Gene Expression Profiling Scores Research Articles

Evidence for Radiation Therapy in Stage III Locoregionally Advanced Cutaneous Melanoma in the Post-Immunotherapy Era: A Literature Review.

In the landscape of Stage III locoregionally advanced cutaneous melanoma treatment, the post-immunotherapy era has sparked a number of questions on the management of the nodal basin. However, much of the available literature is not focused on radiation therapy as an adjuvant therapy. This literature review aims to illuminate the evidence surrounding radiation therapy's potential to mitigate regional recurrences in the adjuvant setting for melanoma. Additionally, it seeks to identify adjunct systemic therapy options and explore the synergy between systemic therapy and radiation. Despite strides in surgical techniques and systemic therapies, controlling regional Stage III melanoma remains a formidable clinical hurdle. While historical data strongly suggest the efficacy of adjuvant radiation therapy in reducing regional recurrence risk, its evaluation predates the advent of MAPK pathway inhibitors and robust immunotherapy options. Notably, clinical trials have yet to definitively demonstrate a survival advantage with adjuvant radiation therapy. Additional research should focus on refining the definition of high risk for regional recurrence through gene expression profiling or tumor immune profiling scores and elucidate the optimal role of adjuvant radiation therapy in patients treated with neoadjuvant systemic therapy.

Post-ASCT consolidation with EPD or ERD in patients with high-risk multiple myeloma.

7534 Background: Patients (pts) with high-risk (HR) Multiple Myeloma (MM), characterized by at least two cytogenetic abnormalities often experience rapid disease progression and early relapse post-autologous stem cell transplant (ASCT) with median Progression-Free Survival (PFS) of 24 or 12 months with or without lenalidomide maintenance (Panopoulou et al, Blood 2023). This study assesses the efficacy and safety of EPD (Elotuzumab, Pomalidomide, Dexamethasone) or ERD (Elotuzumab, lenalidomide , Dexamethasone) as consolidation therapy post-ASCT in HR MM pts. Methods: In this single-center retrospective study, we reviewed records of MM pts treated with consolidative ERD or EPD from September 2016 to March 2023. We collected baseline MM characteristics, treatment history, adverse events (AEs), and survival outcomes. HR MM pts were defined by 2 or more cytogenetic abnormalities (del 17p, 1q21 gain or amplification, t(14;16), t(14;20), complex karyotype) and/or high-risk gene expression profiling scores (GEP, SKY92). All patients, aged ≥21, underwent ≥1 full cycle of EPD or ERD post-ASCT, administered for a total of four 28-day cycles, following the standard dosing regimen with a rapid dexamethasone taper over the last two cycles. Results: A total of 78 HR MM pts, median age 64 years with 51.2% males and 71.7% caucasians were included. Of them, 62 pts received ERD and 16 received EPD. FISH revealed 62.7% of pts had 2 or more high risk cytogenetic abnormalities. Other pts expressed a combination of cytogenetic abnormalities and/or high-risk GEP or SKY92 scores. The table shows improvement in MRD negativity rates from 21.7% to 26.9% pre and post consolidation, and in VGPR or better from 78.2% to 85.8%. Median PFS for all pts, defined from the time of ASCT to disease progression, was 26.9 months. Most common adverse effects were anemia (71.7%), fatigue (55.1%) and thrombocytopenia (51.2%). Five patients experienced grade 3 adverse events, including 3 with infectious etiology (3.8%) and 2 with immune-mediated toxicity (2.5%). No treatment-related serious adverse events were reported. Conclusions: Four cycles of EPD or ERD consolidation therapy post-ASCT, demonstrated promising efficacy in HR MM pts. The observed median PFS of 26.9 months was comparable to that seen with chronic lenalidomide maintenance and superior to patients who do not receive lenalidomide maintenance. The improvement in PFS was achieved with a fixed duration rather than continuous therapy and without lenalidomide-associated toxicities such as secondary malignancies, chronic diarrhea, and financial toxicity. Further prospective confirmatory studies are needed. [Table: see text]

Prognostic Value of Ferritin in ASCT MM Patients: Integration with GEP Models and ISS Series Systems

