Heterometallic complexes of the composition [(LnL2tmhd)2TM(tmhd)2] were isolated for light lanthanides Ln= La, Pr, Sm, Gd, and L= L1 or L2. By single crystal XRD, it has been established that heterometallic compounds containing La, Pr, Cu, Co, and Ni are isostructural linear coordination polymers of alternating mononuclear transition metal complexes and binuclear heteroleptic lanthanide complexes, connected by donor–acceptor interactions between oxygen atoms of the methoxy groups and transition metal atoms. A comparison of powder XRD patterns has shown that all heterometallic complexes obtained are isostructural. Havier lanthanides Ln = Tb, Dy, Lu did not form heterometallics. Instead, homometallic complexes Ln(L3)3 were identified for Ln = Dy, Lu as well as for Ln = La. The thermal properties of the complexes were investigated by TG-DTA and vacuum sublimation tests. The heterometallic complexes were found to be not volatile and decomposed under heating to produce inorganic composites of TM oxides and Ln fluorides. In contrast, Ln(L3)3 is volatile and may be sublimed in a vacuum. Results of magnetic measurements are discussed for several heterometallic and homometallic complexes.The possibility of obtaining volatile polynuclear heterometallic complexes containing lanthanides and transition metals bound by methoxy-β-diketonates was studied. New compounds were prepared by cocrystallization of monometallic complexes from organic solvents. Ln(tmhd) There are also works where 4f–3d heterometallic complexes exhibit several unusual properties simultaneously 2) and heterometallic complexes with alkali metals has been reported, where the coordination of metal cations is also stabilized due to bridging bonds involving methoxy groups, about half of which remain free  type are slightly colored depending on the TM. Crystals of the second component TM(tmhd)2 could be separated either by extraction with heptane or by vacuum sublimation. For example, [(La(L1)2tmhd)2Cu(tmhd)2] is nonvolatile and non-soluble in heptane, while dark blue crystals of Cu(tmhd)2 are well soluble in heptane and sublime at Tsubl = 130 °C in dynamic vacuum ~10−2 Torr. TM(tmhd)2 can easily be identified by XRD, MP, and color. The remaining heterometallic product can be recrystallized from toluene. A series of copper–lanthanide complexes [(Ln(L1)2tmhd)2Cu(tmhd)2], where Ln = La (1), Pr (2), Sm (3), Gd (4), are apparently isostructural according to powder XRD data (2)2tmhd)2Cu(tmhd)2] (8) was prepared. According to XRD, 8 is isostructural with the rest of the copper-containing heterocomplexes 1–4. Following the same approach, [(La(L1)2tmhd)2Ni(tmhd)2] (6), [(La(L1)2tmhd)2Co(tmhd)2] (5) were isolated. Crystals suitable for single crystal XRD were not obtained for 3 and 4 and for [(La(L1)2tmhd)2Mn(tmhd)2] (7) complexes, the manganese derivative turned out to be very unstable during recrystallization.Heterometallic complexes were obtained only by reaction 2 for lanthanide atoms C.N. = 10. Distance Ln…Ln is ~3.8 Å. The lanthanide dimer itself is centrosymmetric and is bound by bridging bonds with the participation of oxygen atoms from both chelate and methoxy groups. However, for the two adjacent methoxy groups, only one is coordinated by the lanthanide atom. While the other methoxy-group remains free in one L, and, in the second L, is coordinated by a transition metal atom. Where both methoxy groups are coordinated the Ln-OCH3 distance is shorter for L1 in contrast to L2. At the same time, the TM-OCH3 bridging bond in 8 is longer than in 1, apparently due to the fact that the –CCH3(OCH3)2 substituent in L2 is bulkier than –CH(OCH3)2 in L1. In turn, the chains of the coordination polymer do not form additional contact with each other. For example, in crystals of 2, the shortest distance between the nearest chains is F…H = 2.620 Å (In the binuclear heteroleptic fragment (Ln(L) 2.620 Å .B) for the heavier lanthanides (Ln = Tb-Lu), no heterometallic products formed. In this case, Cu(tmhd)2 and a colorless glassy product, perhaps a mixed-ligand lanthanide complex, formed separately.In route  give isostructural heterometallic complexes with ligand exchange, while for heavier lanthanides (Ln = Tb-Lu) the ligand exchange occurs without the formation of a heterometallic complex.3)2tmhd)2 should contain two methoxy groups involved in bridging bonds and none free; no heterometallic complex would form. Indeed, the verification of systems with L3 according to route (C) of 3)2 and La(tmhd)3 taken in an equimolar ratio does not yield a heterometallic complex, but rather a two-component system of individual complexes of lanthanum and transition metal was obtained in the result of ligand exchange. The resulting mixture was separated using gradient sublimation. There was only one lanthanum-containing fraction. 1H NMR analysis of the lanthanum-containing fraction has shown the presence of signals for the ligands L3 and tmhd in a ratio of 2:1. It is difficult to say whether this is a mixed-ligand complex or a mixture of the lanthanum homoligand complexes. That is why this product is marked by an asterisk in D) where reactants Ln(tmhd)3 and TML2 were taken in 2:3 ratio still did not give heterometallic complexes. As a result of complete exchange of ligands, homoleptic complexes of lanthanum and copper were obtained, which were also separated by vacuum sublimation. 1H NMR analysis of the lanthanum-containing fraction have shown the presence of signals only for the L3 ligands at the same place, and again two sets of signals are visible in a ratio of 1:2. Moreover, in the 19F NMR spectra, two signals have been recorded in a ratio of 1:2. Thus, the NMR study supports the dimeric structure of the lanthanum complex in solution, with the broad lines indicating that dimer 9 has rather a fluctional structure. Structural data also confirm the dimeric structure, with 9 containing two free methoxy groups. The structure contains two crystallography-independent molecules differing in the orientation of the terminal methoxy groups 3]2 has been reported before 2. As for the heavier lanthanides, the structure of Tb, Dy, and Lu derivatives were not determined because they were isolated as glassy substances. For example, the 1H NMR spectrum of 11 is silent and the 19F NMR spectrum looks strange, indicating that this complex is paramagnetic. On the other hand, 1H NMR of 12 shows broad lines but for one set of ligand signals, and an asymmetric multiplet is observed in the 19F NMR spectrum. Thus, it can be said that at least in solution the Ln(L3)3 for Ln = Tb-Lu seem to be low-symmetry monomers. However, they are more volatile, since 11 and 12 can be re-sublimated at ~160 °C, and 9 already at ~200 °C. To determine the nuclearity of the resulting noncrystalline complexes, we additionally used data on magnetic properties, which are discussed in the corresponding section.Thus, it has been established that under the cocrystallization of La(III) dipaloylmethanate with mono-methoxy-substituted transition metal(II) β-diketonates, the formation of heterometallic complexes does not occur. Instead, an exchange of ligands is observed with the isolation of the binuclear homoleptic lanthanum complex. In the structure of 1–6, and 8 are not volatile and decompose when heated in a vacuum. A comparison of the TG-DTA data for 1, 2, 5, and 6 is shown in 3, and LaOF is obtained from [(La(L1)2tmhd)2Cu(tmhd)2] (1) (1)2tmhd)2Ni/Co(tmhd)2]  gives LaF3, LaOF and probably metallic Ni or Co. Thermolysis of [(La(L1)2tmhd)2Mn(tmhd)2] (7) gives solid product containing LaMnO3 and LaF3 (3)3 complexes are volatile and can be re-sublimed in a vacuum.The sublimation test showed that the obtained heterometallic complexes d)2] (1) .  (1), [(Pr(L1)2tmhd)2Cu(tmhd)2] (2), [(Sm(L1)2tmhd)2Cu(tmhd)2] (3), [(Gd(L1)2tmhd)2Cu(tmhd)2] (4), [(La(L1)2tmhd)2Co(tmhd)2] (5), [(La(L1)2tmhd)2Ni(tmhd)2] (6), [(La(L1)2tmhd)2Mn(tmhd)2] (7), [(La(L2)2tmhd)2Cu(tmhd)2] (8). 