Donor-Derived Cell-free DNA for Personalized Immunosuppression in Renal Transplantation.

BACKGROUND: The long-term outcomes of solid organ transplantation remain suboptimal. Therefore, appropriate biomarkers are needed in addition to immunosuppressive drugs and other traditional approaches for graft monitoring to achieve personalized immunosuppression and reduce premature graft loss. METHODS: Donor-derived cell-free DNA (dd-cfDNA) is a minimally invasive biomarker of cell death due to graft injury. It can be quantified using droplet digital polymerase chain reaction and next-generation sequencing. Fractional dd-cfDNA determination can be affected by changes in recipient cfDNA, such as those caused by leukopenia or infection, leading to false-positive or false-negative results, respectively. Absolute quantification of dd-cfDNA helps in overcoming this limitation. RESULTS: Overall, there is sufficient evidence of the clinical validity of dd-cfDNA. It detects rejection episodes early at an actionable stage and reflects the severity of graft injury without being rejection-specific. Owing to its high negative predictive value, dd-cfDNA is very useful for ruling out graft injury. Dd-cfDNA complements histological findings and can help in avoiding unnecessary biopsies. It indicates a response to rejection treatment and detects underimmunosuppression. CONCLUSIONS: Monitoring changes in dd-cfDNA over time may be helpful in adapting immunosuppression to prevent graft rejection. Moreover, serial dd-cfDNA determination may increase the effectiveness of transplant recipient surveillance and facilitate personalized immunosuppression when combined with other relevant clinical and diagnostic findings.

Time-Dependent Apparent Increase in dd-cfDNA Percentage in Clinically Stable Patients Between One and Five Years Following Kidney Transplantation.

BACKGROUND: Donor-derived cell-free DNA (dd-cfDNA) is reportedly a valuable tool for graft surveillance following kidney transplantation (KTx). Possible changes in dd-cfDNA(%) reference values over time have not been evaluated. For long-term monitoring after KTx, changes in host cfDNA might represent a biasing factor in dd-cfDNA(%) determinations. METHODS: Plasma samples were obtained (n = 929) 12-60 months after engraftment in a cross-sectional cohort of 303 clinically stable KTx recipients. Total cfDNA(copies/mL), dd-cfDNA(%), and dd-cfDNA(copies/mL) were determined using droplet-digital PCR. Stability of threshold values in these stable KTx recipients over time was assessed by 80th, 85th, and 90th quantile regression. RESULTS: Upper percentiles of total cfDNA showed a significant decline of -1902, -3589, and -4753 cp/mL/log(month) (P = 0.014, <0.001, and 0.017, respectively), resulting in increasing dd-cfDNA(%) percentiles by 0.25, 0.46, and 0.72%/log(month) (P = 0.04, 0.001, and 0.002, respectively), with doubling of the 85th percentile value by 5 years. In contrast, dd-cfDNA(cp/mL) was stable during the observation period (P = 0.52, 0.29, and 0.39). In parallel increasing white blood cell counts and decreasing tacrolimus concentrations over time were observed. After 5 years, the median total cfDNA was still 1.6-fold (P < 0.001) higher in KTx recipients than in healthy controls (n = 135) and 1.4-fold (P < 0.001) higher than patients with other medical conditions (n = 364). CONCLUSIONS: The time-dependent decrease of host cfDNA resulted in an apparent increase of dd-cfDNA fraction in stable KTx patients. For long-term surveillance, measurement of absolute dd-cfDNA concentrations appears to be superior to percentages to minimize false positive results.

Absolute quantification of donor-derived cell-free DNA as a marker of rejection and graft injury in kidney transplantation: Results from a prospective observational study.

