Coronary calcium scoring as first-line test to detect and exclude coronary artery disease in patients presenting to the general practitioner with stable chest pain: protocol of the cluster-randomised CONCRETE trial.

INTRODUCTION: Identifying and excluding coronary artery disease (CAD) in patients with atypical angina pectoris (AP) and non-specific thoracic complaints is a challenge for general practitioners (GPs). A diagnostic and prognostic tool could help GPs in determining the likelihood of CAD and guide patient management. Studies in outpatient settings have shown that the CT-based coronary calcium score (CCS) has high accuracy for diagnosis and exclusion of CAD. However, the CT CCS test has not been tested in a primary care setting. In the COroNary Calcium scoring as fiRst-linE Test to dEtect and exclude coronary artery disease in GPs patients with stable chest pain (CONCRETE) study, the impact of direct access of GPs to CT CCS will be investigated. We hypothesise that this will allow for early diagnosis of CAD and treatment, more efficient referral to the cardiologist and a reduction of healthcare-related costs. METHODS AND ANALYSIS: CONCRETE is a pragmatic multicentre trial with a cluster randomised design, in which direct GP access to the CT CCS test is compared with standard of care. In both arms, at least 40 GP offices, and circa 800 patients with atypical AP and non-specific thoracic complaints will be included. To determine the increase in detection and treatment rate of CAD in GP offices, the CVRM registration rate is derived from the GPs electronic registration system. Individual patients' data regarding cardiovascular risk factors, expressed chest pain complaints, quality of life, downstream testing and CAD diagnosis will be collected through questionnaires and the electronic GP dossier. ETHICS AND DISSEMINATION: CONCRETE has been approved by the Medical Ethical Committee of the University Medical Center of Groningen. TRIAL REGISTRATION NUMBER: NTR 7475; Pre-results.

Outstanding negative prediction performance of solid pulmonary nodule volume AI for ultra-LDCT baseline lung cancer screening risk stratification.

OBJECTIVE: To evaluate performance of AI as a standalone reader in ultra-low-dose CT lung cancer baseline screening, and compare it to that of experienced radiologists. METHODS: 283 participants who underwent a baseline ultra-LDCT scan in Moscow Lung Cancer Screening, between February 2017-2018, and had at least one solid lung nodule, were included. Volumetric nodule measurements were performed by five experienced blinded radiologists, and independently assessed using an AI lung cancer screening prototype (AVIEW LCS, v1.0.34, Coreline Soft, Co. ltd, Seoul, Korea) to automatically detect, measure, and classify solid nodules. Discrepancies were stratified into two groups: positive-misclassification (PM); nodule classified by the reader as a NELSON-plus /EUPS-indeterminate/positive nodule, which at the reference consensus read was < 100 mm3, and negative-misclassification (NM); nodule classified as a NELSON-plus /EUPS-negative nodule, which at consensus read was ≥ 100 mm3. RESULTS: 1149 nodules with a solid-component were detected, of which 878 were classified as solid nodules. For the largest solid nodule per participant (n = 283); 61 [21.6 %; 53 PM, 8 NM] discrepancies were reported for AI as a standalone reader, compared to 43 [15.1 %; 22 PM, 21 NM], 36 [12.7 %; 25 PM, 11 NM], 29 [10.2 %; 25 PM, 4 NM], 28 [9.9 %; 6 PM, 22 NM], and 50 [17.7 %; 15 PM, 35 NM] discrepancies for readers 1, 2, 3, 4, and 5 respectively. CONCLUSION: Our results suggest that through the use of AI as an impartial reader in baseline lung cancer screening, negative-misclassification results could exceed that of four out of five experienced radiologists, and radiologists' workload could be drastically diminished by up to 86.7%.

Early detection of obstructive coronary artery disease in the asymptomatic high-risk population: objectives and study design of the EARLY-SYNERGY trial.

