect protein functionality due to the disruption of their folded globular structure. Nevertheless, 22% of human disease mutations occur in intrinsically disordered regions (IDRs) of proteins . Therefo989TSPTF993 BRIP1 (Fanconi Anemia Group J Protein) motif. Alterations in these SLiMs that mediate BRCA1–BRIP1 interaction have been associated with various cancer types, including breast cancer (BC), ovarian cancer, and pancreatic cancer [For instance, several studies have shown that SLiMs play an important role in the activity of BRCA1, a critical tumor suppressor regulating cell cycle progression and DNA-damage-induced checkpoint activation. Two regions  of the BRCA1 C-terminus (BRCT) domain interact with the c cancer ,27,28,29489MTPVNPGV497 can inhibit mouse uterine cell proliferation and has shown anticancer effects in MCF-7 breast cancer cells [Emerging evidence also suggests that specific SLiMs derived from or encompassing the alpha-fetoprotein (AFP), a well-known embryo-specific and cancer-related protein, may have antitumor effects in several types of cancer  ,31,32. Ier cells . In addier cells ,32. In aer cells .Interestingly, MDMX-mediated pharmacological inhibition of the tumor suppressor P53 is also dependent on a SLiM-related mimicry effect . MDMX, aOverall, these data highlight the emerging biological relevance of SLiMs in modulating driver interactions in tumorigenesis and tumor progression, which is further corroborated by the number of different studies and in silico methodologies that have been developed to define a cancer-specific and SLiM-based molecular signature suitable for precision medicine approaches ,36.A thorough understanding of PPI networks is crucial for a systems-level interpretation of the vast quantities of molecular data that are made available to researchers. Experimental detection of protein interactions is complicated by a wide variety of binding affinities, and different assays have varying strengths and weaknesses in overcoming these challenges .For instance, based on the evidence that physical interaction between two proteins requires complementary three-dimensional structures, and considering that it is crucial to identify in which conformation a certain interaction can occur, Halakou et al. described a step-by-step method for incorporating the various conformations of proteins into each PPI. In particular, they showed how a PPI network can be refined by docking alternative three-dimensional conformations of each protein participating in binary interactions, thereby unifying the network view and structural perspective of the involved interactions .Computational methods are a powerful tool to bridge the gaps in the experimental approaches used to investigate PPIs. In particular, they can help transfer our understanding of complex interactions from one species to another, integrate our knowledge scattered across databases covering different types of interactions, provide functional insights into the discoveries made from the analysis of omics datasets, and assemble individual interactions into higher-order functional units, i.e., protein complexes and signaling pathways .In this scenario, several in silico strategies using indirect evidence or based on the ability to learn from existing interaction data gathered by experimental techniques, such as two-hybrid and affinity purification coupled with mass spectrometry, have been devised to build screening tools allowing researchers to predict and/or identify PPIs ,37,38,39The above-described strategies provide a huge amount of data and information on PPIs in different species, which are annotated in specific databases. As a result, our current understanding of PPIs is fragmented across multiple sets of data that cover a variety of interaction types across multiple species. The ability to systematically query the databases of interest is crucial for the incorporation of PPI data into effective downstream analysis techniques. Various tools efficiently address this challenge.The interpretation of high-throughput molecular profiling datasets can be supported by PPI networks. A collection of differentially expressed genes found by RNA-seq, for instance, can have their function clarified by finding enriched biological pathways or by examining their relationships in a larger network context ,39.As indicated in . In the last decade, several in silico algorithms able to extract SLiMs from interaction data have been developed. Surprisingly, emerging evidence suggests that these regions mediate a greater proportion of interactions than was previously anticipated. Moreover, it has been shown that SLiM-mediated interactions can be pharmacologically targeted with specific compounds, which may be used to interfere with or disrupt cancer-related PPI networks [As indicated above, SLiMs are short, conserved stretches of amino acids that mediate PPIs. SLiMs are difficult to identify experimentally due to their extremely small size and poor folding. Based on SLiM-recognition domains, numerous computational and bioinformatics tools have been created to analyze PPI networks, especially focusing on cancer-related pathways networks . TherefoThe various in silico resources listed in CRC tumorigenesis is initiated by spontaneous mutations, environmental mutagens, and genetic or epigenetic alterations that trigger the transformation of normal colorectal epithelial cells into tumor cells. Mutations in crucial factors, such as APC and WNT-β catenin (CTNB1) pathway effectors, along with cytokines, chemokines, and growth factors from the tumor microenvironment, cause the hyperproliferation of initiated cells and the subsequent formation of an aberrant crypt and adenoma . Next, tA few functionally relevant SLiMs have been identified in proteins playing a role in major signaling cascades involved in CRC onset and progression. These include the Wnt/CTNB1, the EGFR/MAPK, and the BCL-2 pathways ,50,57,58Wnt signaling hyperactivation promotes tumor cell proliferation and is required for tumor growth . After bThe proto-oncogene C/EBP homologous protein (CHOP) is a dominant-negative inhibitor of C/EBPs, and its disordered N-terminal region contains crucial SLiMs that are essential for CHOP oligomerization, interactions, and biological activity . In part32SGIHS37 has been well characterized and is therefore reported in the ELM database (ELM ID: ELMI001302). This motif is directly involved in the aberrant proteasomal degradation of CTNB1 [32SGIHS37 SLiM and is routed to proteasomal degradation [Conversely, the CTNB1 SLiM Dof CTNB1 ,79. In pof CTNB1 . CTNB1 pradation . Point mradation .The ELM database reports other SLiMs in CRC regulatory proteins that are recognized by SCF  complexes via repeat domains of associated F-box proteins (FBPs) and are therefore routed to subsequent ubiquitin-mediated degradation . For exaThe EGFR/MAPK signaling pathway is also directly related to CRC oncogenic processes and plays a critical role in tumor growth and disease progression . EGFR isThe proliferative signal associated with the cascade activation of MAPKs depends specifically on 3 SLiM clusters: two clusters enriched in positively charged amino acids surrounding a central cluster enriched in hydrophobic amino acids. These motifs, consisting of a series of positive–hydrophobic–positive amino acids, represent the modular structure of the docking PPIs involved in MAPK signaling. Therefore, point mutations in these crucial amino acids may alter the activation of MAPK signaling cascade .Apoptosis is a crucial cell-death process that is frequently dysregulated in various malignancies, including CRC ,119. ApoThe BH3 motif has been identified as a relevant SLiM in CRC tumorigenesis ,74. It i163NLTKRIDSLPLTE176) of APC nuclear export signal (NES) [Pathological variants of crucial SLiMs have also been observed in inherited CRC syndromes. In particular, familial adenomatous polyposis (FAP) is an autosomal dominant disorder characterized by a predisposition to colorectal polyposis, which in some cases will evolve into colorectal carcinoma if not surgically treated. A variant of FAP is the Gardner syndrome, in which multiple adenomas of the colon and rectum occur with desmoid tumors and osteomas . This coal (NES) ,97.Recent data from our group are consistent with the crucial role played by SLiMs in the PPI network of SET and MYND domain containing 3 (SMYD3) in different cancer study models, including BC, CRC, and other gastrointestinal tumors ,124. In Next, to gain insight into novel SMYD3 cancer-related activities, we focused on the whole-proteome distribution of the P1-P19 tripeptides and assessed the biological function