This study profiled the somatic genes mutations and the copy number variations (CNVs) in cerebrospinal fluid (CSF)-circulating tumor DNA (ctDNA) from patients with neoplastic meningitis (NM).A total of 62 CSF ctDNA samples were collected from 58 NM patients for the next generation sequencing. The data were bioinformatically analyzed by  DAVID software.TP53 , followed by EGFR , PTEN , CDKN2A , APC (27/62: 43.55%), TET2 , GNAQ , NOTCH1 , VHL , FLT3 , PTCH1 , BRCA2 , KDR , KIT , MLH1 , ATM , CBL , and DNMT3A . The mutated genes were enriched in the PI3K-Akt signaling pathway by the KEGG pathway analysis. Furthermore, the CNVs of these genes were also identified in these 62 samples. The mutated genes in CSF samples receiving intrathecal chemotherapy and systemic therapy were enriched in the ERK1/2 signaling pathway.The most common mutated gene was This study identified genes mutations in all CSF ctDNA samples, indicating that these mutated genes may be acted as a kind of biomarker for diagnosis of NM, and these mutated genes may affect meningeal metastasis through PI3K-Akt signaling pathway. Towards this end, a recent study [EGFR)-mutant non-small cell lung cancer (NSCLC). Therefore, characterization of gene mutations in CSF ctDNA samples can provide valuable clinical guidance for precision medicine.Neoplastic meningitis (NM) refers to the dissemination of malignant cells to the leptomeninges, and is associated with very poor survival of patients . The print study  geneticaIn the present study, a total of 58 patients with NM, who received lumbar puncture for the CSF cytology examination between October 2014 and June 2018 in the Department of Neurology, The Second Hospital of Hebei Medical University , were enrolled. The diagnosis of these 58 NM patients was entirely according to the clinical signs and symptoms, positive CSF cytology, and/or neuroimaging findings, such as contrast-enhanced brain magnetic resonance imaging (MRI) or computed tomography (CT), and the results from the CSF ctDNA. The NM clinical signs and symptoms included headache, nausea, vomiting, convulsion, lower back pain, cranial nerve paralysis, paresthesia, gait disturbances, vertigo and defects. The positive CSF cytology was defined by the distinctive pattern of the neoplastic cell morphology. That is, cells that had an irregular size and shape, and contained large and polymorphic nuclei with a lobulated state and malformed buds. The chromatin size increased with the basophilic coarse particles, and the mitotic activity was enhanced with aberrant mitosis. The nuclear membrane was usually thickened with a saw-tooth-shaped and wear edge. In addition, the positive neuroimaging revealed the presence of leptomeningeal enhancement. The clinical data are presented in Table The present study was approved by the Ethics Committee of the Second Hospital of Hebei Medical University , and a written informed consent was obtained from each patient or their legal surrogates.About 0.3 ml CSF was centrifuged at 750 r/min for 4 min . After naturally drying on the slide, the deposit was dyed with May–Grünwald–Giemsa liquid  and Alcian blue staining , according to the manufacturer’s protocol, and observed under a light microscope . The determination of the positive result was as follows: cells that had an irregular size and shape, and contained large and polymorphic nuclei with a lobulated state and malformed buds. The chromatin size increased with the basophilic coarse particles, and the mitotic activity was enhanced with aberrant mitosis. The nuclear membrane was usually thickened with a saw-tooth-shaped and wear edge. Analysis was done using a cell medical image analysis system .g. The pellet was stored at − 20 °C, while the supernatant was centrifuged at 10,000 g for an additional 30 min, according to a previous study [The CSF samples were collected from each patient and placed into EDTA tubes, according to our hospital routine protocols. Then, these were centrifuged for five minutes at 1000 us study . The supThe ctDNA samples were subjected to preparation of the Ion Proton library and DNA sequencing, according to the methodologies from previous studies –12. BrieThe barcoded samples were sequenced using the Ion Proton System with Ion PI v2 Chips (Life Technologies) for 100 cycles, while the Ion PI Sequencing 200 Kit v3 (Life Technologies) was used for the sequencing reactions. Then, the SV-OCP143-ctDNA panel  was used to detect the somatic mutations of 143 cancer-related genes. Since the CSF ctDNA samples contained short DNA fragments, the amplicons in the panel were specially designed for the efficient amplification of ctDNA, and the total read counts were more than 25 million to ensure an average base coverage depth over 10,000 folds. In addition, strict quality control criteria were used to ensure that the average uniformity of the base coverage is no less than 95.5% for the reliability of the DNA sequencing and mutation detection.P-value is < 0.01. The second step was to eliminate ant potential DNA strand-specific errors after visual examination of the gene mutations using the Integrative Genomics Viewer  or Samtools (http://samtools.sourceforge.net) software. For the third step, the total amplicon read counts from the Coverage Analysis Plugin were utilized to identify the copy number variants (CNVs). Then, the read counts per amplicon of each sample was normalized to the total number of reads for a given sample, and divided by normalized counts from a composite normal male genomic DNA sample, which yielded a copy number ratio after correcting for GC content. The gene-level copy number was estimated through the determination of the coverage-weighted mean of the GC-corrected per-probe ratio, which was corrected with the expected error, according to the probe-to-probe variance [The raw DNA sequencing data were processed and analyzed using the Ion Proton platform-specific pipeline (Torrent Suite v5.0) with a specific plug-in , which included the readouts of the raw DNA sequences, the trimming of the adapter sequences, and the filtering and removal of poor signal sequences according to previous studies –12. Thesvariance . AfterwaEGFR mutations in lung cancer tissues were obtained from patient’s medical record. Lung cancer tissues of most patients were sequenced before this study by various methods.Information of https://david.ncifcrf.gov/tools.jsp). Then, the data were clasisified as the functional annotation and the KEGG pathway enrichment. A P-value of < 0.05 was set to be statistically significant.Next, the mutated genes were bioinformatically analyzed using the gene ontology (GO) terms  and the P-value of < 0.05 was considered statistically significant.The statistical significance in gene mutation frequency between the two groups was analyzed using Fisher exact test. Pearson correlation analysis was used for correlation analysis. Nonparametric test was used for two independent samples that did not meet the normal distribution. Statistical analyses were performed using SPSS version 19, and a two tailed Clinical characteristics are shown in Table A total of 62 CSF samples were collected from these 58 NM patients, in which three CSF samples were collected from a single patient, while two CSF samples were collected from other two patients at distinct time points. Furthermore, among the 62 CSF samples, 30 CSF samples were collected from 28 patients who received intrathecal chemotherapy and systemic radiotherapy, chemotherapy, and/or molecule-targeted therapy, 11 CSF samples were obtained from 11 patients who received intrathecal chemotherapy, and 12 CSF samples were obtained from 12 patients who received systemic therapy. The remaining nine CSF samples were collected from nine patients who did not receive any anticancer therapy.TP53 , followed by EGFR , PTEN , CDKN2A , APC , TET2 , GNAQ , NOTCH1 , VHL , FLT3 , PTCH1 , BRCA2 , KDR , KIT , MLH1 , ATM , CBL , and DNMT3A   of the 143 cancer-associated genes analyzed in the present study had mutations in at least one NM CSF sample, and 62 (100%) of the NM CSF samples carried at least one mutated gene. The most commonly mutated gene was P < 0.001, Fig. Furthermore, the variant allele frequency was divided into five groups: ≥50, 30–50%, 10–30%, 1–10% and 0.2–1%, respectively. Then, the variant allele frequency was associated with the detectable tumor cells in the CSF samples, and it was found that the frequency of variant allele frequency (≥1%) was higher in the group with detectable tumor cells than that in the group without detectable tumor cells . In the increase in CDK4 copy number that occurred in five lung adenocarcinomas, four of these exhibited an increase in MDM2 copy number. In addition, an increase in MDM2 copy number was also detected from another lung adenocarcinoma patient. Two CSF ctDNA samples had a gain of ERBB2 (HER2) copy number from a parotid carcinoma patient, while an increase in CD44 copy number was identified in three patients, in which each patient has breast cancer, gastric cancer and unknown cancer, respectively. In addition, an increased EGFR copy number occurred in three lung adenocarcinoma patients. Other CNVs of tumor-associated genes were detected in five patients (six positive CSF ctDNA samples) with decreased AR copy numbers, five patients had decreased CD274 copy numbers, three patients each has a decreased PDCD1LG2 copy number, two patients each has an increase in FGFR2, CCNE1, or NKX2–1 copy numbers, respectively, and one patient had increased TIAF1, GAS6, or IL6 copy numbers, or reduced CSNK2A1, JAK2, MED12, or SMAD4 copy numbers.Data on the CNVs in these CSF ctDNA samples were also obtained, and it was found that high CNVs occurred in 22 of 58 NM patients. Among these 22 patients, the primary tumors were 15 lung cancers , four gastric cancers, and one each of breast cancer, parotid carcinoma, and unknown primary cancer. The deletion of the The data on these unique mutated genes were summarized for each group of patients, followed by a gene-annotation enrichment analysis. It was found that the ERK1/2 pathway was mostly enriched by the GO analysis in patients who received both intrathecal chemotherapy and systemic therapy , followed by TP53 , PTEN , TET2 , APC , CDKN2A , GNAQ , and NOTCH1 . A number of gene mutations previously reported with lung cancer were identified in CSF with NM, while EGFR, TP53, PTEN, TET2, APC, CDKN2A, GNAQ, NOTCH1, FLT3, VHL, BRCA2, PTCH1, CBL, MLH1, BRAF, NRAS, TSC2, CSF1R, KIT, MAP2K1, MSH2, TSC1, HRAS, IFITM1 and BCL9 mutations were statistically more common in the present cohort of NM, when compared to the lung cancer noted in the COSMIC database (https://cancer.sanger.ac.uk) , chronic myeloid leukemia , endometrial cancer , bladder cancer , melanoma , glioma , prostate cancer , and non-small cell lung cancer , while the related signaling pathways were the ErbB signaling , VEGF signaling , MAPK signaling , p53 signaling , and m-TOR signaling  pathways , EGFR , PTEN , CDKN2A , APC , and TET2 . These mutated genes enriched by the KEGG pathway analysis was the PI3K-Akt signaling pathway. The ERK1/2 signaling pathway was significantly activated in NM patients who received intrathecal chemotherapy and systemic therapy, indicating that intrathecal chemotherapy and systemic therapy might induce novel gene mutations in NM patients. The present study also identified the variation of gene copy numbers in these 62 samples. In conclusion, the data obtained from the present study demonstrates the following: (1) ctDNA is detectable in all CSF