BACKGROUND: Coronary artery disease (CAD) burden for society is expected to steeply increase over the next decade. Improved feasibility and efficiency of preventive strategies is necessary to flatten the curve. Acute myocardial infarction (AMI) is the main determinant of CAD-related mortality and morbidity, and predominantly occurs in individuals with more advanced stages of CAD causing subclinical myocardial ischemia (obstructive CAD; OCAD). Unfortunately, OCAD can remain subclinical until its destructive presentation with AMI or sudden death. Current primary preventive strategies are not designed to differentiate between non-OCAD and OCAD and the opportunity is missed to treat individuals with OCAD more aggressively. METHODS: EARLY-SYNERGY is a multicenter, randomized-controlled clinical trial in individuals with coronary artery calcium (CAC) presence to study (1.) the yield of cardiac magnetic resonance stress myocardial perfusion imaging (CMR-MPI) for early OCAD diagnosis and (2) whether early OCAD diagnosis improves outcomes. Individuals with CAC score ≥300 objectified in 2 population-based trials (ROBINSCA; ImaLife) are recruited for study participation. Eligible candidates are randomized 1:1 to cardiac magnetic resonance stress myocardial perfusion imaging (CMR-MPI) or no additional functional imaging. In the CMR-MPI arm, feedback on imaging results is provided to primary care provider and participant in case of guideline-based actionable findings. Participants are followed-up for clinical events, healthcare utilization and quality of life. CONCLUSIONS: EARLY-SYNERGY is the first randomized-controlled clinical trial designed to test the hypothesis that subclinical OCAD is widely present in the general at-risk population and that early differentiation of OCAD from non-OCAD followed by guideline-recommended treatment improves outcomes.

The Diagnostic and Prognostic Value of Coronary Calcium Scoring in Stable Chest Pain Patients: A Narrative Review.

BACKGROUND: Non-contrast computed tomography (CT) scanning allows for reliable coronary calcium score (CCS) calculation at a low radiation dose and has been well established as marker to assess the future risk of coronary artery disease (CAD) events in asymptomatic individuals. However, the diagnostic and prognostic value in symptomatic patients remains a matter of debate. This narrative review focuses on the available evidence for CCS in patients with stable chest pain complaints. METHOD: PubMed, Embase, and Web of Science were searched for literature using search terms related to three overarching categories: CT, symptomatic chest pain patients, and coronary calcium. The search resulted in 42 articles fulfilling the inclusion and exclusion criteria: 27 articles (n = 38 137 patients) focused on diagnostic value and 23 articles (n = 44 683 patients) on prognostic value of CCS. Of these, 10 articles (n = 21 208 patients) focused on both the diagnostic and prognostic value of CCS. RESULTS: Between 22 and 10 037 patients were included in the studies on the diagnostic and prognostic value of CCS, including 43 % and 51 % patients with CCS 0. The most evidence is available for patients with a low and intermediate pre-test probability (PTP) of CAD. Overall, the prevalence of obstructive CAD (OCAD, defined as a luminal stenosis of ≥ 50 % in any of the coronary arteries) as determined with CT coronary angiography in CCS 0 patients, was 4.4 % (n = 703/16 074) with a range of 0-26 % in individual studies. The event rate for major adverse cardiac events (MACE) ranged from 0 % to 2.1 % during a follow-up of 1.6 to 6.8 years, resulting in a high negative predictive value for MACE between 98 % and 100 % in CCS 0 patients. At increasing CCS, the OCAD probability and MACE risk increased. OCAD was present in 58.3 % (n = 617/1058) of CCS > 400 patients with percentages ranging from 20 % to 94 % and MACE occurred in 16.7 % (n = 175/1048) of these patients with percentages ranging from 6.9 % to 50 %. CONCLUSION: Accumulating evidence shows that OCAD is unlikely and the MACE risk is very low in symptomatic patients with CCS 0, especially in those with low and intermediate PTPs. This suggests a role of CCS as a gatekeeper for additional diagnostic testing. Increasing CCS is related to an increasing probability of OCAD and risk of cardiac events. Additional research is needed to assess the value of CCS in women and patient management in a primary healthcare setting. KEY POINTS: · A CCS of zero makes OCAD in patients at low-intermediate PTP unlikely. · A CCS of zero is related to a very low risk of MACE. · Categories of increasing CCS are related to increasing rates of OCAD and MACE. · Future studies should focus on the diagnostic and prognostic value of CCS in symptomatic women and the role in primary care. CITATION FORMAT: · Koopman MY, Willemsen RT, van der Harst P et al. The Diagnostic and Prognostic Value of Coronary Calcium Scoring in Stable Chest Pain Patients: A Narrative Review. Fortschr Röntgenstr 2022; 194: 257 - 265.

