Identifying genuine cortical stimulation-elicited electroencephalography (EEG) is crucial for improving the validity and reliability of neurophysiology using transcranial magnetic stimulation (TMS) combined with EEG. In this study, we evaluated the spatiotemporal profiles of single-pulse TMS-elicited EEG response administered to the left dorsal prefrontal cortex (DLPFC) in 28 healthy participants, employing active and sham stimulation conditions. We hypothesized that the early component of TEP would be activated in active stimulation compared with sham stimulation. We specifically analyzed the (1) stimulus response, (2) frequency modulation, and (3) phase synchronization of TMS–EEG data at the sensor level and the source level. Compared with the sham condition, the active condition induced a significant increase in TMS-elicited EEG power in the 30–60 ms time interval in the stimulation area at the sensor level. Furthermore, in the source-based analysis, the active condition induced significant increases in TMS-elicited response in the 30–60 ms compared with the sham condition. Collectively, we found that the active condition could specifically activate the early component of TEP compared with the sham condition. Thus, the TMS–EEG method that was applied to the DLPFC could detect the genuine neurophysiological cortical responses by properly handling potential confounding factors such as indirect response noises. Recent technological advances in combining transcranial magnetic stimulation (TMS) and electroencephalography (EEG) have enabled direct measurement of cortical responses elicited by TMS, resulting in visualization of neural responses at the stimulation site and their propagation to other areas as time series EEG signals ,2. SimulWhile the TMS–EEG method has shown great potential as a neuroscientific tool to investigate the pathophysiology of neuropsychiatric disorders ,11, therSeveral TMS–EEG studies thus far have been conducted using the sham condition, in which the coil position and angle were shifted from the target site on the scalp to address these confounding factors ,26,27. HAgain, in the sham stimulation condition with peripheral electrical stimulation to the scalp, the magnetic field generated by an active TMS coil is not produced in principle. Thus, the elicited EEG activity generated by the conventional sham condition should be fundamentally different from TEP with active condition. In other words, the limitation of the conventional sham coil (or condition) was that the coil design (or condition) did not account for the effects of the fluctuating magnetic field generated around the coil.To date, a few studies on M1 used sham stimulation conditions that exposed the scalp to a weak and shallow magnetic field, indicating that the sham stimulation conditions induced clearly different TEP components compared with a standard active coil stimulation ,29. On tIn this study, we aimed to investigate and compare the differences between active and sham stimulation conditions with respect to the biophysical characteristics of TEP in terms of (1) amplitude , (2) frequency , and (3) phase , with a particular focus on the early and late components of TEP. Therefore, we hypothesized that TEP in the left dorsolateral prefrontal cortex (DLPFC) generated by the active coil would be more activated than the TEP elicited by the special sham coil in terms of these aspects. Given that the late component of TEP is susceptible to noise derived from peripheral stimulation ,21, we sA total of 28 healthy participants : 33.9 ± 11.0 years old) who met the following criteria participated in the study: (1) between ages 18 and 65 at the time of obtaining consent; (2) no history of neuropsychiatric disorders, as assessed by the Mini-International Neuropsychiatric Interview; (3) normal cognitive function, as assessed by the Mini Mental State Examination (MMSE)   over the left M1 was confirmed. Next, the resting motor threshold (RMT) was determined as the minimum stimulus intensity that produced MEPs of 50 µV or more in the target muscle over 50% of the stimulus trials to the left M1 with the EEG cap in place .In this TMS–EEG experiment, single-pulse TMS was applied to the left DLPFC of each participant. TMS pulses were administered with 500 ms jittering, thus the intertrial interval was set at 4.5–5.5 s. The DLPFC site  was identified by an MRI-guided navigation system  using the individual MRI for each participant see the . The stiFurthermore, participants were blinded to the stimulus conditions applied in each TMS experiment to exclude psychological effects of the participants on the EEG data during the experiment. To minimize the auditory-evoked potentials elicited by TMS click sounds, white noise was applied to participants using an earplug sound stimulation system . The volFinally, to ensure that the active and sham conditions were appropriately blinded for the participants, we asked each participant if they felt any differences during the TMS examination in the intensity of TMS stimulation sensations, the range of head stimulation, and the loudness of the TMS coil click sounds between each stimulation session at the end of the experiment. In this regard, we used Yes/No questions (closed questions) to avoid ambiguous answers, and if the answer was “yes” , we asked them to respond in detail about how they felt the difference.In this study, a monophasic TMS stimulator  and a figure-of-eight butterfly coil with 2 × 70 mm diameter windings  were used to conduct single-pulse TMS experiments. As for the sham condition, a special sham coil which is indistinguishable from an active stimulation coil  was used. Specifically, the sham coil used in this study generates no maximum magnetic flux in the focal region of the figure-of-eight coil. As a result, no effective current is induced in the cortical region just beneath the coil focus. However, this sham coil produces effective magnetic flux equivalent to the active stimulation in the region peripheral to the coil away from the focal area. Thus, the magnetic flux distribution outside the focal area  by sham TMS to the cortex is supposed to stimulate like the active stimulation . The detEEG data were processed offline using the EEGLAB v2021.0 and customized scripts running on MATLAB software  ,36. We uGlobal cortical activity was assessed by averaging the global mean field power (GMFP) ,41 of eaCustom scripts based on the FieldTrip toolbox  were useTo estimate the functional connectivity between brain regions for each stimulus condition at the sensor level, we used the weighted phase lag index (wPLI), which has been shown to be robust to noise in previous studies ,46,47. TThe modulation index (MI) , which rFor the TEP signal source estimation, MNE software  was usedFor the GMFP and LMFP analysis, a paired t-test  was applied to determine the area of the early component 30–60 ms) and late component (100–200 ms) after stimulation between both stimulus conditions . The fal ms and lp = 0.033). In contrast, no significant difference was observed for the later component of the LMFP between the two conditions (see p = 0.007); (2) No significant differences were identified in the late component (100–200 ms).The GMFP analysis of the TEP in active and sham conditions showed no significant differences in either the early component or late component. On the other hand, in the LMFP analysis focused on the stimulated area, the LMFP in the active condition was higher than that in the sham condition in the early component (30–60 ms)  in the active condition compared with the sham condition in θ- and β-bands in the time interval of approximately 30–200 ms post-stimulation  between the active and sham conditions in the DLPFC at the stimulation site, indicating that active stimulation significantly enhanced the θ–γ coupling compared with the sham condition  and the late component (100–200 ms) (p = 0.042) at the brain regions corresponding to the stimulated areas  = 0.55, p = 0.46); (2) feeling of spread of the stimulus site  = 0.03, p = 0.85); and (3) feeling of loudness of the coil click sound  = 0.87, p = 0.35). Of note is that the results of the questionnaire showed no significant difference between active and sham stimulation with respect to each element of stimulus sensation.The results of the differences in stimulus sensation  between active and sham stimulation coils are as follows: (1) feeling of stimulus intensity  is significantly affected by AEP and SSEP . Ross etIn this context, our results show no significant difference in the late component between the two conditions in the sensor-level analysis see . FurtherThe sensor-based, as well as source-based, total power analyses showed no significant difference in TMS-elicited total power between the active and sham stimulation conditions. In contrast, the ITPC , which rThe effects of active stimulation to the left DLPFC on EEG were also evaluated by graph theory-based analysis. In particular, BC in the left fronto-parietal region in the θ- and β-bands were increased, indicating that network efficiency through this region may be enhanced by the ipsilateral stimulation. Furthermore, since more robust increases in the network efficiency in the θ- and β-bands were observed in active stimulation compared with sham stimulation, it is likely that active stimulation is causing a genuine TMS-evoked response by directly stimulating the cortex rather than spurious EEG changes. Moreover, compared with sham stimulation, active stimulation resulted in higher hubness in β-band defined by the four indexes used in the graph theory-based analysis. In addition, active stimulation significantly increased θ-phase and γ-amplitude coupling at the left fronto-central area  with the active condition compared with the sham condition. Synchronization of oscillations with specific frequency bands is considered to represent the mechanism by which information is processed in the network ,70. The There are some limitations in the present study. First, the time–frequency analysis in this study was restricted to the stimulation area of TMS. Therefore, future comprehensive analysis of spatiotemporal information other than the stimulation area may provide further new insights into EEG changes induced from the DLPFC stimulation using TMS–EEG. In our sham coil study, we demonstrated that the coil click sounds produced by the sham coil were 5 dB louder than those by the active coil . HoweverThe present study demonstrated that the combined TMS–EEG technique could detect genuine neurophysiological cortical responses by appropriately addressing potential confounding factors that could be the sources of noise. In recent years, TMS–EEG studies have noted that it is difficult to evaluate genuinely cortex-derived TEPs because TEPs can be contaminated with mechanical and myoelectric noises from TMS, as well as the brain activities derived from peripheral stimulation . However, our results indicate that the TMS–EEG modality can measure and evaluate the cortex-derived TEPs that are specific to the active TMS, although there are certain limitations. The application of this modality to various TMS–EEG neurophysiological paradigms accelerates the clinical application of this modality from healthy participants to patients with neuropsychiatric disorders.