samples; (2) gene mutations are detectable in all CSF samples; (3) the gene copy number varies in all CSF samples; (4) the PI3K-Akt and ERK1/2 signaling pathways are the most altered signaling pathways for these mutated genes; (5) novel gene mutations are induced by intrathecal chemotherapy and systemic therapy in NM patients; (6) lung cancer  is the major primary tumor in the present cohort of NM patients. Future studies would investigate the usefulness of the CSF and ctDNA for the early detection of NM patients, and target these mutated genes for the therapy of NM patients or even patients with these primary tumors.In the present study, NM patients were enrolled for the detection of CSF ctDNA, gene mutations and copy number variations. Furthermore, the present cohort of NM patients revealed that the large majority of primary cancers was lung adenocarcinoma, and 10 patients had NM as the first clinical manifestation, although seven of these 10 patients were clarified for their primary tumor. Afterwards, 62 CSF samples were acquired from 58 NM patients, and all samples contained detectable ctDNA, indicating that detection of CSF ctDNA is a sensitive biomarker for NM patients, since the ctDNA may not originate from benign tumors and non-neoplastic conditions, according to previous studies . Indeed,TP53, EGFR, PTEN, KIT and KDR, could be crucial or at least partially crucial in mediating primary cancer for meningeal metastasis. In particular, numerous isoforms and/or spliced variants of PI3Ks participate in the regulation of various cell processes, such as cell cycle progression, cell polarization, migration, survival and metabolism, as well as tumor angiogenesis [Dioscorea bulbifera-induced apoptosis in human colorectal carcinoma cells [Indeed, the PI3K-Akt signaling pathway, including but is not limited to ogenesis . Furtherogenesis , and theogenesis . For exaogenesis . In humaogenesis –22. Prevogenesis , ovarianogenesis , nasophaogenesis , prostatogenesis , colorecogenesis , and gasogenesis . The prema cells . Conversma cells . This spCDKN2A, CDK4 and MDM2. As it is known, the deletion of tumor suppressor CDKN2A was associated with melanoma and pancreatic neuroendocrine tumors metastasis, and the reduced survival rate of patients [CDK4 and MDM2 mutations occurred in melanomas and liposarcoma, while ERBB2 mutations occurred in breast cancer, EGFR mutations occurred in astrocytoma, and MYCN mutations occurred in neuroblastoma [CD44 expression was associated with the aggressive clinicopathological characteristics of various human cancers [TP53 mutations in the CSF samples of NM patients with lung cancer, in which the COSMIC database also confirmed that these were the most frequently mutated genes in the CSF of NM patients, when compared to primary lung cancer. Furthermore, a higher mutational rate was found from EGFR, TP53, PTEN, TET2, APC, CDKN2A, GNAQ, NOTCH1, FLT3, VHL, BRCA2, PTCH1, CBL, MLH1, BRAF, NRAS, TSC2, CSF1R, KIT, MAP2K1, MSH2, TSC1, HRAS, IFITM1 and BCL9 genes in NM with lung cancer, when compared to lung cancer patients without NM. A previous study revealed that EGFR mutations predisposed to the leptomeningeal metastases of EGFR-mutant non-small-cell lung cancer [In addition, the present study also identified the CNVs of different genes in the CSF ctDNA samples, and the most affected ones were patients , 31. Indpatients . Furtherblastoma . Altered cancers . In addig cancer , and theEGFR activating mutations between primary lung cancer and NM, the present data shows that the EGFR activating mutations in the CSF samples were roughly consistent with those of primary lung cancer. For example, the T790M in two CSF samples were sequentially collected during the TKI therapy. Furthermore, according to the gene set enrichment analysis, it was found that the ErbB, VEGF, MAPK, and m-TOR signaling pathways were significantly enriched in the 45 CSF samples obtained from NM patients with lung cancer, suggesting that the alterations of these signaling pathways might promote NM in lung cancer patients. In particular, the ErbB signaling pathway can regulate cell proliferation, migration, differentiation and apoptosis through the crosstalk with the PI3K-Akt, MAPK, or other signaling pathways [After associating these pathways .EGFR 19Del mutation in the CSF sample, and thereby received icotinib. Thereafter, the CSF sample revealed the EGFR 19Del and T790M mutations. Hence, the treatment was switched to AZD9291, which is a third generation EGFR TKI agent. A high response rate was exhibited by patients with tumors harboring the EGFR T790M mutation, as well as a high capacity to penetrate into the CSF by crossing the blood-brain barrier [EGFR T790M mutation mediates the acquired resistance to EGFR TKI [EGFR T790M resistance mutation [Lastly, the representative patient in the present study provided a unique showcase. The genetic profiling of the CSF ctDNA clearly reflected the dynamic changes in these identified driver genes and treatment responses. That is, this patient was detected to have the  barrier , 37. TheEGFR TKI , 39, andmutation .However, the present study does have some limitations. For example, the cohort has a relatively small number of patients, and the different treatment options could not be separated to analyze the association of gene mutation, and the changes in gene copy number with treatment responses in these patients. Thus, future studies with a larger sample size from multiple institutions could help to solve these issues. Though we have found that mutated genes detected in CSF are enriched in the PI3K-Akt and ERK1/2 signaling pathways, it is hard for us to explain the detected mutations are gain of function mutations or loss of function mutations in such pathways. Thus, we will perform the functional analysis in the following studies. Due to the target amplicon sequencing, we could not address the whole genome alterations and yield the comprehensive landscape of NM. So there are any other area we will focus on to make more depth analyses in the future studies.This study identified gene mutations in all CSF ctDNA samples, indicating that these mutated genes may be acted as a kind of biomarker for diagnosis of NM, and these mutated genes may affect meningeal metastasis through PI3K-Akt signaling pathway.Additional file 1. Table


Several options for granulocyte colony-stimulating factor (G-CSF) prophylaxis of chemotherapy-induced febrile neutropenia are available to patients worldwide. We have developed a novel patient-reported outcome measure, the Satisfaction and Experience Questionnaire for G-CSF (SEQ-G-CSF), to help understand patients’ perspectives of and satisfaction with different G-CSF options.Three oncology nurses and 40 adult oncology patients in the United States were enrolled and participated in focus group discussions to develop and refine the SEQ-G-CSF. Nurses had ≥ 5 years of experience treating oncology patients and were currently involved in the management of oncology patients receiving G-CSF prophylaxis. The patients had breast cancer, lung cancer, non-Hodgkin lymphoma, or prostate cancer (10 patients in each group) and were receiving G-CSF prophylaxis via injection or the on-body injector (OBI) device. The preliminary SEQ-G-CSF contained an item relevance questionnaire and three SEQ modules . Twenty-one patients  discussed their experience and satisfaction with G-CSF. Their most common experiences were G-CSF effectiveness, convenience and benefits of the OBI, and relationships with healthcare providers. Side effects and having to undergo additional treatment were also reported. Satisfaction with aspects of G-CSF included the OBI and effectiveness of G-CSF treatment; dissatisfaction included inconvenience (having to return to the clinic the next day and administration of the injection) and the insurance approval process. The SEQ-G-CSF was finalized after three rounds of cognitive interviews and includes five domains related to general satisfaction (one item), treatment burden (four items), travel burden (two items), time burden (four items), and treatment compliance (two items).The SEQ-G-CSF is a novel instrument that quantifies a patient’s experience and satisfaction with different G-CSF options using 13 easy-to-understand items. This study provides evidence for the content validity of SEQ-G-CSF. Although further psychometric testing is required, the SEQ-G-CSF may be a useful addition to clinical trials, observational studies, and clinical practice.The online version contains supplementary material available at 10.1186/s41687-020-00277-8. One of the side effects of chemotherapy is febrile neutropenia (FN), a condition associated with fever and low numbers of certain white blood cells. Granulocyte colony-stimulating factors (G-CSF) are recommended as a preventive treatment for chemotherapy-induced FN for many types of cancer. With different types of G-CSFs as well as different administration modes of G-CSFs, and currently no available valid method to quantify patients’ experience with G-CSF, the study objective was to develop an instrument to assess the chemotherapy patient’s satisfaction and experience with these different G-CSF treatment options.The recommendation of G-CSF use by physicians and hospitals varies based on preference, and there is currently no valid method to quantify patients’ experience with G-CSF. Thus, it is important to have an established metric for measuring satisfaction in patients who have received G-CSF treatment, to help understand their perspective.We developed a novel questionnaire (SEQ-G-CSF) to quantify a patient’s experience and satisfaction with different G-CSF options, using 13 easy-to-understand items. The questionnaire was developed with input from patients and nurses to ensure that it fully captures their experience.This study has provided evidence of content validity for the SEQ-G-CSF, and our results indicate that patients understand the items in the questionnaire. While additional testing of other forms of validity and reliability is required, the SEQ-G-CSF may be a useful addition to clinical trials, observational studies, and clinical practice. Results of studies including this new tool will provide information to help patients and providers make an educated decision on their treatment, when there are multiple options for preventing FN.Chemotherapy-induced febrile neutropenia (FN) is associated with negative outcomes such as extended hospital stays, higher medical costs, dose reductions, dose delays, poorer treatment response, and higher risk of mortality . CurrentAlthough pegfilgrastim is administered once per chemotherapy cycle, offering increased convenience to patients and caregivers, it should be administered the day after chemotherapy . It is oFactors that could influence use of G-CSF include clinical guidelines, hospital protocol, inclusion of G-CSF on drug formularies and physician and patient preference  as well The SEQ-G-CSF instrument was developed using an iterative process Fig. . The conPrior to initiation of the study, we were unable to identify a fit-for-purpose PRO measure relevant for understanding patients’ experience and satisfaction with different G-CSF options. We therefore developed a de novo measure based on desk research with a preliminary conceptual framework that adapted relevant concepts from several existing treatment satisfaction tools –23. PrelOncology nurses were screened for eligibility and recruited through a third-party recruitment vendor to participate in focus group discussions conducted