Screening for cardiovascular disease risk using traditional risk factor assessment or coronary artery calcium scoring: the ROBINSCA trial.

AIMS: Screening for a high cardiovascular disease (CVD) risk followed by preventive treatment can potentially reduce coronary heart disease-related morbidity and mortality. ROBINSCA (Risk Or Benefit IN Screening for CArdiovascular disease) is a population-based randomized controlled screening trial that investigates the effectiveness of CVD screening in asymptomatic participants using the Systematic COronary Risk Evaluation (SCORE) model or coronary artery calcium (CAC) scoring. This study describes the distributions in risk and treatment in the ROBINSCA trial. METHODS AND RESULTS: Individuals at expected elevated CVD risk were randomized into screening arm A (n = 14 478; SCORE, 10-year fatal and non-fatal risk); or screening arm B (n = 14 450; CAC scoring). Preventive treatment was largely advised according to current Dutch guidelines. Risk and treatment differences between the screening arms were analysed. A total of 12 185 participants (84.2%) in arm A and 12 950 (89.6%) in arm B were screened. In total, 48.7% were women, and median age was 62 (interquartile range 10) years. SCORE screening identified 45.1% at low risk (SCORE < 10%), 26.5% at intermediate risk (SCORE 10-20%), and 28.4% at high risk (SCORE ≥ 20%). According to CAC screening, 76.0% were at low risk (Agatston < 100), 15.1% at high risk (Agatston 100-399), and 8.9% at very high risk (Agatston ≥ 400). CAC scoring significantly reduced the number of individuals indicated for preventive treatment compared to SCORE (relative reduction women: 37.2%; men: 28.8%). CONCLUSION: We showed that compared to risk stratification based on SCORE, CAC scoring classified significantly fewer men and women at increased risk, and less preventive treatment was indicated. TRIAL REGISTRATION NUMBER: NTR6471.

Negative Predictive Value of Non-Echo-Planar Diffusion Weighted MR Imaging for the Detection of Residual Cholesteatoma Done at 9 Months After Primary Surgery Is not High Enough to Omit Second Look Surgery.

OBJECTIVES: To evaluate non echo-planar diffusion weighted magnetic resonance imaging (non-EP DW MRI) at 9 months after primary surgery to rule out residual cholesteatoma in patients scheduled before second-look-surgical exploration. STUDY DESIGN: Prospective observational study. SETTING: Secondary teaching hospital. PATIENTS/INTERVENTIONS: Patients who were scheduled for second-look-surgery after primary canal wall up repair of cholesteatoma underwent 1.5 T MRI including non-EP DWI and high-resolution coronal T1 and T2-FS SE sequences. MAIN OUTCOME MEASURES: Imaging studies were evaluated for the presence of cholesteatoma by three independent observers. Intraoperative observations were regarded the standard of reference. Ear, nose, throat (ENT) surgeons were blinded for imaging findings. The primary outcome was the negative predictive value (NPV) of MR imaging, secondary outcomes were sensitivity, specificity, and positive predictive value. RESULTS: Thirty-three patients underwent both MRI and surgery, among whom 22 had a cholesteatoma. Mean time between primary surgery and MRI was 259 days (standard deviation [SD] 108). NPV of non-EP DW MRI in detecting recurrent cholesteatoma was 53% (95% CI: 32-73%). Sensitivity and specificity were 59% (39-77%) and 91% (62-98%), respectively. The positive predictive value was 93% (69-99%). In five out of nine false-negative cases, recurrent cholesteatoma measured 3 mm or less. Using a 3 mm detection threshold, NPV increased to 79%. CONCLUSION: Non-EP DW MRI cannot replace second look surgery in ruling-out residual cholesteatoma at 9 months after primary surgery. It could be used in a follow-up strategy in low risk patients. Further research is needed which types of residual cholesteatoma are not revealed by MRI.