1-[(La(L1)2tmhd)2Cu(tmhd)2] Anal. Calc. for C72H108O24F12CuLa2, (%): C, 44.9; H, 5.7; F, 11.8. Found: C, 44.2; H, 5.7; F, 12.1. MP(DTA) 216 °C. 2–[(Pr(L1)2tmhd)2Cu(tmhd)2] Anal. Calc. for C72H108O24F12CuPr2, (%): C, 44.8; H, 5.6; F, 11.8. Found: C, 44.5; H, 5.3; F, 11.2. MP(DTA) 201 °C. 3–[(Sm(L1)2tmhd)2Cu(tmhd)2] Anal. Calc. for C72H108O24F12CuSm2, (%): C, 44.8; H, 5.5; F, 11.7. Found: C, 44.1; H, 5.5; F, 11.5. MP(Kofler plate) 170 °C. 4-[(Gd(L1)2tmhd)2Cu(tmhd)2] Anal. Calc. for C72H108O24F12CuGd2, (%):C, 44.1; H, 5.5; F, 11.6. Found: C, 44.0; H, 5.4; F, 11.4. MP(Kofler plate) 138 °C. 5-[(La(L1)2tmhd)2Co(tmhd)2] Anal. Calc. for C72H108O24F12CoLa2, (%): C, 45.0; H, 5.6; F, 11.9. Found: C, 44.9; H, 5.7; F, 11.8; MP(Kofler plate) 185 °C. 6-[(La(L1)2tmhd)2Ni(tmhd)2] Anal. Calc. for C72H108O24F12NiLa2, (%): C, 45.0; H, 5.6; F, 11.9. Found: C, 44.9; H, 5.7; F, 11.8; MP(Kofler plate) 178 °C. 8-[(La(L2)2tmhd)2Cu(tmhd)2] Anal. Calc. for C76H116O24F12CuLa2, (%): C, 46.1; H, 5.9; F, 11.5. Found: C, 46.4; H, 5.7; F, 11.5; MP(Kofler plate) 225 °C.TMLd before ,23,24,25was used . For the3)3]2 (9), Tb(L3)3 (10), Dy(L3)3 (11), Lu(L3)3 (12), were obtained by the same technique using a molar ratio of Ln(tmhd)3 to TM(L3)2 equal to 2:3. The obtained compounds are colorless solids; only 9 is crystalline. Anal. Calc. for C24H30O9F9La (9), (%):C, 37.3; H, 3.9; F, 22.1. Found: C, 36.5; H, 4.1; F, 22.5. 1H NMR : δ 6.10 , 5.51 , 3.61 , 3.20 , 1.31 2) 19F NMR : δ 75.78 and 76.22 in 1:2 ratio for internal and peripheral -CF3 groups. MP (Kofler plate/DTA) 225–229/224 °C. Anal. Calc. for C24H30O9F9Tb (10) (%):C, 36.4; H, 3.8; F, 21.6. Found: C, 35.6; H, 4.0; F, 21.5. Anal. Calc. for C24H30O9F9Dy (11) (%):C, 36.2; H, 3.8; F, 21.5. Found: C, 35.4; H, 4.1; F, 21.4. Anal. Calc. for C24H30O9F9Lu (12) (%):C, 35.7; H, 3.7; F, 21.2. Found: C, 35.5; H, 4.1; F, 21.5. 1H NMR : δ 6.3 , 3.18 , 1.19 2) 19F NMR : δ 87.5–90.5 multiplets.New monometallic complexes 2 (9) or Lu(L3)3 (12) in contrast to [Ln(L1)2tmhd]2. In addition, the more rigid structure of TM complexes also does not allow them to adjust to it. Nevertheless, a number of unique heterocomplexes with interesting structures were obtained, which will be useful for the further design of polynuclear compounds with the required properties.This study has shown that the use of fluorinated methoxy-substituted diketones for the synthesis of volatile heterometallic complexes containing lanthanides and typical transition metals still requires a further search for methods and approaches to achieve the desired result. It has been found so far for these systems that during the cocrystallization of monometallic complexes there is the exchange of ligands by redistributing the latter following the principle of concentrating fluorine-containing ligands around lanthanide cations. Apparently, this is due to the fact that the electronegativity of lanthanides is lower than that of transition metals, and they tend to form complexes with a more polar coordination bond. In addition, large coordination numbers lead to the fact that the available methoxy groups are also coordinated by lanthanides without alternatives so that a homometallic aggregate is formed rather than a heterometallic one. The peculiarity of lanthanides is also manifested in that the set of products split into two series for La-Gd and Tb-Lu, which often occurs in their coordination chemistry. In particular, the former tend to form dimers in contrast to the latter. For dimethoxy diketonates LIn addition, a reliable way to synthesize anhydrous lanthanide complexes with fluorinated diketonates has been found.


Suboptimal medication adherence is a significant problem for patients with serious mental illness. Measuring medication adherence through subjective and objective measures can be challenging, time-consuming, and inaccurate.The primary purpose of this feasibility and acceptability study was to evaluate the impact of a digital medicine system (DMS) among Veterans (patients) with serious mental illness as compared with treatment as usual (TAU) on medication adherence.This open-label, 2-site, provider-randomized trial assessed aripiprazole refill adherence in Veterans with schizophrenia, schizoaffective disorder, bipolar disorder, or major depressive disorder. We randomized 26 providers such that their patients either received TAU or DMS for a period of 90 days. Semistructured interviews with patients and providers were used to examine the feasibility and acceptability of using the DMS.We enrolled 46 patients across 2 Veterans Health Administration sites: 21 (46%) in DMS and 25 (54%) in TAU. There was no difference in the proportion of days covered by medication refill over 3 and 6 months . The DMS arm had 0.85 (SD 0.20) proportion of days covered during the period they were engaged with the DMS . Interviews with patients (n=14) and providers (n=5) elicited themes salient to using the DMS. Patient findings described the positive impact of the DMS on medication adherence, challenges with the DMS patch connectivity and skin irritation, and challenges with the DMS app that affected overall use. Providers described an overall interest in using a DMS as an objective measure to support medication adherence in their patients. However, providers described challenges with the DMS dashboard and integrating DMS data into their workflow, which decreased the usability of the DMS for providers.There was no observed difference in refill rates. Among those who engaged in the DMS arm, the proportion of days covered by refills were relatively high . The qualitative analyses highlighted areas for further refinement of the DMS.ClinicalTrials.gov NCT03881449; https://clinicaltrials.gov/ct2/show/NCT03881449 Suboptimal medication adherence is a significant problem for patients with serious mental illness (SMI), including those with schizophrenia, bipolar disorder, posttraumatic stress disorder, and major depressive disorder. Suboptimal adherence among these individuals may lead to symptom exacerbation, relapse, and hospital readmissions . MoreoveA digital medicine system (DMS), consisting of a drug-device combination, is a way to obtain objective treatment adherence data. DMS may enable patients with SMI to measure and report ingestion of atypical antipsychotic medications, most of which have broad therapeutic indications . The colThe ABILIFY MYCITE System is an example of a DMS developed to track adherence to oral aripiprazole, an atypical antipsychotic. This system enables patients and their mental health providers the opportunity to view real-time adherence data. Specifically, the ABILIFY MYCITE System is a drug-device combination product (aripiprazole tablets with a sensor) that comprises 4 separate components that enable the monitoring of treatment adherence by a patient and the patient’s provider . The fouOnce a participant swallows the tablet, the ingestible sensor transmits an electrical signal that is detected and then recorded by software within the patch that is worn by the participant on the left rib cage. Using Bluetooth, the patch then transmits the aripiprazole ingestion data to the participant’s smartphone, which is then saved to the secure, cloud-based MYCITE System dashboard. Participants have the ability to view this medication data each day on their smartphone, while the patient’s provider, study team, and selected caregivers are able to view the data on the MYCITE System dashboard via the cloud-based server .Minimal research exists on the real-world comparison between a DMS and treatment as usual (TAU) with regard to medication adherence for individuals with SMI. Therefore, before testing a DMS in a large randomized controlled trial, we wanted to examine the feasibility and acceptability of a DMS among patients with SMI. As the Veterans Health Administration (VHA) is the largest health care provider for individuals with SMI in the United States ,8, it prThis was an open-label, 2-site, provider-randomized, prospective, 2-arm (DMS vs TAU) clinical trial. Patients assigned to the intervention group by provider randomization were enrolled in the DMS for a period of 90 days with the option to continue use for up to 9 additional months. Participants assigned to the control arm (TAU) continued to receive care as recommended by their mental health provider, which included their continued use of aripiprazole. Study duration for both groups was up to 12 months or study closeout, whichever came