Application of TruGraf v1: A Novel Molecular Biomarker for Managing Kidney Transplant Recipients With Stable Renal Function.

TruGraf v1 is a laboratory-developed DNA microarray-based gene expression blood test to enable proactive noninvasive serial assessment of kidney transplant recipients with stable renal function. It has been previously validated in patients identified as Transplant eXcellence (TX: stable serum creatinine, normal biopsy results, indicative of immune quiescence), and not-TX (renal dysfunction and/or rejection on biopsy results). TruGraf v1 is intended for use in subjects with stable renal function to measure the immune status as an alternative to invasive, expensive, and risky surveillance biopsies. MATERIALS AND METHODS: In this study, simultaneous blood tests and clinical assessments were performed in 192 patients from 7 transplant centers to evaluate TruGraf v1. The molecular testing laboratory was blinded to renal function and biopsy results. RESULTS: Overall, TruGraf v1 accuracy (concordance between TruGraf v1 result and clinical and/or histologic assessment) was 74% (142/192), and a result of TX was accurate in 116 of 125 (93%). The negative predictive value for TruGraf v1 was 90%, with a sensitivity 74% and specificity of 73%. Results did not significantly differ in patients with a biopsy-confirmed diagnosis vs those without a biopsy. CONCLUSIONS: TruGraf v1 can potentially support a clinical decision enabling unnecessary surveillance biopsies with high confidence, making it an invaluable addition to the transplant physician's tool kit for managing patients. TruGraf v1 testing can potentially avoid painful and risky invasive biopsies, reduce health care costs, and enable frequent assessment of patients with stable renal function to confirm the presence of immune quiescence in the peripheral blood.

Investigator Assessment of the Utility of the TruGraf Molecular Diagnostic Test in Clinical Practice.

BACKGROUND: TruGraf v1 is a well-validated DNA microarray-based test that analyzes blood gene expression profiles as an indicator of immune status in kidney transplant recipients with stable renal function. METHODS: In this study, investigators assessed clinical utility of the TruGraf test in patient management. In a retrospective study, simultaneous blood tests and clinical assessments were performed in 192 patients at 7 transplant centers, and in a prospective observational study they were performed in 45 subjects at 5 transplant centers. RESULTS: When queried regarding whether or not the TruGraf test result impacted their decision regarding patient management, in 168 of 192 (87.5%) cases the investigator responded affirmatively. The prospective study indicated that TruGraf results supported physicians' decisions on patient management 87% (39/45) of the time, and in 93% of cases physicians indicated that they would use serial TruGraf testing in future patient management. A total of 21 of 39 (54%) reported results confirmed their decision that no intervention was needed, and 17 of 39 (44%) reported that results specifically informed them that a decision not to perform a surveillance biopsy was correct. CONCLUSIONS: TruGraf is the first and only noninvasive test to be evaluated for clinical utility in determining rejection status of patients with stable renal function and shows promise of providing support for clinical decisions to avoid unnecessary surveillance biopsies with a high degree of confidence. TruGraf is an invaluable addition to the transplant physician's tool kit for managing patient health by avoiding painful and invasive biopsies, reducing health care costs, and enabling frequent assessment of patients with stable renal function to confirm immune quiescence.

Orthogonal Comparison of Molecular Signatures of Kidney Transplants With Subclinical and Clinical Acute Rejection: Equivalent Performance Is Agnostic to Both Technology and Platform.

We performed orthogonal technology comparisons of concurrent peripheral blood and biopsy tissue samples from 69 kidney transplant recipients who underwent comprehensive algorithm-driven clinical phenotyping. The sample cohort included patients with normal protocol biopsies and stable transplant (sTx) function (n = 25), subclinical acute rejection (subAR, n = 23), and clinical acute rejection (cAR, n = 21). Comparisons between microarray and RNA sequencing (RNA-seq) signatures were performed and demonstrated a strong correlation between the blood and tissue compartments for both technology platforms. A number of shared differentially expressed genes and pathways between subAR and cAR in both platforms strongly suggest that these two clinical phenotypes form a continuum of alloimmune activation. SubAR is associated with fewer or less expressed genes than cAR in blood, whereas in biopsy tissues, this clinical phenotype demonstrates a more robust molecular signature for both platforms. The discovery work done in this study confirms a clear ability to detect gene expression profiles for sTx, subAR, and cAR in both blood and biopsy tissue, yielding equivalent predictive performance that is agnostic to both technology and platform. Our data also provide strong biological insights into the molecular mechanisms underlying these signatures, underscoring their logistical potential as molecular diagnostics to improve clinical outcomes following kidney transplantation.