Fibromyalgia is a condition associated with widespread musculoskeletal pain, fatigue and sleep problems. Fibromyalgia treatment guidelines recommend non-pharmacological interventions and the development of self-management skills. An example of a programme that fits these guidelines is the Fibromyalgia Self-management Programme (FSMP) which consists of one 2.5-hour weekly session over six successive weeks and includes education about fibromyalgia, goal setting, pacing, sleep hygiene and nutritional advice. The FSMP is currently provided in a secondary care hospital setting and co-delivered by a multidisciplinary team. Delivery in a primary care setting has the potential to improve the accessibility of the programme to people with fibromyalgia. Therefore, this feasibility study aimed to determine the practicality and acceptability of conducting a future definitive randomised controlled trial of the FSMP in a community setting.An exploratory, parallel-arm, one-to-one, randomised controlled trial. Participants were recruited from general practices across South West England, and the FSMP was co-delivered by physiotherapists and occupational therapists across two community sites. To determine the outcome measures for a future definitive trial several were tested. The Revised Fibromyalgia Impact Questionnaire, Arthritis Self-Efficacy Scale-8, Chalder Fatigue Scale, Short form 36, 5-Level EQ-5D version and Jenkins Sleep Scale were collected at baseline, 6 weeks and 6 months. Semi-structured interviews were conducted with patient participants, occupational therapists and physiotherapists to explore the acceptability and feasibility of delivering the FSMP in a community setting.n = 38) or control arm (n = 36). Attrition from the trial was 42% (31/74) at 6 months. A large proportion of those randomised to the intervention arm  failed to attend any sessions with six of the 13 withdrawing before the intervention commenced. The proportion of missing values was small for each of the outcome measures. Three overarching themes were derived from the interview data; (1) barriers and facilitators to attending the FSMP; (2) FSMP content, delivery and supporting documentation; and (3) trial processes.A total of 74 participants were randomised to the FSMP intervention (It is feasible to recruit people with fibromyalgia from Primary Care to participate in a randomised controlled trial testing the FSMP in a community setting. However, improvement in trial attrition and engagement with the intervention is needed.The trial is registered with ISRCTN registry and was assigned on 29/04/2019. The registration number is ISRCTN10824225. Fibromyalgia (FM) is a common condition that affects over 5% of the UK population , 2 and hMembers of the Rheumatology Therapies team at the Royal United Hospitals Bath (RUHB) developed the Fibromyalgia Self-Management Programme (FSMP). The programme is a non-pharmacological, multidisciplinary exercise and education group intervention that aims to provide condition-specific, patient-centred education and exercise advice and support the development of self-management skills. The FSMP is delivered in 2.5 hour sessions over six successive weeks and includes sessions on education about FM, goal setting, pacing, sleep hygiene and nutritional advice. Using Michie Behaviour Change Taxonomy , a recenUntil recently, the FSMP has been provided in a secondary care hospital setting and co-delivered by a multidisciplinary team. Local audit data reports good patient satisfaction and improvements in self-efficacy to manage FM symptoms. However, this has yet to be formally evaluated using robust quantitative research methods. The UK government plans to increase investment in community-based healthcare services to deliver care for those living with long-term conditions closer to home. Therefore, some services that were once provided in acute hospitals will now be transferred to the community , 20. A rThis feasibility study aimed to determine the practicality and acceptability of conducting a future definitive randomised controlled trial (RCT) of the clinical and cost-effectiveness of a community-based FSMP.Determine the feasibility of training Band 6 PTs and OTs to deliver the FSMP in the community;Explore the feasibility of recruiting adults with FM to the trial from primary care;Assess the feasibility of collecting a range of outcome data to identify the primary outcome for a future trial;Assess the feasibility and acceptability of collecting health economic data;Determine the recruitment rate and calculate the sample size for a full trialDetermine the safety of delivering the FSMP in a community settingUnderstand the patient and health professional acceptability of delivering the FSMP in the community.The specific objectives were to:This feasibility randomised controlled trial followed a pre-specified protocol  (ISRCTN Two patient Research Partners (RP) were recruited and contributed to the study conception, design and interpretation of findings. The PPI perspective was represented at all trial management meetings, and both RPs gave invaluable guidance and support to the research team throughout the development and delivery of the study.Participants were recruited from research-active general practices (GP) across SW England. The practice manager, research nurse or general practitioner at consenting GP sites, using GP read codes, conducted a database search for patients diagnosed with FM and aged 18 and over. From the identified potential trial participants, a member of the GP team screened for eligibility and suitability . The GP then sent an information pack by mail, which included; an invitation letter from the GP; a detailed participant information leaflet (PIL); the contact details of the research team; a reply slip, and a prepaid envelope. Interested participants returned the reply slip or contacted the research team by telephone or e-mail.Potential trial participants were then screened over the telephone by the Chief Investigator (CI) (JP) for further eligibility criteria and were excluded if they had a General Anxiety Disorder Assessment (GAD-7) score > 15 , had prehttps://www.versusarthritis.org/about-arthritis/conditions/fibromyalgia [The intervention consisted of a 6 week, FM condition-specific group programme delivered by a PT and an OT. The intervention was delivered four times at two selected community sites in SW England . Trial site A was situated in a community GP in a city, and Trial site B was located in a rural community hospital. The FSMP comprised a 2.5 hour weekly group session over six consecutive weeks. Each week, the course focused on supporting individual self-management skills by increasing knowledge and understanding of the condition, medication, goal setting, pacing, dietary advice, sleep hygiene, relaxation, and exercise. All participants attended the exercise sessions but could opt-out at any time if they needed a rest. Participants also received a booklet that contained information on the programme, information about FM and self-management strategies, online links to other relevant resources  and Fibomyalgia )) and woThe FALCON training programme was developed by the research team and delivered by RUHB clinical lead occupational therapist (SD) and physiotherapist (JR), both of whom were instrumental in creating the existing secondary-care FSMP at the RUHB. Four physiotherapists and two occupational therapists attended a two-day training programme at the RUHB in February and March 2019. Two additional therapists were trained to ensure that the intervention could be delivered if one or more therapists were absent. The training consisted of an overview of the study and the FSMP, including documentation and administration, strategies when managing groups, and all content included in the FSMP . To offer further support and to ensure intervention fidelity JD or SD attended two of the six FSMP community sessions. If the delivering therapists had any concerns, they could contact either facilitator by telephone or e-mail.Participants continued to receive usual care from their GP, they were not discouraged from seeking additional healthcare for their FM symptoms but were advised to contact the research team if they were referred to a pain self-management programme. Once data collection was complete, participants randomised to the control arm were sent an FM information leaflet designed by the research team. The leaflet provided information on FM, current treatments and information about local support groups.The outcomes that were of particular relevance for the feasibility of a future trial included the number of patients identified with FM in primary care, percentage of FM patients deemed eligible, recruitment to the feasibility trial as a percentage of those contacted, number of analysable completed patient-reported questionnaires, attendance at the FSMP and number of patients who drop out of the FSMP.To determine the outcome measures for a future definitive trial, several outcomes were tested. All clinical outcome measures were patient-reported. Participants randomised to the intervention arm returned the outcome measures by post at baseline and 6 months but completed the 6 week outcome measures on-site at the end of the intervention. Those who did not attend were sent the questionnaires