first. At the study conclusion, we used a descriptive qualitative analysis design and rapid qualitative analysis procedures to examine the feasibility and acceptability of using the DMS. All study procedures were approved by both sites’ respective institutional review boards and research and development committees. The trial is registered at ClinicalTrials.gov (NCT03881449), and the sponsor requested for the study to be concluded prematurely.Study participants were recruited from 2 VHA Medical Centers in Durham, North Carolina, and Houston, Texas. Eligible patients were aged ≥18 years and met the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, criteria for schizophrenia, schizoaffective disorder, bipolar I disorder, or major depressive disorder. Additional study eligibility included (1) an active prescription for oral aripiprazole and (2) approval to participate in the study from their mental health provider. Exclusion criteria included (1) a current neurocognitive disorder that would affect the patient’s ability to complete the trial ; (2) the patient’s mental health provider determining that the patient was not fit to participate; (3) the patient being currently enrolled in an investigational drug trial, a medication management study, or program or participation in an investigational drug trial 30 days before trial enrollment; (4) the patient being pregnant, planning on becoming pregnant during the trial, or breastfeeding; (5) the patient failing an initial cognitive screener; (6) the patient having a known allergy to adhesive tape or any pertinent components of the DMS patch; (7) the patient not having skin on the anterior chest just above the lower edge of the rib cage, having dermatologic conditions, such as dermatitis or open wounds, in the location where the patch would be placed, or unwilling to refrain from the use of topical products on the skin patch sites; and (8) the patient having <20% proportion of days covered (PDC) with aripiprazole in the 6 months before enrollment.. Qualifying patients were sent an introductory letter in mail, describing the study and inviting them to contact the study team for more information and further eligibility screening by telephone. The study used an opt-out recruitment strategy that entailed contacting participants approximately 7 to 10 business days after a recruitment letter was sent, unless participants contacted the study team to indicate that they were not interested in participating. Once patients were confirmed eligible and were interested in participating, they were seen at a scheduled in-person baseline appointment. At the baseline study visit, written informed consent was obtained, smartphone compatibility was verified or a study-owned smartphone was provided if needed, and baseline assessments and surveys were completed  for technical assistance related to the DMS.As part of the 12-month trial, participants in the DMS arm used the DMS tablet for 90 days. Participants then decided whether to continue beyond the initial 90 days. If participants discontinued the DMS tablet, they restarted their oral aripiprazole as prescribed by their provider. We continued to follow these individuals and obtained pharmacy refill data. Follow-up visits for both the DMS and TAU arms occurred at 3, 6, and 12 months. Early termination visits were completed for DMS arm participants only, per the study sponsor.Participants randomized to the TAU arm continued to receive usual care as provided by their mental health provider. Participants assigned to the TAU arm completed all required study visits and data collection surveys.All study measures were collected by trained study staff at both sites.A research assistant collected demographic data ; clinical data ; and data regarding the use of mobile health devices. Diagnoses were obtained from medical records and the following International Classification of Diseases codes were used: F33.0 (major depressive disorder), F31.0 (bipolar I disorder), F25.0 (schizoaffective disorder), and F20.0 (schizophrenia).The primary outcome was medication refill based upon the number of days covered from baseline to 6 months. Using 2 approaches, we measured adherence using PDC , a leadiGuided by rapid qualitative analysis procedures , we compInterview questions inquired about the feasibility of and facilitators of and barriers to the DMS. Questions for the patients focused on medication experience (pre-enrollment), onboarding to DMS, system usability, satisfaction with support, and feedback. Questions for the providers inquired about the prescriber experience (prestudy), DMS dashboard account setup, system usability, satisfaction with support, and feedback. We used probes  to obtain greater detail and clarify responses. Interviews were completed by a trained research assistant and included a notetaker who recorded responses via a structured note form. After the interview, the research assistant and notetaker debriefed and reviewed interview responses in the context of other interviews. Interviews were conducted via the telephone and were recorded but not transcribed. Patient and provider interviews lasted for 43 (SD 12) minutes on average.Oral aripiprazole refill was measured by the number of days covered from baseline to 3 and 6 months using PDC in both intention-to-treat and DMS-engaged analyses. Owing to the small number of participants, we conducted descriptive analyses rather than a model-based approach. The intention-to-treat analyses used all data from baseline to 3 or 6 months, depending on the outcome, while the DMS-engaged analysis censored participants in the DMS arm at system discontinuation.We followed rapid analysis procedures for data analysis and Microsoft Excel (version 2002) to support coding and analysis. Two authors (AAL and SG) reviewed all notes and debrief notes taken during interviews with patients and providers. These authors used thematic analysis  and the This study was approved by the Durham Veterans Affairs Medical Center Institutional Review Board (ID number 02188) on January 19, 2019.. We issued 6 study-owned smartphones to patients to facilitate enrollment in the study. The sample was on average aged 53 (SD 13.3) years and mostly male , 52% (24/46) self-reported as Black, and 15% (7/46) had a high school education or less. The clinical diagnoses breakdown for enrolled patients included 54% (25/46) with major depressive disorder, 7% (3/46) with schizophrenia, 11% (5/46) with schizoaffective disorder, and 28% (13/46) with bipolar I disorder. Before study enrollment, half  of the enrolled patients had downloaded a health app onto their mobile phone, and approximately one-third  of the participants had used a wearable tech device such as a fitness tracker or smartwatch . Of the eligible patients from participating providers, a total of 46 patients consented and enrolled in the trial , with 21 (46%) participating in the DMS arm and 25 (54%) participating in the TAU arm . We issuartwatch ..In the intention-to-treat analyses, PDC over 3 and 6 months was 0.82 (SD 0.24) and 0.75 (SD 0.26) in the DMS arm and 0.86 (SD 0.19) and 0.82 (SD 0.21) in the TAU arm, respectively. In the DMS-engaged analysis, the DMS arm had 0.85 (SD 0.20) PDC over the period .Patients in the DMS arm stayed engaged with the DMS for 144  days on average. Among the participants in the DMS arm, 5 (24%) stopped using the system by 3 months because of skin irritation adverse events from the use of the DMS patch.Qualitative interviews were completed with 14 patients and 4 providers, with half  of each group from each of the 2 sites. Patient respondents were mostly male 86% (12/14), while 64% (9/14) were Black, 29% (4/14) were White, and 7% (1/14) were Asian. Overall, 86% (12/14) of patient respondents used the DMS for ≥90 days; 43% (6/14) used the DMS for ≥180 days. Of the providers, 50% (2/4) identified as male, 75% (3/4) were providers, and 25% (1/4) were prescribing pharmacist.We identified 5 themes when analyzing the patient data and 5 themes for the providers’ data that described the feasibility and acceptability of using the DMS. Patient themes included pre-enrollment adherence strategies and interest in the DMS, positive