of each putative SMYD3 interactor to identify the most important candidates associated with cancer hallmarks . SurprisA significant benefit of this strategy is that our tripeptides can be used to develop pharmacological inhibitors of SMYD3 oncogenic PPIs in order to modify the composition of relevant multiprotein complexes associated with cancer driver proteins. Similar to other approaches devised in the last few years ,134,135,This methodology is schematically outlined in Growing evidence suggests that microbes represent relevant players in CRC tumorigenesis . Thus, tF. nucleatum–human cross-talk in the context of gastrointestinal diseases [Interestingly, SLiM-based molecular mimicry is investigated as a significant aspect of microbiome-host relationships, which are of crucial relevance in CRC tumorigenesis and inflammatory bowel diseases . In partdiseases .Taken together, these data confirm the biological significance of SLiMs in cancer-related pathways and suggest that they may play a role in PPI networks involved in CRC. In addition, they could be used to devise pharmacological strategies to interfere with the main signaling cascades driving tumorigenesis and cancer progression.The biological relevance of PPIs in key cancer-related processes, including cell growth, proliferation, differentiation, and signal transduction, is corroborated by the evidence that PPIs are altered in various cancer types ,141. ThiThere are different structural caveats to the pharmacological targeting of PPIs, including their large interfaces, which are frequently flat or shallow. Historically, drug design and discovery have given rise to the belief that PPIs are difficult to target because the compounds should be small enough to enter the cell and also be able to affect the large and often shallow PPI interaction sites . DespiteFurthermore, current advances in molecular profiling of tumor samples as a result of high throughput techniques such as NGS and mass spectrometry enable the rapid detection of alterations in genes encoding oncoproteins, providing a large number of potential therapeutic targets and offering opportunities for the design of new drugs. Notably, based on different studies carried out by pharmacological companies and academic groups on SMYD3 cancer-related activity, novel and more efficient SMYD3 inhibitors have been generated ,148,149.The development of novel anticancer drugs is complicated by limitations related to the specific chemical characteristics of potential therapeutic molecules and the structural requirements of druggable targets . The cheMost drug targets are molecules with binding sites for low-molecular-weight compounds. These compounds are then used as starting points for the development of small chemical analogs that bind to the same site and can function as competitive or irreversible inhibitors . If a peAs a matter of fact, SLiM-based small molecules are still in their early days. For instance, BH3-mimetics are SLiM-based engineered peptide inhibitors able to bind to the hydrophobic groove of the corresponding anti-apoptotic proteins ,158. In One of the first examples of short peptides tested for their anticancer effects are the parasporin (PS) peptides isolated from the gram-positive bacterium Bacillus thuringiensis . A totalVarious studies support the use of short peptide aptamers as promising pharmacological inhibitors of signal transducer and activator of transcription 3 (STAT3). STAT3 is a transcription factor that is activated by tyrosine kinase phosphorylation and has a well-defined domain structure ,153,156.Cordyceps Militaris (CM) proteome using various machine learning-based ACP prediction servers, i.e., AntiCP, iACP, and MLACP. The purpose of the authors was to select CM-derived ACPs to be used as an alternative or adjunct treatment for CRC, hence minimizing the need for chemotherapy. To confirm their anticancer effects, CM-biomimetic peptides were tested in vitro in a non-metastatic colon cancer cell line in comparison and in combination with doxorubicin, a typical chemotherapeutic agent for the treatment of colon cancer. Overall, the results demonstrated that the selected biomimetic peptide C-ori improved the efficacy of doxorubicin treatment [Anticancer peptides (ACPs) are an emerging class of naturally occurring or synthetic anticancer compounds. They show greater selectivity for cancer cells and less propensity for drug resistance . In a rereatment .2) in their hydrophilic domains exhibit more selective anticancer activity, whereas peptides containing arginine (R)  display strong toxicity in normal cells. Furthermore, it was shown that ACP anticancer activity depends on their helical composition and hydrophobicity [2) increased their anticancer activity by boosting their hydrophobicity and helical content. The higher ACP toxicity detected in cancer cells compared to normal cells was due to better penetration into the negatively charged cancer cell membranes, resulting in greater cellular uptake, and their cytotoxic effect was primarily exerted by damaging mitochondrial membranes, resulting in apoptosis [Another group performed an in silico analysis to generate a new library of short cationic amphiphilic α-helical ACPs with specific cytotoxicity against colorectal and cervical cancer . The α-hhobicity . Indeed,poptosis .Overall, these data confirm that short peptides and SLiMs exhibit promising pharmacokinetic characteristics in vivo.Several short peptides have been recognized as PPI modulators and therefore investigated in clinical trials for CRC treatment , which rA major advantage of short peptides is their high targeting specificity; however, the number of terminated clinical trials suggests that new nanotechnologies are needed to enhance the chemical, physical, and biological stability of these compounds .https://clinicaltrials.gov/; accessed on 5 September 2022) [https://clinicaltrials.gov/; accessed on 5 September 2022) [New therapeutic opportunities may come from highly specific small-peptide inhibitors called BH3 mimetics, which have been developed to target anti-apoptotic BCL-2 proteins by mimicking the action of BH3-only proteins . By inteer 2022) . Notablyer 2022) . Thus, ger 2022) .Currently available treatments for colon cancer are based on specific combinations of surgery, radiation, chemotherapy, and targeted therapy. Nevertheless, following surgical resection and intensive chemotherapy, 50 percent of CRC patients have disease recurrence. In addition, the occurrence of chemoresistance hampers the efficacy of chemotherapeutic agents . TherefoMoreover, it is crucial to improve the clinical management of CRC patients in order to prevent relapse. This should be combined with the search for novel and more efficient approaches to obtain additional diagnostic and prognostic information.An efficient approach to identify cancer drivers, as well as diagnostic markers for accurate tumor staging and prediction of clinical outcomes, involves in silico studies to detect differentially expressed genes across different phenotypes, followed by the analysis of the relevant pathways with specific bioinformatics tools evaluating the enrichment of altered oncoproteins .In the last decade, efforts in cancer research have increasingly focused on the development of integrated strategies combining in silico, in vitro, in cellulo, and in vivo approaches to provide in-depth knowledge of cancer-related pathways, PPI networks, and the biological implications underlying a multifactorial disease such as cancer . Indeed,Recent advances in high-throughput biotechnology applications enable large-scale analysis of disease-associated genes and proteins involved in critical molecular processes. The biological and clinical significance of the resulting data needs careful evaluation. Characterization of the critical SLiMs that mediate PPIs between key players in CRC tumorigenesis and progression is expected to support the development of more efficient and specific pharmacological approaches based on small-molecule anticancer drugs, which may be used in combination with chemotherapeutic agents or pharmacological inhibitors . For instance, the association between pharmacological inhibitors such as ralimetinib (a MAPK14 inhibitor) and SLiM-based small molecules interfering with MAPK signaling activation may prove an effective therapeutic approach to hamper MAPK-dependent CRC development and progression . FurtherFurther studies are required to gain deeper insight into the therapeutic potential of SLiM-based small molecules in CRC and to address the main limitations related to their stability, bioavailability, and pharmacokinetics.