by post. Participants in the control arm were sent the outcome measures by post for all three time-points.To identify a suitable and feasible primary outcome measure for the definitive trial, the research team collected a range of FM symptom-based, quality of life (QoL) and self-management specific outcome measures. To assess the impact of FM symptoms, the Revised Fibromyalgia Impact Questionnaire (FIQR), a validated outcome measure to evaluate FM, was used . As fatiTo assess the feasibility of collecting health economic data, the Client Service Receipt Inventory (CSRI) was adapted for FM to collect health and social care use , 43. It To account for loss-to-follow, missing data and estimate parameters such as the participation or completion rates, and those required to derive the sample size for the main trial with enough precision. The trial aimed to recruit a total sample size of 70 participants, with a minimum of 35 in each arm.Participants were randomised to either the control arm or the FSMP using a parallel 1:1 study design. Randomisation was stratified by Trial site and cohort. Randomisation was conducted independently by the Research and Development (R&D) team at the RUHB using online software . To presParticipants in the intervention arm attended the FSMP, therefore participant blinding was not possible. The therapists delivering the FSMP, the research associate (JC) and the data analyst (SP) were also unblinded to participant allocation.A nested qualitative study was conducted to understand the acceptability of FSMP and whether it was feasible to be delivered in a community setting. After completing the programme, all patient participants randomised to the intervention and the therapists delivering the programme were invited to take part in semi-structured interviews to share their experiences of the FSMP. Of those patients who responded, participants were purposively selected based upon key characteristics including Trial site, age, gender, the severity of FM and attendance at the FSMP. The semi-structured interviews were conducted by JC .Quantitative descriptive analysis included the number and percentages of participants approached, recruited and retained in the study and the completion of the intervention and outcome data. The final data included reasons for non-participation, withdrawal, missing data and noncompliance with the protocol with the emphasis on how these may impact on the full-scale trial. These rates are presented by trial arms to investigate any differences requiring particular attention in the design of the main trial. Deviations from the protocol were recorded and reported by relevant categories to identify areas requiring particular attention during the design of the main trial. Descriptive statistics, including means and standard deviations, were used to analyse the patient-reported outcome measures. Sample size calculations for a full RCT were performed using G*Power (version 3.1.9.4), 90% power, and a two-sided alpha of 5% . The datn = 3) doubled coded (JP) to ensure all data was captured and the interpretation was refined. The codes were then grouped into categories and thematically analysed [The qualitative research was underpinned by a qualitative description approach . All intanalysed , 49 to dSix of the FSMP sessions were audio-recorded for fidelity purposes. A coding framework was developed by the research team and used as a fidelity assessment tool , 51. TheAll data were managed in accordance with the 2018 Data Protection Act . All sern = 74) were randomised to either the intervention (n = 38) or the control arm (n = 36). Baseline data for six participants from the intervention arm and two from the control arm were not received, leaving n = 66 available for quantitative data analysis (n = 32 intervention and n = 34 control).Between April and August 2019, 20 General Practices across two sites in SW England invited 1414 patients with an FM diagnosis and aged 18 or over to participate in the study. A total of 217 patients (15%) responded, 198 (14%) were screened for eligibility and 77 (5%) consented to take part. Of the 19 participants who were not screened for eligibility, 42% were unable to contact, 5% were no longer interested in participating in the study, and 53% wanted to participate but there were insufficient places available at Trial Site A. Data were not collected for those who were invited but either declined or did not respond. Three participants withdrew prior to randomisation . The remaining participants  at 6 months. Attrition was higher in the intervention arm compared to the control arm (32% versus 22% at 6 weeks and 53% versus 31% at 6 months). See Fig. /74 at 6 A large proportion of those randomised to the intervention arm  failed to attend any sessions (six of the 13 withdrew before the intervention commenced). This was particularly noticeable in cohort 2 in Trial site B, where 7/9 participants did not attend any sessions  (Table n = 66).The control arm  were on average older than the intervention arm . CSRI data were available for 100% of participants who returned questionnaires at each time point .Findings from this feasibility study and published literature were used to inform decisions about how much change in each outcome measure might be considered clinically relevant. These changes were calculated at 6 weeks and 6 months using data from the feasibility study control arm (patients randomised to the standard care arm only) and derived sample size calculations for a full size trial/RCT accordingly. Further details are presented in Table The minimum clinically important difference (MCID) for FIQR is 14% improvement . This MCn = 32) and four therapists delivering the programme were invited to take part in semi-structured interviews. Of the 32 patient participants invited, 19 responded. Thirteen patient participants were selected and consented to an interview. The qualitative interviews were conducted by JC between September 2019 and January 2020. Patient participant interviews (n = 3) took place via telephone (n = 6) or face-to-face at the participants home (n = 4) or in an interview room at the University (n = 3). Therapist interviews (n = 4) took place at the University (n = 2) or individuals’ workplace (n = 2). The duration of interviews ranged from 8 minutes (an interview with a participant who did not attend the intervention) to 127 minutes (mean: 58 minutes). Tables . Three overarching themes describing the experience of the FSMP from both the patients’ and therapists’ perspectives emerged and included: barriers and facilitators to attending the FSMP; FSMP content, delivery and supporting documentation; and Trial processes.All patient participants who were randomised to the intervention arm and were actively involved in the study  or participated in a pain management programme would already have the skills to self-manage. Therefore, it is proposed that a full trial includes participants who attended a pain management programme > 12 months previously. Individuals with Rheumatoid Arthritis were excluded from the feasibility study, but those with other inflammatory rheumatic diseases such as, Ankylosing Spondylitis and Lupus were included. As FM is prevalent in several inflammatory rheumatic diseases and appears to affect disease severity , it is pThe study successfully recruited patients diagnosed with FM from primary care. As intended, the FSMP programme was delivered twice across two sites in SW England. This feasibility study has shown that it is possible to successfully train non-specialist therapists to deliver the FSMP in a community setting. Finally, the nested qualitative research provided an understanding of trial processes, including why some patients failed to engage with the FSMP and the acceptability of the FSMP from patient and therapist perspectives.A limitation of the study was the high attrition rate (42%). Further to this, it was impossible to blind participants and treating therapists in this feasibility study. Blinding the participants and clinical staff will be challenging in further study. Blinding the data analyst would help to minimise the potential for bias in a full RCT .Although the COVID-19 pandemic did not affect the delivery of this study, it may have delayed the return of the 6-month questionnaire data. Due to UK national lockdown, the study team received outcome data up to 4 months after the 6-month outcome data was initially sent. Additionally, although the medical record review was planned to be conducted by the RA in person, this was conducted via e-mail which impacted the timing of this study and may have impacted the data quality.It is feasible to recruit people living with FM from primary care to participate in an RCT testing clinical and cost-effectiveness of the FSMP delivered in a community setting. However, improvement in trial attrition and engagement with the intervention is needed. In addition, trial inclusion criteria should be refined to include those with inflammatory rheumatic diseases and those who have attended pain self-management programmes more than 12 months previously. An initial assessment by a therapist before attending the FSMP is also warranted to ensure patient readiness. Finally, it is suggested that a future trial incorporates an investigation of virtual delivery of the FSMP.