impact on medication adherence, system usability challenges, support needs, and suggested improvements to system design and functionality. Provider themes included prestudy concerns for patient medication adherence and interest in the DMS, concerns with the DMS (prestudy), DMS dashboard usability issues and support, challenges in impact of the DMS, and suggestions to increase provider use.Patients described the following pre-enrollment medication adherence strategies: maintaining a routine and setting up reminders through environmental cues . Regarding their pre-enrollment medication adherence strategy, a patient stated that the “hardest part for me is sometimes I can’t remember if I took my meds that day, so that’s why I try to take them first thing in the morning when I wake up...” Patients were interested in using the DMS because they felt that the DMS would help them with their medication adherence. A patient shared the following statement:...I have issues with taking medication on time, not remembering to take it, skipping doses, so the way that the [DMS] would remind you...that would be helpful to someone.Many patients expressed interest in the technology used in the DMS, including a patient who stated the following:I was interested because of the new technology...having the [DMS] app so you can see what your daily activities are.Patients reported the DMS made them more mindful of taking their medication. For example, a patient shared the following statement:[The DMS] made me more cognizant of what time my dosages were and definitely to make sure that I took [my medications] daily...when I opened my phone, I would see the DMS app and remember, oh, I have to take my medicine in the morning.The visual reminder of seeing their DMS patch and the DMS app on their phone acted as a prompt to take their medication. In addition, the DMS app’s medication notifications to log missed doses into the DMS app reinforced the habit of taking their medication consistently. A patient stated the following:The [DMS app] itself, it asks you a series of questions why you forgot to take it and so when you’re going through that it just feels like it’s telling you to not forget it again. I definitely thought it was helpful...it reinforced that good habit of remembering to take your medication every day.Many patients shared that they experienced problems with DMS patch connectivity and skin irritation. A patient described the challenges they experienced as follows:If the [DMS] patch wasn’t paired and I wasn’t paying attention to the [DMS] app, I would take my medication and it wouldn’t register and that was very frustrating...I [also] had problems with the DMS patch adhering and when it wouldn’t adhere, it would unpairwith the DMS appA patient who experienced skin irritation from the DMS patch stated the following:It seemed the more that I changed the DMS patch, the more irritated my skin got with the DMS patch...no matter where I put it.Patients also expressed that the DMS patch was uncomfortable to wear, as the DMS patch caught on things when working in tight spaces and fell off when the patient sweated because of weather or physical activity.Patients experienced challenges with the DMS app during DMS tablet registration . A patient stated the following:I had a lot of issues with the [DMS] app also, the [DMS] app was freezing up on me and I would have to call [DMS support] and we would have to walk through it, we’d have to uninstall it and reinstall it to get the [DMS] app to not be frozen anymore so that it would download the [DMS] patch and pair the [DMS] patch.Overall, despite assistance from study staff, patients felt the DMS was complicated to learn because of the numerous steps needed to complete each process, and many patients shared that they discontinued use because of recurring challenges.Patients expressed that their onboarding to the DMS was helpful because of the in-person, one-on-one training with their local study staff members. Patients who called DMS support felt that DMS support was professional and knowledgeable in describing step-by-step solutions. However, patients reported that DMS support was not always able to resolve the patient’s issues and provide the patient with a long-term fix for their challenges with using the DMS. A patient described this as follows:I continued to have the same issue and that’s why I discontinued using the DMS...[DMS support]’s recommendation to change the patch and re-pair it wasn’t a long-term fix.Patients preferred in-person support for resolving issues with the DMS because study staff members could see their smartphone and DMS app in real time. When asked about support for the DMS, a patient stated the following:Since the study team was local, I was more inclined to call them if I needed help with something...just being there and having them explain to me face-to-face and answer my questions right away, it was helpful...and also that they were able to see my DMS app and see what was going on.Patient feedback for the DMS included two main suggestions: (1) developing a smaller DMS patch or an alternative way to track ingestion that did not involve wearing a DMS patch and (2) improving the usability and functionality of the DMS app as well as the reliability and accuracy of the DMS as a whole.Regarding patch improvement suggestions, a female patient shared the following:If I could have placed the DMS patch somewhere else on my body, I might have continued...if it could be worn somewhere discreetly on the body that would be ideal...where it had to be at the top of my torso, that was problematic for me...I don’t think it would be problematic for a man or someone with a smaller chest.Several patients described how the DMS patch was cumbersome to wear, wearing the DMS patch for long periods led to skin irritation, or that they experienced issues with the DMS patch sticking to their skin. Another patient shared the following:I did not like the DMS patch...it gave me a rash...if we could figure out some other way of doing it without the DMS patch involved that would be wonderful.A patient who experienced DMS patch adherence issues stated the following:I think having to have it on your torso is a problem...how active I am, it just couldn’t stay on me. Maybe if it was moved to an extremity...even if it was a fitness tracker, that would have made that part of the whole process much easier.Regarding suggested improvements to the DMS app usability and functionality, a patient said the following:When you log into the DMS app, it logs you off rather quickly...the login information wasn’t saved, so it was time consuming to access the DMS app.In addition, another patient suggested the following:If your old DMS patch doesn’t upload information [to the DMS app]...having a mechanism to bypass that and record it and send it to DMS support, so you can continue going through the pairing process with the new DMS patchwould be helpfulFinally, some patients suggested that DMS support be more proactive . A patient stated the following:For people like us that’s not as savvy with technology as others are...if [DMS support] have called me and checked, maybe [using the DMS] would have been easier and simpler.In addition, patients described an interest in receiving further training and material on strategies for addressing potential challenges to the DMS, including technical issues with the DMS app, the DMS, DMS patch connectivity, and DMS patch contact. A patient expressed that they would have liked to receive written material, as this patient wanted to refer back to information during the study.Providers shared a common concern for patient compliance with medication and acknowledged several challenges to medication adherence . A provider explained, “It’s hard to know, by patient report, how consistently they’ve been taking the medication...” A provider described challenges patients have in refilling medications as follows:It’s very daunting to get the refills and if the refills are done, they often times...get them late or they forget to order it...it’s not like they don’t want to intentionally not take their medicine.Overall, providers were interested in the DMS, as this system could