INTRODUCTION Ferritin is associated with myeloma poor prognosis, but it was never included into current myeloma prognostic models. Here, we conducted an analysis of individual data from 3446 consecutive patients with multiple myeloma (MM) who received autologous stem cell transplantation (ASCT) therapy. The median follow-up time was 87.2 months (Q1: 44.1 months, Q3: 137.2 months). Using multivariate Cox regression analysis, we determined that serum ferritin was an independent risk factor for MM. We further investigated the prognostic effect of ferritin in the context of gene expression profiling (GEP) models and the different ISS series systems (including ISS, R-ISS, and R2-ISS). Our study showed that 39.5% of MM patients had high (above the normal range) serum ferritin levels. High serum ferritin was an independent poor prognostic factor regardless of age, gender, race, treatment modality, all GEP models and most of the ISS series system stages. ISS I&II patients with high ferritin should have a higher risk score, while R-ISS and R2-ISS III patients with normal ferritin should have a lower risk score than in their original scoring systems. Furthermore, ferritin's prognostic role was also explored in a dataset comprising 708 non-transplanted MM patients. METHODS We conducted an analysis of individual data from 3446 consecutive patients with multiple myeloma (MM) who received autologous stem cell transplantation (ASCT) therapy. Data analyzed included serum ferritin and other parameters such as gender, race, age, albumin (ALB), beta-2-microglobulin (B2M), lactate dehydrogenase (LDH), triple color interphase fluorescence in-situ hybridization (TriFISH), and gene expression profiling (GEP) scores. Serum ferritin was included in a multivariate Cox regression analysis and was identified as an independent prognostic factor. We further investigated the prognostic effect of ferritin on the different stages of the ISS (including ISS, R-ISS, and R2-ISS) and GEP models. Furthermore, we validated ferritin's prognostic role in a dataset comprising 708 non-transplanted MM patients. RESULTS Our study showed that 39.5% of MM patients had high serum ferritin levels. High serum ferritin (≥ 336mg/L for males and ≥ 306mg/L for females) had a hazard ratio (1.90, 95%CI: [1.75, 2.07] for overall survival and 1.74, 95%CI: [1.69, 1.81] for progression-free survival. The group with high ferritin levels had a median progression-free survival (PFS) of 40.5 months, whereas the normal ferritin group had a median PFS of 87.5 months. High serum ferritin was an independent poor prognostic factor regardless of age, gender, race, treatment modality, most of the ISS series system stages, and all GEP models. ISS I&II patients with high ferritin should have higher risk score, while R-ISS and R2-ISS III patients with normal ferritin should have lower risk score. And in a 708 non-transplant MM cohort, ferritin's prognostic role was also explored combine with demographic variables and ISS system. CONCLUSIONS Serum ferritin is a reliable prognostic factor for patients with MM and, therefore, should be routinely performed. Based on our findings, we recommend incorporation of ferritin into the ISS series models

The utilization of molecular microscope in management of heart transplant recipients in the era of noninvasive monitoring.

Monitoring for graft rejection is a fundamental tenet of post-transplant follow-up. In heart transplantation (HT) in particular, rejection has been traditionally assessed with endomyocardial biopsy (EMB). EMB has potential complications and noted limitations, including interobserver variability in interpretation. Additional tests, such as basic cardiac biomarkers, cardiac imaging, gene expression profiling (GEP) scores, donor-derived cell-free DNA (dd-cfDNA) and the novel molecular microscope diagnostic system (MMDx) have become critical tools in rejection surveillance beyond standard EMB. This paper describes an illustrative case followed by a review of MMDx within the context of other noninvasive screening modalities for rejection. We suggest MMDx be used to assist with early detection of rejection in cases of discordance between EMB and other noninvasive studies.

Immune response, phenotyping and molecular graft surveillance in kidney transplant recipients following severe acute respiratory syndrome coronavirus 2 vaccination.