KDM4 proteins are a subfamily of histone demethylases that target the trimethylation of lysines 9 and 36 of histone H3, which are associated with transcriptional repression and elongation respectively. Their deregulation in cancer may lead to chromatin structure alteration and transcriptional defects that could promote malignancy. Despite that KDM4 proteins are promising drug targets in cancer therapy, only a few drugs have been described as inhibitors of these enzymes, while studies on natural compounds as possible inhibitors are still needed. Natural compounds are a major source of biologically active substances and many are known to target epigenetic processes such as DNA methylation and histone deacetylation, making them a rich source for the discovery of new histone demethylase inhibitors. Here, using transcriptomic analyses we determined that the KDM4 family is deregulated and associated with a poor prognosis in multiple neoplastic tissues. Also, by molecular docking and molecular dynamics approaches, we screened the COCONUT database to search for inhibitors of natural origin compared to FDA-approved drugs and DrugBank databases. We found that molecules from natural products presented the best scores in the FRED docking analysis. Molecules with sugars, aromatic rings, and the presence of OH or O- groups favor the interaction with the active site of KDM4 subfamily proteins. Finally, we integrated a protein-protein interaction network to correlate data from transcriptomic analysis and docking screenings to propose FDA-approved drugs that could be used as multitarget therapies or in combination with the potential natural inhibitors of KDM4 enzymes. This study highlights the relevance of the KDM4 family in cancer and proposes natural compounds that could be used as potential therapies. The met lysines ; for fur lysines ;  that mainly target the trimethylation of H3K36 and H3K9, which are associated with active transcription and heterochromatin , respectively . The KDMp53 in colorectal cancer  were downloaded from Xena Browser . RSEM exp value <0.05. COXPH estimate <0 was labeled as “good prognosis” and COXPH estimate >0 as “bad prognosis”. Kaplan Meier plot and Log Rank Test were performed using patients with KDM expression < Q1 (Low-KDM) and patients with KDM expression > Q3 (High-KDM). Difference in overall survival between groups was considered significant with p value <0.05.Event and time-to-event information was used to evaluate the association between expression of the KDM4 subfamily genes and Overall Survival of patients using the Survival v3.2-11 package . For COX∧ ‐1) and used as input for DESeq2 v1.32.0 to compare neoplastic vs. non-neoplastic samples , and COCONUT . The accession numbers and references for all the models used are available in 2+ and Zn2+ as cofactors in the active site; crystallographic reports have also shown the presence of Ni2+ as a cofactor, with no significant differences in KDM conformation. Since the development of a competitive inhibitor must consider the effect of the metal in the active site, in this work, Zn2+ was considered as the representative metal for the functional activity of KDM4. For the docking study, the representation of coordination bonds between metals and the active site is not necessary, so we consider this as a good representation of the electrostatic potentials for the metal in the force field used. The conserved residue GLU190 of the active site was defined as the anchor point for the search docking box. Amber ff94 force field was used for protein and Zn2+ partial charges calculation.The KDM4 structures were fitted by structural alignment to maintain the same active site orientation. Different reports have established that KDM4 is active in the presence of Fe2+ cofactors in the active site of each KDM4. For the APO system, any metal cofactor was removed for the analysis. Each cured chemical database was docked to the receptor in APO as GLU190 residue as reference of binding site, while for the HOLO form we used GLU190 and Zn2+ as reference for the FRED program from OpenEye Scientific software. The Chemgauss4 scoring function was used, and the top 100 scoring molecules for each case were considered possible hits.Two systems were implemented to understand the metal influence over KDM4 proteins’ active site, HOLO and APO. The HOLO form included the ZnFlexophore descriptor. Two compounds were considered similar if their similarity relationship surpassed a threshold of 95%. For each protein and database evaluated, the top 100 scoring compounds were analyzed. As nine different PDB structures were used for KDM4D, a random sample of 100 compounds from each database were chosen.The 3D-pharmacophore similarity analysis between compounds was performed with DataWarrior  using thPBSA) of the ligands. Since this approach requires a large amount of computational resources, we performed the ΔGPBSA calculation for only a subset of molecules according to the following criteria:a) Molecules that bind only to one KDM4b) Molecules that bind to some or all the significantly overexpressed KDM4s in a cancer type.c) The best 10 molecules for each KDM4 according to their FRED/Chemgauss4 score, regardless of the source database.Molecular docking methods are efficient tools for large database screening; however, their main limitation is inaccurate binding energy estimations. The molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) method was used to estimate the absolute binding energy  vs. low-KDM (first quartile). The log2 Fold Change (log2FC) of each KDM4 protein and the number of differentially expressed genes (DEGs) in each comparison are shown in Since we observed that KDM4 proteins are deregulated in several neoplasies and that their expression is related to several processes associated with cancer, we next used molecular docking to screen for potential inhibitory compounds. In order to explore the scaffold for inhibition specificity, we docked a total of 418,912 compounds from three different databases  against the active sites of each KDM4; the protein targets used are disclosed in For docking analysis, both HOLO and APO forms of 13 KDM4 structures were prepared; thus, a total of 26 structures were sampled for FRED/Chemgauss4 docking. The 100 best-scored results were selected, recording a total of 7,800 protein-ligand interactions . Figure Next, we seek to further explore whether structural similarities exist among the compounds predicted to bind the KDM4 subfamily members; such findings could be important for understanding the molecular signatures involved in the protein-ligand interactions and for the future development of KDM4 subfamily inhibitors. To address this idea, we evaluated the chemical scaffold of the top 100 hits for each KDM4 from FDA, DrugBank, and COCONUT databases  using a similarity flexophores map. This graphical method tests whether the conformational flexibility of a molecule plays a significant role as a potential inhibitor of proteins . Usually2+, Ni2+ or Fe2+; thus, in drug design, the inclusion of sugars and phenols represents an advantage for the achievement of a competitive inhibitor.The molecules’ distribution by library is shown in GPBSA) was calculated with the MM-PBSA method for a representative subset of molecules . Because performing molecular dynamics with HOLO systems represents a computational challenge  are the most flexible areas. It is also noticeable that the ligand is vibrating inside the protein’s active site . KDM4A’sotential . Due to 2+, Ni2+, or Fe2+). It was also observed that, although the E and D residues in the catalytic site do not directly interact with the ligands, they do contribute to the overall negative microenvironment of the cavity. For example, epigallocatechin gallate (EGCG), a molecule belonging to cluster number 3, establishes one coordination bond with KDM4A metal through the flavonoid group, meanwhile, the secondary catechol bends in the opposite direction of the metal due to repulsive forces effect between them. Additionally, compounds in clusters 14 and 15 exhibit a favorable orientation of the OH groups of the sugar on the flavonoid that allows the formation of 2–3 coordination bonds with the metal  which not only stabilize the binding site but also contribute to the protein-ligand binding through pi stacking interactions with other aromatic groups. The residues that most frequently interact with the ligands are I71, Q84, N86, Y132, A134, D135, G170, V171, Y175, Y177, F185, H188, E190, D191, S196, N198, W208, L241, and S288. he metal ; althougPBSA energy distribution through the protein, we calculated the per residue binding energy contribution of the KDM4A-CNP0371131 complex. We observed that the binding energy is mainly driven by long-range electrostatic interactions and it is distributed along all the residues, not only the ones present in the cavity  and the nonpolar interaction energy (ΔEnonpolar), while an unfavorable penalty is applied by the polar interaction energy (ΔEpolar), mainly due to the solvation effect of both the ligand and the active site , facilitate interpretation and explore the applicability of the results, we constructed a protein-drug-disease network containing the five KDM4s and the top seven hits for each KDM4 from the three databases evaluated. We also included the neoplasms related to each enzyme; a neoplasm was included if a KDM4 was overexpressed or if it was of bad prognosis in any of the two survival analyses. When integrating these data we observed that according to their KDM4 expression pattern, a different drug set for each neoplasm can be found ; for exaA detailed example of the usefulness of this analysis is the network extracted for uterine corpus endometrioid carcinoma , which sThis analysis also allows us to observe that KDM4 proteins, when overexpressed, trigger expression changes that affect genes involved in various cellular