Mortality Risk Factors Among Patients With Cirrhosis and a Low Model for End-Stage Liver Disease Sodium Score (≤15): An Analysis of Liver Transplant Allocation Policy Using Aggregated Electronic Health Record Data.

Although the Model for End-Stage Liver Disease sodium (MELD Na) score is now used for liver transplant allocation in the United States, mortality prediction may be underestimated by the score. Using aggregated electronic health record data from 7834 adult patients with cirrhosis, we determined whether the cause of cirrhosis or cirrhosis complications was associated with an increased risk of death among patients with a MELD Na score ≤15 and whether patients with the greatest risk of death could benefit from liver transplantation (LT). Over median follow-up of 2.3 years, 3715 patients had a maximum MELD Na score ≤15. Overall, 3.4% were waitlisted for LT. Severe hypoalbuminemia, hepatorenal syndrome, and hepatic hydrothorax conferred the greatest risk of death independent of MELD Na score with 1-year predicted mortality >14%. Approximately 10% possessed these risk factors. Of these high-risk patients, only 4% were waitlisted for LT, despite no difference in nonliver comorbidities between waitlisted patients and those not listed. In addition, risk factors for death among waitlisted patients were the same as those for patients not waitlisted, although the effect of malnutrition was significantly greater for waitlisted patients (hazard ratio 8.65 [95% CI 2.57-29.11] vs. 1.47 [95% CI 1.08-1.98]). Using the MELD Na score for allocation may continue to limit access to LT.

Changes in Urinary Microbiome Populations Correlate in Kidney Transplants With Interstitial Fibrosis and Tubular Atrophy Documented in Early Surveillance Biopsies.

An unbalanced microbiome may lead to disease by creating aberrant immune responses. A recent association of cellular rejection with the development of interstitial fibrosis and tubular atrophy (IFTA) suggests the role of immune-mediated tissue injury. We hypothesized that developing IFTA correlates with altered urinary tract microbiomes (UMBs). UMBs at two serial time points, 1 and 6-8 months posttransplant, were assessed by 16S microbial ribosomal gene sequencing in 25 patients developing biopsy-proven IFTA compared to 23 transplant patients with normal biopsies and excellent function (TX) and 20 healthy nontransplant controls (HC). Streptococcus, the dominant genera in HC males, was lower in IFTA and TX males at 1 month compared to HCs. At 6-8 months, Streptococcus was further decreased in IFTA males, but normalized in TX. IFTA males and females had increases in number of genera per sample at 6-8 months. UMB composition varied substantially between individuals in all groups. Despite the wide variation in UMBs between individuals, IFTA was associated with a loss in dominant resident urinary microbes in males, and a parallel increase in nonresident, pathogenic bacteria in males and females. UMB changes may contribute to IFTA development by alteration of the host immune response.

Gene Expression in Biopsies of Acute Rejection and Interstitial Fibrosis/Tubular Atrophy Reveals Highly Shared Mechanisms That Correlate With Worse Long-Term Outcomes.

Interstitial fibrosis and tubular atrophy (IFTA) is found in approximately 25% of 1-year biopsies posttransplant. It is known that IFTA correlates with decreased graft survival when histological evidence of inflammation is present. Identifying the mechanistic etiology of IFTA is important to understanding why long-term graft survival has not changed as expected despite improved immunosuppression and dramatically reduced rates of clinical acute rejection (AR) (Services UDoHaH. http://www.ustransplant.org/annual_reports/current/509a_ki.htm). Gene expression profiles of 234 graft biopsy samples were obtained with matching clinical and outcome data. Eighty-one IFTA biopsies were divided into subphenotypes by degree of histological inflammation: IFTA with AR, IFTA with inflammation, and IFTA without inflammation. Samples with AR (n = 54) and normally functioning transplants (TX; n = 99) were used in comparisons. A novel analysis using gene coexpression networks revealed that all IFTA phenotypes were strongly enriched for dysregulated gene pathways and these were shared with the biopsy profiles of AR, including IFTA samples without histological evidence of inflammation. Thus, by molecular profiling we demonstrate that most IFTA samples have ongoing immune-mediated injury or chronic rejection that is more sensitively detected by gene expression profiling. These molecular biopsy profiles correlated with future graft loss in IFTA samples without inflammation.