provide an objective measure of adherence instead of self-report. A provider verbalized as follows:I’m always looking for ways to help my patients...take ownership of their own care and to improve quality of care. Those are the two things that drew me to the DMS.Providers also shared that a common issue can be reconciling what a patient says compared with medication data in the electronic health record. A provider said the following:I was drawn to something like technology that would help both on the provider’s end and the patient’s end to overcome that kind of barrier.Providers expressed concerns with the implementation of DMS in their patient populations, specifically those who experience paranoia and would be apprehensive of using the DMS to track medication adherence. A provider said the following:My initial reaction was paranoid, psychotic patients might be a bit concerned that they’re taking a pill that has a sensor...I was thinking that they might not be willing to take such a medication where one would know [provider or family member] whether they are taking it or not.An additional concern was around the use of the DMS patch because of the provider’s experience with skin sensitivity to adhesives.Providers experienced frustration with the multistep process to log into the DMS dashboard via the notification email. This frustration led them to not check notification emails or log into the DMS dashboard. Improvement suggestions included streamlining the log-in process for the DMS dashboard  and including additional details  in the notification email to encourage them to log into the DMS dashboard more frequently. Notably, none of the providers interviewed called DMS support for assistance with the DMS dashboard, rather these providers called the study staff because of their accessibility.Providers stated that the objective medication adherence data for each patient in the DMS was helpful. However, owing to numerous usability challenges , these providers did not use the DMS dashboard, rather they relied on the study team for information on their patient’s medication data.Provider feedback for DMS centered on (1) improving accessibility to the DMS dashboard and (2) change in DMS data management to improve workflow. Providers suggested a streamlined process for receiving notifications such as including additional information in the notification email and embedding the DMS log-in into the electronic health record  to make it easier to integrate into their regular workflow. A provider stated the following:If there was some way we could incorporate [ingestion data] in our templated notes...while we are writing, documenting the notes, we could click the link to the DMS dashboard...that might work.Finally, many providers recommended that the DMS dashboard and data should instead be managed by an individual in the clinic , as this individual can summarize patient data for the provider. A provider said the following:...If the DMS could be created in a way that there will be a go-between, an intermediary between the provider and the patient on the DMS, who would be monitoring, more closely, the ingestion data...[and summarize] this is number of days of adherence, number of days of not adhering...so that the prescriber has that information right in front of them, even before seeing the patient.This open-label, 2-site, 12-month, provider-randomized trial assessed aripiprazole refill adherence in patients with SMI and examined patient and provider perspectives on the feasibility and acceptability of DMS for this population in the VHA health care system. Our study showed that there was no notable difference in the refill rates between the DMS and TAU arms in intention-to-treat analyses. Among the users of the DMS, the PDC by refill rates were 0.85 (SD 0.20) for those patients when engaged in the DMS. Qualitative findings indicated several challenges to the feasibility and acceptability of the DMS for patients and providers. These challenges included technical issues and contact issues with the DMS patch, including skin irritation and adherence on the skin, and affected the length of user participation as well as overall confidence and interest in using the DMS as a reliable and accurate method of tracking medication ingestion. Both patients and providers discussed recommendations to improve the patient-provider experience and overall satisfaction with the DMS. Notably, patients and providers commented on how to increase confidence in using the DMS for patients with SMI, particularly with patients who may not have extensive experience with smartphones, Bluetooth technology, and health app use.Our results indicated extended patient use of the DMS to manage their SMI. However, our results should be interpreted with caution, as the DMS did not appear to outperform TAU in this study, and enrollment was only a fraction of our original goal. Patients were asked to use the DMS for 90 days, at which time they, along with their provider, could decide whether to continue using the DMS beyond 90 days and up to the full 12 months of study participation. Compared with previous studies using a DMS with an ingestible sensor pill for a period of 8  or 12 1 weeks, tIn another trial using the same DMS  as in ouOur study’s qualitative findings highlighted concerns regarding DMS patch use and contact issues (skin irritation and adherence to skin). In a recent study that evaluated patient responses to using a comparable digital medicine program with an ingestible sensor coencapsulated with antiretroviral therapy medication, patients reported similar issues as seen in our qualitative findings, including issues with patch adherence to skin and overall frustration with using the digital medicine program patch . As detaSeveral considerations should be acknowledged. First, enrollment and data collection for the study ended early per the study sponsor’s request, presumably before more null or potentially negative findings could emerge. Second, technology issues may have affected the continued use of the DMS and engagement in the intervention. Third, a limitation for consideration with regard to adherence is the potential of the Hawthorne effect related to the observation of both participant groups (DMS and TAU) in a trial. This may have affected adherence rates as measured by PDC.In addition, while qualitative interviews were conducted with 14 VHA patients who used the DMS during their study participation, only 4 providers were interviewed. Overall, provider engagement with the DMS dashboard was limited—some providers were unable to share feedback on their experience with the DMS dashboard and the impact that the use of the DMS had on their patients and on patient-provider communication. Providers who consented to participate in the qualitative interviews shared their level of interaction with the DMS dashboard, and interview questions were designed to capture potential barriers to use and to gather feedback on how to increase provider use. Individuals enrolled had a relatively high rate of refill at baseline; future studies using DMS may want to focus on individuals who are having greater challenges with refill adherence. Finally, some patients were interviewed about their DMS experience several months or sometimes up to 1 year after enrollment, which could have affected how well the patients remembered specific details about their experience.Despite the aforementioned limitations, we collected data on the impact of DMS use in a specific patient population, while also gathering detailed feedback on patient and provider experiences, and suggested modifications and considerations for DMS improvements. Evaluation of this novel DMS in the VHA health care system provided insight into real-world use for increased use in community-based, private practice, and public health care settings.Our study of a DMS in patients with SMI did not demonstrate a detectable improvement in medication adherence. Our findings indicate critical issues to consider in improving the feasibility and acceptability of a DMS for patients and providers. Specifically, our study highlights the importance of conducting a feasibility and acceptability trial and collecting quantitative and qualitative data to further refine and improve the DMS for long-term adherence.