processes. For example, the network detected for pheochromocytoma and paraganglioma shows that the proteins are involved in the positive regulation of gene expression, which is also closely related to the negative regulation of H3K9 methylation function found in networks adrenocortical cancer (KDM4A overexpression), lung adenocarcinoma (KDM4A overexpression) and uterine corpus endometrioid carcinoma. Furthermore, we found that some druggable processes are related to monoamine GPCRs or closely related to cocaine addiction pathways, in thyroid and uterine carcinomas , this isEpigenetic processes play an important role in the regulation of gene transcription. The discovery of histone demethylases has contributed to understanding the dynamic process of histone marks establishment where the deregulation of these enzymes can contribute to the development of several diseases including cancer . These tSince our main interest is to propose natural KDM4 inhibitors, we used the COlleCtion of Open Natural Products (COCONUT), which gathers 406,744 natural products from over 50 different databases, where nearly half the compounds come mainly from plants, fungi, bacteria, and to a lesser extent, from animal or marine origins . Most ofHowever, a challenge faced during the development of specific inhibitors is the resemblance of JMJC family members’ catalytic sites. Since these proteins share a catalytic mechanism, their active sites have a high resemblance, which complicates the design of ligands that could be specific for a single enzyme . The KDM4 subfamily active site consists of a TIM-barrel fold , which is a usual structural pattern in proteins that allows a wide assortment of functions . Thus, tCamellia sinensis) that contains catechol and whose effect has been studied in various epigenetic processes (Since the KDM4 subfamily is a promising therapeutic target for drug design, a wide number of synthetic and nature-inspired molecules have been explored. Among them, it has been proposed that catechol and flavonoids as structural scaffolds, these kinds of molecules have gained attention because of their high content of OH with redox capacity that can also act as free radical regulators . Such furocesses . EGCG isrocesses . Anotherrocesses .de novo molecules for cancer treatment. One of the limitations of our study is that it is not supported by experimental assays, but its strength is that this work is the first step towards an experimental approach that could contribute to the treatment of different neoplasms.In addition to the contribution of coordination bonds that favor specificity, there are other non-covalent binding forces that can also have an impact on specificity and binding affinity such as salt bridges, hydrophobic interactions, hydrogen bonds, and pi-stacking interactions. In particular, pi-stacking interactions among aromatic rings are an important factor in the protein-ligand complex formation; in such interactions, the geometric orientation of the rings change the dipole attraction forces among them as well as the hydrophobic and van der Waals forces rearrange . The preOur results also show that members of the KDM4 subfamily are promising drug targets for the development of therapeutic alternatives in different types of cancer. Since specificity is hard to achieve for KDM inhibitors, we aimed to use this to our advantage searching for ligands that could target all the KDM4s relevant for a specific neoplasm without altering the others. We highlight the importance of natural compounds against KDM4 subfamily members, not only because of their high potential as inhibitors but also because these compounds could contribute to an integrative cancer treatment. As shown in this work the identified molecules could have an amplified therapeutic effect by modulating, not only KDM4 functions but entire cellular processes, by modifying the activity of proteins involved in the same pathways. This mechanism of action has been proposed for other diseases and protein targets before ; however


The recommended schedule for killed oral cholera vaccine (OCV) is two doses, 2 weeks apart. However, during vaccine campaigns, the second round is often delayed by several months. Because more information is needed to document antibody responses when the second dose is delayed, we conducted an open-label, phase 2, noninferiority clinical trial of OCV. One hundred eighty-six participants were randomized into three dose-interval groups (DIGs) to receive the second dose 2 weeks, 6 months, or 11.5 months after the first dose. The DIGs were stratified into three age strata: 1 to 4, 5 to 14, and > 14 years. Inaba and Ogawa vibriocidal titers were assessed before and after vaccination. The primary analysis was geometric mean titer (GMT) 2 weeks after the second dose. Data for primary analysis was available from 147 participants . Relative to the 2-week interval, groups receiving a delayed second dose had significantly higher GMTs after the second dose. Two weeks after the second dose, Inaba GMTs were 55.1 190.3, and 289.8 and Ogawa GMTs were 70.4, 134.5, and 302.4 for the three DIGs respectively. The elevated titers were brief, returning to lower levels within 3 months. We conclude that when the second dose of killed oral cholera vaccine was given after 6 or 11.5 months, vibriocidal titers were higher than when given after the standard period of 2 weeks. This provides reassurance that a delayed second dose does not compromise, but rather enhances, the serological response to the vaccine. Vibrio cholerae, is introduced into the area. The intestinal infection with V. cholerae causes diarrhea, which in severe cases, leads to severe, life-threatening dehydration. Deaths can be prevented with rehydration using oral or intravenous rehydration, but outbreaks may occur suddenly, before facilities are prepared to provide adequate treatment or when patients are not able to reach the health facility. This contributes to higher case fatality ratios early in outbreaks or in areas without access to treatment.,–Cholera is an infectious disease that continues to cause outbreaks with a high burden in countries of Asia, Africa, and, until recently, in Hispaniola.,,,Although OCV is effective against cholera, it must be used strategically and be given to people at highest risk. This includes 1) preventive use for people living in geographic areas deemed to be cholera hotspots, 2) preventive and emergency use for people in a humanitarian crisis in areas with risk of cholera, and 3) emergency use to control an outbreak.If there is to be a delay in administering the second round of vaccine, the acceptable interval for this delay is not known. Changing the interval to 1 month was not inferior to 2 weeks,,The present study was conducted to compare the vibriocidal antibody titers when the two doses of OCV were given at longer intervals; thus, participants were randomized to receive the second dose of OCV after 2 weeks, 6 months, or 11.5 months after the first dose. We then determined the geometric mean titers (GMT) 2 weeks after the second dose, as well as additional follow-up periods in the different groups. We hypothesized that the serum vibriocidal GMTs in serum obtained 2 weeks after the second dose would not be inferior to the recommended interval of 2 weeks if the second dose was extended to 6 months or 11.5 months. Additional aims of the study included an evaluation of 1) age-group-specific serum vibriocidal GMTs during the same time intervals, 2) vibriocidal seroconversion rates in the participants overall and by age group, and 3) persistence of elevated vibriocidal GMTs during follow-up for several months. Because protection from OCV is age dependent,This study was approved by the Cameroon National Research Ethics Committee (2018/02/975/CE/CNERSH/CE) and by the Institutional Review board of the Johns Hopkins Bloomberg School of Public Health (IRB No. 8114). Potential study participants received full information regarding study objectives, procedures, duration, and their rights to decide to participate or not to participate. Adults gave signed informed consent and parents of young children (1–12 years) provided signed parental permission. Parental permission and participant assent was obtained for participants aged 13 to 20 years, The trial was monitored by a local monitor, and the protocol was registered at Clinical Trials.gov (NCT03719066).This was an open label, randomized, phase 2, noninferiority clinical trial. Participants were randomly allocated by stratified random sampling to three dose-interval groups (DIGs). Each DIG received two doses of OCV with the first dose given on day 0. The second dose was given 2 weeks later for DIG1, at 6 months for DIG2, and at 11.5 months DIG3. Each DIG was divided into three age strata: 1 to 4 years, 5 to 14, and > 14 years. Vibriocidal titers were assessed at different time points before and after vaccination. Acceptable time windows were defined for the dose intervals and blood draws.The study was conducted at the Soboum health area situated in the Nylon health district in the large port city of Douala, Cameroon. This is a crowded urban area with working-class families. Most people living in this area are involved in commercial and urban transport activities, and a smaller fraction working in business and as civil servants.The first dose was administered on October 26, 2018, and the field phase of the study ended on February 27, 2020. Households of the health area were visited by trained community volunteers to invite potential participants to the study site. Enrollment was done at the Soboum Medical Center, which is the main health facility for delivering primary health care in the Soboum health area. Although this area is considered at risk for cholera, there were no cases reported in Douala since a large outbreak in 2010–2012. OCV had never been used in the study area before or during the study