through an online platform. Eligible oncology nurses had ≥ 5 years of experience with treating oncology patients and were currently involved in the management of oncology patients receiving G-CSF prophylaxis. The 60-min nurse focus group discussion took place via a webcam link. A semi-structured, standardized discussion guide was used in the focus group discussions with oncology nurses. The purpose of the focus groups was to assess the patient experience and satisfaction with G-CSF. Topics included their background and experience as well as their assessment of patient experience and satisfaction with oncology treatment and G-CSF prophylaxis Table . Nurses Breast cancer, lung cancer, non-Hodgkin lymphoma, and prostate cancer patients were screened for eligibility and recruited through a third-party recruitment vendor to participate in focus group discussions on an online platform.Oncology patients were required to submit proof of a physician’s verification of cancer diagnosis and chemotherapy medication. Other eligibility criteria included: patients aged ≥ 18 years old with a self-report of breast cancer, lung cancer, non-Hodgkin lymphoma, or prostate cancer as well as current or recent (< 6 months from screening date) chemotherapy and prophylaxis with G-CSF. In addition, patients were required to be US residents, have access to the internet, have sufficient proficiency in the English language to complete the consent process and participate in focus group discussions, and be willing to be audio-recorded during discussions.n = 10 in each group) and two subgroups . The target quotas were to recruit ten patients per cancer type: five who were OBI-naive and five who were OBI-experienced. The goal was to conduct up to three rounds of focus groups; a minimum of two to three focus groups per round.Based on the types of concepts being explored, it was estimated that a minimum of 10 patients were required per group to achieve saturation . Quota sThe study protocol was reviewed and approved by Quorum Review Institutional Review Board , and all participants provided their written informed consent before the start of each interview discussion.The objective of the concept elicitation portion of the patient focus group was to elicit patients’ experience with G-CSF prophylaxis Table . Semi-stFocus group discussions took place via a webcam link; 90 min for Round 1 and 2 participants and 60 min for Round 3 and individual interviews. Each focus group discussion was audio- and video-recorded, and transcripts from audio data were de-identified and coded. All participants were compensated for their time based on an assessment of the fair market value and length of focus group discussion.Based on results from round 1 concept elicitation focus group discussions, the initial version of the SEQ-G-CSF was revised, and additional hybrid concept elicitation and cognitive interview focus groups were planned as needed to support attainment of concept saturation.The objective of the cognitive interview portion of the patient focus group was to elicit patients’ experience with G-CSF prophylaxis and assess comprehension whether the instructions, items, response options, and terminology in the SEQ-G-CSF were understandable and relevant Table . The iniAfter the cognitive interview portion, patients completed three additional SEQ modules, including an SEQ-G-CSF item relevance questionnaire . The iteSaturation was defined as the point at which no substantially new themes, descriptions of a concept, or terms were introduced as additional rounds of focus group discussions were being conducted. The concepts arising from the concept elicitation sessions were evaluated for saturation as the discussions were being conducted. No further additional interviews were considered if saturation was achieved.During focus group discussions, structured counts were taken for each treatment experience and satisfaction concept to document saturation using a saturation grid that displayed the first mention of emerging concepts. For the concept elicitation portion, a coding framework was developed related to the meaning of treatment experience and satisfaction. The preliminary conceptual framework was imported into ATLAS.ti v8 to code the concept elicitation portion of the focus group transcripts. A content analysis approach was used to analyze and interpret the concept elicitation–coded data . For theQuantitative data from case report forms were descriptive ; analyses were done using SAS version 9.4 .Four oncology nurses were screened and were eligible to participate in the focus group discussions; however, one nurse was unable to participate on the scheduled date. On average, each of the three nurses who participated in the study saw 118 oncology patients per week across the four cancer groups, had experience administering G-CSF , and had provided the OBI to patients in each of the four cancer groups (an average of six patients per week).n = 21/40) participated in the concept elicitation portion of the focus groups.A total of 40 patients who were receiving G-CSF, 19 OBI-experienced and 21 OBI-naive, were enrolled. Between December 06, 2018, and January 17, 2019, 10 focus group discussions were conducted with 36 patients, and individual interviews were conducted with four patients. All 40 patients completed the questionnaires and participated in the cognitive interview portion; data were collected from three rounds of focus groups. Fifty-three percent of the participants  years. Overall, 40% of patients had metastatic cancer, 83% had completed chemotherapy within 6 months of enrollment, 75% had received their last G-CSF treatment within 6 months of enrollment, and 48% had received pegfilgrastim for febrile neutropenia prophylaxis.Oncology nurses described patients’ satisfaction and tolerability with chemotherapy and G-CSF treatment based on their own clinical practice experience. Nurses reported that patients receiving chemotherapy and G-CSF derive satisfaction from experiencing a treatment response (an indication that the treatment is working), minimal hospital stays and co-pays, and good communication with their cancer care team. Nurses also stated that patients tend to tolerate treatment side effects if they see a treatment response or if their quality of life is not significantly impacted.The concept elicitation portion of the patient focus groups identified patients’ experience and satisfaction with chemotherapy and G-CSF prophylaxis, including side effects, treatment effect, and the treatment process and setting.Eighteen patients  discussed their experience and satisfaction with chemotherapy. The most common positive aspects related to patients’ experience with chemotherapy were treatment effectiveness (50%), relationships with healthcare providers (39%), and having supportive relationships (22%). The most common negative aspects were side effects of chemotherapy (56%); emotional burden, including fear and feelings of vulnerability (33%); time burden due to frequent treatment visits, follow-up visits, and time spent waiting for and receiving treatment (33%); concerns over drug safety (33%); and treatment burden (22%).Satisfaction with aspects of chemotherapy included positive interactions with healthcare providers (33%), treatment effectiveness (28%), minimal time burden (17%), and having insurance and minimal paperwork (11%). Dissatisfaction with aspects of chemotherapy were health outcomes after chemotherapy (33%), disease course (22%), treatment burden (22%), and side effects (17%).Twenty-one patients  discussed their experience and satisfaction with G-CSF. The most common positive experiences were G-CSF effectiveness (90%), convenience , relationships with healthcare providers (38%), and benefits of the OBI (29%), including convenience, ease of use, available support, and reduced travel and time burden  and effectiveness of G-CSF treatment (47%). Dissatisfaction with aspects of G-CSF were inconvenience  and dose regulation  (Table n = 15), four focus group discussions in round 2 (n = 15), and two focus group discussions (n = 6) and four individual interviews in round 3. Of the 40 patients who participated, 65% provided their overall impression of the SEQ-G-CSF item relevance questionnaire, 20% stated that questions should be removed, 18% stated that questions should be changed, 13% stated that the length of the questionnaire was appropriate, and 13% stated that additional questions were needed.A total of 10 focus group discussions with 36 patients and individual interviews with four patients were conducted: four focus group discussions in round 1  that are not scored. The conceptual framework was revised to link individual items to hypothesized PRO domains and overall summary scores (as applicable) based on findings from the focus group discussions. The final conceptual framework includes five domains related to general satisfaction (one item), treatment burden (four items), travel burden (two items), time burden (four items), and treatment compliance (two items) (Fig. The SEQ-G-CSF was developed and tested using a systematic, iterative process to confirm content validity based on accepted best practices and guidance for PRO development –27. The Several treatment satisfaction instruments were available prior to development of the SEQ-G-CSF, including those that assess satisfaction with medication delivery via injections –34 or saThe economic burden of clinic visits to receive G-CSF therapy after chemotherapy is substantial . In addiIn our study, patients most commonly rated G-CSF effectiveness, convenience and benefits of the OBI, and relationships with healthcare providers as positive experiences, whereas side effects, undergoing additional treatment, injection discomfort, and sleep interruption were the most common negative experiences. Surprisingly, economic burden did not factor into patients’ satisfaction with G-CSF treatment. The SEQ-G-CSF can help clinicians assess the quality of G-CSF treatment in terms of process, which is one of the three pillars of healthcare quality . In addiOur study has several limitations. The overall small sample size makes it difficult to generalize patients’ treatment experience and satisfaction to the broader oncology patient population. In addition, there was heterogeneity in G-CSF prophylaxis experiences; participants received several types of G-CSF prophylaxis with varying modes of delivery, frequency, and duration of treatment. Furthermore, in the event of time limitations during each focus group discussion, the interviewer prioritized the review of SEQ-G-CSF items for which participants had proposed changes in prior discussions.With the availability of different options for administering G-CSF prophylaxis, it is important to support decision making with patients’ perspectives of and satisfaction with their treatment experience. The SEQ-G-CSF is a novel PRO measure that quantifies a patient’s experience and satisfaction with different G-CSF options, using 13 easy-to-understand items. While additional testing is required, this study provides evidence of content validity. The use of SEQ-G-CSF in further studies may provide an opportunity for further psychometric testing, including item reduction, scoring, reliability, and construct validity assessment, which is necessary to support the use of the SEQ-G-CSF in scientific studies and clinical practice. After additional testing, the SEQ-G-CSF may be a useful addition to clinical trials, observational studies, and clinical practice.Additional file 1.