Donor-derived cell-free DNA (dd-cfDNA) is a noninvasive biomarker for comprehensive monitoring of allograft injury and rejection in kidney transplantation (KTx). dd-cfDNA quantification of copies/mL plasma (dd-cfDNA[cp/mL]) was compared to dd-cfDNA fraction (dd-cfDNA[%]) at prespecified visits in 189 patients over 1 year post KTx. In patients (N = 15, n = 22 samples) with biopsy-proven rejection (BPR), median dd-cfDNA(cp/mL) was 3.3-fold and median dd-cfDNA(%) 2.0-fold higher (82 cp/mL; 0.57%, respectively) than medians in Stable Phase patients (N = 83, n = 408) without rejection (25 cp/mL; 0.29%). Results for acute tubular necrosis (ATN) were not significantly different from those with biopsy-proven rejection (BPR). dd-cfDNA identified unnecessary biopsies triggered by a rise in plasma creatinine. Receiver operating characteristic (ROC) analysis showed superior performance (P = .02) of measuring dd-cfDNA(cp/mL) (AUC = 0.83) compared to dd-cfDNA(%) (area under the curve [AUC] = 0.73). Diagnostic odds ratios were 7.31 for dd-cfDNA(cp/mL), and 6.02 for dd-cfDNA(%) at thresholds of 52 cp/mL and 0.43%, respectively. Plasma creatinine showed a low correlation (r = 0.37) with dd-cfDNA(cp/mL). In a patient subset (N = 24) there was a significantly higher rate of patients with elevated dd-cfDNA(cp/mL) with lower tacrolimus levels (<8 µg/L) compared to the group with higher tacrolimus concentrations (P = .0036) suggesting that dd-cfDNA may detect inadequate immunosuppression resulting in subclinical graft damage. Absolute dd-cfDNA(cp/mL) allowed for better discrimination than dd-cfDNA(%) of KTx patients with BPR and is useful to avoid unnecessary biopsies.

Circulating Cell-Free DNA-Diagnostic and Prognostic Applications in Personalized Cancer Therapy.

Genomic analyses in oncologic care allow for the development of more precise clinical laboratory tests that will be critical for personalized pharmacotherapy. Traditional biopsy-based approaches are limited by the availability of sequential tissue specimens to detect resistance. Blood-based genomic profiling ("liquid biopsy") is useful for longitudinal monitoring of tumor genomes and can complement biopsies. Tumor-associated mutations can be identified in cell-free tumor DNA (ctDNA) from patient blood samples and used for monitoring disease activity. The US Food and Drug Administration approved a liquid biopsy test for EGFR-activating mutations in patients with non-small-cell lung cancer as a companion diagnostic for therapy selection. ctDNA also allows for the identification of mutations selected by treatment such as EGFR T790M in non-small-cell lung cancer. ctDNA can also detect mutations such as KRAS G12V in colorectal cancer and BRAF V600E/V600K in melanoma. Chromosomal aberration pattern analysis by low-coverage whole genome sequencing is a new, broader approach. Genomic imbalances detected in cell-free DNA (cfDNA) can be used to compute a copy number instability (CNI) score. In clinical studies, it was demonstrated that the change in CNI score can serve as an early predictor of therapeutic response to chemotherapy/immunotherapy of many cancer types. In multivariable models, it could be shown that the CNI score was superior to clinical parameters for prediction of overall survival in patients with head and neck cancer. There is emerging evidence for the clinical validity of ctDNA testing regarding identification of candidates for targeted therapies, prediction of therapeutic response, early detection of recurrence, resistance mutation detection, measuring genetic heterogeneity, tumor burden monitoring, and risk stratification. Improvement of sensitivity to detect tumors at very early stages is difficult due to insufficient mutant DNA fraction of ≤0.01%. Further developments will include validation in prospective multicenter interventional outcome studies and the development of digital platforms to integrate diagnostic data.

From Research to the Bedside: Challenges for Pediatric Academic Researchers.

BACKGROUND: Although improving, development of drugs and devices for children is still less effective than for adults. Pediatric academicians play an important role in the bench-to-bedside research process, but much remains to be done to improve their contributions. OBJECTIVE: To provide a non-comprehensive review of selected literature based on my own personal experience as a U.S. based academic researcher who has spent over 4 decades doing pediatric drug and device development. METHODS: This commentary presents a summary of a talk given at a recent pediatric drug development conference. The observations and conclusions reached were based on the author's (largely US) experience and review of past history, the role of academicians in this process, some successful models of public-private collaboration, available funding, and barriers that remain to be overcome. RESULTS: Pediatric-specific legislation and more available funding have increased participation from and successes of US academicians in the pediatric drug and device development process. Incentive based public-private collaborations have been particularly successful. However, academicians still face both attitude and practical barriers to success. CONCLUSIONS: Changes are needed if academicians are to maximize their involvement in pediatric drug and device development.