period other than through this study.Residents of the Soboum health area aged ≥ 1 year were eligible to participate. We included clinically healthy participants who assured the study team that they would be available during the study period and consented to participate. At the time of recruitment and before the second dose, we excluded pregnant women, those who received OCV from other sources or who received an investigational product (within 30 days before vaccination). We also excluded those with a history of diarrhea during the 7 days before first dose of vaccine (defined as ≥ 3 unformed loose stools in 24 hours); history of chronic diarrhea (lasting for more than 2 weeks in the past 6 months); current use of laxatives, antacids, or other agents to lower stomach acidity; planning to become pregnant in the next 2 years; presence of a significant medical or psychiatric condition. Examples include reported diagnosis and treatment of tuberculosis or HIV, renal insufficiency, hepatic disease, oral or parenteral medication known to affect the immune function such as corticosteroids or other immunosuppressant drugs, or behavioral or memory issues. Participants who at one point could not been reached or did not present to the health facility for their second dose with no definitive reason were considered lost to follow-up and excluded from the analysis.V. cholerae O1 and O139 in single-dose glass vial containing 1.5 mL of vaccine. The entire vial of OCV was administered orally to each participant irrespective of their ages. The vaccine vial was shaken to ensure mixing and was inspected by the study nurse before administration. To monitor for safety, all participants were observed for 30 minutes after taking the vaccine. The timing of the follow-up vaccinations and blood draws along with the acceptable visit windows for each event are shown on The cholera vaccine used in the study was Shanchol (Lot 2SCNO33A17), which consists of a mixture of killed The participants were followed for 12 months for DIG1 and for 15 months for the other two DIGs. The primary outcome was the vibriocidal GMT measured 2 weeks after the second doses of OCV for each DIG. Secondary outcomes included 1) vibriocidal antibody seroconversion rates  2 weeks after the second dose, 2) age-group-specific serum vibriocidal GMTs and seroconversion rates 2 weeks after the second dose in each DIG. Additional outcomes compared the GMTs at other time points to determine persistence of the vibriocidal titer changes and the geometric mean titer fold increase ratio.For adults, 5 mL of whole blood was collected by venipuncture. For children (1–4 years), 350 µL capillary blood was collected using a capillary tube and diluted 1:5 in normal saline. The blood samples were transported with ice packs to the field laboratory at the Congo medical center located at about 5 km away from the Soboum Medical Center. The samples were centrifuged for serum extraction and stored at –20°C and were then transported to the central laboratory based in Yaoundé, ∼240 km from Douala for serologic analyses.,V. cholerae strains used as the target vibrio strains were originally isolated in Cameroon (Inaba FO14-2018-EN and Ogawa CPC-BACT-01). Briefly, the strains were incubated with serially diluted, heat-inactivated serum and exogenous guinea pig complement  on a shaker (50 rev/min) at 37°C for 1 hour.The serum vibriocidal responses were measured as previously described,The sample size was calculated to determine noninferiority of the vibriocidal GMTs of DIG2 and DIG3 compared with DIG1 2 weeks after the second dose. The margin of noninferiority was set to 15%. The true difference between the GMTs was assumed to be 0.000. The data assumed a population with a standard deviation of 7.770 for both groups. With alpha = 0.025, beta 0.10, one-sided, two-sample mean, a sample size of 11 in each group was required. Assuming a 10% high baseline titer and 20% dropout over the study period, a sample size of 20 per age strata was calculated, totaling 180 subjects in the study.https://www.sealedenvelope.com) to create a randomization list following the fixed block randomization method. The random assignment numbers were placed in sealed envelopes in sequential order and participants were given their DIG assignment based on their sequence of enrollment. Because the study was designed to include a given number of participants in each age stratum, recruitment continued to reach the planned number of participants, although in some cases a few additional participants were included for logistical reasons.Participants in each age strata were randomized into three DIGs using a block randomization of 6 per block. Before study initiation, three randomization tables for each of the age strata were generated using “Sealed Envelope” . Data was excluded from participants if they vomited one of the doses. Vomiting was defined as the expulsion of stomach contents through the mouth within 10 minutes after administration of the vaccine. None of the subjects in the study experiencing mild or transient spitting up when dosed. We observed each subject to ensure that each child swallowed the entire dose. If they missed one of the other scheduled blood draws, their available data were included when analyzing the “additional outcomes.”t test for quantitative variables. Because of the imbalance of baseline titers, especially the higher baseline titers for DIG3, we calculated the geometric mean titer fold increase ratio by comparing the geomeans of the fold increase between baseline and follow-up sera. The 95% confidence interval of Geometric Mean Responses was constructed through 100,000 bootstraps. All analyses were conducted using R (version 4.0.4).All analyses were done between DIGs for overall and for each age group considering DIG1 as the reference group. Baseline characteristics and study outcomes were compared using χParticipants were engaged in the study from October 26, 2018, until February 27, 2020. Of 217 participants who came to the field study site to consider joining the study, 18 did not consent to be screened and 13 were excluded. One hundred and eighty-six (186) participants were enrolled, with 62, 61, and 63 randomly assigned to DIG1, DIG2, and DIG3, respectively. Two children in DIG2 and one child in DIG3 vomited a first dose of vaccine, and their data were excluded. As shown on the consort flow diagram, others did not take a second dose or missed a primary blood draw. Serum was available from 147 participants who took two full doses of vaccine and had blood samples 14 days after the second dose including 54, 44, and 49, respectively, in DIG1, DIG2, and DIG3. Data from these participants were included in the primary analysis. Details of participants flow are presented in the consort flow diagram of the study in A comparison of participants’ characteristics per study DIG is presented in As shown in The Inaba and Ogawa GMTs for the DIGs during the entire span of the study are shown in As expected, the vibriocidal GMTs 2 weeks after the second dose were all significantly higher than baseline GMTs; however, the increase in GMTs was greater for both DIG2 and DIG3 compared with DIG1. The vibriocidal GMTs before the first dose and 2 weeks after the second dose and response rates after the second dose are shown in The age-specific vibriocidal GMTs are shown in This study from Cameroon found that extending the dose interval to either 6 or 11.5 months elicited a higher vibriocidal GMT 14 days after the second dose compared with the standard 14-day interval. After a single dose of OCV, the GMT increased among participants in DIT1, but the second dose given 2 weeks later did not increase the GMT further. By contrast, when the second dose was delayed for 6 or 11.5 months, the increase in vibriocidal GMT was significantly higher than when the second dose was given after 2 weeks. Furthermore, by delaying the second dose, there were two periods of the year, one after each dose, with an elevated vibriocidal titer; however, each of these periods with an elevated titer were relatively brief. Three months after vaccination, for each of the groups, the GMTs fell to lower levels, but the titers were still significantly higher that the baseline GMT. Thus, the study showed that the vibriocidal GMTs 2 weeks after the second dose were not inferior to the standard interval and in fact were superior to the standard dose interval. In addition to the higher GMTs with the delayed second dose, the vibriocidal response rate was also higher if the second dose was delayed.The recommendation for a second dose after 2 weeks was based on results of the pivotal study in Kolkata that used this 2-week dose interval and found this strategy to be protective for 5 years.A dose-interval study with similar design conducted in rural Zambia showed that extending the interval between doses to 6 months stimulated GMT vibriocidal titers that were not inferior to that stimulated by the 2-week schedule.There is a potential immunological explanation for the higher vibriocidal response after a delayed second dose. The first dose of vaccine stimulates an intestinal immune response, which is maximal at about 9 to 14 days after vaccination.The study did not have sufficient power for age-stratified analysis due to a small sample size, but higher vibriocidal responses were observed in the DIG2 and DIG3 groups among participants aged 1 to 4 years. Although not statistically significant, the 1 to 4 years age group had a lower GMT and lower seroconversion rate at 14 days post–dose 1 or dose 2 compared with the other two groups. The higher response rates with the delayed second dose for youngest children is of potential importance because vaccine efficacy and protection is lower in the age group < 5 years. The potential benefit of delaying the second dose for young children needs to be balanced against the longer interval between doses, which