Understanding immunogenicity and alloimmune risk following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination in kidney transplant recipients is imperative to understanding the correlates of protection and to inform clinical guidelines. We studied 50 kidney transplant recipients following SARS-CoV-2 vaccination and quantified their anti-spike protein antibody, donor-derived cell-free DNA (dd-cfDNA), gene expression profiling (GEP), and alloantibody formation. Participants were stratified using nucleocapsid testing as either SARS-CoV-2-naïve or experienced prior to vaccination. One of 34 (3%) SARS-CoV-2 naïve participants developed anti-spike protein antibodies. In contrast, the odds ratio for the association of a prior history of SARS-CoV-2 infection with vaccine response was 18.3 (95% confidence interval 3.2, 105.0, p < 0.01). Pre- and post-vaccination levels did not change for median dd-cfDNA (0.23%vs. 0.21% respectively, p = 0.13), GEP scores (9.85vs. 10.4 respectively, p = 0.45), calculated panel reactive antibody, de-novo donor specific antibody status, or estimated glomerular filtration rate. SARS-CoV-2 vaccines do not appear to trigger alloimmunity in kidney transplant recipients. The degree of vaccine immunogenicity was associated most strongly with a prior history of SARS-CoV-2 infection.

Advances and Controversies in Our Understanding of Guttate and Plaque Psoriasis.

Acute guttate psoriasis (AGP) is considered an uncommon variant of psoriasis (PsO), characterized as a widespread eruption of erythematous, psoriasiform papules, and plaques on the trunk, extremities, and scalp. Predisposing factors include a family history of PsO, variation in the main PsO susceptibility gene HLA-Cw*0602, and previous infection with viruses or acute β-hemolytic Streptococcus A program focused on controversies and recent advances in understanding AGP was presented at the Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) 2022 annual meeting. Topics included an overview of clinical presentation and natural history, predisposing genetic and environmental factors, and the recent molecular profiling that supports classification of AGP as a form of PsO. Early molecular profiling studies using proteomic signatures have suggested similarities between AGP and contact dermatitis, but recent studies using gene expression profiling and gene set enrichment scores demonstrate that AGP is more similar to chronic PsO. The expression of regulatory immune pathways seen with AGP suggests potential for early and sustained remission if the disease is suppressed by targeted treatments. Published case reports documenting clinical improvement of AGP with biologics that antagonize interleukin (IL)-12/23, IL-23, and IL-17 support the role of the IL-23/IL-17 axis in AGP, similar to that in PsO. Data supporting the use of antibiotics and other therapeutic agents for AGP are lacking, and randomized controlled trials are needed. Trial design for AGP is challenged by the low incidence, tendency for spontaneous remission, lack of validated end points, and the need for long-term follow up.

Real world validation of an adjunctive gene expression–profiling assay for melanoma diagnosis and correlation with clinical outcomes at an academic center

A commercially available diagnostic gene expression profiling (GEP) assay (MyPath™) reportedly has high sensitivity and specificity in distinguishing nevi from melanoma based on manufacturer-conducted studies. However, data regarding the performance of this GEP assay in routine clinical practice are lacking. The purpose of this study was to better assess the real-world performance of GEP in a large academic practice. Retrospective review of GEP scores were compared with final histomorphologic interpretation on a wide spectrum of melanocytic lesions demonstrating some degree of atypia. In a sample of 369 lesions, the sensitivity (76.1%) and specificity (83.9%) of the GEP test as compared with final dermatopathologist-rendered diagnosis in our dataset was appreciably lower than that reported in the prior manufacturer-conducted validation studies. Limitations of this study were that it was a single-center study, its retrospective nature, nonblinded nature of GEP test result, concordance of only two pathologists, and limited follow-up time.The sensitivity and specificity of a commercially available GEP diagnostic assay for melanoma may be lower in routine clinical practice, where melanocytic lesions typically exhibit some degree of histomorphologic atypia. Reported cost effectiveness of GEP testing is questionable if all ambiguous lesions that undergo such testing are re-excised in clinical practice.

Primary Graft Dysfunction Is Associated With Development of Early Cardiac Allograft Vasculopathy, but Not Other Immune-mediated Complications, After Heart Transplantation.