Cell-Free Plasma DNA for Disease Stratification and Prognosis in Head and Neck Cancer.

BACKGROUND: Clinicians face many challenges in disease stratification and outcome prediction in head and neck squamous cancer cell (HNSCC) patients. Given the limitations of currently used clinical scoring, repetitive biopsies, and imaging techniques, liquid biopsy approaches may provide valuable additional diagnostic and prognostic information. METHODS: A noninterventional, single-center observational study was performed with clinical data and plasma samples from HNSCC patients. Cell-free tumor DNA-derived copy number aberrations (CNAs) were determined in 116 patients by low-coverage next-generation sequencing (NGS). Significant CNAs were combined in a genome-wide copy number instability score (CNI), which was evaluated with respect to conventional clinical staging and patient outcome. RESULTS: Receiver-operating characteristic (ROC) curve analysis comparing the presurgery CNI in patients (n = 103) with that in tumor-free controls (n = 142) yielded an area under the ROC curve of 87.2% (95% CI, 79.4%-93.3%). At a specificity of 95%, the sensitivity to detect tumors varied between 46% (pT1) and 94% (pT4). A CNI above the median (i.e., >72) had a positive predictive value of 90% (95% CI, 79%-96%) for lymph node involvement (LNI), while the negative predictive value was 57% (95% CI, 43%-70%). For a CNI >72, overall survival (OS) was worse (hazard ratio, 4.89; 95% CI, 1.39-17.17; P = 0.01) with 62% and 90% survivors 3 years after surgery for a CNI >72 and ≤72, respectively. In multivariable models, the CNI was a superior predictor of OS compared to established disease features, including LNI. CONCLUSIONS: The CNI may assist in predicting LNI and prognosis in HNSCC with direct therapeutic implications concerning the need for neck dissection or more aggressive treatment.

Using circulating cell-free DNA to monitor personalized cancer therapy.

High-quality genomic analysis is critical for personalized pharmacotherapy in patients with cancer. Tumor-specific genomic alterations can be identified in cell-free DNA (cfDNA) from patient blood samples and can complement biopsies for real-time molecular monitoring of treatment, detection of recurrence, and tracking resistance. cfDNA can be especially useful when tumor tissue is unavailable or insufficient for testing. For blood-based genomic profiling, next-generation sequencing (NGS) and droplet digital PCR (ddPCR) have been successfully applied. The US Food and Drug Administration (FDA) recently approved the first such "liquid biopsy" test for EGFR mutations in patients with non-small cell lung cancer (NSCLC). Such non-invasive methods allow for the identification of specific resistance mutations selected by treatment, such as EGFR T790M, in patients with NSCLC treated with gefitinib. Chromosomal aberration pattern analysis by low coverage whole genome sequencing is a more universal approach based on genomic instability. Gains and losses of chromosomal regions have been detected in plasma tumor-specific cfDNA as copy number aberrations and can be used to compute a genomic copy number instability (CNI) score of cfDNA. A specific CNI index obtained by massive parallel sequencing discriminated those patients with prostate cancer from both healthy controls and men with benign prostatic disease. Furthermore, androgen receptor gene aberrations in cfDNA were associated with therapeutic resistance in metastatic castration resistant prostate cancer. Change in CNI score has been shown to serve as an early predictor of response to standard chemotherapy for various other cancer types (e.g. NSCLC, colorectal cancer, pancreatic ductal adenocarcinomas). CNI scores have also been shown to predict therapeutic responses to immunotherapy. Serial genomic profiling can detect resistance mutations up to 16 weeks before radiographic progression. There is a potential for cost savings when ineffective use of expensive new anticancer drugs is avoided or halted. Challenges for routine implementation of liquid biopsy tests include the necessity of specialized personnel, instrumentation, and software, as well as further development of quality management (e.g. external quality control). Validation of blood-based tumor genomic profiling in additional multicenter outcome studies is necessary; however, cfDNA monitoring can provide clinically important actionable information for precision oncology approaches.

Graft-derived cell-free DNA, a noninvasive early rejection and graft damage marker in liver transplantation: A prospective, observational, multicenter cohort study.