Positron emission tomography (PET) probes derived from anti-programmed death ligand-1 (PD-L1) Adnectins, including 18F- and 68Ga-labeled BMS-986192, are recently developed for the prediction of patient response to immune checkpoint blockade. The 68Ga-labeled BMS-986192, in particular, is an attractive probe for under-developed regions due to the broader availability of 68Ga. However, the pharmacokinetics and biocompatibility of 68Ga-labeled BMS-986192 are still unknown, especially in non-human primates, impeding its further clinical translation.Adnectin is a protein family derived from the 10th type III domain of human fibronectin  as the radionuclide–chelator. The resultant probe, 68Ga-NODAGA-BMS986192, was evaluated in terms of targeting specificity using a bilateral mouse tumor model inoculated with wild-type B16F10 and B16F10 transduced with human PD-L1 (hPD-L1-B16F10). The dynamic biodistribution and radiation dosimetry of this probe were also investigated in non-human primate cynomolgus.We developed a variant of 68Ga-NODAGA-BMS986192 was prepared with a radiochemical purity above 99%. PET imaging with 68Ga-NODAGA-BMS986192 efficiently delineated the hPD-L1-B16F10 tumor at 1 h post-injection. The PD-L1-targeting capability of this probe was further confirmed using in vivo blocking assay and ex vivo biodistribution studies. PET dynamic imaging in both mouse and cynomolgus models revealed a rapid clearance of the probe via the renal route, which corresponded to the low background signals of the PET images. The probe also exhibited a favorable radiation dosimetry profile with a total-body effective dose of 6.34E-03 mSv/MBq in male cynomolgus.68Ga-NODAGA-BMS986192 was a feasible and safe tool for the visualization of human PD-L1. Our study also provided valuable information on the potential of targeted PET imaging using Adnectin-based probes.The online version contains supplementary material available at 10.1186/s13550-022-00906-x. Overexpiomarker . Howeveriomarker .10Fn3), Adnectins have comparable binding affinities with whole antibodies, but are smaller in size and simpler in structure [Derived from the 10th type III domain of human fibronectin  [68Ga-labeled BMS-986192 also produced high-contrast images to differentiate hPD-L1-positive tumors from hPD-L1-negative tumors in NSG mice [68Ga-BMS-986192, such as the unknown pharmacokinetics and biocompatibility in non-rodent species. In addition, compared to that of 1,4,7,10-tetraazacyclododecane-N,N',N,N'-tetraacetic acid (DOTA), the coordination cavity formed by 1,4,7-triazacyclononane-N-glutamic acid-N′,N″-diacetic acid (NODAGA) chelates 68Ga with a higher stability [68Ga labeling of BMS-986192 using a bifunctional chelator (BFC) maleimide-NODA-GA, and evaluated the binding affinity and stability of the resultant probe, 68Ga-NODAGA-BMS986192. Mice bearing hPD-L1-B16F10 tumors and healthy male cynomolgus were used to test the feasibility of 68Ga-NODAGA-BMS986192 for the in vivo imaging of PD-L1 expression and its safety profile.Currently, an Adnectin ADX_5322_A02)-based anti-human PD-L1 (hPD-L1) probe,  (NSCLC) . AnotherNSG mice . However_A02-base68Ga/68Ge generator was purchased from ITG Isotope Technologies Garching Gmbh (Germany). PD-10 columns were purchased from GE Healthcare . Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry was performed on a Microflex LT/LRF system . High-performance liquid chromatography (HPLC) /size exclusive chromatography (SEC) was performed on a Waters e2695 system equipped with a Superdex 200 Increase 10/300 GL column . Instant thin-layer chromatography (iTLC) was performed on Eckert & Ziegler Mini-Scan/FC . The samples for binding affinity and biodistribution were measured with a gamma counter .The customized anti-PD-L1 Adnectin ADX_5322_A02 was prepared by ChinaPeptides Co. Ltd . The sequencing report, protein expression, and sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) analysis are shown in Additional file 68Ga-NODAGA-BMS986192 is shown in Fig. 68Ga (370 to 450 MBq) was eluted from a 68Ga/68Ge generator using 0.05 M HCl, mixed with the NODAGA-BMS986192 in sodium acetate, and then incubated at 37 °C for 10 min. The final product, 68Ga-NODAGA-BMS986192, was purified using a PD-10 column using saline as the eluent.The synthesis scheme of 68Ga-NODAGA-BMS986192 was measured by the radio-iTLC. The radiochemistry purity (RCP) was measured by the radio-HPLC/SEC chromatography using 0.01 M phosphate buffer (pH 7.4) as the mobile phase at a flow rate of 1 mL/min. In vitro stability was tested by incubation in fresh human serum at 37 °C for 2 h. Cold 69GaCl3 solution was used for the synthesis of 69 Ga-NODAGA-BMS986192 as the standard reference.The radiochemistry yield (RCY) of 2.Human glioblastoma cell line LN-229 was purchased from BcNa Culture Collection Co. Ltd . Mice melanoma cell lines B16-F10 and hPD-L1-gene transfected B16-F10 (hPD-L1-B16F10) were obtained from Ubigene Biosciences Co. Ltd . The real-time qPCR results confirmed that the hCD274 (hPD-L1) RNA expression of hPD-L1-B16F10 is over 5000 times higher than that of the B16-F10 wild type  as well as the free radioligand was measured in a gamma counter .Binding affinity was tested using a PD-L1 positive human glioblastoma cell line LN-229 . Briefly6) and B16-F10 cells (1 × 106) [3 in size [Bilateral mouse models were established by subcutaneous inoculation of hPD-L1-B16F10 cells (1 × 101 × 106)  at contr in size . Healthyn = 4/group) were intravenously injected with 3.7–5.6 MBq (100–150 μci) 68Ga-NODAGA-BMS986192. Dynamic PET scans were recorded from 2 to 120 minutes post-injection. For blocking groups, cold ADX_5322_A02 (10 mg/kg) was simultaneously injected with 68Ga-NODAGA-BMS986192. Data were reconstructed in 16 time frames  using an ordered subsets expectation maximization (OSEM) algorithm with scattering, attenuation, and decay corrections applied. Three-dimensional regions of interest (ROIs) were drawn over tumors and major organs, and the radioactivities in the regions were measured and quantified as the percentage injected dose per gram (% ID/g).All scans were performed on an uBioEXPLORER PET/CT scanner . Mice (n = 5/group) were intravenously injected with 3.7 MBq 68Ga-NODAGA-BMS986192. At 1 h and 2 h post-injection, tumor, blood, and selected organs were collected, washed, and weighed, their radioactivities measured on a gamma counter. The counts per minute (CPM) values for each sample were converted to percent of injected dose per gram of tissue (% ID/g). Data were decay-corrected to injection time and shown as means ± standard deviation (SD). To further improve the rigor of our study, a supplementary biodistribution study of blocking (n = 3/group) was performed by co-administration of unlabeled ADX_5322_A02 (10 mg/kg) with 3.7 MBq 68Ga-NODAGA-BMS986192, tissues were collected at 1 h post-injection.For ex vivo biodistribution, mice bearing bilateral tumors (68Ga-NODAGA-BMS986192 at a dose of 5.6 MBq (~ 150 μci)/kg was injected via the cephalic vein. Dynamic images were collected from the calvarium to the hypogastrium (A Field of View = 484 mm) for two hours, and reconstructed in 58 time frames . Three-dimensional ROIs were drawn on major organs and the mean standardized uptake values (SUVmean) were computed to form time-activity curves (TACs). Serial whole-body images were acquired by eight consecutive PET scans (15 min per scan) using a continuous bed motion method , for up to 120 min post-probe injection.Cynomolgus were first sedated by intramuscular injection of atropine and Shumianling II, and then anesthetized with a mixture of isoflurane and oxygen through endotracheal intubation. Vital parameters were monitored using a portable electrocardiograph . 