High-pitch versus sequential mode for coronary calcium in individuals with a high heart rate: Potential for dose reduction.

BACKGROUND: To determine the impact of high-pitch spiral acquisition on radiation dose and cardiovascular disease (CVD) risk stratification by coronary artery calcium (CAC) assessment with computed tomography in individuals with a high heart rate. METHODS: Of the ROBINSCA trial, 1990 participants with regular rhythm and heart rates >65 beats per minute (bpm) were included. As reference, 390 participants with regular heart rates ≤65 bpm were used. All participants underwent prospectively electrocardiographically(ECG)-triggered imaging of the coronary arteries using dual source CT at 120 kVp, 80 ref mAs using both high-pitch spiral mode and sequential mode. Radiation dose, Agatston score, number of positive scores, as well as median absolute difference of the Agatston score were determined and participants were stratified into CVD risk categories. RESULTS: A similar percentage of participants with low heart rates and high heart rates had a positive CAC score in data sets acquired in high-pitch spiral (low heart rate: 57.7%, high heart rate: 55.8%) and sequential mode (58.0%, 54.7%, p = n.s.). The median absolute difference in Agatston scores between acquisition modes was 14.2% and 9.2%, for the high and low heart rate groups, respectively. Excellent agreement for risk categorization between the two data acquisition modes was found for the high (κ = 0.927) and low (κ = 0.946) heart rate groups. Radiation dose was 48% lower for high-pitch spiral versus sequential acquisitions. CONCLUSION: Radiation dose for the quantification of coronary calcium can be reduced by 48% when using the high-pitch spiral acquisition mode compared to the sequential mode in participants with a regular high heart rate. CVD risk stratification agreement between the two modes of data acquisition is excellent.

Coronary Artery Calcium Imaging in the ROBINSCA Trial: Rationale, Design, and Technical Background.

RATIONALE AND OBJECTIVES: To describe the rationale, design, and technical background of coronary artery calcium (CAC) imaging in the large-scale population-based cardiovascular disease screening trial (Risk Or Benefit IN Screening for CArdiovascular Diseases [ROBINSCA]). MATERIALS AND METHODS: First, literature search was performed to review the logistics, setup, and settings of previously performed CAC imaging studies, and current clinical CAC imaging protocols of participating centers in the ROBINSCA trial were evaluated. A second literature search was performed to evaluate the impact of computed tomography parameter settings on CAC score. RESULTS: Based on literature reviews and experts opinion an imaging protocol accompanied by data management protocol was created for ROBINSCA. The imaging protocol should consist of a fixed tube voltage, individually tailored tube current setting, mid-diastolic electrocardiography-triggering, fixed field-of-view, fixed reconstruction kernel, fixed slice thickness, overlapping reconstruction and without iterative reconstruction. The analysis of scans is performed with one type and version of CAC scoring software, by two dedicated and experienced researchers. The data management protocol describes the organization of data handling between the coordinating center, participating centers, and core analysis center. CONCLUSION: In this paper we describe the rationale and technical considerations to be taken in developing CAC imaging protocol, and we present a detailed protocol that can be implemented for CAC screening purposes.

CD4+ T-cell alloreactivity toward mismatched HLA class II alleles early after double umbilical cord blood transplantation.