BACKGROUND: Graft-derived cell-free DNA (GcfDNA), which is released into the blood stream by necrotic and apoptotic cells, is a promising noninvasive organ integrity biomarker. In liver transplantation (LTx), neither conventional liver function tests (LTFs) nor immunosuppressive drug monitoring are very effective for rejection monitoring. We therefore hypothesized that the quantitative measurement of donor-derived cell-free DNA (cfDNA) would have independent value for the assessment of graft integrity, including damage from acute rejection. METHODS AND FINDINGS: Traditional LFTs were performed and plasma GcfDNA was monitored in 115 adults post-LTx at three German transplant centers as part of a prospective, observational, multicenter cohort trial. GcfDNA percentage (graft cfDNA/total cfDNA) was measured using droplet digital PCR (ddPCR), based on a limited number of predefined single nucleotide polymorphisms, enabling same-day turn-around. The same method was used to quantify blood microchimerism. GcfDNA was increased >50% on day 1 post-LTx, presumably from ischemia/reperfusion damage, but rapidly declined in patients without graft injury within 7 to 10 d to a median <10%, where it remained for the 1-y observation period. Of 115 patients, 107 provided samples that met preestablished criteria. In 31 samples taken from 17 patients during biopsy-proven acute rejection episodes, the percentage of GcfDNA was elevated substantially (median 29.6%, 95% CI 23.6%-41.0%) compared with that in 282 samples from 88 patients during stable periods (median 3.3%, 95% CI 2.9%-3.7%; p < 0.001). Only slightly higher values (median 5.9%, 95% CI 4.4%-10.3%) were found in 68 samples from 17 hepatitis C virus (HCV)-positive, rejection-free patients. LFTs had low overall correlations (r = 0.28-0.62) with GcfDNA and showed greater overlap between patient subgroups, especially between acute rejection and HCV+ patients. Multivariable logistic regression modeling demonstrated that GcfDNA provided additional LFT-independent information on graft integrity. Diagnostic sensitivity and specificity were 90.3% (95% CI 74.2%-98.0%) and 92.9% (95% CI 89.3%-95.6%), respectively, for GcfDNA at a threshold value of 10%. The area under the receiver operator characteristic curve was higher for GcfDNA (97.1%, 95% CI 93.4%-100%) than for same-day conventional LFTs (AST: 95.7%; ALT: 95.2%; γ-GT: 94.5%; bilirubin: 82.6%). An evaluation of microchimerism revealed that the maximum donor DNA in circulating white blood cells was only 0.068%. GcfDNA percentage can be influenced by major changes in host cfDNA (e.g., due to leukopenia or leukocytosis). One limitation of our study is that exact time-matched GcfDNA and LFT samples were not available for all patient visits. CONCLUSIONS: In this study, determination of GcfDNA in plasma by ddPCR allowed for earlier and more sensitive discrimination of acute rejection in LTx patients as compared with conventional LFTs. Potential blood microchimerism was quantitatively low and had no significant influence on GcfDNA value. Further research, which should ideally include protocol biopsies, will be needed to establish the practical value of GcfDNA measurements in the management of LTx patients.

Graft-Derived Cell-Free DNA as a Marker of Transplant Graft Injury.

Although short-term success after solid organ transplantation is good, long-term graft and recipient survival are both not satisfactory. Despite therapeutic drug monitoring (TDM) of immunosuppressive drugs (ISDs), both excessive and insufficient immunosuppression still do occur. There is a need for new biomarkers that, when combined with TDM, can be used to provide more effective and less toxic, personalized immunosuppression to improve long-term survival. Currently used methods are insufficient to rapidly, cost-effectively, and directly interrogate graft integrity after solid organ transplantation. However, because organ transplants are also genome transplants, measurement of graft-derived circulating cell-free DNA (GcfDNA) has shown promise as a way to improve both graft and recipient outcomes after solid organ transplantation through the early detection of severe graft injury, enabling an early intervention. A newly developed droplet digital polymerase chain reaction (ddPCR) method has advantages over expensive high-throughput sequencing methods to rapidly quantify GcfDNA percentages and absolute amounts. This procedure does not require donor DNA and therefore can be applied to any organ donor/recipient pair. The droplet digital polymerase chain reaction method allows for the early, sensitive, specific, and cost-effective direct assessment of graft integrity and can be used to define individual responses to ISDs including the minimal ISD exposures necessary to prevent rejection. This is especially important in patients undergoing ISD switches due to ISD toxicity, infections, or malignancies. Although prospective, multicenter clinical trials in liver, heart, and kidney transplantation have not been completed, early results suggest that GcfDNA can be combined with TDM to guide changes in immunosuppression to provide more effective, and less toxic treatment. Personalized immunosuppression will shift emphasis in transplantation from reaction to prevention and could improve outcome at lower health care costs.