Monocytes are recruited into the cerebrospinal fluid (CSF) of patients with neurosyphilis, suggesting abnormal chemokine expression. We aimed to investigate the aberrant expression of chemokines in the CSF of these patients.® Human Chemokine Antibody Array C1. CCL24 and CXCL7 levels in the patients’ CSF and serum were further measured using RayBio® CCL24 and CXCL7 ELISA kits.CSF and serum samples were collected from patients with neurosyphilis between July 2017 and June 2019 in the Dermatology Department, Second Affiliated Hospital of Zhejiang University. Differences in the expression of 38 chemokines between patients with and without neurosyphilis were detected using RayBioP = .0185), CXCL7 (P < .0001), CXCL13 (P < .0001), CXCL10 (P < .0001), and CXCL8 (P < .0001) were significantly higher in patients with than without neurosyphilis. ELISA confirmed significantly higher CCL24 and CXCL7 levels in the CSF of patients with than without neurosyphilis . Increased CCL24 and CXCL7 expression was seen throughout all neurosyphilis stages, had moderate diagnostic efficiency for neurosyphilis, and correlated poorly with CSF cell count and Venereal Disease Research Laboratory titer. CSF CCL24 levels also correlated poorly with CSF protein concentration.Ninety‐three CSF and serum samples of patients with syphilis were collected. Antibody array analysis showed that the CSF levels of CCL24 (Abnormally high CSF chemokines levels may play a role in the pathogenesis of neurosyphilis. In patients with neurosyphilis, the CSF shows both pleocytosis (mainly lymphocyte accumulation) and mildly increased protein concentration.New potential biomarkers are warranted for discriminating neurosyphilis from other diseases. MicroRNAs (miRNAs) seem to be potential candidate biomarkers, due to their low immunity, good transmission, and ability to cross the blood‐brain barrier. Chen et al reported that miR‐590‐5p, miR‐570‐3p, and miR‐570‐5p are upregulated, while miR‐93‐3p is downregulated in the CSF of patients with neurosyphilis.In this study, we investigated the abnormal expression of chemokines in the CSF of patients with neurosyphilis in more detail, by using a semi‐quantitative chemokine array that includes overlapping components with that used by Wang et al, but also tests for additional chemokines, such as CCL24 and CXCL7, which have not been previously tested.22.1Cerebrospinal fluid and serum samples of patients with syphilis who were referred to the Department of Dermatology, Second Affiliated Hospital of Zhejiang University, School of Medicine, Hangzhou, China, were consecutively collected between July 2017 and June 2019. This study was approved by the Ethics Committee, and written informed consent was obtained from all participants.https://stacks.cdc.gov/view/cdc/31403). Non‐neurosyphilis refers to patients with syphilis, including primary, secondary, and tertiary syphilis, who do not meet the diagnostic criteria of neurosyphilis. Different stages of neurosyphilis were subdivided according to the clinical features and associated laboratory test results.The diagnostic criteria of neurosyphilis complied with the Sexually Transmitted Diseases Treatment Guidelines, 2015, of the US Department of Health and Human Services Centers for Disease Control and Prevention , according to the manufacturer's instructions. The full chemokine array paradigm is illustrated in Figure TM MP Imaging system; Bio‐Rad). Signal intensities  curves. Spearman correlation analysis was performed to evaluate the relationship between the levels of CSF chemokines and CSF total protein concentration, cell count, VDRL titer, and serum rapid plasma reagent (RPR) titer. All statistical calculations were performed using GraphPad Prism 7.0 (GraphPad Software), with statistical significance set at P < .05.Variations in mean values are expressed as standard errors, and variations in medians are expressed as interquartile range (IQR). Differences between groups were analyzed by Student's 33.1Cerebrospinal fluid and serum samples from 93 patients with syphilis were available for this study. The characteristics of the overall study population are shown in Table P = .0185), CXCL7 (P < .0001), CXCL13 (P < .0001), CXCL10 (P < .0001), and CXCL8 (P < .0001) in the CSF of neurosyphilis patients were higher than those in the CSF of patients without neurosyphilis  Table  were sels Figure .3.2P = .0037; Figure P = .0118; Figure We then validated whether the levels of CCL24 and CXCL7 were elevated in the CSF of patients with neurosyphilis, as compared to the levels in those without neurosyphilis, by performing ELISA measurements. The CSF levels of CCL24 and CXCL7 were both statistically significantly higher in patients with neurosyphilis than in those without  of 0.791 .Fig S1Click here for additional data file.Supplementary MaterialClick here for additional data file.


Remote cerebellar hemorrhage (RCH) is a rare complication following cranial or spinal neurosurgical procedures. Traditionally, RCH has been associated with frontal or frontotemporal craniotomy with supine patient positioning. Though the exact etiology is unknown, theories have described patient positioning and excessive cerebrospinal fluid (CSF) drainage intra-operatively as contributing factors to cerebellar displacement (cerebellar sag), obstruction of venous flow, and pathogenesis of RCH. We report two cases of RCH following a prone, suboccipital craniotomy-C1 laminectomy and a temporal burr hole evacuation of a subdural hygroma. In each case, a large volume of CSF was rapidly evacuated intra-operatively. To the best of our knowledge, both instances represent relatively rare settings for RCH. Additionally, we conducted a comprehensive literature review of PubMed, EMBASE, and Web of Science for all cases of RCH in which peri-operative CSF leakage was explicitly detailed.Although RCH is thought to be a rare complication of frontotemporal and frontal craniotomies, this case report signifies that RCH may occur in the setting of sub-occipital craniotomy or even after minimally invasive burr hole procedures. For these procedures, careful symptomatic monitoring and follow-up imaging remain essential in diagnosis. Controlled CSF drainage may be useful in mediating dramatic alterations in intracranial pressure (ICP) and cerebellar sag contributing to RCH. Remote cerebellar hemorrhage (RCH) is a rare intracerebral hemorrhage isolated from surgical sites following cranial or spinal procedures that may result in significant peri-operative neurologic morbidity and mortality . DiagnosCase 1A 55-year-old female presented to our clinic with several months history of right-sided facial numbness and left upper extremity numbness and tingling that progressed to the right upper extremity. She was referred by an outside neurologist following MRI that demonstrated a large, right-sided foramen magnum lesion compressing the spinal cord and brainstem. Imaging revealed a homogenously enhancing, extramedullary, intradural mass extending from the level of the foramen magnum to the mid-C2 vertebral body with an associated dural tail Figures , 1B.Significant mass effect, compression, and displacement of the corticomedullary junction and upper cervical cord were identified. A working diagnosis of foramen magnum meningioma was made and the patient was scheduled for resection of the lesion. A sub-occipital craniotomy and C1-C2 laminectomy were performed with the patient in the prone position. During the exposure, standard arachnoid dissection facilitated CSF egress permitting a maximal safe resection of the tumor with minimal cerebellar retraction. On a post-operative day (POD) one, the patient developed nausea, vomiting, and positional headache exacerbated while lying flat. CT showed an interval increase in ventricular size with new cerebellar hyperdensity concerning hemorrhage and effacement of the fourth ventricle  was placed. The RCH stabilized, hydrocephalus resolved, and EVD was removed on POD 10; the patient was discharged on POD 14. At one-month follow-up, the patient displayed a complete resolution of symptoms following physical rehabilitation; strength was 5/5 in bilateral upper and lower extremities with a Karnofsky Performance Status (KPS) of 90.Case 2A 79-year-old man with a history of hypertension, diabetes, hyperlipidemia, and aspirin use presented to our clinic with a three-day history of worsening generalized weakness and headache. The patient denied any symptoms other than weakness and slight tremor. Prior head CT displayed a large right-sided, hollow hemispheric hygroma with approximately 1 cm midline shift. The patient denied prior head trauma and no other focal neurologic deficits were observed. After five days of aspirin cessation, the patient underwent hygroma evacuation. Two burr holes were made and, following the dural opening, high-pressure CSF was immediately evacuated Figures , 3B.A Jackson-Pratt (JP) drain was placed over the posterior burr hole followed by closure. The patient awoke on POD one able to follow commands but over subsequent hours, decompensated and had a tonic-clonic seizure. Head CT revealed a new cerebellar hemorrhage and the patient was taken for emergent suboccipital decompression Figures , 3D. On A comprehensive literature review was performed in PubMed, EMBASE, and Web of Science databases to identify RCH cases with perioperative CSF leakage. Keywords included: remote, cerebellar, and hemorrhage. All abstracts were reviewed, and relevant articles were collected using the following inclusion and exclusion criteria. All articles in the English language presenting cases of RCH with perioperative CSF leakage were included. Review articles, letters and commentary, and cases without specific mention of perioperative CSF loss were excluded.Including our own, twelve cases of RCH following craniotomy with peri-operative CSF leakage were identified -10. The Seven cases of RCH following spinal surgery with peri-operative CSF leak were identified -16. The Remote cerebellar hemorrhage is a rare complication following cranial and spinal surgeries with an estimated incidence of 0.08-0.6% . RCH likSeveral symptoms have been described in patients who develop RCH following supratentorial craniotomy including altered level of consciousness, motor deficits, gait ataxia, and seizures . ConcernInterestingly, our review yielded only one asymptomatic case identified incidentally on follow-up imaging. It is possible minor intra-operative or post-operative CSF leaks present with milder symptoms due to less dramatic intracranial pressure changes. Konig et al. identified higher frequencies of RCH following resection of large supratentorial tumors and hypothesized that resection of larger masses confer greater reduction of intracranial pressure (ICP) and increased transmural venous pressure, increasing RCH risk . HoweverThe role of patient positioning in RCH pathogenesis is uncertain. In craniotomy cases with position reported, patients were commonly placed supine with the head rotated opposite the craniotomy . InteresOur study highlights the importance of avoiding rapid excessive CSF loss during surgery, to reduce the risk of RCH peri-operative complications. The use of controlled CSF drainage techniques, including small controlled dural openings, provides favorable strategies to regulate peri-operative CSF drainage rates. If large losses of CSF are observed, patients should be carefully monitored and imaged post-operatively. Placing patients in reverse Trendelenburg can be performed if there is a concern for RCH after high-risk procedures, although no data supporting this method has been shown. For symptomatic patients with altered mental status and reduced consciousness, RCH secondary to hemorrhagic venous infarction should be suspected.Our cases illustrate that RCH is not limited to supratentorial craniotomies. Our case following burr hole surgery represents a rare neurosurgical setting for post-operative RCH. To our knowledge, only two previous cases of RCH following burr hole evacuation of subdural hematomas have been described ,20. The Although RCH is largely considered a frontotemporal and frontal craniotomy complication, we report the first known case of RCH following suboccipital craniotomy. Additionally, we report one of the first cases of RCH following burr holes for subdural hygroma evacuation, demonstrating the risk of RCH even after less-invasive procedures. Although traditional descriptions of RCH etiology detail intra-operative over drainage of CSF, we hypothesize it is the rate of CSF drainage, rather than volume, that contributes to RCH pathogenesis. Controlled CSF drainage may prevent this occurrence from happening. Similarly, after burr holes for subdural hygromas, it may be wise to place JP drains to gravity rather than bulb suction.