leaves young children more vulnerable until they receive the second dose. For preventive campaigns designed to prevent future outbreaks (but not an ongoing outbreak), this issue is less important. However, during an outbreak, when active cholera transmission is ongoing, a shorter interval between doses may be favored.In a single-dose efficacy study and its follow-up study in Dhaka, Bangladesh, one dose of OCV was not efficacious for children ≤ 5 years.These findings should be considered when considering logistical and practical implications of delaying the second dose. When vaccine campaigns are carried out, the campaign attempts to provide vaccine to all persons ≥ 1 year residing in the designated area. Thus, the two vaccine rounds may reach somewhat different individuals in the two rounds if there is movement of people in and out of the area. With a longer delay, there is a higher chance that some persons will have moved away and others will have moved into the area. This increases the number of people who will receive at least one dose, but also increases the number who will receive only a single dose. For extremely mobile populations , the control program will need to consider the movement of people and may have to consider a third round.Escherichia coli was seen in subjects who had received the vaccine 1 to 2 years earlier.Haemophilus influenzae Type B, and hepatitis BOCV responded more vigorously if they had received OCV 3 years earlier.The study had several strengths that should be highlighted. First, the participants were studied over a long period with multiple follow-up serum specimens allowing for evaluation of the persistence of vibriocidal titers. Second, the study was conducted in an area at high risk for cholera, but unlike areas in Asia where similar studies have been carried out, no cases were reported in recent years, and none were seen during the conduct of the study when such cholera infections might have confounded the results. Third, because age is an important variable, this study included a similar number of subjects in the different age strata.,There were also some limitations. The sample size was determined based on a comparison of the primary outcomes between the three DIGs but was not sufficient to conduct this comparison for the age-specific strata. Second, we did not collect serum from participant in DIG2 and DIG3 2 weeks after the first dose because we wanted to limit the number of blood collections and assumed the randomization would provide data to define the response 2 weeks after the first dose. Also, the slightly lower proportion of male subjects in DIG3 was noted. Some studies have found a sex difference in immune responses to other injected vaccines,,There are several implications from this study. For example, in regions with a predictable cholera seasonality (e.g., BurundiThis study found that extending the dose interval to 6 or 11.5 months elicits higher vibriocidal antibody titers 14 days after the second dose compared with the standard 14-day interval, and this should allow increased flexibility regarding timing of the second dose for programs implementing vaccine campaigns. Future studies are needed, using case–control or test-negative designs, to monitor vaccine effectiveness of delayed dose intervals in countries where the second doses were delayed. Second, future studies are needed with larger groups of young children to evaluate whether a longer dose interval will improve immune responses in this vulnerable age group. Finally, if these findings are further validated, a recommendation for a change in dose schedule may be considered for OCV.


Research has consistently found that people with mental illness (known as consumers) experience lower levels of participation in meaningful activities, which can limit their opportunities for recovery support. The aim of this study was to describe the outcomes of participation in a group program designed to address all stages of activity participation, known as Pathways to Participation (P2P). A descriptive longitudinal design was utilized, collecting data at three time points. Outcomes were measured by the Camberwell Assessment of Need Short Appraisal (CANSAS), Recovery Assessment Scale—Domains and Stages (RAS-DS), Behavior and Symptom Identification Scale (BASIS-24), Living in the Community Questionnaire (LCQ), and time-use diaries. All data were analyzed using descriptive statistics and Chi-square analyses. A total of 17 consumers completed baseline data, 11 contributed post-program data, and 8 provided follow-up data. Most were female (63.64%) and had been living with mental illness for 11.50 (±7.74) years on average. Reductions in unmet needs and improvements in self-rated recovery scores were reported, but no changes were identified in either time use or psychosocial health. The findings indicate that the P2P program may enable consumers to achieve positive activity and participation outcomes as part of their personal recovery. The link between participation in meaningful activity and health or well-being has been well established through rigorous research studies across age and diagnostic groups . For peoHowever, a considerable body of evidence demonstrates that consumers participate in lower levels of activity than members of the general community. An Australian study of consumers living in the community  found thDecreased participation in meaningful activity has multiple, cumulative effects on consumer health and well-being, increasing the risk of both physical and psychiatric morbidity ,10. PeopGiven the serious consequences of decreased participation, several interventions have been developed to enable consumers to engage in meaningful activities. To date, these interventions have mostly focused on vocational participation. A survey of Australian mental health consumers found that 76% were unemployed , with boInterventions that address the earlier phases of activity participation during recovery have begun to emerge in the past decade. The Action Over Inertia (AOI)  program The Balancing Everyday Life (BEL) intervention  is a 12-The preliminary evidence for both AOI and BEL provides promising data regarding their impact on consumer participation. However, none of the available interventions address a complete continuum of activity participation from initial engagement through to workforce participation as part of personal recovery. Participants in these programs may therefore need access to additional interventions to meet their vocational needs, which may not provide a coherent or well-articulated pathway to their ultimate recovery goals. The aim of this study was to describe the outcomes of a group program designed to address all stages of activity participation during recovery, known as Pathways to Participation (P2P).The study utilized a descriptive pilot longitudinal design to investigate outcomes for consumers participating in a single community mental health service. The study received ethical approval to proceed from the relevant Human Research Ethics Committee (HREC 2017.074).The P2P intervention is a hybridized program that aims to enable consumers to engage in meaningful activities at all stages of recovery. The P2P program combines two evidence-based interventions, the Action Over Inertia (AOI) program and The WORKS program. In addition to the overview of AOI presented above, the program emphasizes gradual momentum towards personal recovery goals and aims to increase engagement, enhance activity balance, promote social interaction and community engagement, and improve overall health and well-being. The WORKS framework  was deveThe developers of both the AOI and WORKS programs encourage modifications for local conditions, and the first author undertook adaptations based on her previous experience of delivering these interventions in community mental health services. The model of co-facilitation by occupational therapists and peer support workers was retained from previous iterations of the WORKS, given the positive feedback provided by stakeholders of that program . ConsultPrior to the commencement of the P2P program at each site, facilitating occupational therapists and peer support workers were provided with intervention training based on the manual content. This training was instituted based on previous findings , which iA single iteration of the P2P program was delivered at each of the four catchment areas of the adult mental health service. Each co-facilitated session lasted for two hours, with the first four weeks based on AOI content and the final six weeks based on The WORKS content. An information session was provided to consumers prior to P2P commencement, ensuring they understood the commitment required before agreeing to attend. Consumers were encouraged to attend the P2P program every week; however, it was not compulsory, and anyone missing a session was provided with extra contact from the facilitators to support them to rejoin and catch up on content. An overview of the content of each session is provided below in A total of five key outcomes were identified for investigation in this study: consumer identified needs, time use, self-rated recovery, psychosocial health, and community participation. Consumer identified needs were measured using the Camberwell Assessment of Need Short Appraisal (CANSAS) , which wTime use was measured via a 24 h time diary, in which consumers listed the activities they participated in, where it was performed and whom it was performed with at hourly intervals for the previous day. This method of gathering time use data is practiced across a range of disciplines, although it can be susceptible to recall bias . Data rePsychosocial health was measured using the Behavior and Symptom Identification Scale (BASIS-24) , which iIn combination, the outcome measures used in this study took approximately 15 min to complete. Outcomes were measured at