Mycophenolic acid reverses TGF beta-induced cell motility, collagen matrix contraction and cell morphology in vitro.

The aim of this study was to elucidate functional and molecular effects of mycophenolic acid (MPA) on non-lymphatic, kidney epithelial cells treated with transforming growth factor (TGF). MPA effects were studied using HK2 cells incubated with EGF and TGF. The reversibility of these effects was verified using guanosine and 8-aminoguanosine. The following assays were applied: cell proliferation, viability, collagen matrix contraction, scratch wound closure, spindle index, FACS with anti-CD29 and anti-CD326, promoter demethylation of RAS protein activator like 1 (RASAL1), as well as gene expression of RASAL1, integrin 1ß (ITGB1) (CD29) and epithelial cell adhesion molecule (EpCam) (CD326). Cell proliferation was inhibited by increasing concentrations of MPA, whereas neither apoptosis nor cytotoxicity was detected. Stimulation with EGF and/or TGF led to a significant collagen matrix contraction that was successfully inhibited by MPA. In addition, scratch wound closure was inhibited by incubation with TGF alone or with EGF. Under the same conditions, cell morphology (spindle shape) and molecular phenotype (ITGB1(High)EpCam(Low)/ITGB1(Low)EpCam(High)) were both significantly changed, suggesting an epithelial to mesenchymal transformation. Cell morphology and motility, as well as molecular phenotype, were reversible after MPA treatment with TGF transformation in both presence/absence of EGF, thereby suggesting a correlation with the previously described antifibrotic effects of MPA. Dysregulation of TGF signal transduction appears to be related to progression of fibrosis. A TGF-transformed kidney epithelial cell line derived from human proximal tubules was used to study whether the immunosuppressive drug: MPA possesses any functional or molecular antifibrotic effects. Functional and morphological in vitro changes induced by both the TGF and epithelial-growth-factor were reversible by treatment with MPA. An inhibitory effect of MPA on the TGF pathway appears to be responsible for the previously described antifibrotic effects of the MPA in the COL4A3-deficient mouse model of renal fibrosis.

Effects of mycophenolate mofetil on kidney function and phosphorylation status of renal proteins in Alport COL4A3-deficient mice.

BACKGROUND: We investigated the effects of mycophenolate mofetil (MMF) on kidney function and on protein phosphorylation in a mouse model for the human Alport syndrome. METHODS: COL4A3-deficient (COL4A3-/-) mice were randomly allocated to receive a placebo (PLC COL4A3-/-) or MMF treatment (MMF COL4A3-/-). Wild type mice (WT) were used as controls. Changes in serum creatinine, total protein and blood urea nitrogen (BUN), concentrations of mycophenolic acid (MPA) and its glucuronide metabolite (MPAG), serum protein electrophoresis, urine dipstick chemistry and sediment were measured. Changes in the phosphorylation status of renal proteins and histology were analyzed. RESULTS: MMF influenced kidney function and protein phosphorylation. Serum creatinine and BUN were lower in MMF treated compared to PLC treated COL4A3-/- mice. Serum albumin and alpha-1 globulins were significantly decreased while serum creatinine, alpha-2 globulins, urine dipstick protein, leukocyte esterase, hemoglobin and red blood cells were all increased in both COL4A3-/- groups compared to WT. Differential 2DE-gel analysis identified six phosphorylated kidney protein spots that were significantly altered by MMF. CONCLUSIONS: These data suggest that the MMF treatment in this murine model moderately improved kidney function and reversed the phosphorylation status of six renal phosphoprotein spots to that seen in WT mice.

Use of graft-derived cell-free DNA as an organ integrity biomarker to reexamine effective tacrolimus trough concentrations after liver transplantation.