68Ga-NODAGA-BMS986192 scans at 10, 25, 40, 70, and 115 min after probe injection, respectively. Source organs were chosen based on the highest probe uptake and previously published reports, which consisted of kidneys, bladder, heart, liver, spleen, adrenals, and the remainder of the body [Data for radiation dosimetry was acquired from whole-body the body . ROIs ovthe body .n = 3) were fixed, embedded in paraffin, and cut into a 4 μm thick sections. The sections were then deparaffinized to retrieve antigen using 10 mM citrate buffer (pH 6.0). Slides were treated with 3% H2O2 for 10 min, blocked with 5% goat serum for 1 h, and then incubated with anti-hPD-L1 mouse monoclonal antibody MIH1  or anti-mPD-L1 rat monoclonal antibody MIH5   at 4 °C overnight. Subsequently, slides were washed, incubated with biotinylated anti-mouse IgG, and visualized by adding DAB chromogen. Sections were counterstained with hematoxylin.Harvested tumors , behavior patterns , and signs of injury, pain, and illness. Blood samples were collected for routine blood test as well as cardiac, hepatic, and renal functions. Heart, liver, spleen, lung, and kidney organs were also collected for histopathological examination.Toxicity tests were performed in accordance with the Organization of Economic Co-operation and Development (OECD) guidelines for testing chemicals with minor modifications. A total of 12 healthy C57BL/6 mice  were randomly assigned into saline control, acute toxicity (24 h post-injection), and subchronic toxicity (7 d post-injection), in which the treated groups were injected with a single dose (37 MBq) of p value < 0.05 was considered to be statistically significant. Differences between groups were evaluated using Student's t test or one-way analysis of variance followed by post hoc Tukey multiple comparisons.All data were presented as mean ± standard deviation from three to five independent replicates. Statistical analyses were performed using GraphPad Prism 5.0 , and a 69Ga-NODAGA-BMS986192 were 11162.168, 14212.190, and 14273.838, respectively, as characterized by mass spectroscopy . Blocking with an excess of unlabeled ADX_5322_A02 (10 mg/kg), on the other hand, reduced the probe uptake by 65 ± 7% in the hPD-L1-B16F10 tumors, confirming the target specificity of the probe. There was a considerable variation in the peak radioactivity within the tumor microenvironment, as detected by TACs of PET imaging. This variation can be attributed to the extent of tumor blood flow and the massive excretion from the kidneys. Nevertheless, the tumor uptake of radioactivity decreased gradually over the first hour post-injection, and then slowly plateaued for the later time points. In comparison, the muscular uptake of radioactivity increased from 0 ~ 5 min post-injection, and then decreased steadily, yielding better contrast for images at later time points  Fig. . PET ROITable Murine PD-L1 was expressed in about 20% of the B16F10 cells regardless of transfection  is much higher than that of 68Ga-DOTA (log K = 21.33). The coordination cavity of DOTA, consisted of four nitrogen atoms and four oxygen atoms, is too big for 68Ga3+, whereas only four nitrogen atoms and two oxygen atoms contribute to the actual chelation. In comparison, the coordination cavity of NODAGA chelates 68Ga3+ with a higher stability [This study reported an optimized  log K = .98 is mutability . Compare log K = .33. The 68Ga-NODAGA-BMS986192 toward hPD-L1. In comparison, no blocking effect was observed in the B16-F10 wild-type tumors (murine PD-L1 positive) and other immune organs [68Ga-NODAGA-BMS986192 toward murine PD-L1 or non-specific accumulation.The PET imaging and ex vivo biodistribution studies demonstrated the rapid and homing of the radioprobe toward hPD-L1 positive tumors Fig. . The bloe organs , confirm68Ga-NODAGA-BMS986192 to assess tumor PD-L1 expression and stratify patients that can potentially benefit from anti-PD-L1 treatment. In addition, no excess unlabeled precursor was needed during PET probe administration due to the low overall background, which result in a feasible and costless medical examination than antibodies [68Ga-FAPI-46 (7.80E−03 mSv/MBq) [68Ga-PSMA-11 (1.08E−02 mSv/MBq) [68Ga-NODAGA-BMS986192 to major organs. Therefore, this probe may have a better safety profile for further clinical translation. Moreover, the excretion of radioprobe from patients can be accelerated by drinking and urination which can further improve imaging quality and reduce radiation exposure.The PET images and TACs of NHPs Figs. , 5 showetibodies . NotablymSv/MBq)  or 68Ga-mSv/MBq) . Togethe68Ga labeling method and obtained 68Ga-labeled BMS-986192 with higher stability. However, the uptake of the radiotracer in the hPD-L1+ tumor was lower than that in the previous reports, probably because the murine PD-L1 expressed in wild-type B16F10, albeit with low affinity, still trapped some of the tracers from the blood circulation. Immunocompetent murine models have been widely used in Immuno-SPECT/PET imaging [68Ga-NODAGA-BMS986192 compared to immune-deficient Balb/c and NSG mice. Furthermore, radiation dosimetry analysis in NHPs provided vital information to make clinical transitions of 68Ga-NODAGA-BMS986192 move forward.Compared to reported preclinical studies based on ADX_5322_A02 , 21, all imaging , 33, 36. imaging . Therefo68Ga-NODAGA-BMS986192, has shown very promising results as imaging agents for their favorable physicochemical properties, such as small size, high stability, high affinity and specificity, rapid tumor uptake, and normal tissue clearance. Moreover, the mild radiolabeling conditions and rapid imaging after 1 h p.i as well as the outstanding biosafety and radio-safety profile make it potential for clinical application of PD-L1 imaging. Collectively, these results are highly encouraging for the application of 68Ga-NODAGA-BMS986192 to screen the beneficiaries of immunotherapy and predict the efficacy of anti-PD-L1 treatment in clinical application.In brief, the Adnectin-based PET probe, 2 field of view. Although this scanner provided an excellent total-body imaging of large animals, the image resolution was sub-optimal for tumor-bearing mice. Third, only male cynomolguses were enrolled for the radiation dosimetry study, which may lead to gender bias, and cautions should be taken when extrapolating the results. Fourth, a second tumor model with endogenously low expression of PD-L1 would help further validate the performance of this probe.Our study has some limitations. First, PET imaging of hPD-L1 was performed in the hPD-L1-B16F10 bearing C57BL/6 mice model instead of a transgenic mice model with humanized PD-L1 . Althoug68Ga-NODAGA-BMS986192) offering hPD-L1 visualization in moderate and flexible radiolabeling conditions. Evaluation of the developed anti-hPD-L1 radioprobe in immune-activity mice provided effective scrutiny of the interaction between radioprobe and the immune microenvironment. Coupling with the dosimetry assay in NHPs with the advantage of similarities to human beings, our findings could facilitate the transition of 68Ga-NODAGA-BMS986192 for clinical PET imaging of hPD-L1.We developed an optimized Adnectin-based anti-hPD-L1 probe  The 10th type III domain of human fibronectin, BC, DE, and FG loops represent variable domains; (B) The core sequence of ADX_5322_A02 (underlined part representing the binding site of hPD-L1). Figure S2 (A) DNA sequence report of the ADX_5322_A02;  SDS–PAGE results of Adnectin expression and purification. Figure S3 hPD-L1-B16F10 cell line RT-qPCR results indicated that the human CD274 RNA expression rate is 556047% overexpressed compared to B16F10 wild type. Figure S4  The accurate molecular weight of obtained ADX_5322_A02, NODAGA-Adnectin, and the standard reference 69 Ga-NODAGA-BMS986192 determined by MALDI-TOF mass spectrometry. Figure S5 Representative iTLC result of 68Ga-NODAGA-BMS986192. Figure S6 (A) HPLC analysis of 68Ga-NODAGA-BMS986192. The UV-chromatogram retention time was 11.627 min, and the radio-chromatogram 12.071 min; (B) In vitro serum incubation of 68Ga-NODAGA-BMS986192 up to 2 h determined by Radio-HPLC demonstrated a satisfactory stability of RCP > 99%. Figure S7 The growth curves of mice (n = 4). Subchronic group (37 MBq 68Ga-NODAGA-BMS986192), control group . Figure S8 Representative images of H&E eosin staining . Neither noticeable organ impairment nor obvious inflammation or necrosis was observed for all groups. Table S1 The results of blood routine test (n = 4). Table S2 The results of biochemical analyses (n = 4).