the beginning of the P2P program (T1), immediately post P2P program (T2), and 3 months post P2P program (T3). Demographic data were also collected for all consumers at recruitment; at subsequent time points, a further question regarding any other vocational interventions was included. Participating consumers were reimbursed for their time completing data collection tasks with shopping vouchers. Initially, presentations about the P2P program were provided to multidisciplinary teams at each local service. Posters and brochures were also distributed to multidisciplinary team members to provide to potential participants and were displayed in the waiting room of each service to enable consumer self-referral. Multidisciplinary team members were not expected to recruit consumers, only to provide them with information and support the consumer to complete a brief registration form if required. Participation in the P2P program was not contingent on participating in this study. All local community services within the catchment of the mental health service were included in this study, comprising Continuing Care Units, Prevention and Recovery Care Services, and Community Mental Health Teams. For inclusion in the study, consumers needed to be aged between 18 and 65 years old and be receiving care from one of the participating services at the time of recruitment. Other inclusion criteria were (1) generally stable health ; (2) formal diagnosis with a serious mental illness or mental disorder with associated significant levels of disturbance and psychosocial disability; (3) spoken English to a level sufficient to participate in the group program without an interpreter; (4) capacity to independently provide informed consent, and (5) considered to present low to moderate risk by their key clinician. As co-morbidity was so prevalent amongst consumer populations , those wOnce registered for the P2P program, the consumer was provided with information regarding this study from the first author at an information session for all group attendees prior to the program commencing. Individual, specific written consent was obtained for every consumer participant in this study. Consumers were provided with a plain language statement prior to consent being sought and were encouraged to ask questions from their supporters prior to confirming their intention to participate. Participating consumers could choose extended P2P session attendance, a separate clinic appointment, or a telephone interview to complete outcome measures at each time point. Descriptive analysis was undertaken for all outcome measures collected in this study. Interval variables were analyzed using means, standard deviations, and ranges, while categorical variables were analyzed using frequencies and proportions. Where descriptive analysis indicated a large change in outcomes, analytical analysis was deployed using the Chi-square test. n = 1), moving away (n = 2), and choosing not to continue (n = 6). Most consumers were female . Mood disorders, anxiety disorders, and schizophrenia were the most prevalent psychiatric diagnoses, which consumers had been living with for 11.50 years (±7.74) on average. They also experienced other health conditions , which were also long-standing . Around half of the sample  had co-morbidities in addition to their mental illness. A total of 17 consumers were recruited for this study; however, 6 were lost to follow-up prior to the end of the P2P program. The following analysis focuses on the 11 consumers for whom repeat measures were available at T2 and the 8 consumers who provided data at T3. Reasons for attrition from this study included gaining employment  were singled or divorced, gained income from welfare, had completed high school education, and did not have children. A similar proportion  were living with family members and undertaking additional vocational interventions to the P2P program during the study. Most consumers  experienced a decrease in unmet needs over the course of this study. There was a significant change in the proportion of identified needs between T1 and T2, with met needs increasing and unmet needs decreasing, χ2  = 4.55, p = 0.033. Further decreases in both met and unmet needs were reported at T3, but these were not statistically significant. Information about conditions or treatment  was the most commonly unmet need at T1, along with self-care . Money was frequently identified at T1  and T2 , while physical health  and daytime activities  were highlighted T2  and T3 . Distress  and psychotic symptoms  were only prevalent at T3. Most consumers (n = 3), productivity activities (n = 2), and leisure activities (n = 4) during this study. Both positive and negative changes in activity participation times were recorded across the study, and there was considerable variability between individual consumers. The amount of time consumers spent in the community remained steady from T1 , to T2 , and T3 . As shown below in While time used for sleep remained stable for most, some consumers increased or maintained participation in activities of daily living  experienced an increase in total recovery scores between T1 and T2, while 5 consumers (62.5%) experienced further increase or maintenance of their total recovery scores between T2 and T3.There were no statistically significant changes in total or subscale BASIS-24 scores over time for any participating consumers. Psychosocial health remained constant for consumers throughout the study, with any changes observed being small fluctuations in scores.The proportion of consumers rating their social participation as ‘about right’ increased by 46% between T1 and T2, with smaller increases reported for caring participation (17%) and work participation (17%). A further 25% of consumers rated their participation in unpaid work as ‘about right’ at T3; however, , having a say in the community (30%), and hopefulness (21%). However, fewer consumers felt positive about having a say within families, having opinions respected, happiness, goal achievement, and belonging after 3 months than at T2. This study has described the outcomes of consumers participating in a group program designed to address all stages of activity participation during recovery, known as Pathways to Participation (P2P). While this pilot study can only provide a limited perspective on effectiveness, the findings presented here indicate the program did achieve some positive outcomes for the consumers who took part. P2P appeared to have the most positive impact on decreasing unmet needs for consumers and on social recovery and participation. The specific mechanisms by which consumers were able to meet their needs more effectively were not part of the data collection for this study but should be qualitatively investigated in future research. However, an Israeli study of the needs of people with serious mental illness  asserts However, some of the improvements reported by consumers between baseline and the end of the program were not sustained over the following three months. The evaluation of the long-term effects of occupational interventions is important to understand whether they have had a meaningful impact on the consumer, particularly as a fading of reported effectiveness over time is reported in many therapeutic studies . Given tThis aspect of the finding also raises questions about whether a program like P2P is most appropriately delivered by occupational therapists employed by state-based statutory mental health services. The complexity of factors that influence the activity and participation of consumers indicates that longer-term psychosocial rehabilitation is needed to enable recovery. In recent years, rehabilitation has had less of a presence in Australian mental health policy, with much of the public discourse focused on the tension between resourcing acute and community-based services . HoweverWhile some positive outcomes were short-lived, no significant changes in other outcomes were identified during this study. Both time use and psychosocial health outcomes remained steady across all time points, which suggests the P2P program made no impact on either of them. However, the proportion of consumers rating their social participation and unpaid work as ‘about right’ increased, as did the number of participants identifying their well-being as good to excellent. The AOI program is a time-use intervention , and so The findings of this study also highlight that recovery is a very individualized and personal process for consumers. Recovery is not a linear process, which is why recovery scales do not have ‘clinically significant’ change thresholds . The traThis study has several limitations that must be considered when interpreting the findings. The small sample size as a pilot study means that effectiveness could not be comprehensively assessed, and the participating consumers were not representative of any larger population. It was also completed within a limited geographical area and did not reflect prevailing changes to practice required since the COVID-19 pandemic. Aside from the methodology, there was also significant attrition from the study and loss of follow-up over time. This issue may be particularly problematic for mental health studies, as participants are experiencing a range of issues that may interfere with their ability to participate . DespiteIn conclusion, this study has described the outcomes experienced by consumers participating in pilot iterations of the Pathways to Participation (P2P) program. Some positive outcomes were identified regarding unmet needs and social participation, but no significant improvements were recorded in time use or psychosocial health. The findings presented here provide a basis for ongoing research into the effectiveness of the P2P program, which should include qualitative and mixed methodologies to better capture consumer experiences of the program.