Background:Ex situ donor liver machine perfusion is a promising tool to assess organ viability prior to transplantation and platform to investigate novel therapeutic interventions. However, the wide variability in donor and graft characteristics between individual donor livers limits the comparability of results. We investigated the hypothesis that the development of a split liver ex situ machine perfusion protocol provides the ideal comparative controls in the investigation of machine perfusion techniques and therapeutic interventions, thus leading to more comparable results.Methods: Four discarded human donor livers were surgically split following identification and separation of right and left inflow and outflow vessels. Each lobe, on separate perfusion machines, was subjected to normothermic perfusion using an artificial hemoglobin-based oxygen carrier solution for 6 h. Metabolic parameters as well as hepatic artery and portal vein perfusion parameters monitored.Results: Trends in hepatic artery and portal vein flows showed a general increase in both lobes throughout each perfusion experiment, even when normalized for tissue weight. Progressive decreases in perfusate lactate and glucose levels exhibited comparable trends in between lobes.Conclusion: Our results demonstrate comparability between right and left lobes when simultaneously subjected to normothermic machine perfusion. In the pre-clinical setting, this model provides the ideal comparative controls in the investigation of therapeutic interventions. Each lobe has their own inflow and outflow vessels as well as bile drainage, during Four donor livers were included in this study, resulting in a total of eight perfusions. All donor livers included in this study were originally retrieved with the primary intention for transplantation as per the policy of the National Health Service Blood and Transplant (NHSBT). The organs were subsequently rejected for transplantation by all UK liver transplant centers and, following that, were offered nationally for research by NHSBT. The authors had no influence in the process of declining donor livers. This was done by the transplant surgeons at each center. Informed consent for research use of donor organs was obtained by specialist nurses in organ donation from the donor's next of kin during informed consent for organ donation. Authorisation for research was mediated by each centre's specialist nurse in organ donation. All methods described were performed in accordance with NHSBT guidelines and regulations. Study ethical approval was obtained from the London-Surrey Borders National Research Ethics Service (Reference Number 13/LO/1926) and the NHSBT ethics committee (Reference Number 06/Q702/61). No organs used in this study originated from executed prisoners.Donor liver exclusion criteria for this study were: gross macroscopic appearance indicative of moderate or severe steatosis; asymmetric or poor perfusion demonstrated during organ retrieval; presence of hepatic malignancy; organ subjected to machine perfusion prior to being discarded.During transportation to our center, the organs were preserved in University of Wisconsin fluid at hypothermic temperatures in ice as per current standard clinical practice for static cold storage in the United Kingdom. Donor livers were split while in cold storage prior to commencement of perfusion in order to ensure that both lobes were subjected to similar ischaemic times and assessed simultaneously when placed in the perfusion devices. Upon receipt by our center, the liver was initially cleared of excess tissue in order to assess the quality of the organ. This also allowed for assessment of the organ's outflow, with respect to the hepatic veins and post-hepatic inferior vena cava, as well as the inflow, in terms of the hepatic arterial tree and portal vein . Since tOnce the vena cava was opened, the liver hilum was cleared further of excess tissue in order to identify the left and right branches of the portal vein and hepatic artery, respectively . ParticuIf vessel length and diameter was deemed to be inadequate for safe cannulation of either lobe, particularly with respect to the isolated branches, then a decision was made to reconstruct the vessels to enable proper positioning of the cannulas for perfusion. This was done by fashioning a conduit using native vessels for extra length in one of two ways: (1) resecting the distal portion of the main arterial or venous trunk retained by either lobe, which was then anastomosed onto the branch of the other; (2) utilizing the length of donor iliac vessels for the reconstruction  . Which tBoth perfusion devices were primed with our previously described  perfusioThe pre-perfusion weight of each lobe was recorded post-splitting. The arterial and portal venous supplies were cannulated with each lobe positioned inside the organ reservoir such that the open drainage from the hepatic veins could be visualized directly. Perfusion was commenced at 36–37°C with oxygenated pulsatile flow and non-pulsatile flow in the hepatic artery and PV, respectively. Perfusion of each lobe was commenced within five to 15 min of each other . PerfusiData analysis was carried out using Prism 7 . Continuous data at each timepoint was compared using Wilcoxon signed-rank test. Statistical significance was set at p <0.05. Data is presented as per lobar mass as well as following normalization of pre-perfusion lobar weight per gram of tissue.Four livers were included in the study. Information relating to donor characteristics and liver characteristics can be found in . Right lLactate levels were comparable at the start of perfusion, T0, and decreased significantly in all lobes up until the end of the perfusion experiment, T6 . Trends Hepatic artery and portal venous flows were relatively consistent in both left and right lobes with a trend toward an increase in flow rate as the perfusion experiment progressed . DiffereOverall histological analysis showed that both liver lobes behaved in a similar manner throughout the perfusions. Lobar architecture was well-preserved in both pre- and post-perfusion biopsies. Some variable centrilobular necrosis was observed at both time points. However, this was present to the same degree before and during NMLP. All biopsies were PAS positive with staining showing a range of intensity from mild to strong and ranging from even distribution throughout the parenchyma to patchy. Whatever the variability between liver, each of the paired lobes showed strong similarity, with no change throughout the perfusion. In addition, patterns of histology did not correlate with any parameters of functional assessment. Only one liver showed evidence of significant macrovesicular steatosis which did not change between lobes or throughout the perfusion .Several studies have demonstrated that NMLP enables the functional recovery of discarded donor livers and provides a window of opportunity for viability assessment prior to transplantation. However, results are compounded by small sample size as a consequence of the limited availability of discarded donor livers for research, further compounded by the inherent differences between donor livers. This limits comparability between groups of individual livers.ex situ end-ischaemic NMLP, therefore, has the potential to provide a platform for suitable comparative controls for the investigation and assessment of therapeutic interventions prior to being subjected to the rigor of a clinical trial. Research published previously by our group allowed for an initial four-hour window of NMLP before a full viability assessment of the perfused organ is carried out  did not require any reconstruction as the blood was freely drained and collected in the device reservoir. As such, hepatic veins anatomy and inferior vena cava accessories did not influence the surgical technique or the perfusion parameters. The situation would be different if a closed circuit was being used or if clinical use of these segmental grafts was under consideration.The incorporation of NMLP to the split liver model has the benefit of enabling liver functional assessment under near-physiological conditions. The use of an artificial acellular hemoglobin-based perfusion fluid also has its advantages. Unlike blood products (derived from various donors), it has low immunogenicity and may therefore be of greater value in the study of liver-specific immune cell populations and mechanistic studies . SecondlIt must be noted that there are limitations to the design of this perfusion model. First, splitting of the liver requires considerable surgical expertise and increases the cold ischaemia time beyond what is usual for the preparation of a whole liver for machine perfusion. Secondly, due to the liver's anatomy, the right lobe is invariably larger than the left lobe and therefore they cannot be split into entities of equal mass without compromising blood supply. Furthermore, the shorter and smaller caliber of hepatic artery and portal vein branches makes them much more difficult to cannulate. We attempted to circumvent this issue by retaining the main trunk of the vessel with one lobe while performing vascular reconstructions on the other, using donor iliac vessels or larger caliber sections from the proximal celiac trunk or splenic artery (if included in the specimen). In all experiments this provided the additional length for safe cannulation. However, this technique further increases the cold ischaemia time of the liver and would also require the appropriate surgical expertise for a robust reconstruction that would not compromise lobar perfusion. All these aforementioned factors may lead to differences in the perfusion characteristics of each lobe and must be considered. Finally, not all livers can be split. This is often a result of variations in arterial anatomy due to branching patterns which would compromise the inflow of a segment of the lobe if divided.We acknowledge that the data presented in this manuscript is from a small number of human donor livers. Nevertheless, the results of this proof-of-concept study indicate that trends in the functional recovery and metabolic parameters between lobes were very similar and may therefore prove useful in the assessment of responses to therapeutic interventions in the pre-clinical normothermic setting. Additionally, this model addresses the variability that exists between individual donor livers by enabling the same donor liver to be placed in the treatment and control groups. This also maximizes use of resources as one donor liver for pre-clinical studies.Other benefits observed during these experiments came from the use of a perfusion machine with an open circuit. These included: facilitated manipulation of the graft for direct visualization, direct visualization of and access to graft outflow and superior access to graft for direct therapeutic interventions when compared to closed circuit alternatives.Liver splitting is a feasible model for providing comparative controls for pre-clinical normothermic machine perfusion research. Further work should involve the optimisation of this protocol to minimize the need for surgical expertise when preparing the liver for machine perfusion as well as its application to other perfusion modalities, namely, hypothermic and sub-normothermic perfusion. This novel split liver model can be tested for cellular therapies to investigated cellular phenotype and lineage changes and future pharmacological interventions of donor liver before implantation.