We investigated associations between primary graft dysfunction (PGD) and development of acute cellular rejection (ACR), de novo donor-specific antibodies (DSAs), and cardiac allograft vasculopathy (CAV) after heart transplantation (HT). A total of 381 consecutive adult HT patients from January 2015 to July 2020 at a single center were retrospectively analyzed. The primary outcome was incidence of treated ACR (International Society for Heart and Lung Transplantation grade 2R or 3R) and de novo DSA (mean fluorescence intensity >500) within 1 y post-HT. Secondary outcomes included median gene expression profiling score and donor-derived cell-free DNA level within 1 y and incidence of cardiac allograft vasculopathy (CAV) within 3 y post-HT. When adjusted for death as a competing risk, the estimated cumulative incidence of ACR (PGD 0.13 versus no PGD 0.21; P = 0.28), median gene expression profiling score (30 [interquartile range, 25-32] versus 30 [interquartile range, 25-33]; P = 0.34), and median donor-derived cell-free DNA levels was similar in patients with and without PGD. After adjusting for death as a competing risk, estimated cumulative incidence of de novo DSA within 1 y post-HT in patients with PGD was similar to those without PGD (0.29 versus 0.26; P = 0.10) with a similar DSA profile based on HLA loci. There was increased incidence of CAV in patients with PGD compared with patients without PGD (52.6% versus 24.8%; P = 0.01) within the first 3 y post-HT. During the first year after HT, patients with PGD had a similar incidence of ACR and development of de novo DSA, but a higher incidence of CAV when compared with patients without PGD.

(1031) Distribution of Gene Expression Profiling Scores in Pediatric Heart Transplant Recipients

(1031) Distribution of Gene Expression Profiling Scores in Pediatric Heart Transplant Recipients

230.5: Antibody Response and Molecular Graft Surveillance in Kidney Transplant Recipients Following SARS-CoV-2 Vaccination

Introduction: Preliminary studies suggest that kidney transplant recipients (KTRs) show diminished humoral responses to SARS-CoV-2 vaccination. Although reports of allograft rejection around the time of SARS-CoV-2 vaccination have been rare, there is no recommended framework for monitoring for potential vaccine-related allograft injury. Here, we describe an approach for longitudinal assessment of immunogenicity and safety of SARS-COV-2 vaccination in KTRs. Methods: KTRs eligible for SARS-CoV-2 vaccination were identified through medical records, beginning March 12, 2021. Baseline and weekly blood samples were collected for routine assessment, SARS-CoV-2 spike protein antibody titers, dd-cfDNA (AlloSure, CareDx) and gene expression profiling (GEP) (AlloMap, CareDx) for 12 weeks. HLA DSA testing was performed at baseline, 2 weeks after completion of vaccine doses and at week 12. Antibody response was defined as a 10-fold increase in total binding IgG titers. Results: 49 KTRs were identified for analysis. Patient demographics are summarized in Table 1. 10 patients (20.4%) demonstrated a spike antibody response following completion of the vaccine series. Patients with a prior history of COVID-19 were more likely to mount a spike antibody response (n=8, 53.3%) compared to those with no reported history of COVID-19 (n=2, 5.8%). The odds ratio was 18.3 (95% CI 3.2, 105.0, p=0.0005). Median dd-cfDNA levels did not differ between pre- and post-vaccination (0.23% versus 0.21% respectively; Table 2). There was no significant difference between pre- and post-vaccination GEP scores (9.85 versus 10.4 respectively; Table 2). No patients developed clinically significant DSA, eGFR decline or allograft rejection following vaccination. Conclusions: Quantitative antibody responses were strongly associated with a diagnosis of prior SARS-CoV-2 infection. Stability of eGFR, dd-cfDNA, GEP profiles and lack of allosensitization reinforce the safety profile of SARS-CoV-2 vaccination in KTRs. Further studies are needed to better understand immunogenicity in SARS-CoV-2 naïve individuals, including whether cellular responses are protective in the absence of humoral responses.

Is Absolute Change in AlloMap More Informative Than Absolute Value?