Applying the WHO conceptual framework for the International Classification for Patient Safety to a surgical population.

OBJECTIVE: Efforts to improve patient safety are challenged by the lack of universally agreed upon terms. The International Classification for Patient Safety (ICPS) was developed by the World Health Organization for this purpose. This study aimed to test the applicability of the ICPS to a surgical population. DESIGN: A web-based safety debriefing was sent to clinicians involved in surgical care of abdominal organ transplant patients. A multidisciplinary team of patient safety experts, surgeons and researchers used the data to develop a system of classification based on the ICPS. Disagreements were reconciled via consensus, and a codebook was developed for future use by researchers. RESULTS: A total of 320 debriefing responses were used for the initial review and codebook development. In total, the 320 debriefing responses contained 227 patient safety incidents (range: 0-7 per debriefing) and 156 contributing factors/hazards (0-5 per response). The most common severity classification was 'reportable circumstance,' followed by 'near miss.' The most common incident types were 'resources/organizational management,' followed by 'medical device/equipment.' Several aspects of surgical care were encompassed by more than one classification, including operating room scheduling, delays in care, trainee-related incidents, interruptions and handoffs. CONCLUSIONS: This study demonstrates that a framework for patient safety can be applied to facilitate the organization and analysis of surgical safety data. Several unique aspects of surgical care require consideration, and by using a standardized framework for describing concepts, research findings can be compared and disseminated across surgical specialties. The codebook is intended for use as a framework for other specialties and institutions.

Nonchimeric HLA-Identical Renal Transplant Tolerance: Regulatory Immunophenotypic/Genomic Biomarkers.

We previously described early results of a nonchimeric operational tolerance protocol in human leukocyte antigen (HLA)-identical living donor renal transplants and now update these results. Recipients given alemtuzumab, tacrolimus/MPA with early sirolimus conversion were multiply infused with donor hematopoietic CD34(+) stem cells. Immunosuppression was withdrawn by 24 months. Twelve months later, operational tolerance was confirmed by rejection-free transplant biopsies. Five of the first eight enrollees were initially tolerant 1 year off immunosuppression. Biopsies of three others after total withdrawal showed Banff 1A acute cellular rejection without renal dysfunction. With longer follow-up including 5-year posttransplant biopsies, four of the five tolerant recipients remain without rejection while one developed Banff 1A without renal dysfunction. We now add seven new subjects (two operationally tolerant), and demonstrate time-dependent increases of circulating CD4(+) CD25(+++) CD127(-) FOXP3(+) Tregs versus losses of Tregs in nontolerant subjects (p < 0.001). Gene expression signatures, developed using global RNA expression profiling of sequential whole blood and protocol biopsy samples, were highly associative with operational tolerance as early as 1 year posttransplant. The blood signature was validated by an external Immune Tolerance Network data set. Our approach to nonchimeric operational HLA-identical tolerance reveals association with Treg immunophenotypes and serial gene expression profiles.

Unintended Consequences of the New National Kidney Allocation Policy in the United States.

The new national Kidney Allocation System of the Organ Procurement and Transplantation Network (OPTN), effective as of December 4, 2014, was designed to improve the chances of transplanting the most highly sensitized patients on the waitlist, those with calculated panel reactive antibody values of 98%, 99% and 100%. Recently, it was suggested that these highly sensitized patients will experience inequitable access, given the reported high prevalence of antibodies to HLA-DP, and the fact that only about 1/3 of deceased donors are typed for HLA-DP antigens. Here we report that 320/2948 flow cytometric crossmatches performed for the Northwestern transplant program over the past 28 months were positive solely due to HLA-DP donor-specific antibodies (11%; 16.5% of patients with HLA antibodies-sensitized patients). We further show that 58/207 (12%) HLA-DR serologically matched donor-recipient pairs had a positive B cell flow crossmatch due to donor-specific HLA class II antibodies, and 2/34 (6%) serologic zero-HLA-A-B-DR mismatch had a positive flow crossmatch due to HLA-DSA. We therefore provide information regarding the necessity and importance of complete donor HLA typing including both chains of the HLA-DP antigen (encoded by HLA-DPA1 and HLA-DPB1) at the time of organ offer.

Optimization and critical evaluation of decellularization strategies to develop renal extracellular matrix scaffolds as biological templates for organ engineering and transplantation.