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the London-Surrey Borders National Research Ethics Service (Reference Number 13/LO/1926) and the NHSBT Ethics Committee (Reference Number 06/Q702/61). Informed consent for research use of donor organs was obtained by specialist nurses in organ donation from the donor’s next of kin during informed consent for organ donation.JA, YB, LW, and SA designed the study and established the methodology. JA, LW, DO-B, and VR conducted the experiments. JA and LW collected the data. DM and HM supervised the liver splitting procedure. JA performed the data analysis and drafted the manuscript. SA and GR reviewed the histological specimens. YO, MP, HM, and DM contributed to critical revision and editing of the final manuscript. All authors approved the final version of this manuscript.JA is a clinical research fellow at the Queen Elizabeth Hospital in Birmingham and employed by University Hospitals Birmingham. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Diagnostic evaluation of febrile young infants is challenging. Empirical antimicrobial treatment is therefore common practice in this setting despite high percentage of causative viral infections. The objective of this study was to investigate the impact of rapid enterovirus cerebrospinal fluid polymerase chain reaction (CSF EV PCR) test on hospital length of stay (LOS) and antimicrobial treatment duration in young febrile infants.Retrospective observational study comparing duration of antimicrobial treatment and hospital LOS before  and after  the introduction of rapid CSF EV PCR testing in infants < 90 days of age presenting with fever and CSF pleocytosis at the University Children’s Hospital Zurich. Additionally, the same variables were compared after test introduction between CSF EV PCR positive and negative children.n = 42) compared to both EV negative (n = 28)  and untested patients (n = 58) , and also in tested compared to untested group patients . Hospital LOS was significantly shortened in EV positive compared to EV negative patients , while an overall reduction was not observed between tested and untested group patients.One hundred twenty-eight children were enrolled in the study, 58 before and 70 after the introduction of rapid CSF EV PCR testing. Duration of antimicrobial treatment was significantly shortened in EV positive (In this study we demonstrate that antimicrobial treatment duration could be significantly shortened in neonates and young infants < 90 days of age with aseptic meningitis after the introduction of a rapid CSF EV PCR test compared to untested patients before test introduction. The diagnostic evaluation of febrile young infants (< 90 days of age) is challenging. Clinical and laboratory findings at presentation do not allow to reliably identify and treat only those with serious bacterial infections –3. NeonaThe objective of this study was to investigate the impact of a rapid CSF EV PCR with time to result of 3 to 6 h on hospital LOS and duration of antimicrobial treatment in children < 90 days of age with aseptic meningitis.We performed a retrospective observational study at the University Children’s Hospital of Zurich. Demographic, clinical, and laboratory data including hospital LOS and duration of antimicrobial treatment were extracted from medical records and the microbiology laboratory database.S. epidermidis, viridans group streptococci, Bacillus spp., Propionibacterium spp., Corynebacterium spp., Aerococcus spp. and Micrococcus spp.) in blood or CSF cultures or growth of 2 or more bacteria in urine culture.Previously healthy infants < 90 days of age (i.e. children without any underlying condition) presenting at our emergency department with fever  were eligible for inclusion if they underwent standardized full sepsis evaluation  prior to antimicrobial treatment between May 1, 2014 and June 30, 2017. In house rapid CSF EV PCR testing was introduced June 1, 2015. The test was not routinely applied to all febrile infants under the age of 90 days. The attending physician decided whether to perform the test based mainly on exposure history, seasonality and the presence of CSF pleocytosis. Infants presenting with fever and CSF pleocytosis, defined as the presence of ≥5 white blood cells (WBC) per μl, after test introduction  were included in the tested group if they had CSF EV PCR performed at admission, and those presenting within 13 months before its introduction  in the untested group. Although the reported thresholds for definition of CSF pleocytosis in children < 8 weeks may be higher , 23, theCSF EV PCR was performed using the commercially available Xpert® EV method . The assay runs on GeneXpert® system with integrated sample processing, viral RNA extraction, reverse transcription (RT) and nucleic acid amplification, as well as detection of the target sequence by real-time PCR, in a single reaction cartridge. The following EV species and subtypes are detected: Species A , Species B , Species C , Species D  and Poliovirus (Poliovirus 1–3). The test has a running time of less than 2.5 h and was available every day between 07:00 AM and midnight in the in-house microbiology laboratory. During these 17 h, treating physicians were informed immediately about the test results with a time to result of 3 to 6 h. CSF samples awaiting processing were kept at 4 °C until testing.U test was used to compare medians between groups. For the comparison of more than two groups Pearson Chi-squared and Kruskal-Wallis tests were used for categorical and continuous data respectively. Subsequent post hoc testing by pairwise comparisons was performed in case of statistical significance in order to identify the groups with significant differences. All analyses were performed with IBM SPSS Statistics version 24. Two-tailed p-values smaller than 0.05 were considered to be statistically significant.Continuous data were described using median and interquartile range (IQR), whereas categorical data were described using frequency and percentage. Mann-Whitney p < 0.001).During the three-year study period, a total of 228 previously healthy infants < 90 days of age presented with fever and underwent full sepsis evaluation Fig. . After ip < 0.001) and untested group patients   Fig. a. Furthep < 0.001 as shown in Fig. p = 0.537; Fig. Hospital LOS was significantly shorter in CSF EV PCR positive compared to negative patients .Bacterial contamination rates detected in cultures from blood, CSF and urine did not differ between CSF EV PCR positive, negative and untested group patients Table . The anaIn this observational study we demonstrate that antimicrobial treatment duration could be significantly shortened in neonates and young infants < 90 days of age with aseptic meningitis at our institution after the introduction of a rapid CSF EV PCR test compared to untested patients before test introduction. This is mostly due to the effect of the significant reduction of antimicrobial treatment duration by more than 60% to a median length of treatment of less than 24 h in EV positive compared to EV negative patients . Additionally, a significant reduction of median hospital LOS by at least one day for febrile infants < 90 days of age tested CSF EV PCR positive was achieved compared to those tested negative (3 vs 4 days). These findings confirm previous studies reporting a reduction of antimicrobial treatment duration between 30 and 50% , 15, 19.The majority of neonates and young children < 90 days of age presenting with fever are hospitalized in good general condition and empirically treated for potential sepsis, even if unlikely, awaiting negative culture results from blood, CSF and urine for 48–72 h . As the In our study the effect of EV testing in CSF was not yet strong enough to demonstrate a significant reduction of hospital LOS before and after test introduction. Possible explanations for these findings are the not yet systematic introduction of the test (i.e. the attending physician decided whether or not to perform the test), the lack of rapid implementation (i.e. discharge of the patient by the responsible team of physicians at bedside as soon as the test result was available) and to a lesser extent the non-availability of the test during the night. It remains to be seen whether systematic testing, immediate reporting of the test result and its subsequent implementation with cessation of antimicrobial treatment in EV positive infants, if in good general condition and with normal urine analysis, may allow to achieve a significant overall reduction of hospital LOS.Significantly higher CSF leucocyte levels have been described for CSF EV PCR positive patients  and coulTherefore, the fact that pleocytosis was the main indication to perform the test is a limitation of our study. Other limitations of our work are its retrospective design, the small sample size and the unsystematic use of the CSF EV PCR test in febrile infants < 90 days of age.The use of rapid CSF EV PCR testing in febrile infants < 90 days of age undergoing full sepsis evaluation leads to a significant reduction of antimicrobial treatment duration and hospital LOS in EV positive compared to negative patients but also to an overall reduction of antimicrobial treatment in patients with aseptic meningitis. These benefits of rapid CSF EV PCR testing in febrile neonates and infants on duration of treatment and hospital stay may be increased if systematic testing is introduced , time to reporting of the test result is optimized, and medical staff is instructed to rapidly implement the positive test result on patient management.