Although double umbilical cord blood transplantation (dUCBT) in adult patients may be associated with less graft failure compared with single UCBT, hematopoietic recovery generally originates from a single cord blood unit (CBU). CBU predominance is still incompletely understood. We recently showed that blood CD4+ T-cell numbers rapidly increase after dUCBT, and early CD4+ T-cell chimerism predicts for graft predominance. Given the frequent HLA class II allele mismatches between CBUs in dUCBT, we hypothesized that alloreactive HLA class II-specific CD4+ T cells from the "winning" CBU may contribute to rejection of the "loser" CBU. We evaluated whether CD4+ T cells originating from the predominant (PD)-CBU would recognize HLA class II allele mismatches, expressed by the nonengrafting (NE)-CBU. Alloreactive effector CD4+ T cells toward 1 or more mismatched HLA class II alleles of the NE-CBU were detected in 11 of 11 patients, with reactivity toward 29 of 33 (88%) tested mismatches, and the strongest reactivity toward DR and DQ alleles early after dUCBT. Mismatched HLA class II allele-specific CD4+ T cells recognized primary leukemic cells when the mismatched HLA class II allele was shared between NE-CBU and patient. Our results suggest that cytotoxicity exerted by CD4+ T cells from the PD-CBU drives the rapid rejection of the NE-CBU, whose alloreactive effect might also contribute to graft-versus-leukemia.

Hip Resurfacing Implant Orientation Analysis: A Comparison of a Computer-Added Design Technique and Computed Tomography.

BACKGROUND: Accurate acetabular component orientation in hip resurfacing is mandatory. The aim of this study is to analyze if interpretation of pelvic radiographs with computer-added design (CAD) software is comparable to computed tomography (CT) in measurement of acetabular anteversion and inclination of a Birmingham Hip Resurfacing (BHR) hip. METHODS: A consecutive series of 49 patients (50 hips) who underwent hip resurfacing arthroplasty between 2005 and 2007 with the BHR system were retrospectively included. The surgical procedure was performed by 1 orthopedic surgeon in the beginning of his learning curve. Computer-added design software was used to measure acetabular component orientation on an anteroposterior pelvic radiograph. These measurements were compared with CT measurements. We calculated the correlation between the CAD software and CT analysis. The degree of underestimation or overestimation was determined, and a Bland-Altman plot was created to visualize the agreement between CAD software and CT results. RESULTS: We analyzed 50 BHR hips with mean inclination of 54.6° and 55.6° and mean anteversion of 24.8° and 13.3° measured by CT and CAD, respectively. Pearson correlation coefficient for inclination was 0.69 (P < .001) and for anteversion 0.81 (P < .001). Computer-added design showed a mean underestimated anteversion of 11.6° (P < .001). There was no significant underestimation or overestimation of inclination with CAD analysis compared to CT measurements. CONCLUSION: The CAD software is useful to assess acetabular inclination in hip resurfacing but underestimates anteversion.

[Referral for mammography often not according to guideline]. / Verwijzing voor mammogram vaak niet volgens richtlijn.

OBJECTIVE: To analyse the percentage of women with a family history of breast cancer referred by general practitioners (GPs) for a screening mammography in accordance with the Dutch Breast Cancer Guideline produced by the Netherlands Comprehensive Cancer Centre (IKNL). DESIGN: Prospective cohort study. METHOD: Women referred by their GP between December 2011 and December 2012 for mammography, with the indication "family history of breast cancer", were invited to take part in this study. A trained radiology laboratory assistant carried out a structured questionnaire to assess their risk on the basis of the categories of the 2008 IKNL guideline "Family history of breast/ovarian cancer". Based on the presence of certain risk factors, the women were allocated to one of the following groups: "referral for mammography", "referral to a clinical geneticist" or "no referral indicated". RESULTS: 242 women were referred by their GPs to the Radiology Department for mammography on the basis of family history; we included 210 women in our study. Their ages ranged from 25 to 77 years (mean age: 48 years). Forty-five patients (21%) were referred for mammography in accordance with the guideline. Twenty-two patients (10%) should have been referred to a clinical geneticist according to the guideline, whereas 143 patients (68%) did not meet the criteria for a screening mammography outside the screening programme. CONCLUSION: In only 21% of patients referred by their GPs for a screening mammography, with "family history" given as the reason, this referral was in accordance with the standard of the Dutch College of General Practitioners (NHG) or the IKNL guideline. Screening outside the breast cancer screening programme was not indicated according to the guideline for the majority of the women. Referral of 10% of the women referred should have been to a clinical geneticist; this figure rises to as many as 20% using the 2012 IKNL guideline.