BACKGROUND: Immunosuppressant therapeutic ranges for transplant patients have traditionally been established by indirect clinical means. However, "liquid biopsy" methods measuring graft-derived cell-free DNA (GcfDNA) in blood directly interrogate donor organ integrity. This study was performed to determine whether GcfDNA quantification could be used to reexamine minimally effective trough tacrolimus (Tacro) concentrations in liver transplantation (LTx) patients. METHODS: As part of a large prospective study to demonstrate the ability of GcfDNA to identify early graft rejection, 10 adult white LTx patients [8 men, 2 women, 3 hepatitis C virus (HCV) positive; mean ± SD age (years) = 56 ± 9.4] had simultaneous GcfDNA and whole-blood trough Tacro concentrations measured between days 5 and 30 after LTx. Samples were analyzed using droplet digital polymerase chain reaction for GcfDNA and liquid chromatography tandem mass spectrometry for Tacro. GcfDNA and trough Tacro concentrations were then compared to identify Tacro concentrations associated with intact graft integrity. RESULTS: Although there were large individual differences, there was a highly significant (Fisher P = 0.00002) segregation between whole-blood Tacro concentrations of ≥8 µg/L and normal (≤10%) GcfDNA percentages. The best discrimination in this population between effective and ineffective trough Tacro concentrations was estimated to be at 6.8 µg/L (P < 10(-7)). Compared with HCV- patients (n = 7), the 3 HCV+ patients had more variable associations between GcfDNA percentages and Tacro concentrations. CONCLUSIONS: Direct measurement of graft integrity using GcfDNA was useful to confirm the lower limit of the therapeutic ranges for trough Tacro concentrations after LTx. It would probably be useful to do so also for other immunosuppressant drugs and after other solid organ transplants. The method might be especially useful to detect graft injury during immunosuppressant dose minimization strategies.

Pharmacokinetics of meropenem in critically ill patients with severe infections.

BACKGROUND: Meropenem is an effective ß-lactam antibiotic that is frequently used to treat serious infections in both intensive care unit (ICU) and febrile neutropenic hematology/oncology (Hem/Onc) patients. Studies suggest that to be effective, meropenem concentrations must be maintained above the inhibitory concentrations for the majority of a dosing interval. However, the pharmacokinetics (PK) of meropenem seem to differ in critically ill patients compared with healthy or less ill subjects used to select labeled dosing regimens. OBJECTIVES: This study was designed to investigate meropenem PK in critically ill patients and to see how often standard dosing regimens produced adequate plasma concentrations. A secondary objective was to investigate how achieved concentrations were related to outcomes (morbidity and mortality) in these patients. METHODS: Meropenem plasma concentrations over time were measured using a high pressure liquid chromatography assay in febrile Hem/Onc and ICU patients who were treated with standard meropenem dosing schedules. Outcomes such as fever control and survival were assessed in these patients and compared with individual meropenem PK data and with recommended target concentrations. RESULTS: A total of 25 subjects including 10 febrile Hem/Onc and 15 ICU patients with a variety of serious illnesses and baseline renal function were studied. Mean peak concentrations were less variable than were pre-dose concentrations. Post peak and trough concentrations were often below recommended minimum inhibitory concentrations. Both clearance and volumes of distribution were greater than reported in less ill subjects, only in part explained by increased renal clearance. Therefore, serum concentrations often did not exceed recommended concentration targets even for moderately sensitive organisms. Inadequate concentrations were especially common in the mostly ill, febrile neutropenic Hem/Onc subjects and seemed to explain at least some therapeutic failures. Conversely, drug accumulation occurred in ICU subjects with decreased renal function. CONCLUSIONS: Standard meropenem dosing regimens were inadequate in many critically ill febrile, neutropenic Hem/Onc, and septic ICU patients. These data suggest a role for meropenem concentration monitoring in such patients.

Personalized Minimal Effective Concentration Therapy.