BRAF mutations. BRAF-MEK inhibitor drugs improve survival in the approximately 50% of patients with melanoma that harbor BRAF mutations. As BRAF mutation detection in tissue often takes days to weeks, it is not always possible or timely to obtain BRAF status in tissue using immunohistochemistry or next generation sequencing. Plasma-derived circulating tumor DNA (ctDNA) is a potential alternative analyte in such treatment settings. We present a case of metastatic melanoma that was treated in an emergent setting using therapy supported by rapid PCR-based detection of ctDNA positive for a BRAF V600 mutation. In this rapidly deteriorating 53-year-old male with diffuse melanoma metastases and unknown BRAF mutation status requiring hospital admission, a plasma-based BRAF mutation detection supported treatment with targeted therapy, dabrafenib and trametinib. Same-day initiation of therapy resulted in swift amelioration allowing discharge within a week, followed by substantial clinical improvement over the following weeks. In cases requiring urgent clinical decision making, a plasma-based, near point-of-care detection system is useful in supporting targeted therapy decisions without the need for invasive and time-consuming biopsy.Treatment of metastatic melanoma includes the option of targeted therapy in patients with driver  Circulating tumor DNA (ctDNA) is a useful disease monitoring analyte in many cancers . DiagnosBRAF mutation status in melanoma offers the option for specific targeted therapy agents. Specifically, the combination of BRAF and MEK inhibitors in melanoma are highly effective in improving survival and often precipitate rapid symptom resolution (BRAF mutation status by tissue immunohistochemistry (IHC) or molecular diagnostic techniques, which is accomplished by next generation sequencing (NGS). Tissue-based BRAF detection may take days to weeks, depending on an institution’s capabilities and whether tests are typically “sent out” to a reference laboratory. In patients where the primary site is occult, and sites of metastases are not accessible for tissue sampling, BRAF mutation status may be difficult to obtain and/or remain unknown. In advanced melanoma, ctDNA is generally higher in concentration compared to earlier stages , a PCR-based, near-point of care, rapid testing platform could be useful to detect BRAF variants in plasma-derived ctDNA  performed on the biopsy tissue is the gold standard for detecting BRAF mutation in patients diagnosed with melanoma. Tissue immunohistochemistry (IHC) is a sensitive and relatively quicker alternative routinely employed to identify the presence of BRAF V600E mutations  was employed to detect BRAF variants in plasma-derived ctDNA  followed by dabrafenib/trametinib (Dab/Tram) to the converse sequence in treatment-naive BRAFV600-mutant patients with ECOG performance status 0 or 1 suggests that the treatment sequence beginning with the combination of Ipi/Nivo results in superior OS (BRAF mutant status supported the decision to initiate targeted therapy with dabrafenib and trametinib. Of note, the patient also underwent an inpatient biopsy procedure on the same day as the peripheral blood sample for ctDNA testing was drawn. After 16 days, BRAF variant identification was indeterminate due to insufficient tumor in the liver biopsy. Ultimately, the ctDNA result was the only modality available to support the treatment decision, which allowed patient discharge and marked clinical improvement over the next 100 days.As was the case with this patient, the clinical decision on the ability to use the preferred early therapeutic, ie, BRAF/MEK inhibition, was dependent on rapid acquisition of the patient’s erior OS . The treBRAF variant detection has been reported to be a prognostic marker in patients with brain metastasis (BRAF variant in the patient’s plasma supported subsequent further imaging that revealed brain metastases three months after admission. The detection of new brain metastases precipitated initiation of immunotherapy with ipilimumab and nivolumab.Further, ctDNA-based tastasis . PersistIn conclusion, a rapid real-time PCR-based evaluation of peripheral blood could serve as a non-invasive, rapid tool to aid prompt treatment decision making in advanced melanoma in emergent situations where BRAF mutation detection utilizing NGS or IHC of the biopsy tissue is not feasible or could potentially delay clinical decision making.The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding author.The studies involving human participants were reviewed and approved by Cleveland Clinic ethics committee. Written informed consent was obtained from the next of kin for the publication of any potentially identifiable images or data included in this article.TB, JS, JK, DE, JA, BG, DF and PF contributed to conception and writing, DF and PF contributed to finalize the manuscript. All authors approved the submitted version.This work and TB were partially supported by Gross Family Melanoma Registry.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.


Telehealth and telecare are particularly important and beneficial to long-term care facilities due to care demands, workforce, and the unique environment. Stemming from the recent findings on telehealth utilisation in residential aged-care facilities in Australia, this commentary seeks to identify lessons and perspectives learned during the Covid-19 pandemic from multiple users, including patients, physicians, nurses, and healthcare workers in long-term care (LTC) settings.From patients’ perspectives, older adults residing in LTC settings often opt not to use virtual care, with the majority preferring in-person visits. This is despite residents expressing their willingness to use telehealth, and virtual care has advantages in LTC settings or in remote areas. Additionally, hearing, vision, or cognitive impairment can limit residents’ ability to use information technology to access care, so their preferences for phone or video consultations depend on the health conditions or care requirement. From physicians’ perspectives, most healthcare practitioners have a positive attitude toward using telehealth. However, telephone consultations tended to be the dominant mode during the early period of the Covid-19 pandemic. Physicians also raised several major concerns, including technical and equipment-related issues, expanded roles, or additional workloads of LTC staff that could negatively affect clinical decision-making and unequal access in rural, older, and cognitively impaired patients. Most nurses and healthcare workers perceived telehealth positively as a way to enhance patients’ care access. However, the majority had concerns about acquiring appropriate knowledge of using the technology for themselves and their patients. In remote areas, nurses expressed higher efficiency and higher care quality when utilising telehealth in caring for older patients than in the regular in-person care mode.Since the beginning of the Covid-19 pandemic, telehealth has continued as an alternative platform in clinical services. However, as a healthcare platform that offers flexibilities of time, location, and improved efficiency, changing the traditional mindset is essential to shift the paradigm to use telehealth when appropriate. Importantly, telehealth needs substantial support in rural or remote long-term care facilities. Doing so will contribute to the reduction of healthcare inequity in long-term care facilities in remote settings and those with social disparities. Telehealth and telecare will likely be part of the routine care delivery methods post the Covid-19 pandemic. A significant innovation, telehealth ensured timely care access during the pandemic in the general population as well as residents living in long-term care facilities (LTC). For the latter, this is particularly important and beneficial due to the high care demands of LTC residents, the limited workforce, and the unique environment of a long-term care facility where residents have limited mobility.We previously reported a retrospective study of 27,980 residents aged 65 years or over in LTC settings  during the Covid-19 pandemic regarding the patterns of telephone or videoconference usage in general practice from March 2020 to August 2021 in New South Wales and Victoria in Australia. Furthermore, we identified residents’ socioeconomic characteristics of telehealth consultations to illustrate the barriers to using telehealth in the LTC setting [Our comprehensive analyses show that telehealth visits significantly increased around the months when outbreaks occurred, ranging from the highest usage of 25% in August 2020 (strict lockdowns occurred during July—September in Victoria) to the lowest of 9% in April 2021 (schools and workplaces opened during November 2020 to May 2021). These data indicate that the telehealth utilisation rates correspond to the severity of the pandemic situations. However, over 90% of the telehealth visits were conducted via telephone, with residents who received government pensions (an indicator of low-income status) and rural residents  being more likely to use telehealth (particularly phone) relative to face-to-face consultations, as compared to their counterparts. Additionally, video utilisation was evident with significantly lower usage than phone consultation in rural residents. These results were similar when stratified by different Covid surge, suggesting the consistency of findings of residents’ socioeconomic status in association with telehealth utilisation during the pandemic.Our findings imply that social disparity during the Covid-19 pandemic further divided care access. Those who were financially sound or lived in urban areas were more likely to see a general practitioner (GP) in person. For example, when a facility is more equipped to control the spread of Covid infection, such as having sufficient care workers, protection measures, and equipment, residents are more likely to have more doctor’s visits than phone consultations. Moreover, the dire use of virtual care via videoconferencing indicates health disparity in information technology, such as slow or intermittent internet connection, lack of equipment, poor information and communication technology (ICT) skills, and low ICT literacy. These factors can significantly hamper the optimal use of virtual care in remote facilities, even though residents living in these areas would benefit the most from such technology-assisted care.While our analysis provides comprehensive epidemiological data, it cannot unfold the voices of telehealth users, including patients, physicians, nurses, and care workers. Understanding these users’ perspectives would aid the improvement of care quality and efficiency in telehealth that centre on patients’ needs in LTC in an ongoing effort.It is no doubt that patients are at the centre of quality care. Among older patients aged 55 years and over, a study found that patients’ habits played the most critical role in using telehealth during the Covid-19 pandemic among various factors . In addiIt should be noted that most studies on older patients’ preferences for phone or video visits were conducted in general clinical settings. While patients generally would be willing to use the video modality, older patients still prefer in-person visits over telehealth , 8. For The good news is that most healthcare practitioners have a positive attitude toward using telehealth –14. GeneIt is expected that some physicians had not used telehealth until the Covid-19 pandemic when the government expanded telehealth broadly in the healthcare system due to the pandemic . In lineInterestingly, in a U.S. study among 814 clinicians of the Department of Veterans in New England, primary care clinicians (18.8%) and specialists (31.7%) had a similar preference for using telephone or video care during the Covid-19 pandemic. However, both raised the challenges of using video care in conducting a physical examination. By contrast, mental health clinicians (49.5%) preferred using videoconferencing over phone for new or existing patients remotely, and expressed higher quality and efficiency of using video in mental health consultations .While most of the studies provided physicians' perspectives, understanding the perspectives of nurses and other healthcare workers is equally essential, as they are the backbone of the care workforce to ensure routine care schedules and logistics flow are on track.In an Australian survey, twenty-five primary healthcare nurses acknowledged that telephone consultation was primarily used in care delivery during the first few months after the beginning of the pandemic in 2020. However, while most nurses and healthcare workers perceived telehealth positively as a way to enhance patients’ care access, the majority had concerns about acquiring appropriate knowledge of using the technology for themselves and patients .On the other hand, health professional workers in LTC in France managing residents’ neuropsychiatric conditions perceived telehealth as a great platform to improve the quality of and access to care, particularly in remote areas . These hTelehealth in rural or remote areas has a particular advantage in serving residents in LTC settings. A body of evidence before the Covid-19 pandemic has demonstrated the benefits of delivering care using telehealth in rural areas, particularly under conditions where healthcare professionals, services, and resources are limited. For example, nurses could handle a few more patients using telehealth than regular care when serving remote areas .Perspectives from different users mentioned above illustrate that overcoming technology limitations and increasing user training are probably the most critical strategies to facilitate telehealth in rural or remote facilities. Decisions around phone versus video, the frequency and length of telehealth versus in-person consultation or care should be primarily based on patients’ clinical severity or  stability of the conditions, and the scheduling of physicians’ routine visits.Since the beginning of the Covid-19 pandemic, telehealth has continued as an alternative platform in clinical services. Therefore, telehealth will likely stay as part of routine clinical services beyond the pandemic because of its advantages and benefits mentioned above , 22. In Additionally, in LTC settings, virtual platforms and internet technology can be introduced beyond clinical care, such as family/friend visits, group exercise, and entertainment, such as dancing and singing among residents. Indeed, some e-health services have already been available in commercial space . A shiftTelehealth offers flexibilities of time, location, and improved healthcare efficiency. However, its optimal utilisation in long-term care settings requires continuing government funding for suitable technology and devices, transparent billing codes, and policies protecting patients’ privacy and security. Importantly, telehealth needs substantial support in rural or remote long-term care facilities. Doing so will contribute to reducing healthcare inequity in remote settings and those with social disparities.