Accuracy and interobserver agreement between MR-non-expert radiologists and MR-experts in reading MRI for suspected appendicitis.

OBJECTIVE: To compare accuracy and interobserver agreement between radiologists with limited experience in the evaluation of abdominal MRI (non-experts), and radiologists with longer MR reading experience (experts), in reading MRI in patients with suspected appendicitis. METHODS: MR imaging was performed in 223 adult patients with suspected appendicitis and read independently by two members of a team of eight MR-inexperienced radiologists, who were trained with 100 MR examinations previous to this study (non-expert reading). Expert reading was performed by two radiologists with a larger abdominal MR experience (>500 examinations) in consensus. A final diagnosis was assigned after three months based on all available information, except MRI findings. We estimated MRI sensitivity and specificity for appendicitis and for all urgent diagnoses separately. Interobserver agreement was evaluated using kappa statistics. RESULTS: Urgent diagnoses were assigned to 147 of 223 patients; 117 had appendicitis. Sensitivity for appendicitis was 0.89 by MR-non-expert radiologists and 0.97 in MR-expert reading (p=0.01). Specificity was 0.83 for MR-non-experts versus 0.93 for MR-expert reading (p=0.002). MR-experts and MR-non-experts agreed on appendicitis in 89% of cases (kappa 0.78). Accuracy in detecting urgent diagnoses was significantly lower in MR-non-experts compared to MR-expert reading: sensitivity 0.84 versus 0.95 (p<0.001) and specificity 0.71 versus 0.82 (p=0.03), respectively. Agreement on urgent diagnoses was 83% (kappa 0.63). CONCLUSION: MR-non-experts have sufficient sensitivity in reading MRI in patients with suspected appendicitis, with good agreement with MR-expert reading, but accuracy of MR-expert reading was higher.

Comparison of imaging strategies with conditional contrast-enhanced CT and unenhanced MR imaging in patients suspected of having appendicitis: a multicenter diagnostic performance study.

PURPOSE: To compare the diagnostic performance of imaging strategies with magnetic resonance (MR) imaging and computed tomographic (CT) imaging in adult patients suspected of having appendicitis. MATERIALS AND METHODS: Institutional review board approval was obtained prior to study initiation, and patients gave written informed consent. In a multicenter diagnostic performance study, adults suspected of having appendicitis were prospectively identified in the emergency department. Consenting patients underwent ultrasonography (US) and subsequent contrast-enhanced CT if US imaging yielded negative or inconclusive results. Additionally, all patients underwent unenhanced MR imaging, with the reader blinded to other findings. An expert panel assigned final diagnosis after 3 months. Diagnostic performance of three imaging strategies was evaluated: conditional CT after US, conditional MR imaging after US, and immediate MR imaging. Sensitivity and specificity were calculated by comparing findings with final diagnosis. RESULTS: Between March and September 2010, 229 US, 115 CT, and 223 MR examinations were performed in 230 patients (median age, 35 years; 40% men). Appendicitis was the final diagnosis in 118 cases. Conditional and immediate MR imaging had sensitivity and specificity comparable to that of conditional CT, which resulted in 3% (three of 118; 95% confidence interval [CI]: 1%, 7%) missed appendicitis, and 8% (10 of 125; 95% CI: 4%, 14%) false-positives. Conditional MR missed appendicitis in 2% (two of 118; 95% CI: 0%, 6%) and generated 10% (13 of 129; 95% CI: 6%, 16%) false-positives. Immediate MR missed 3% (four of 117; 95% CI: 1%, 8%) appendicitis with 6% (seven of 120; 95% CI: 3%, 12%) false-positives. Conditional strategies resulted in more false-positives in women than in men (conditional CT, 17% vs 0%; P = .03; conditional MR, 19% vs 1%; P = .04), wherease immediate MR imaging did not. CONCLUSION: The accuracy of conditional or immediate MR imaging was similar to that of conditional CT in patients suspected of having appendicitis, which implied that strategies with MR imaging may replace conditional CT for appendicitis detection.

Long-term stability of T-cell activation and transduction components critical to the processing of clinical batches of gene-engineered T cells.