There are few treatment options for patients with leptomeningeal metastases (LM).We report a case series of patients with breast cancer and LM treated with intra‐CSF topotecan (TOPO). Outcome was assessed by clinical exam and MRI at baseline, at end of induction (4‐5 weeks), then every 3 months; CSF cytology was determined at baseline and with each treatment.Thirty‐one women  received treatment. At baseline, 68% had positive CSF cytology, and 90%, leptomeningeal enhancement on MRI. 84% of patients also received focal RT (not during TOPO) and 77% received concomitant systemic hormonal or chemotherapy. Median number of TOPO treatments was 14.5 ; median duration of treatment, 11 weeks (1‐176); and median OS, 6.9 months . Patients remaining progression‐free during 4‐6 weeks of induction (81%) had a median OS of 11.5 months . Overall neurologic PFS at 6, 12, and 24 months was 39%, 26%, and 6%, respectively. Clearing of CSF malignant cells for >3 consecutive samples occurred in 10/21 (48%) patients with positive CSF cytology at baseline, remaining clear for a median duration of 15.9 months . Grade 3 adverse events included headache or vomiting (3pts), T2 hyperintensity surrounding the ventricular catheter (2 pts), and meningitis (2 pts).Intra‐CSF TOPO, with focal RT as needed for symptomatic areas of enhancement produced durable clearing of CSF malignant cells in 48% of patients positive at baseline, with promising median PFS and OS. In this case series, intra‐CSF topotecan treatment of women with leptomeningeal metastases and breast cancer was associated with durable CSF clearing of malignant cells in 48% of patients, median OS of 6.9 months, and 11.5 months in those completing 4‐6 weeks induction. Thirty‐two percentage of patients remained free of neurologic progression at 12 months. A method for treatment of patients with VP shunts and programmable valves is also described. LM was defined as: clinical symptoms and/or exam findings consistent with LM; and either positive CSF cytology for malignant cells at baseline, or evidence of leptomeningeal enhancement on MRI of brain and/or spine performed at baseline. All patients had a histologic diagnosis of breast cancer, confirmed by review of official pathology reports. All MRIs, clinical assessments and CSF examinations were performed at our institution, and radiographic evidence of LM was confirmed by review of actual MRIs and reports.Prior to treatment, all patients had baseline CSF cytologic examination and enhanced MRI neuroimaging of brain and spine. All patients were treated using a similar treatment regimen and schedule of assessments, modified from a prior reported studyPatients received TOPO 0.4 mg twice weekly for 4‐6 weeks (induction period), then were assessed for clinical benefit by clinical examination, CSF cytology, and repeat MRI of all brain and/or spine sites which showed LM enhancement at baseline. At end of induction, patients were considered to be deriving “clinical benefit” and continued on intra‐CSF TOPO if they did not have clinical neurologic decline; unacceptable adverse events; new or progressive meningeal enhancement on MRI; or had converted from previously negative CSF cytology to positive. A small number of patients who were otherwise deriving clinical benefit but with positive CSF cytology were treated beyond induction, until either worsening of symptoms, radiographic progression or persistent positive cytology for more than 3‐4 more weeks. After induction, maintenance TOPO was administered as detailed in Table TOPO treatment was administered to all patients via an intraventricular reservoir. If patients required removal of the reservoir for clinical reasons, treatment was continued by lumbar puncture, until a new reservoir was placed. All patients requiring ventriculo‐peritoneal (VP) shunts had programmable valves placed in the system. The procedure for treatment in shunted patients was as follows: immediately prior to dosing, the valve was programmed to the highest position . Treatment was then administered, with the valve setting at the high position for 1‐2 hours, after which the valve was restored to the pretreatment setting. The decision to utilize 1‐2 hours was based on the CSF T ½ of 1.3 hours for TOPO and active lactone metabolite,Clinical records detailing symptoms and neurologic exam findings, CSF cytopathology reports, and MRI imaging studies and reports were reviewed on all patients. Neurologic response was retrospectively defined, based on the best status achieved  as compared with the baseline assessment. “Improved” was defined as improvement of neurologic symptoms, combined with objective improvement or stability on neurologic examination, and in the absence of CSF cytologic and/or radiographic progression. “Stable” was defined as stable neurologic symptoms and signs, in the absence of CSF cytologic or radiographic progression. Worsening was defined as any other status.MRI response was categorized by the best status achieved , as compared with the baseline scans, as described on the official neuroradiology reports. A designation of “improved” required that the report specifically mentioned reduction in leptomeningeal enhancement as compared to baseline, and in the absence of new areas of enhancement. “Stable” was defined as no progression or new areas of leptomeningeal enhancement. “Worse” was defined as any other status.CSF was considered positive if the official cytopathology report described suspicious or malignant cells, and negative if containing no malignant cells, or only atypical or degenerated cells. CSF cytology was considered evaluable for response if CSF malignant cells were present at baseline, prior to TOPO treatment. Time to cytologic response (clearing) was defined as time from the first TOPO treatment to the time of the last of three consecutively negative CSF cytologic determinations. The duration of cytologic response was defined as the time from the third consecutively negative CSF cytology to the time of recurrent positive CSF cytology, or to time of cessation of TOPO for any reason.Neurologic progression free survival (NPFS) was assessed by retrospective review of serial clinical assessments as documented, in combination with the MRI findings. Neurologic progression was defined as any of the following: worsening of neurologic examination and/or LM‐related neurologic symptoms; patient or physician decision to discontinue treatment; appearance of positive CSF cytology after durable prior negative cytology; persistently positive cytology for three consecutive determinations ; or progressive enhancement on MRI. Neurologic progression‐free survival (NPFS) was defined as the time from first intra‐CSF TOPO dose to development of neurologic progression, death due to any cause or loss of follow‐up. Overall survival (OS) was defined as the time from first intra‐CSF TOPO dose to death from any cause or loss of follow‐up.All time‐to‐event endpoints were analyzed using the Kaplan‐Meier method, survival distributions were compared using the logrank test, and hazard ratios (HR) were calculated using Cox Proportional Hazards models. Additional exploratory analyses were performed to compare the outcomes of patients based on HER2 status (Negative vs Positive) and presence of programmable VP shunt (Not Present vs Present).All patients signed consent for placement of intraventricular devices, and informed written consent for the intra‐CSF TOPO treatment. This case series review was approved by the Institutional Review Board.3A total of 31 women with breast cancer and LM received treatment with intra‐CSF TOPO. Patient demographics, ER/PR/Her‐2 marker status, and pre‐TOPO treatments were summarized  evaluable patients with positive CSF cytology at baseline, 10 (48%) cleared CSF of malignant cells for at least three consecutively negative CSF specimens, meeting our definition for CSF response. The median time from start of TOPO to conversion of CSF cytology from positive to negative was 4 weeks . In the patients clearing CSF of malignant cells, the median duration of CSF cytologic clearing was 15.9 months . Two patients initially cleared CSF of malignant cells for 69 and 48 weeks, respectively, but later relapsed; one during treatment interruption for replacement of a ventricular device, and one after entering the every 3‐month maintenance period. These patients were re‐challenged with intraventricular TOPO with induction dosing, and both cleared CSF of malignant cells again, at 6 and 1 weeks, respectively, with CSF remaining negative for malignant cells on TOPO for 105 and 49 additional weeks, respectively.CSF CA 15‐3 levels were measured in three patients at baseline; all had positive CSF cytology also at baseline. In all three, CSF cleared of malignant cells during TOPO treatment and serial CA15‐3 levels decreased below the limits of detection of the assay (<8 U/ml).P = .002), as well as in patients with programmable VP shunts, compared to those without shunts , though this latter comparison was underpowered due to small sample size. OS6, OS12, and OS24 are detailed in Table P < .001) and in patients with programmable VP shunts , though this latter comparison was underpowered due to small sample size.Survival outcome is presented in Figure Adverse events included reversible headache (3), vomiting (2) in the first 2 days following treatment. For subsequent cycles these patients were premedicated with antiemetics and dexamethasone, TOPO dose reduced to 300 micrograms, without headache or vomiting.. Two (6%) patients developed bacterial meningitis requiring removal of the intraventricular reservoir; after recovery, treatment was resumed by intralumbar approach. One patient developed incisional dehiscence at the site of the intraventricular device. Two patients developed increasing T2 hyperintensity around the intraventricular catheter, and treatment was subsequently administered intralumbar. Two additional patients discontinued treatment due to unrelated adverse events . We did not identify hematologic toxicity attributable to intra‐CSF TOPO.4Topotecan has shown activity in treatment of breast cancer.Following intra‐CSF treatment of patients with LM and breast cancer, survival has typically ranged from 2.9‐5.4 months,In our study, most patients (94%) had advanced stage disease and had received multiple prior systemic treatment regimens at time of LM diagnosis. Despite this, nearly one‐half of patients with initially positive CSF cytology cleared CSF of malignant cells, which was reasonably durable . CSF clearing was observed relatively early on, at a median of 4 weeks. The observed 45% CSF cytologic response rate compares favorably with that previously reported (21%) with TOPO treatment of patients with LM and unselected malignancies.P = .0059). Of possible relevance was that only two of the four Her‐2+ patients were receiving systemic Her‐2 inhibitors.The majority (52%) of our patients with LM had ER+ PR+, Her‐2‐negative neoplasms, 16% were Her‐2 positive, and 26% were triple‐negative, which is similar to the proportions reported by others.The authors recognize the inherent limitations of a retrospective case series review with a modest sample size. It is conceivable that our study population included a disproportionate number of patients harboring tumors with indolent biological behavior; notable was that the median time from initial breast cancer diagnosis to LM diagnosis was 7.0 years in our population. Since this parameter has not typically been reported in prior series, it is difficult to make comparisons. It is likely that local‐field radiation therapy at baseline for bulky or symptomatic disease (74%), concomitant hormonal (50%) or systemic chemotherapy (46%) might have influenced outcome; however, it is noted that these concomitant therapies have been provided in conjunction with intra‐CSF therapy in previously reported studies.5Intra‐CSF TOPO, alone or in combination with focal RT has activity in breast cancer patients with LM, producing durable clearing of CSF malignant cells in nearly one‐half of patients, notable neurologic PFS6 and median OS, and a number of long‐term survivors. Intra‐CSF TOPO was generally well tolerated. The results support further testing within the context of a prospective trial.The authors declare no conflicts of interest regarding work involved in this study or manuscript.Kurt A. Jaeckle involved in conceptualization, methodology, validation, investigation, data curation, writing, review, and editing of the manuscript. Jesse Dixon and Stephen Keith Anderson involved in methodology, data analysis, writing, review, and editing of the manuscript. Kathy Hebenstreit carried out investigation, data curation, and review of the manuscript. Alvaro Moreno‐Aspitia carried out investigation, review, and editing of the manuscript. Geraldo Colon‐Otero, Samarth L. Reddy, Tejal Patel, and Edith Perez contributed to investigation and review of the manuscript.