There is no doubt that nanotechnology has the potential to enhance healthcare systems by simplifying and improving cancer diagnostics and treatment. Throughout this review, we summarize and highlight recent developments in nanotechnology applications for lung cancer in diagnosis and therapy. Moreover, the prospects and challenges in the translation of nanotechnology-based diagnostic and therapeutic methods into clinical applications are also discussed.As a highly invasive thoracic malignancy with increasing prevalence, lung cancer is also the most lethal cancer worldwide due to the failure of effective early detection and the limitations of conventional therapeutic strategies for advanced-stage patients. Over the past few decades, nanotechnology has emerged as an important technique to obtain desired features by modifying and manipulating different objects on a molecular level and gained a lot of attention in many fields of medical applications. Studies have shown that in lung cancer, nanotechnology may be more effective and specific than traditional methods for detecting extracellular cancer biomarkers and cancer cells  Lung cancer is a heterogeneous disease that is broadly classified into non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC), with NSCLC accounting for nearly 85% of all cases . CurrentThe development of nanotechnology and its application in the field of medicine has provided a new strategy with great potential for the early diagnosis and accurate and effective treatment of lung cancer. Nanotechnology includes a variety of complex theories, methods, techniques, and tools developed at the nanoscale, and the application of numerous nanomaterials. In the past few decades, researchers have discovered/created a variety of multifunctional nanomaterials to improve our quality of life. Some of these nanomaterials are further explored for a wide range of biomedical applications, known as “nanomedicine”, to meet increasingly stringent clinical requirements –7. Todayin vivo , computed tomography (CT), magnetic resonance imaging (MRI), sputum cytology, and bronchial biopsy, are heavily dependent on the size and progression of the tumor and may require extra specific medical equipment . The mosin vivo Figure 1in vivo , 15. As in vivo .Limited options for curative treatment at the time of diagnosis are also the major reason for the poor prognosis of lung cancer. Depending on the stages and subtypes of lung cancer, treatment strategies are different, but surgical therapy, radiotherapy, and chemotherapy remain the top choice for lung cancer. Combined with the molecular type of gene mutations, targeted therapy or immunotherapy gradually become excellent options . HoweverIn this review, we summarized and analyzed the potential applications of nanotechnology in improving the early diagnosis and precision treatment of lung cancer, intended to provide an adequate theoretical framework for promoting new diagnosis and treatment options. Moreover, we also discussed current limitations and obstacles that hinder the successful translation of nanomedicines from research to clinical use and tried to analyze how to better chart a course of nanomedicine to enter the clinic and make beneficial differences to lung cancer patients.As a biological molecule that can be detected and measured in blood, body fluids, or other tissues, cancer biomarkers including various types of proteins (secreted proteins or cell surface proteins), nucleic acids , and carbohydrates usually act as a sensitive indicator of cancer occurrence and progression , 24. HowThe detection of cancer-specific genetic aberrations in Circulating tumor DNA  released from primary tumors or circulating tumor cells facilitates efficient early diagnosis of lung cancer even before any sign occurs . It is rin vitro diagnosis 22 nanoparticles, a nanomaterial effective against several types of cancer, as a model nanomaterial to study the native protein fingerprints of personalized protein crowns formed in 10 human lung squamous cell carcinoma patients. Results analysis revealed that a specific biomarker in the corona of lung cancer personalized protein, the complement component C1q, was massively bound to Gd@C82(OH)22 nanoparticles  effect. In addition to being specific to tumor tissue, these nanoparticle probes also usually exhibit long circulation times, as well as low toxicity to nearby healthy tissue, allowing them to be used for imaging and diagnosing cancer in vivo whole-body imaging techniques because NIR light penetrates the body more deeply than light in the visible spectrum. Researchers have also applied AuNPs to in vivo tumor imaging through passive targeting. Lai et al. reported that mercaptoundecanoic acid-coated AuNPs could identify and track cancer cells at the inoculation sites in mice. Furthermore, these particles detected tumor-associated microvasculature in detail. In some cases, chitosan nanoparticles have been used for in vivo imaging through the EPR effect (Quantum dots (QDs) are nanometer-size luminescent semiconductor crystals (2~100 nm) characterized by obvious photostability, tunable emission, and high quantum yield, contributing to their wide application in tumor tissue imaging through passive accumulation dependent on the EPR effect. Compared with organic dye molecules, one of the more exclusive properties of QDs is the bright fluorescence and high photochemical endurance, which renders them optimal fluorophores for biomedical imaging . Near-inR effect . Nam et R effect .in vivo imaging of cancer, current related studies have focused on nanoparticle probe accumulation in tumor tissue for diagnosing cancer in animal models, generally mouse models, and there is still a long way to go for real clinical applications due to safety concerns.Although nanotechnology has shown great potential for Surgical resection has always been considered the best curative option for eradication or palliative treatment of lung cancer. As a means of local treatment, surgery mainly focuses on the primary lesion and local infiltration . It can via NIR-II photothermal effects (Intraoperative near-infrared (NIR) fluorescence imaging is a novel technique that allows real-time visualization of tumor tissue structures for surgical guidance using a fluorescent contrast agent in conjunction with a dedicated near-infrared camera system . In the  effects . Similar effects .Robotic surgical systems, such as the da Vinci system, allow the surgeon to translate hand movements into smaller, precise movements of tiny instruments located inside patients’ bodies, and are widely used in minimally invasive surgery procedures . It has Radiotherapy is another essential local treatment for lung cancer including external beam radiotherapy and internal radioisotope therapy that delivers high-intensity ionizing radiation with high accuracy to the tumor tissue. It is likely that the appropriate dose of radiation is sufficient to kill tumor cells with little harm to surrounding normal tissues, since mitotically active tumor cells are generally slightly more sensitive to ionizing radiations than healthy cells . HoweverNanomaterials can act as novel radiosensitizers to enhance the sensitivity of tumor cells to various radioisotopes, thereby promoting the therapeutic effect of lung cancer . Gao et in vivo targeted delivery of radioisotopes specifically to tumors and avoid systemic radio-toxicity to normal tissues , which can effectively increase tumor oxygenation by triggering decomposition of tumor endogenous H2O2 and significantly inhibit the tumor growth in company with radiotherapy  . Moreover, activation of the apoptosis pathway can also be caused by PepTs2 in H1299 cells, DOX-loading PepTs2 have high killing efficiency toward H1299 cells due to the combined chemo-photodynamic therapy   by co-assembling ligand-tagged tube-forming peptoids that contain folic acid , dansyl , and meso-tetrakis(4-carboxy phenyl)porphyrin  functionalized with linkers that are specifically cleaved by matrix metalloproteinase 9 , allowing for controlled release of chemotherapeutics from the MSNs in high MMP9-expressing tumor areas .With the rapid development of comprehensive molecular characterization of lung cancer, the understanding of the cellular origins and molecular pathways affected in each subtype has been gradually enhanced . These fBy targeting the tyrosine kinase site and binding to the tyrosine kinase receptor domain covalently and irreversibly, epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) can block the signaling pathway that activated in various neoplasms, resulting in especially effective treatment for cancer . Given tTo achieve longer circulation in the blood and an enhanced permeability and retention effect in tumors, Lu et al. encapsulated AFT in liposomes (LPs). Concomitantly, Cetuximab  was also designed to bind to LPs, which also results in the promotion of tumor cell selectivity and therapeutic activity . HoweverKim et al. synthesized water-soluble erlotinib (NUFS-sErt) by using a nano-particulation platform technology utilizing fat and supercritical fluid (NUFS™) to resolve the low solubility problem that typically prevents the creation of injectable forms of EGFR-TKIs. The effects of NUFS-sErt were similar to those of conventional erlotinib in terms of inhibiting the proliferation of EGFR-mutant lung cancer cells and suppressing EGFR signaling . By prolAs an emerging cancer treatment, immunotherapy has shown great potential to target and eradicate cancer cells by orchestrating immune systems, which is likely to result in durable antitumor responses and reduction of tumor metastasis and recurrence . It has + T cells in multifocal lung metastases (in vivo (The extent of antigen presentation by antigen presenting cells (APCs), which phagocytize tumor-associated antigens (TAA) in the tumor site and migrate to tumor draining lymph nodes (TDLN) for the activation of T cells, is very crucial for successful cancer immunotherapy (CIT). Liu et al. reported an phosphatidylserine (PS) coated inhalable liposome loaded with STING agonist cyclic guanosine monophosphate–adenosine monophosphate (NP-cGAMP) enables targeted delivery of immunostimulants to pulmonary intratumoral APCs to enhance anticancer immunity against lung metastases. PS on the surface of NP-cGAMP can be recognized and engulfed by macrophages and dendritic cells (DCs). After ingestion by APCs, cGAMPs can be cytosolic released in response to low endosomal pH, stimulating STING signaling and type I interferons production in APCs, resulting in the formation of pro-inflammatory tumor microenvironment and recruitment of cytotoxic CD8tastases . Im et a(in vivo .)  