BACKGROUND AIMS: The generation of gene-modified T cells for clinical adoptive T-cell therapy is challenged by the potential instability and concomitant high financial costs of critical T-cell activation and transduction components. As part of a clinical trial to treat patients with metastatic renal cell cancer with autologous T cells engineered with a chimeric antigen receptor (CAR) recognizing carboxy-anhydrase-IX (CAIX), we evaluated functional stability of the retroviral vector, T-cell activation agent Orthoclone OKT3 (Janssen-Cilag, Beerse, Belgium) monoclonal antibody (mAb) and the transduction promoting agent RetroNectin (Takara, Otsu, Japan). METHODS: Carboxy-anhydrase-IX chimeric antigen receptor retrovirus-containing culture supernatants (RTVsups) were generated from two packaging cell lines, Phoenix-Ampho (BioReliance, Sterling, UK) and PG13, and stored at -80°C over 10 years and 14 years. For Orthoclone OKT3 and RetroNectin, aliquots for single use were prepared and stored at -80°C. Transduction efficiencies of both batches of RTVsups were analyzed using the same lots of cryopreserved donor peripheral blood mononuclear cells, Orthoclone OKT3 and RetroNectin over time. RESULTS: We revisit here an earlier report on the long-term functional stability of the RTVsup, observed to be 9 years, and demonstrate that this stability is at least 14 years. Also, we now demonstrate that Orthoclone OKT3 and RetroNectin are functionally stable for periods of at least 6 years and 10 years. CONCLUSIONS: High-cost critical components for adoptive T-cell therapy can be preserved for ≥10 years when prepared in aliquots for single use and stored at -80°C. These findings may significantly facilitate, and decrease the financial risks of, clinical application of gene-modified T cells in multicenter studies.

Double umbilical cord blood transplantation: a study of early engraftment kinetics in leukocyte subsets using HLA-specific monoclonal antibodies.

Single cord blood unit (CBU) predominance is usually established within the first month after double umbilical cord blood transplantation (UCBT). However, the kinetics of engraftment of the different leukocyte subsets and the mechanism of graft predominance is largely unknown. To investigate whether a differential engraftment might reveal a specific subset that could play a key role in the mechanism of graft predominance, we studied early engraftment kinetics of different leukocyte subpopulations by flow cytometry using human monoclonal antigen-specific human leukocyte antigen antibodies, directed against mismatched human leukocyte antigen-A or -B antigens between recipient and CBUs. Twenty-two patients, who had received a double UCBT preceded by a reduced-intensity conditioning regimen, were evaluated at days +11, +18, +25, and +32 posttransplantation. Single CBU predominance in the various leukocyte subsets was established within 18 days posttransplantation. CD4+ T cells of the dominant CBU showed early peripheral blood expansion. Moreover, chimerism in CD4+ and CD8+ T cell and natural killer cell subsets at day +11 was predictive of ultimate graft predominance. These findings show that engraftment kinetics of the various leukocyte subsets vary considerably after double UCBT and may suggest an important role for CD4+ T cells in a presumed alloreactive graft-versus-graft rejection.

KRAS and BRAF mutation status in circulating colorectal tumor cells and their correlation with primary and metastatic tumor tissue.

Although anti-EGFR therapy has established efficacy in metastatic colorectal cancer, only 10-20% of unselected patients respond. This is partly due to KRAS and BRAF mutations, which are currently assessed in the primary tumor. To improve patient selection, assessing mutation status in circulating tumor cells (CTCs), which possibly better represent metastases than the primary tumor, could be advantageous. We investigated the feasibility of KRAS and BRAF mutation detection in colorectal CTCs by comparing three sensitive methods and compared mutation status in matching primary tumor, liver metastasis and CTCs. CTCs were isolated from blood drawn from 49 patients before liver resection using CellSearch™. DNA and RNA was isolated from primary tumors, metastases and CTCs. Mutations were assessed by co-amplification at lower denaturation temperature-PCR (Transgenomic™), real-time PCR (EntroGen™) and nested Allele-Specific Blocker (ASB-)PCR and confirmed by Sanger sequencing. In 43 of the 49 patients, tissue RNA and DNA was of sufficient quantity and quality. In these 43 patients, discordance between primary and metastatic tumor was 23% for KRAS and 7% for BRAF mutations. RNA and DNA from CTCs was available from 42 of the 43 patients, in which ASB-PCR was able to detect the most mutations. Inconclusive results in patients with low CTC counts limited the interpretation of discrepancies between tissue and CTCs. Determination of KRAS and BRAF mutations in CTCs is challenging but feasible. Of the tested methods, nested ASB-PCR, enabling detection of KRAS and BRAF mutations in patients with as little as two CTCs, seems to be superior.