The ability to generate patient-specific cells through induced pluripotent stem cell (iPSC) technology has encouraged development of three-dimensional extracellular matrix (ECM) scaffolds as bioactive substrates for cell differentiation with the long-range goal of bioengineering organs for transplantation. Perfusion decellularization uses the vasculature to remove resident cells, leaving an intact ECM template wherein new cells grow; however, a rigorous evaluative framework assessing ECM structural and biochemical quality is lacking. To address this, we developed histologic scoring systems to quantify fundamental characteristics of decellularized rodent kidneys: ECM structure (tubules, vessels, glomeruli) and cell removal. We also assessed growth factor retention--indicating matrix biofunctionality. These scoring systems evaluated three strategies developed to decellularize kidneys (1% Triton X-100, 1% Triton X-100/0.1% sodium dodecyl sulfate (SDS) and 0.02% Trypsin-0.05% EGTA/1% Triton X-100). Triton and Triton/SDS preserved renal microarchitecture and retained matrix-bound basic fibroblast growth factor and vascular endothelial growth factor. Trypsin caused structural deterioration and growth factor loss. Triton/SDS-decellularized scaffolds maintained 3 h of leak-free blood flow in a rodent transplantation model and supported repopulation with human iPSC-derived endothelial cells and tubular epithelial cells ex vivo. Taken together, we identify an optimal Triton/SDS-based decellularization strategy that produces a biomatrix that may ultimately serve as a rodent model for kidney bioengineering.

Molecular classifiers for acute kidney transplant rejection in peripheral blood by whole genome gene expression profiling.

There are no minimally invasive diagnostic metrics for acute kidney transplant rejection (AR), especially in the setting of the common confounding diagnosis, acute dysfunction with no rejection (ADNR). Thus, though kidney transplant biopsies remain the gold standard, they are invasive, have substantial risks, sampling error issues and significant costs and are not suitable for serial monitoring. Global gene expression profiles of 148 peripheral blood samples from transplant patients with excellent function and normal histology (TX; n = 46), AR (n = 63) and ADNR (n = 39), from two independent cohorts were analyzed with DNA microarrays. We applied a new normalization tool, frozen robust multi-array analysis, particularly suitable for clinical diagnostics, multiple prediction tools to discover, refine and validate robust molecular classifiers and we tested a novel one-by-one analysis strategy to model the real clinical application of this test. Multiple three-way classifier tools identified 200 highest value probesets with sensitivity, specificity, positive predictive value, negative predictive value and area under the curve for the validation cohort ranging from 82% to 100%, 76% to 95%, 76% to 95%, 79% to 100%, 84% to 100% and 0.817 to 0.968, respectively. We conclude that peripheral blood gene expression profiling can be used as a minimally invasive tool to accurately reveal TX, AR and ADNR in the setting of acute kidney transplant dysfunction.

Opportunities for shared decision making in kidney transplantation.

Health researchers and policy-makers increasingly urge both patient and clinician engagement in shared decision making (SDM) to promote patient-centered care. Although SDM has been examined in numerous clinical settings, it has received little attention in solid organ transplantation. This paper describes the application of SDM to the kidney transplantation context. Several distinctive features of kidney transplantation present challenges to SDM including fragmented patient-provider relationships, the time-sensitive and unpredictable nature of deceased organ offers, decision-making processes by transplant providers serving as both organ guardians (given the organ scarcity) versus advocates for specific patients seeking transplantation, variable clinical practices and policies among transplant centers, and patients' potentially compromised cognitive status and literacy levels. We describe potential barriers to and opportunities for SDM, and posit that SDM is feasible, warranting encouragement in kidney transplantation. We propose strategies to promote and overcome obstacles to SDM in kidney transplantation. We contend that engagement in SDM can be facilitated by re-organization of clinical care, communication and education of providers and patients.

Outcomes and native renal recovery following simultaneous liver-kidney transplantation.