The burden of cardiovascular disease (CVD) on the current aging society in China is substantial. Climate change, including extreme temperatures and humidity, has a detrimental influence on health. However, epidemiological studies have been unable to fully identify the association between climate change and CVD among older adults. Therefore, we investigated the associations between temperature and relative humidity and CVD among older adults in China.We used cohort data from the China Longitudinal Health and Longevity Survey (CLHLS) conducted in 2002, 2005, 2008, 2011, 2014, and 2018. A total of 39,278 Chinese adults 65 years and older participated in the analyses. The average annual temperatures and relative humidity during 2001 and 2017 (before the survey year) at the city level in China were used as the exposure measures. We selected patients with hypertension, heart disease, and stroke to create a sample of CVD patients. The associations between temperature and relative humidity and CVD were analyzed using the generalized estimation equation (GEE) model. Covariates included sociodemographic factors, health status, lifestyle, and cognitive function.The average annual temperature was negatively correlated with the prevalence of CVD. Every 1°C increase in the average annual temperature reduced the rates of hypertension by 3% , heart disease by 6% , and stroke by 5% . The results of the analyses stratified by sex, urban/rural residence, and educational level were robust. The average annual relative humidity was inversely associated with the likelihood of CVD among older adults. Every 1% increase in the average annual relative humidity reduced the rates of hypertension by 0.4% , heart disease by 0.6% , and stroke by 0.08% . However, the effects were more obvious with higher humidity levels (>70).Our findings suggest that higher temperatures and relative humidity may reduce the risk of CVD among older adults. Cardiovascular disease (CVD) is the leading cause of death and poses a critical public health challenge worldwide , 2. SincIn the context of steep climatic shifts, environmental risk factors have emerged as a major public health concern; therefore, an increasing number of studies have investigated the effects of temperature and humidity on CVD. CVD is highly sensitive to the weather of various environments worldwide . RegardiThe role of humidity, which is another important environmental factor, in human health outcomes has attracted increasing attention as well , 20. SomExtreme temperature events occur frequently with climate warming. China is the largest developing country in the world; therefore, its climate changes and environmental pollution may become more serious. Additionally, older adults are at relatively high risk for CVD , and theThis research used data from the 2002–2018 waves of the Chinese Longitudinal Healthy Longevity Survey (CLHLS). The CLHLS is a nationwide survey conducted in 631 randomly selected cities and counties in 22 provinces, covering about 85% of the total population of China. More than 8,500 participants aged 80+ were interviewed at the 1998 baseline survey, and re-interviewed face-to-face every 2 or 3 years. Overall response rate is about 90% at each wave. In each follow-up survey, the CLHLS study re-interviewed older adults who participated in the previous wave and additionally included new recruitments to replace deceased and lost-to-follow-up respondents with the same sex and age. A complete description of the CLHLS is given elsewhere . The PekSince 2002, the CLHLS adopted a wider sample design by interviewing approximately three randomly selected nearby elders aged 65–79 of predefined age and sex in conjunction with every two centenarians. Thus, we used data collected in waves 2002, 2005, 2008, 2011, 2014, and 2018, a total of 16 years. After excluding those samples that only participated in the survey for one wave, the final analysis comprised 39,728 older individuals. The flowchart illustrated data cleansing and inclusion and exclusion criteria .http://data.cma.cn/). To evaluate temperature and relative humidity exposure for each CHLHS participant, hourly data was averaged yearly or seasonally using administrative codes corresponding to the cities where CLHLS samples were taken (non-publicly available information). For those cities without meteorological monitoring points, the missing values of temperature or humidity would be filled in by that of the most adjacent cities. The temperature and humidity exposures for this investigation were: (1) average relative humidity in the past year; (2) average temperature in the past year; (3) average relative humidity during the period from 1990 to the year of survey; (4) average temperature during the period from 1990 to the year of survey. To avoid seasonal bias, we additionally assessed the participants' exposure to temperature using the following: (5) average temperature during cold months; (6) average temperature during warm months. As in earlier publications, the warm months were defined as May through October, and the average temperature for these 6 months was defined as the average temperature for these months (Hourly city-level relative humidity (%) and temperature (°C) data in 2002, 2005, 2008, 2011, 2014, and 2018 were obtained from China Meteorological Data Network  Having hypertension or not . An older adult is viewed as “Having hypertension” if his/her systolic ≥ 140 or diastolic ≥ 90 in the physical examine of the survey. A person without hypertension is defined if his/her systolic < 140 and diastolic < 90. (2) Having heart disease or not . (3) Having Stroke or cerebrovascular disease or not . Whether the participant had the last two diseases was self-reports based on the medical record or the doctor's diagnosis. The samples would be removed if they reported to have heart disease or Stroke/cerebrovascular disease but have neither medical record nor the doctor's diagnosis.To limit the influence of possible confounders, we analyzed current research from PubMed published within the previous decade to identify factors as covariates, including frequent CVD predictors. Sex , age, geographical location , residence , marital status [single  or married], education , and economic status [logarithm of per capita income = ln(per capita household income + 1)] comprised the variables. Per capita household income in CNY denotes the total annual household income divided by the number of family members. A restriction in any activity of daily life, including bathing, dressing, using the restroom, indoor transferring, continence, or eating, is a handicap. Cognitive functioning was assessed based on the Chinese version of the Mini Mental State Examination (MMSE), one of the most widely used instruments for measuring cognitive health in older persons and recording cognitive shifts , 34.This research studied the impact of temperature and relative humidity on the incidence of CVD. The participants' baseline attributes were summarized according to CVD existence or otherwise. We reported data for continuous variables as means and standard deviations (SD), and for categorical variables as frequencies and percentages. The generalized estimating equation (GEE) was used to examine the associations between temperature, relative humidity, and CVD. By examining the data correlational structure, GEE allows us to develop efficient and objective regression parameters . Based oThis study included a total of 39,278 Chinese older adult observations. The mean age of the older adults in the sample was 83.99 ± 11.0 years; the gender composition was generally balanced (46.5% men). The prevalence of CVD was 58.13%, the proportions of hypertension, heart disease and stroke in the total sample are 51.3, 12.1, and 7.6% respectively. Approximately one-quarter (24.5%) of the older adults lived in urban areas. The geographical distribution of samples was relatively balanced, nearly half (46%) lived in east China. In general, the education of the Chinese older adult is low, 53% of the participants were illiterate. The marriage samples accounted for 40.6%. The average number of surviving children of the older adults is 3.54 ± 1.88. The logarithm of the average income of the older adults is 8.21 ± 1.47. 20% of the older adults suffered from disability in activities of daily living, and 16.3% of them had cognitive impairment .The average temperature over the past year was negatively correlated with the prevalence of CVD. In the past year, every 1°C increase in temperature would reduce the prevalence of hypertension by 3% , heart disease by 6% , and stroke by 5% . Every 1% increase in average relative humidity in the past year would reduce the prevalence of hypertension by 0.4% , heart disease by 0.6% , and stroke by 0.08% , 0.94 (95% CI: 0.92–0.95), and 0.95 (95% CI: 0.94–0.97), respectively. The negative relationship between relative humidity and CVD was significant at a level of 10%, as shown by some subgroup analyses. Furthermore, we found that the effects of temperature and humidity on CVD were non-linear. As the temperature increased, the impact on CVD increased. Additionally, the influence of humidity on CVD only occurs at high levels. These associations indicate that lower temperatures and humidity may be linked to a higher incidence of CVD among older individuals.We found that exposure to lower temperatures and humidity was associated with an increased risk of CVD. In terms of the influence of temperature on CVD, recent reviews provided evidence that both cold and heat would increase mortality after deviating from the optimal temperature; however, the epidemiological studies were conducted in different areas where cold effects were stronger than heat effects , 36, 37.The mechanism by which temperature affects CVD remains undetermined; however, it may involve changes in vascular tone, the autonomic nervous system response, and oxidative stress . Saeki eRegarding humidity, as suggested by most related studies, the negative correlation between the average annual relative humidity and the risk of CVD in GEE models is generally not stable , 26. HowCompared to other studies of the effects of temperature and humidity on CVD in China, this study had two unique advantages. First, the samples included in previous studies were mostly limited to a single region or some large cities , 49, 50.This study had several limitations. First, because of the lack of detailed biomedical indicators in the questionnaire data, we were unable to conduct a more in-depth analysis of the effects of temperature and humidity on CVD. Second, the impact of other environmental factors, in addition to temperature and humidity, on CVD should be explored because some may interfere with the effects of temperature and humidity on CVD. For example, northern China uses coal for heating in winter; therefore, cold temperatures are often accompanied by high pollution rates, which may have affected the estimation results of this study. Furthermore, personal exposure varies with indoor ventilation and indoor activity; therefore, it may be appropriate to measure indoor temperature and relative humidity.In conclusion, we found that the average annual temperature was negatively associated with the CVD likelihood and the inhibitory effect of the annual moderate or higher relative humidity on the prevalence of heart disease and stroke among older adults. Our findings indicate that extreme cold and low humidity are independently associated with a higher likelihood of CVD among older adults.https://agingcenter.duke.edu/CLHLS.Publicly available datasets were analyzed in this study. This data can be found at: Ethical approval for the CLHLS was obtained from the Research Ethics Committees of Peking University and Duke University, and all participants provided written informed consent. All methods were performed in accordance with the relevant ethical guidelines and regulations. The patients/participants provided their written informed consent to participate in this study.HC contributed to perform the data analyses and the design and conception of the study. XZ contributed to write the manuscript. All authors contributed to the article and approved the submitted version.