targeting programmed death 1 (PD-1) and PD-ligand 1 (PD-L1) have made a significant breakthrough in the management of lung cancer. However, only a minority of NSCLC patients respond to ICIs, superior immunotherapies are urgently needed. Reda et al. reported a nano-immunotherapy termed ARAC (Antigen Release Agent and Checkpoint inhibitor) for co-delivery of a PLK1 inhibitor (volasertib) and PD-L1 antibody, which is built upon a polymer-coated mesoporous silica nanoparticle (NP) platform that can effectively achieve the therapeutic synergy of the drug combination  . Mao et immunity .in vitro and in the enhancement of imaging in vivo. By providing better encapsulation and bioavailability, controlled release, and lower toxicity, nanotechnology-based therapeutics are able to overcome the inherent limitations of traditional lung cancer treatment, including drug resistance, systemic toxicity, rapid clearance rate, etc  of nanomaterials. Although some studies have shown that some nanomaterials can maintain structural integrity for a long time in vivo (in vivo. The potential immune stimulatory characteristics and adverse immune reactions caused by nanomaterials should also be explored in depth in the process of research. Therefore, any nanomaterials should undergo systematic experiments, verify their safety and undergo strict review by relevant departments before being used in clinical practice. In general, mouse xenograft cancer models are widely used to perform in vivo mimics of nanodrugs, but it is difficult to accurately simulate the immune landscape of an orthotopic tumor in a subcutaneous model of the tumor. In addition, the heterogeneity of human diseases affects the biodistribution of nanomedicine, which is also difficult to simulate and replicate in animal models. Therefore, the safety of nanomedicine is still the biggest challenge in the process of clinical transformation of nanotechnology. Finally, the cost of producing some nanomedicines is a concern. For practical biomedical applications, we need to be able to produce functional nanomaterials with high purity, which often means an increase in cost (A growing number of nanomedicines are gaining regulatory approval and have great potential for clinical translation. However, the application of nanotechnology in clinical practice still faces some challenges and requires further in-depth research. First, we currently know very little about the pharmacokinetics ( in vivo . However in vivo . However in cost . At presYL and YZ designed this study. YD drafted the manuscript. CS performed drawing and organization of figures. YL, YZ, YD and CS revised the manuscript. All authors contributed to the article and approved the submitted version.


Technologies and innovations are critical for addressing the future food system needs where genetic resources are an essential component of the change process. Advanced breeding tools like “genome editing” are vital for modernizing crop breeding to provide game-changing solutions to some of the “must needed” traits in agriculture. CRISPR/Cas-based tools have been rapidly repurposed for editing applications based on their improved efficiency, specificity and reduced off-target effects. Additionally, precise gene-editing tools such as base editing, prime editing, and multiplexing provide precision in stacking of multiple traits in an elite variety, and facilitating specific and targeted crop improvement. This has helped in advancing research and delivery of products in a short time span, thereby enhancing the rate of genetic gains. A special focus has been on food security in the drylands through crops including millets, teff, fonio, quinoa, Bambara groundnut, pigeonpea and cassava. While these crops contribute significantly to the agricultural economy and resilience of the dryland, improvement of several traits including increased stress tolerance, nutritional value, and yields are urgently required. Although CRISPR has potential to deliver disruptive innovations, prioritization of traits should consider breeding product profiles and market segments for designing and accelerating delivery of locally adapted and preferred crop varieties for the drylands. In this context, the scope of regulatory environment has been stated, implying the dire impacts of unreasonable scrutiny of genome-edited plants on the evolution and progress of much-needed technological advances. Climate change, population growth, pandemic, conflicts and rising socio-economic disparities are putting considerable pressure on the already stressed food systems. While the precise impacts of climate change are still unclear, unpredictable climate changes are expected to impact several vulnerable regions more adversely. This requires informed approaches to address sustainability issues to meet the future food needs. While staple crops have limited resilience to changing climate in the drylands, locally grown underutilized crops despite being vital for diverse nutrient and local adaptations are grown in low-input conditions. Underutilized crops  (5′-NGG-3′) and cleaves target DNA just three to four bases upstream of a PAM sequence to create blunt-end DSBs  and 33 sub-types . While iWhile the evolution of CRISPR as a tool is remarkable and each shortcoming has been overcome with even more novel editing technologies, achieving precise single base DNA editing is an arduous task. Recently developed editing tools such as base editing (BE) ensure precise single base changes without the involvement of DSBs, HDR and donor DNA templates for selected irreversible nucleotide base substitutions at target sites. Technically, the base editing system mainly comprises of catalytically impaired Cas protein, guide RNA, and a nucleobase deaminase domain.Arabidopsis genomes  method is a “search-and-replace” system can alter the new genetic information directly at the targeted site without any DSBs or template DNA . Prime eALS2, and PDS1 was achieved at a frequency of 6.7% and 3.4%, respectively  histone H3 lysine 4 (H3K4) demethylase domain to a locus of interest, the APX2 gene in this case that showed transcriptional memory after heat stress . This offers potential opportunities for editing crop epialleles for adaptation traits. Nonetheless, this will require combined and multidisciplinary efforts in different areas of plant science and better integration of epigenomic data obtained in different crops.Emerging tools such as epigenetic QTLs or epigenetic single nucleotide polymorphisms tools also offer opportunities for activating or repressing candidate gene(s) or pathway(s) for trait improvement in crops, which could lead to the development of a new, efficient, and transgene-free breeding methods . While tAgrobacterium or particle bombardment possess persistent challenges in crops where efficient transformation systems are not available. While these plant transformation methods are cost-effective, convenient, and easily available in laboratories, their delivery efficiencies remain highly dependent on several factors such as type of explant, Agrobacterium strain, genotype, construction of independent editing reagents in multiple binary vectors etc. To overcome these drawbacks, direct methods of gene delivery, such as protoplast transfection, virus-mediated, RNP-based, meristem induction, lipofection-and PEG-mediated protoplasts and usage of aiding elements such as special peptides and nanoparticles have been developed and adopted , these can give rise to multiple copies of the same gene causing undesirable altered expression . However, the primary disadvantage of this technique lies in its inability to transform all plant species, especially monocots. While DNA-free genome editing methods have been obtained by delivering CRISPR/Cas reagents as pression .lox P sites and DsRed2 expression  have gained pace as delivery vehicles since they can be designed according to the type of tissue and organism of interest . For exapression . NPs arepression .Cas9 RNPs, containing negatively charged gRNA molecules quickly form a complex with cationic lipids. Nucleic acids have been exploited to function as polymeric substrates for Cas9 RNP delivery. However, further modification in the polymeric coating is vital to ensure that degradation through cellular pathways and enzymes does not occur. An alternative to encapsulation is the modification of the protein and nucleic acid. Cell-penetrating peptides (CPPs) are short peptide sequences that can penetrate the cell membrane easily. They can be conjugated with Cas9 protein and gRNA for enhanced delivery. However, these peptides do not protect the protein from protease degradation within the cell and can be