With the increase in patients having impaired renal function at liver transplant due to MELD, accurate predictors of posttransplant native renal recovery are needed to select candidates for simultaneous liver-kidney transplantation (SLK). Current UNOS guidelines rely on specific clinical criteria for SLK allocation. To examine these guidelines and other variables predicting nonrecovery, we analyzed 155 SLK recipients, focusing on a subset (n = 78) that had post-SLK native GFR (nGFR) determined by radionuclide renal scans. The 77 patients not having renal scans received a higher number of extended criteria donor organs and had worse posttransplant survival. Of the 78 renal scan patients, 31 met and 47 did not meet pre-SLK UNOS criteria. The UNOS criteria were more predictive than our institutional criteria for all nGFR recovery thresholds (20-40 mL/min), although at the most conservative cut-off (nGFR ≤ 20) it had low sensitivity (55.3%), specificity (75%), PPV (67.6%) and NPV (63.8%) for predicting post-SLK nonrecovery. On multivariate analysis, the only predictor of native renal nonrecovery (nGFR ≤ 20) was abnormal pre-SLK renal imaging (OR 3.85, CI 1.22-12.5). Our data support the need to refine SLK selection utilizing more definitive biomarkers and predictors of native renal recovery than current clinical criteria.

Complications of living donor hepatic lobectomy--a comprehensive report.

A wider application of living donor liver transplantation is limited by donor morbidity concerns. An observational cohort of 760 living donors accepted for surgery and enrolled in the Adult-to-Adult Living Donor Liver Transplantation cohort study provides a comprehensive assessment of incidence, severity and natural history of living liver donation (LLD) complications. Donor morbidity (assessed by 29 specific complications), predictors, time from donation to complications and time from complication onset to resolution were measured outcomes over a 12-year period. Out of the 760 donor procedures, 20 were aborted and 740 were completed. Forty percent of donors had complications (557 complications among 296 donors), mostly Clavien grades 1 and 2. Most severe counted by complication category; grade 1 (minor, n = 232); grade 2 (possibly life-threatening, n = 269); grade 3 (residual disability, n = 5) and grade 4 (leading to death, n = 3). Hernias (7%) and psychological complications (3%) occurred >1 year postdonation. Complications risk increased with transfusion requirement, intraoperative hypotension and predonation serum bilirubin, but did not decline with the increased center experience with LLD. The probability of complication resolution within 1 year was overall 95%, but only 75% for hernias and 42% for psychological complications. This report comprehensively quantifies LLD complication risk and should inform decision making by potential donors and their caregivers.

Transplant center provision of education and culturally and linguistically competent care: a national study.

Although transplant centers are required to educate patients about kidney transplantation (KT) and living donation (LD), little is known about the educational format, and cultural and linguistic competence necessary for patients to make informed treatment decisions. This study surveyed US transplant administrators about education provided concerning KT and LD and culturally and linguistically competent care. Transplant administrators were invited to participate in an anonymous Internet-based survey about education format, education providers, promoting LD, culturally and linguistically competent care and center characteristics. Most (61%) transplant administrators contacted (N = 280/461) completed the survey. Most administrators (91%) reported that their center provides any type of formal education in their pre-KT evaluation. Education was mostly provided by: nurses (97%), social workers (72%) and surgeons (55%), and predominantly as one-on-one (80%) versus group discussions (60%). Education was primarily delivered through written materials (93%). Written educational materials in Spanish (86%) and the provision of interpreters (82%) were emphasized over educational sessions in Spanish (39%), or employing bilingual (51%) and bicultural staff (39%). Half (55%) promoted LD as the best option. Transplant centers need to take greater efforts to consistently provide appropriate education, promote LD, and provide culturally and linguistically competent care to ensure effective communication with all patients.

US Health Care Reform and Transplantation, Part II: impact on the public sector and novel health care delivery systems.

The Patient Protection and Affordable Care Act passed in 2010 will result in dramatic expansion of publically funded health insurance coverage for low-income individuals. It is estimated that of the 32 million newly insured, 16 million will obtain coverage through expansion of the Medicaid Program, and the remaining 16 million will purchase coverage through their employer or newly legislated insurance exchanges. While the Act contains numerous provisions to improve access to private insurance as discussed in Part I of this analysis, public sector coverage will significantly be affected. The cost of health care reform will be borne disproportionately by Medicare, which faces nearly $500 billion in cuts to be identified by a new independent board. Transplant centers should be concerned about the impact of the reform on the financial aspects of transplantation. In addition, this legislation also utilizes the Medicare Program to drive reform of the health care delivery system, by encouraging the development of integrated Accountable Care Organizations, experimentation with new 'models' of healthcare delivery, and expanded support for Comparative Effectiveness Research. Transplant providers, including transplant centers and physicians/surgeons need to lead this movement, drawing on our experience providing comprehensive multidisciplinary care under global budgets with publically reported outcomes.