It has been recognized for literally centuries that patients should be given only the amount of medication necessary to treat disease(s) or relieve symptoms. It is also well known that this amount can vary greatly between patients or even over time in the same patient. The ability to identify this amount, that is, to "personalize" dosing, requires a reliable measure of a patient's response to treatment. The development of analytical methods for the accurate measurement of pharmacologically meaningful drug concentrations in physiologic fluids, combined with mathematical methods for reliable prediction of how dosing changes affect these concentrations, has led to the development of therapeutic drug management (TDM) for more effective individualization of dosing. Using TDM, clinicians modify dosing to achieve concentrations or exposures (ie, AUC) found to be effective in patients with similar clinical attributes and conditions. These concentrations, called therapeutic (or target) concentrations or exposure ranges (TRs), are specific to both disease/condition and patient population. TDM is routinely used by many clinicians to adjust dosing of a wide range of medications for maximal efficacy and limited toxicity, thereby improving clinical outcomes. Failure to properly perform TDM or to appreciate the limitations of TDM have, however, contributed to the delayed acceptance of TDM by clinicians. This Commentary briefly discusses the limitations and the benefits of TR-guided TDM, and then discusses immunosuppressant drugs and anticancer medications as examples of drugs that require clinicians to change their prescribing practices from giving all patients the same or maximal tolerated doses, to instead adjusting individual doses to achieve minimal effective concentrations identified using circulating tumor- or graft-derived DNA or copy number instability rather than published TRs.

Donor-derived cell-free DNA as a diagnostic tool in transplantation.

There is a need to improve personalized immunosuppression in organ transplantation to reduce premature graft loss. Biomarkers are needed to better detect rejection, asymptomatic graft injury, and under-immunosuppression. Assessment of minimal necessary exposure to guide tapering and prevent immune activation is also important. There is robust clinical evidence from a large number of published studies supporting the role of dd-cfDNA for monitoring graft integrity and detection or exclusion of rejection. Dd-cfDNA indicates graft cell death without being rejection specific. It can be determined in plasma through droplet digital PCR using preselected SNPs or next generation sequencing. Changes in recipient cfDNA (e.g., by infection) can affect the results of dd-cfDNA fractional determination. This limitation can be overcome using absolute dd-cfDNA quantification. The combination of fractional and absolute determination including total cfDNA is recommended for meaningful interpretation of the results. The value proposition for the patient includes earlier transplant injury detection and intervention, less full blown rejection risk, an alternative to invasive biopsies, and personalized immunosuppression with potential for improved long-term outcome. Transplant physicians benefit from better immunosuppressive guidance and having an alternative when biopsies are refused or contraindicated. Further advantages are improved biopsy interpretation, less trial and error changes in immunosuppression, and less time dealing with complications. The laboratory medicine specialist can provide more effective services. Hospital management and insurance companies could benefit from more cost-effective surveillance of transplant recipients. Potential cost savings would result from fewer biopsies as a result of the tests' high negative predictive value, fewer re-transplantations, and less organ failure with return to dialysis. A pathway to implementation and metrics is suggested to measure the effectiveness of dd-cfDNA testing.

Acetaminophen and ibuprofen in the treatment of pediatric fever: a narrative review.

OBJECTIVE: A narrative review of randomized, blinded, controlled studies assessing the antipyretic effect of ibuprofen versus acetaminophen or combined or alternating treatment in children was conducted. METHODS: Searches of the PubMed and Embase literature databases were conducted to identify relevant articles. Selected articles were limited to studies published in English that investigated OTC oral tablet and syrup formulations of acetaminophen and ibuprofen; there were no publication date limits. Open-label studies, nonrandomized studies, and those evaluating intravenous or suppository formulations of acetaminophen or ibuprofen were excluded. Variations in designs, endpoints, methods, and patient populations precluded our ability to conduct a formal systematic review. RESULTS: At physician-directed dosing (acetaminophen 15 mg/kg vs ibuprofen 10 mg/kg), no significant differences in antipyretic effects from 0‒6 h and between 0‒6, ‒12, ‒24, or ‒48 h, with single or multiple-doses, respectively, were observed. Tolerability profiles at physician dosing were similar. In 14 over-the-counter dose comparisons (acetaminophen, 10-15 mg/kg; ibuprofen, 2.5-10 mg/kg), antipyresis favored ibuprofen in 6, was similar between groups in 7, and favored acetaminophen (15 mg/kg vs ibuprofen 5 mg/kg) in 1 comparison. Both medications were well tolerated. Efficacy favored combination over individual components in 3 of 4 studies; alternating use results were mixed. All combination or alternating treatments were well tolerated. CONCLUSIONS: Antipyretic effects of ibuprofen and acetaminophen are similar at physician-directed doses; ibuprofen may be modestly superior at over-the-counter doses.