complexed with other delivery methods. Further, nuclear localization sequences (NLSs) which are sequences synthesized in the cytoplasm for tagging proteins and transported into the nucleus are also being explored. They are poly-arginine/lysine and behave as signal molecules attached to proteins for nuclear transport. As Cas9 needs to be transported into the nucleus, NLS are excellent agents for delivery by synthesizing proteins containing NLS or encoding into the Cas9 construct . While min planta methods like floral dipping (WUSCHEL2(Wus2) and BABY BOOM by using the genome-editing machinery. It has been demonstrated that the ectopic expression of DRs like Wus2, SHOOT MERISTEMLESS(STM) or MONOPTEROS (MP) induces the development of meristem-like structures in Arabidopsis. Additionally, the co-expression of Wus/STM and CRISPR/Cas9 cassette in Nicotiana benthamiana to target phytoene desaturase (PDS) gene was also carried out. The meristems later develop into shoots  mRNA, resulting in mutation of up to 65%–100% in the edited plants. Further, TRV was not detected in the progeny, subsequently protecting the progenies from any viral effects. This in planta gene-editing technique showed tremendous promise as it successfully generated small mutations in the gene. However, the method exhibited certain shortcomings in terms of identifying species-specific effective viral vectors and the gRNA-FT translocation abilities, highlighting the need for identification and characterization of viruses that infect meristematic tissues . The climate change has made imminent the need for adapting climate-smart crops as well as growing more resilient underutilized having low water requirements. A comprehensive overview of the major crops grown in drylands and their area of production is given in To mitigate the challenges of agricultural productivity in the underdeveloped and developing countries and to make agriculture sustainable under diverse climatic conditions, it is critical to develop transformative strategies for breeding pipelines by using the new breeding acceleration techniques. While tweaking selection accuracy and intensity can lead to minor improvements in a breeding context, generating novel and useful genetic variations and rapidly fixing the traits facilitate crop genetic gains can allow faster turnover of improved cultivars for accelerated delivery of improved varieties to farmers. The availability of reference genomes and ever-increasing re-sequencing data has significantly advanced breeding applications and allowed to capture the genomic diversity and its effective mining. This revelation has helped in understanding genes and the mechanisms underlying various biotic and abiotic stress responsiveness, quality, besides nutrition and plant architecture parameters, thereby aiding considerably in developing crop species with adaptive and resilient traits.Genetic resource collections that are deemed to harbor a wealth of undisclosed allelic variants are being unlocked by identifying allelic variation of relevant traits within these collections. The enormous genetic diversity present in wild species or landraces of crops as a source of allele-mining could very well be utilized and translated to elite backgrounds using genome editing tools, thereby potentially expanding the crop germplasm pool. Optimization of these tools in the underutilized food crops like sorghum, millets, groundnut, beans, cowpea, teff, banana, cassava etc. that are primarily cultivated by the poor and resource-poor farmers of the drylands would lead to huge impact in achieving the global food and nutritional security goals. This will not only accelerate the pace of the ongoing research but will potentially enable a disruptive reduction in cost for development of both farmer- and consumer-centric traits/products in these important crops. A comprehensive overview of the potential traits which can be explored in underutilized crops using genome editing has been given in Agrobacterium transformation has been successfully developed for almost all staples, there has not been a great deal of progress in improving the transformation frequencies for a many underutilized crops. Currently, transformation competent methods have been developed for crops such as finger millet  and finger millets that are known to flourish and grow well in East African climatic and soil conditions  and Rht-D1 genes imparted lodging tolerance via dwarf plant development  auxin efflux transporter (The green revolution (GR) evolved from specific requirements in nutrition and yield productivity primarily enabled by vast genetic resources of the gene banks. The transfer or replacement of dwarfing genes into cultivated crops such as rice and wheat resulted in shorter straws allowing diversion of more nutrients into grain, besides making heavier ears that allowed higher yields and better agronomic performance . The GR nditions , where lnditions . Similar quality . Similarelopment . In maiznsporter .dw (1–4) genes  has recently been sequenced and annotated (SiMTL gene that is orthologous to the maize MATRILINEAL/NOT-LIKE-DAD/PHOSPHOLIPASE A (MTL/NLDZmPLA) gene that generated haploids in maize (The dwarfing trait in sorghum has been bred using 4) genes . The pos4) genes . Editing4) genes  The diplnnotated  that couin maize . This stRht, dw3 or Br2 family genes  is a very important crop which is not only vital to food security in tropics and subtropics, but also a predominant raw material of starch industry  and granule bound starch synthase , have been reduce amylose content in cassava root starch  and fonio (Digitaria sp.) which despite their applications in food and feed, high nutrient content and high durability, are among the most under-utilized crop species in the African region  gene that conferred semi-dwarfism and significantly higher resistance to lodging resistance in tef. Similarly, homologs of OsSPL14 (squamosa promoter binding protein-like 14) gene and microRNA “OsmiR397” that have been reported to confer panicle branching trait in rice has potential to be explored and targeted in teff and fonio , a pseudo cereal belonging to Amaranthus family originated in the Andean region grows in the marginal lands is one of the best food choices due to its balanced amino acid profile, vitamins, minerals, ions, and antioxidants, quinoa received a “superfood” status and contributes to the economic and global nutritional security  that grows in some of the most hostile-to-farm landscapes despite its many superior attributes, has an unsolved quality issue of flour rancidity, posing a hindrance to its wider acceptability. Rapid development of off-flavor in pearl millet flour within 5–7 days of milling hinders the commercial use of this crop besides creating additional drudgery for women of the household, as the amount that can be pounded is limited to a few days of household use, thereby necessitating that the grain be milled immediately prior to use. A collaborative effort between the CGIAR and industry outlined a direct mechanism for hydrolytic and oxidative rancidity in millet flour, allelic variation two candidate lipase genes, PgTAGLip1 and PgTAGLip2 were identified, that correlated with the rancidity profile, confirming their function. Mutations in these key TAG lipases in pearl millet have potential in protection of lipids from TAG hydrolysis and fatty acid oxidation, leading to a reduction in off-flavour volatiles  has revolutionized GAB by facilitating development of markers for unique agronomic traits  and haveBeing a short-day legume differential genotypic sensitivity to photoperiod has major implications in adaptation of pigeonpea with respect to latitude, altitude and season. Most of the traditionally grown pigeonpea cultivars and landraces are represented by varieties from the medium- and long-duration maturity groups that mature in 150–280 days. To expand pigeonpea cultivation into new crop improvement programs, the manipulation of flowering time is likely to contribute greatly to crop yields through tailoring of cultivars to specific climates or to changes in climate that are anticipated to occur. Certain SSRs and SNPs have been identified which shed light on the pleiotropic relationship between photosensitivity and flowering time .The manipulation of flowering time is likely to contribute greatly to crop yields through tailoring of cultivars to specific climates or to changes in climate that are anticipated to occur.C. cajan offering clues to the role of PEBP (FT) family genes, based on genome-wide analyses and expression profiling. CcFT6 and CcFT8, were identified as probable Flowering locus T genes that are responsible for the production of florigen in pigeonpea. While CcFT6 upregulates under SD in photoperiod sensitive, MAL3 genotype, CcFT6 and CcFT8 upregulate in photoperiod insensitive genotype (ICP20338) under SD and LD conditions, respectively. The presence of CcFT8 as an additional florigen producing gene, having ability to flower in a photoperiod independent manner under LD conditions provide some clues on its photoperiod insensitive nature  declares that genome editing is almost equivalent to conventional breeding and therefore, does not require any regulatory process within the United States. Some speculations from USDA define that gene-edited plants can be considered as a separate category . In a reArgentina’s regulatory process is in accordance with the Cartagena Protocol on Biosafety and evaluated on case-by-case assessment, irrespective of the method used for product development . If the While in Australia and New Zealand, products developed through CRISPR/Cas9 and other editing tools are excluded from the