The objective of this study was to determine whether neurotoxic kynurenine metabolites, induced by inflammation, in plasma and cerebrospinal fluid (CSF) are associated with symptom severity and nigral pathology in Parkinson's disease (PD).Clinical and MRI data were obtained from 97 PD and 89 controls. We used ultra‐performance liquid chromatography to quantify kynurenine metabolites and high‐sensitivity multiplex assays to quantify inflammation in plasma and CSF. We evaluated group‐wise differences as well as associations between the biomarkers, motor and nonmotor symptoms, and nigral R2* (MRI metric reflecting iron content).PD subjects had >100% higher 3‐hydroxykynurenine and 14% lower 3‐hydroxyanthranilic acid in plasma. The 3‐HK in plasma was closely associated with both symptom severity and disease duration. PD subjects also had 23% lower kynurenic acid in the CSF. Higher CSF levels of the excitotoxin quinolinic acid were associated with more severe symptoms, whereas lower levels of the neuroprotective kynurenic acid were linked to olfactory deficits. An elevated quinolinic acid/picolinic acid ratio in the CSF correlated with higher R2* values in the substantia nigra in the entire cohort. Plasma C‐reactive protein and serum amyloid alpha were associated with signs of increased kynurenine pathway activity in the CSF of PD patients, but not in controls.Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.In PD, the kynurenine pathway metabolite levels are altered in both the periphery and the central nervous system, and these changes are associated with symptom severity. Replication studies are warranted in other cohorts, and these can also explore if kynurenine metabolites might be PD biomarkers and/or are involved in PD pathogenesis. © 2020 The Authors.  Parkinson's disease (PD) is a common, progressive movement disorder that also features debilitating nonmotor symptoms. The neurodegenerative features of PD include the loss of dopaminergic neurons in the substantia nigra pars compacta, the formation of α‐synuclein‐containing Lewy bodies and increased neuroinflammation.2+, which may initiate a range of toxic processes. In addition to its direct action at the NMDA receptor, QUIN can cause increased neuronal glutamate release, decreased glutamate uptake by astrocytes, and activation of microglia and neuroinflammation.ACMSD gene itself have been found in sporadic PD and a familial syndrome with tremor and parkinsonism, respectively. Recent studies have detected activation of the kynurenine pathway in the plasma of PD patients,Increased metabolism of tryptophan via the kynurenine pathway Fig.  is a conThus, in the present study we examined a large and clinically well‐characterized cohort to establish how/if the levels of kynurenine metabolites and inflammatory factors in plasma and CSF differ between PD subjects and age‐ and sex‐matched healthy controls. We hypothesized that PD subjects would exhibit increased activation of the kynurenine pathway with an accumulation of the neurotoxic metabolites QUIN and 3‐HK and that these changes would associate with the severity of clinical symptoms as well as with neuroimaging metrics reflecting PD‐related neuropathology in the substantia nigra.We obtained clinical and MRI data from 100 PD and 90 control subjects who participated in the National Institute of Neurological Disorders and Stroke PD Biomarker Program (NINDS PDBP) at Pennsylvania State University between 2012 and 2015. PD diagnosis was confirmed by movement disorders specialists.The NINDS PDBP used NINDS common data elements for collection of clinical data. We used the UPDRS scores (obtained for PD participants in the “on‐medication” state) to assess nonmotor and motor severity. Montreal Cognitive Assessment (MoCA)am. Fasting blood was obtained in BD Vacutainer Glass Blood Collection Tubes with K3‐ethylenediaminetetraacetic acid , centrifuged at 1700g for 10 minutes, followed by aspiration, ensuring that none of the buffy coat contaminated the plasma. CSF samples were available for a subset (30 controls and 25 PD patients). Lumbar punctures were performed by an experienced neurologist following sterile PDBP procedures that used a 24/22G Sprotte atramatic, introducer, and 22G hypodermic needles, along with a plastic syringe (6 mL) to draw the CSF. Within 15 minutes of collection, the samples were transferred to 15 mL of conical tubes and centrifuged at 2000g for 10 minutes at room temperature. For both plasma and CSF, 1‐mL aliquots were banked at Pennsylvania State University in a −80°C freezer with CO2 backup and 24/7/365 monitoring. For the current project, the samples were stored at −80°C for 4–7 years, then thawed once, when aliquoted for shipment to the Van Andel Institute on dry ice for analysis.Biological samples were drawn in the morning before 10 2H4‐KYN, 2H5‐KA, 2H4‐picolinic acid (PIC), 13C6‐nicotinamide (NTA), 2H3‐QUIN, 13C2.15N‐3HK, 2H3‐3HAA (5000 nM 2H3‐TRP) 1.2% formic acid in Milli‐Q water, vortexed and centrifuged through a COSTAR Spin‐X 0.22‐μm filter tube, and transferred to an amber vial (Agilent) containing a glass insert.Plasma samples were mixed with internal standard diluent and centrifuged. The supernatant was removed and dried under reduced pressure conditions by Savant Speedvac SPD131DDA . Samples were resuspended in 0.6% formic acid in Milli‐Q water, centrifuged through a COSTAR Spin‐X 0.22‐μm filter tube and transferred to an amber vial (Agilent) containing a glass insert. CSF samples were combined 1:2 with a solution containing 500 nM Kynurenine pathway metabolites , tryptophan, and serotonin see Fig.  were quaC‐reactive protein (CRP) and serum amyloid alpha (SAA) levels were analyzed on a Meso Scale Discovery Sector 6000 imager, according to the manufacturer's instructions . All samples were run in duplicate, and the average value of each sample was used for statistical analysis. Intra‐assay CVs for the analytes measured in plasma were: CRP, 5.2%; and SAA, 4.2%. Intra‐assay CVs for the analytes measured in CSF were: CRP, 4.2%; and SAA, 3.9%. Lower limit of quantification (LLOQ) for analytes measured in plasma were: CRP, 1.68 pg/mL; and SAA, 8.98 pg/mL. LLOQ for analytes measured in CSF were: CRP, 2.25 pg/mL; and SAA, 13.80 pg/mL.As part of the NINDS PDBP at Penn State, T1‐weighted, T2‐weighted, and multigradient‐echo magnetic resonance images were acquired on a 3T Siemens scanner for each subject. Detailed scan parameters are described in a previous study.t tests for continuous variables and chi‐square tests for categorical variables. Group‐wise comparisons between PD subjects and controls were performed using the Mann‐Whitney U test on the raw data, visualized in Figure Z scores were generated from linear regression models. As exploratory outcomes, we determined the relationship between plasma and CSF kynurenine metabolites and inflammatory markers, as well as between plasma and CSF biomarkers and disease duration, by Pearson's correlation using the age‐ and sex‐adjusted Z scores. To correct for multiple comparisons, significance was determined using a false discovery rate (FDR) at 10% as described by Benjamini‐Hochberg.P values are presented in Supplemental Tables Z scores are plotted in Figures Statistical analyses were performed using IBM's Statistical Package for the Social Sciences SPSS v.25). Comparisons of demographic variables were made using Student 5. ComparAll participants gave written informed consent in accordance with the Declaration of Helsinki. The study was approved by the Pennsylvania State University Hershey Institutional Review Board and the Van Andel Institute Review Board.All data generated or analyzed during this study are included in this published article.2), years of education (15 vs 15 years), or sex (49% women vs 53% women). As expected, PD subjects had significantly higher UPDRS I–III scores, lower MoCA and UPSIT scores, and higher R2* values compared with controls , an end product of the kynurenine pathway. There were no significant differences in the absolute plasma levels of TRP, KYN, KYNA, QUIN, or PIC between PD and control subjects  in patients (n = 97). The 3‐HK was not associated with symptoms within the controls. The 3‐HAA was not linked to any symptoms in patients or controls.We assessed the capacity of peripheral levels of 3‐HK and 3‐HAA to reflect symptom severity by establishing the association between the age‐ and sex‐adjusted variables with UPDRS I–III symptom severity. Plasma 3‐HK was significantly associated with UPDRS I . Higher scores on these scales indicate more severe symptoms.We next assessed the association between age‐ and sex‐adjusted CSF levels of kynurenines and symptoms. In PD patients (n = 25), higher CSF levels of QUIN were significantly associated with higher total UPDRS II (motor aspects of daily living [ADLs]) and UPDRS III (motor examination) scores  and QUIN/3‐HK  ratios were both strongly associated with increasing severity of ADL symptoms in the PD subjects . The QUIN/KYNA ratio .We have previously shown that elevated R2* in the substantia nigra pars compacta, reflecting iron content, is associated with PD progressionP = 0.001), and there was a trend toward lower SAA in both the CSF (P = 0.07) and plasma (P = 0.08) of PD patients , suggesting a potential connection between peripheral inflammation and central tryptophan metabolism in PD subjects. Higher plasma CRP and SAA were also linked with higher CSF QUIN  and with the QUIN/PIC ratio .Because inflammation is known to activate the kynurenine pathway, we examined the association between inflammatory markers in plasma with the kynurenine metabolites in the CSF of PD subjects (n = 25) and controls n = 30); see Supplemental Table 0; see SuR = 0.33, P = 0.001) as well as the CSF levels of the acute‐phase proteins SAA  and CRP  correlated with disease duration. Adjusting for sex only did not alter these results (not shown). The correlation with disease duration is presented in full in Supplemental Table We investigated the association between PD disease duration and the biomarkers, adjusted for sex and age. Plasma 3‐HK  and QUIN levels in the CSF. Higher levels of neurotoxic kynurenine metabolites in both plasma and CSF correlated with more severe symptoms. Together, these data suggest that kynurenine metabolites may serve as novel biomarkers and/or are involved in the PD pathological process.We observed higher plasma 3‐HK in PD subjects, in line with a recent study that found elevated 3‐HK in a small group of PD subjects with dyskinesias (n = 12).In PD CSF samples, we detected a reduction of the neuroprotective metabolite KYNA.We did not observe elevated levels of acute‐phase proteins in PD, but CSF CRP and SAA levels correlated with disease duration, even when correcting for age, suggesting that inflammation is associated with disease progression. In addition, the acute‐phase reactants correlated closely between plasma and CSF in both PD patients and controls. This is interesting because the communication of the acute‐phase reactants across the blood–brain barrier is still poorly understood. Intriguingly, only in PD patients did the acute‐phase proteins correlate with the CSF QUIN, QUIN/PIC, and KYN/TRP ratios, suggesting a disease‐specific kynurenine pathway activation. The mechanism underlying this is unclear, and we speculate that the kynurenine pathway is more sensitive to activation by inflammation in PD.Our study is the first to describe an association between CSF kynurenine metabolites and PD motor and nonmotor symptoms. We and others previously reported that subjects with depression and suicidality exhibit inflammation and increased QUIN/KYNA ratios.The strengths of our study include a clinically well‐characterized cohort, measures of kynurenine metabolites and inflammatory markers in both plasma and CSF, and imaging data from the substantia nigra. As a limitation, this cross‐sectional study was associative, and therefore causality could not be demonstrated. In addition, the PD subjects were on medication and were evaluated in the ON state, as defined per the PDBP study protocol. Therefore, we could not rule out that the observed effects were impacted by medication. Finally, because of the number of analyses performed, there is always a risk of reporting spurious results. Thus, replication in an independent cohort is necessary, as well as studies of kynurenine metabolites in medication‐free subjects with early PD or in the defined OFF state.In summary, we found evidence of an inflammation‐associated activation of the kynurenine pathway in both plasma and CSF in PD patients. We observed altered absolute levels of 3‐HK, 3‐HAA, and KYNA in PD, together with changes in several neurotoxic/neuroprotective ratios, indicating that tryptophan metabolism is disrupted in PD. In addition, we found that plasma and CSF levels of kynurenine metabolites were closely associated with worsening of symptoms in PD. Further replication studies are warranted, and these can also explore if kynurenine metabolites can serve as novel biomarkers and/or are involved in PD pathogenesis, which might lead to the identification of novel therapeutic targets.SUPPLEMENTAL TABLE 1 Subject demographics.MoCA, Montreal Cognitive Assessment; UPSIT, University of Pennsylvania Smell Identification Test; R2*, relaxation rates in the substantia nigra pars compacta; N/A; not available.1All values are reported as mean ± SD.aStudent t test.bChi‐square test.cANCOVA, with age and sex as covariates.Click here for additional data file.SUPPLEMENTAL TABLE 2 Median levels of metabolites and inflammatory markers. All values are reported as Median (IQR).aMann‐Whitney U test.Click here for additional data file.SUPPLEMENTAL TABLE 3 Correlations over the blood–brain barrier (BBB)P values after adjusting for sex and age.Pearson correlation coefficients and P values are listed in separate columns.FDR‐adjusted Click here for additional data file.SUPPLEMENTAL TABLE 4 Correlation of biological analytes with disease duration in years. Pearson correlation coefficients and P values after adjusting for sex and age. FDR‐adjusted P values are listed in separate columns.Click here for additional data file.