Inflammation and fatigue dimensions in advanced cancer patients and cancer survivors: an explorative study.

BACKGROUND: Inflammation may underlie cancer-related fatigue; however, there are no studies that assess the relation between fatigue and cytokines in patients with advanced disease versus patients without disease activity. Furthermore, the relation between cytokines and the separate dimensions of fatigue is unknown. Here, association of plasma levels of inflammatory markers with physical fatigue and mental fatigue was explored in advanced cancer patients and cancer survivors. METHODS: A total of 45 advanced cancer patients and 47 cancer survivors completed the subscales Physical Fatigue and Mental Fatigue of the Multidimensional Fatigue Inventory. Plasma concentrations of C-reactive protein (CRP), interleukin-1 receptor antagonist (IL-1-ra), interleukin-6 (IL-6), interleukin-8 (IL-8), and neopterin were measured. Nonparametric tests were used to assess differences in fatigue intensity and levels of inflammatory markers and to determine correlation coefficients between the fatigue dimensions and inflammatory markers. RESULTS: Compared with cancer survivors, patients with advanced cancer had higher levels of physical fatigue (median 16 vs 9, P < .001) and mental fatigue (median 11 vs 6, P = .01). They also had higher levels of all cytokines (P < .01). In advanced cancer, CRP (r = 0.49, P = .001), IL-6 (r = 0.43, P = .003), IL-1-ra (r = 0.32, P = .03), and neopterin (r = 0.25, P = .10) were correlated with physical but not with mental fatigue. In cancer survivors, only IL-1-ra was related to both physical fatigue (r = 0.24, P = .10) and mental fatigue (r = 0.35, P = .02). CONCLUSIONS: In advanced cancer, inflammation seems to be associated with physical fatigue, but not to mental fatigue. In cancer survivors, there was no convincing evidence that inflammation plays a major role in fatigue.

Insufficient recovery of thymopoiesis predicts for opportunistic infections in allogeneic hematopoietic stem cell transplant recipients.

BACKGROUND: Recovery of thymopoiesis after allogeneic hematopoietic stem cell transplantation is considered pivotal for full immune competence. However, it is still unclear to what extent insufficient recovery of thymopoiesis predicts for subsequent opportunistic infections and non-relapse mortality. DESIGN AND METHODS: A detailed survey of all post-engraftment infectious complications, non-relapse mortality and overall survival during long-term follow-up was performed in 83 recipients of allogeneic stem cell grafts after myeloablative conditioning. Recovery of thymopoiesis was assessed using analysis of signal joint T-cell receptor rearrangement excision circles. The impact of recovery of thymopoiesis at 2, 6, 9 and 12 months post-transplantation on clinical outcome beyond those time points was evaluated by univariate and multivariate Cox regression analyses. RESULTS: The cumulative incidence of severe infections at 12 months after transplantation was 66% with a median number of 1.64 severe infectious episodes per patient. Patients in whom thymopoiesis did not recover were at significantly higher risk of severe infections according to multivariable analysis. Hazard ratios indicated 3- and 9-fold increases in severe infections at 6 and 12 months, respectively. Impaired recovery of thymopoiesis also translated into a higher risk of non-relapse mortality and outweighed pre-transplant risk factors including age, donor type, and disease risk-status. CONCLUSIONS: These results indicate that patients who fail to recover thymopoiesis after allogeneic hematopoietic stem cell transplantation are at very high risk of severe infections and adverse clinical outcome.