regulatory process , the EurIn India, a recent notification has exempted Site Directed Nuclease (SDN) 1 and 2 types (SDN1 and SDN2) of products which do not carry any vector DNA and are like the products of spontaneous or induced mutations from the transgenic regulation and risk assessment under Rules 1989. Guidelines for the safety assessment of genome edited plants 2022 have been released in May 2022, that define various categories of genome edited plants and determine regulatory requirement for appropriate category and provide the regulatory framework and scientific guidance on data requirement .Similarly, Philippines has moved ahead with, a policy discussion paper under review and consideration on how products of new plant breeding technologies should be treated under existing regulatory regime, the benefits that may be derived and the capacity of the country to utilize such techniques. The policy framework will rely on a case-to-case and crop-to-crop based decision or regulatory pathways which will be the entry point of any genome edited plant products with or without involving the insertion of genes from non-sexually compatible species. However, in regions where the technology and infrastructure has not advanced enough, containment and monitoring measures are expected to be comparatively strict.In South Africa, SDN-1 involving “small, targeted and untargeted inserts or deletions based on non-homologous end joining (NHEJ)” resulting from ZFNs, MNs, TALENs and CRISPR/Cas and considered to be exempt from GM Act . The regWhile CRISPR/Cas9 is an inexpensive and flexible technology, international harmonization of the regulatory frameworks needs to be developed to ensure that these are based on sound science and the community of practices developed around the world . More dehttps://www.avisaproject.org/), Excellence in Breeding (EiB) Platform (https://excellenceinbreeding.org/), Crops to End Hunger (CtEH) (https://www.cgiar.org/excellence-breeding-platform/crops-to-end-hunger/) and various CGIAR Agri-food research programs (https://www.cgiar.org/research/research-portfolio/). These initiatives put major emphasis on modernizing breeding mainly through developing specific regional product profiles, mechanized operations and databases, besides focusing on infra-structure/human capacity development for efficient breeding and seed systems. Nonetheless, crops face intractable problems not easily solved by traditional breeding and hence there is a need for future breakthroughs in global agriculture. These ongoing breeding modernization agendas integrate innovations in advanced breeding tools (ABTs) such as CRISPR/Cas that are increasingly becoming relevant to fill gaps in the pipeline research required to deliver high yielding, nutritious and climate resilient crop varieties as per the regional demands. Integrating the ABTs such as CRISPR, reverse breeding, double haploids etc. in the “modernized crop breeding platforms” will not only provide game changing solutions to some of the most “intractable” traits but may also be used for enhancing the expression of superior alleles and removal of deleterious effect alleles. Furthermore, these tools and methodologies may be deployed to reverse domestication by editing genes related to domestication traits in wild species making superior lines with enhanced stress resistance for crop improvement. However, accomplishing these desired impacts would require having curated crop genotyping data sets integrated with the trait data from various crop germplasm panels to assist the discovery of trait-specific SNPs and haplotypes for further excavation of superior genes/alleles that may be subsequently deployed for gene editing applications. To support these endeavors, adaptive and user-friendly allele mining platforms need to be in place to manage and mine the massive datasets that have been generated by sequencing reference genomes and re-sequencing efforts on hundreds of new accessions and large transcriptome datasets.To achieve sustainable increase in the rate of genetic gains in food crops, transformative strategies for accelerated crop breeding pipelines need to be embraced. Several national agricultural research system (NARS) initiatives are ongoing under several major initiatives for Africa including Modernizing Ethiopian Research on Crop Improvement (MERCI). Accelerated Varietal Improvement and Seed Delivery of Legumes and Cereals in Africa (AVISA) (in vitro regeneration and transformation pose a major challenging obstacle. Moreover, the genotypic effects on plant regeneration and transformation can be very challenging. To overcome the problems posed by tissue culture and low transformation efficiencies in important crop species, several plant transformation systems such as RNP based systems or transformation free systems need to be established to increase the precision and editing efficiency of plant genome editing.CRISPR/Cas technology has made remarkable progress in recent years for its practical applicability for targeted genome editing in plant species including crop plants. However, certain obstacles such as transformation efficiency and off-target mutations still need to be overcome. For underutilized crops that are less researched, There have been continuous efforts in development of the tools and applications which has helped us discover newer technologies with each passing decade. The addition of these advanced tools and technologies in the breeder’s tool kit holds tremendous potential to bring changes precisely and efficiently in the genetic makeup of the ruling elite varieties, significantly reducing the need for long breeding cycles for incremental traits speeding up the rate of genetic gains. In addition to the CRISPR/Cas system, several other recently developed systems such as base editing and prime editing have revolutionized the conventional breeding approaches and provided a new direction to the crop improvement programs. With the advancement in new prediction system for on-target activity such as sgRNA CNN , an arraArchaea and undoubtedly, has emerged as the most powerful technology due to its precision, cost effectiveness, and uniqueness to overcome the shortcomings of crop breeding. While ensuing climate change, exploration and creation of additional genetic diversity of underutilized crops require using these precision genetic tools, to create impact on ground, such efforts need to be framed within a breeding pipeline mindset and should be included in the product design process. Although the regulatory pathway for gene edited products is expected to be less complex than for GMOs in several geographies, issues such as freedom to operate and securing the social license need consideration at the intervention design stage. Although CRISPR has potential to deliver disruptive innovations, the trait prioritization should consider the breeding product profiles and market segments for designing and accelerated delivery of locally adapted and preferred crop varieties for the drylands.In conclusion, the evolution of genome editing tool kit over the decade has been escalating since the discovery of Cas9 from


A particular focus of this RT is placed on longitudinal research, which is still underrepresented compared to the abundantly available cross-sectional studies. Longitudinal studies are, however, urgently needed to understand aging better. A further emphasis is placed on understanding “healthy aging,” thus excluding issues related to dementia, mild cognitive impairments, or clinically relevant limitations in other behavioral domains. Sixteen manuscripts have been accepted for final publication. Seven papers investigated longitudinal changes concerning neuroanatomical, neurophysiological, and psychometric aging issues . Six papers describe training and intervention effects in older adults regarding cognitive abilities and brain activations . The remaining three papers report additional age-related findings, either from cross-sectional or methodological studies .This Research Topic (RT) covers papers presented at the 5th International Conference on Aging & Cognition in Zurich  database project  deviated from this homogeneity and showed an accelerated decline with advancing age. Concerning subcortical regions, change trajectories were highly variable, with some areas showing strong  and others slight decline .First, we shortly describe the longitudinal contributions included in this RT. The papers by Oschwald et al. focused on the white matter microstructure and identified age-related fractional anisotropy declines for selected fiber tracts. Although clear declines were also evident for motor ability measures, these changes were only sparsely related to the anatomical changes. Jockwitz et al. report a cross-validation analysis comprising longitudinal data of the 1000BRAINS project conducted at the Research Center Juelich (Germany) and the LHAB project in Zurich. The authors demonstrate that annual percentage changes in cortical thickness revealed similar results for the two independent samples. Between-sample differences were only marginally present after correction for major covariates. This suggests that the development over time is generalizable over independent samples with similar demographic characteristics when the same methodology is used. Lannen et al. and Wehrle et al. describe study designs of two extremely long-lasting (60 years!) retrospective and prospective longitudinal studies conducted in Zurich (Switzerland). The two reports comprise the applied methods, description of cohorts, statistical analysis, ethical problems, data reconstruction, and data validation.Lacker et al. focus on longitudinal changes over 4 years of steroid hormone concentration , genetic markers, psychological wellbeing, and physical health in men between 40 and 75. They identified that psychological wellbeing and physical health remained stable over time. Further, they revealed that estradiol moderated the course of psychological wellbeing, while the androgen receptor gene polymorphism moderated the course of physical health. These results suggest that the hypothalamic-pituitary-gonadal (HPG) axis is essential for maintaining psychological wellbeing in men. At the same time, physical health depends more on interindividual differences in the androgen receptor gene and testosterone. Wank et al. provide short-term longitudinal data based on ambulatory assessment technologies. They studied real-world conversations over 4 days in a sample of cognitively normal older adults using a smartphone application known as the Electronically Activated Recorder (EAR). Specifically, they extracted instances where the participants shared memories and future thoughts, scoring them for their make-up of episodic and semantic detail. They revealed that older age was associated with reduced real-world sharing of autobiographical episodic and semantic memories. In addition, autobiographical episodic memories of older people appeared to be less detailed.This group of longitudinal studies increases our understanding of how brain anatomy, brain activation, behavior, and endocrinology change in the context of healthy aging. Beyond this, the studies highlight the importance of longitudinal datasets that comprise more than two measurement occasions to enable the advanced analysis of within-person change trajectories. Furthermore, they emphasize the necessity of combining and cross-validating findings across different research groups using different samples  or in performing single tasks (control group). A group of untrained younger adults served as an additional control group. The authors observed training-specific improvements in task switching and training-specific effects at the neuronal level measured with electroencephalography.The next group of papers in this RT addresses training and intervention effects in older adult samples regarding cognitive abilities and associated brain activations. Gajewski et al., participants completed a 16-weeks-lasting training intervention during which they practiced various cognitive functions . Compared to active and passive control groups, cognitive training enhanced performance in several cognitive tests dissimilar to the intervention, thus providing some evidence for transfer. A transfer to daily activities was not observed. Dziemian et al. conducted a longitudinal diffusion tensor imaging study to investigate the effects of an intensive, adaptive 4 week-lasting working memory training on white matter microstructure in young and older adults. The authors quantified the anatomical changes using global and tract-wise mean diffusivity measures. As expected, the behavioral analysis showed increased working memory performance after the working memory training. The neuroanatomical analysis revealed decreased mean diffusivity in the older working memory training group after the training intervention in some brain areas . Thus, this study suggests that the behavioral training gains are associated with training-induced changes in white matter integrity in fiber tracts known to underlie working memory performance.Older adults trained in cue updating while switching showed a reduction in mixing costs indicated by the cue-related P3 component and the associated P3 topography. These results are taken as support for an improvement in preparatory updating processes. However, they did not obtain training-specific improvements in the context updating at the behavioral or neuronal levels, suggesting that there are limitations in the transfer of cognitive training. In the training study of Mikos et al. analyzed whether cognitive training is accompanied by changes in task-induced deactivation within the default mode network (DMN). They analyzed data collected from a group of healthy older adults who completed a 6 week, process-based, object-location memory (OLM) training and an active control group. As a main result, the authors identified that the dorsal DMN was deactivated during the in-scanner task relative to a visual fixation baseline. In addition, this deactivation was more pronounced in the OLM training group relative to the active control group. Many studies have shown that cognitive and motor learning strongly depends on the evaluation of feedback information.de Haan et al. conducted a study to examine the relation between learning-related neural processes and feedback-based learning in two age groups. Their results show that less successful learning in older adults corresponds to less effective processing of behavior feedback. At the neural level, activation following positive and negative feedback was less distinctive in older adults due to a smaller feedback-evaluation response to positive feedback in this group. These results suggest that diminished learning performance with age can be attributed to a reduced evaluation of positive feedback and reduced knowledge updating related to changes in choice behavior. In another branch of cognitive training and intervention research, the focus is placed on cognitive augmentation and rehabilitation using non-invasive brain stimulation. In their paper, Figeys et al. comprehensively reviewed the existing literature on transcranial direct current stimulation (tDCS) on cognitive performance and hemodynamic response patterns measured with near-infrared spectroscopy (NIRS). They applied rigorous literature research to identify eight studies using simultaneous tDCS and NIRS in older adult samples. Based on these studies, the authors concluded that tDCS effects on cognitive performance and functional NIRS metrics are most prominent in young, healthy adults and appear to become less robust with age. At the same time, the authors are cautious in drawing firm conclusions due to the small number of studies.However, the ability to learn from feedback declines with age. Since the neural mechanisms underlying this age-related change in feedback processing are still insufficiently understood, In summary, these training and intervention studies align with pertinent literature . Ramanoël et al. examined how older adults process visuospatial information. In their functional magnetic resonance imaging (fMRI) study, young and healthy older subjects performed a landmark-based navigation task while the associated hemodynamic responses were measured. The authors found that the older adults' navigational abilities were overall diminished compared to young adults. Most importantly, the left inferior temporal gyrus and the left amygdala/hippocampus region were less activated during landmark-based navigation in older compared to younger subjects. This between-group difference only appeared when the landmark condition was contrasted with the control condition. They suggest that in older age, fine-grained information processing in occipital and temporal regions is reduced, thus hindering the capacity to use landmarks adequately for navigation. This work supports a better understanding of the neural dynamics subtending landmark-based navigation and provides new insights on the impact of age-related visuospatial processing differences on navigation capabilities.The final set of contributions for this RT comprises three papers reporting age-related cross-sectional or methodological studies  method for analyzing large-scale datasets. They applied this technique to NIRS data, which measured cerebral oxygenation during standing in a large study cohort . The application of fPCA elegantly reduced a large amount of data to a few parameters, summarizing the inter-participant variation of the obtained data during standing. Thus, this technique has been proven useful for data-driven approaches to high-resolution data analysis in aging research.Modern studies on aging must rely on sufficiently large and complex datasets, particularly when collecting neurophysiological data. Consequently, data analysis becomes more and more challenging. This RT provides a comprehensive overview of the current approaches to studying cognitive, neurophysiological, and neuroanatomical aging. The longitudinal studies show that aging is accompanied by specific neuroanatomic and neurophysiological changes. These studies have also emphasized the importance of collecting longitudinal data across more than two time points. In addition, evidence is provided that the identified changes are robust and consistent across different study samples. The Zurich Longitudinal Studies (ZLS) study protocols, which comprise an outstanding long observation period, promise essential insights into the lifelong cognitive aging processes. The intervention and cross-sectional studies emphasize that the aging brain and the associated behaviors can be influenced by training. Therefore, we hope this RT will prompt a more positive view of the opportunities and resources linked with healthy aging. In addition, we hope that this research area will attract more researchers from related disciplines such as biology, psychology, neurology, psychiatry, and public health. Strengthening interdisciplinary collaboration will improve our understanding of the complex human aging process and facilitate innovation to support a healthy aging population (Cherbuin et al., LJ, MM, CR, and SM writing of the manuscript. All authors contributed to the article and approved the submitted version.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.