<h3>Purpose</h3> Increased variability in the non-invasive gene expression profiling (GEP) (AlloMap®) is associated with an increased risk of future events, but the association of the absolute change of below threshold GEP scores with future events has not yet been evaluated. <h3>Methods</h3> Patients from the Surveillance HeartCare Outcomes Registry (SHORE) with GEP score of < 34 had the change in GEP test calculated. The event of interest was a combination of rejection (ISHLT ≥ 2R and/or AMR ≥ 1), death, and graft dysfunction (LVEF <40% or LVEF change >=25% in a consecutive visit). Patients were divided into three groups: 1: Event - GEP within 14 days of event, 2: Prior - 3 or more GEP tests over 30 days prior to an event, 3: No Event - GEP testing in those with no event. Patients in Group 1 and 2 are paired. The N number in the x-axis label of Figure 1 are the patient number in each group. <h3>Results</h3> Of the 2029 patients in SHORE, there were 34 in the Event, 34 in the Prior to Event and 743 in the No Event groups. Age, race, and sex did not differ significantly between groups. The median GEP was highest in the Event group (34.0), compared to both Prior to Event (30.1) and No Event (30.7) groups, p<0.001. GEP scores 14 days prior to event are significantly elevated compared to GEP scores ≥30 days from event, p<0.001. The GEP scores ≥30 days are equivocal to the non-event control group, p = 0.25. Patients with GEP< 34 are significantly different at the time of event compared to scores prior to the event p=0.0046. (Figure 1) <h3>Conclusion</h3> GEP changes ahead of clinical events. Absolute GEP change may add incremental prognostic information for future events, beyond use of the absolute GEP value, especially below scores of 34. Further analysis is necessary to elucidate what absolute change in AM may be predictive of events.

Racial Disparities in Gene Expression Profiling but Not Donor-Derived Cell-Free DNA after Heart Transplant

<h3>Purpose</h3> Recipient race remains a powerful risk factor for mortality after heart transplantation (HT), with African Americans (AA) having worse survival than Caucasians. The aim of this analysis was to evaluate racial differences in gene expression profiling (GEP) and donor derived cell free DNA (dd-cfDNA) after HT using the Surveillance HeartCare Outcomes Registry (SHORE). <h3>Methods</h3> SHORE is an ongoing registry with >1200 patients enrolled at 60 HT centers in the United States. The registry includes HT recipients aged ≥15 years and ≥55 days post-transplant who had routine GEP and dd-cfDNA testing for rejection surveillance. Since patient sampling is from multiple centers, a linear mixed effects model, an extension of the linear model, was used to accommodate both fixed and random effects in the data. <h3>Results</h3> Among 956 recipients included in the analysis, 198 (20.7%) were AA, median age 57.0 years [0, 74.0] and 73.9% male. Overall, median GEP values demonstrated a borderline difference (p=0.07, Kolmogorov Smirnov test) for AA versus non-AA patients (Figure). This is evident by the cumulative distribution function, where a slightly higher rise in slope is observed for the non-AA cohort. Using linear mixed effects models, there was a statistically significant difference in GEP score when grouped by months post-transplant, demonstrating an early difference between AA and non-AA patients between months 2-5, but not subsequently. For dd-cfDNA analysis, median values were similar between AA (0.12%; Range 0.1-3.9%) and non-AA (0.12%; 0.1-3.5%) patients within the first year post-transplant (p=0.87, 2-sided K-S test) (Figure). <h3>Conclusion</h3> AA race is associated with higher GEP scores early post-transplant. This may be related to differential expression of the MARCH8 and FLT3 genes following HT, as previously described. Further analysis of specific GEP clusters and covariates across racial groups may shed light on these observed differences. In contrast, dd-cfDNA values do not appear to be influenced by race.

Remote Monitoring of Heart Transplant Recipients during the COVID-19 Pandemic

PurposeThe COVID-19 pandemic created significant challenges in monitoring heart transplant (HT) recipients for rejection due to efforts to minimize contact with the hospital setting. The aim of this study was to evaluate the safety and efficacy of transitioning HT patients to home phlebotomy and a monitoring protocol based on gene expression profiling (GEP) and donor derived cell free DNA (ddcfDNA).MethodsA single-center cohort study that prospectively enrolled consecutive HT patients who were transitioned to a remote monitoring protocol employing home phlebotomy and non-invasive surveillance for rejection. Patients were enrolled starting at 2 months post-HT. Positive GEP values were defined as ≥32 (up to 6 months post-HT) and ≥34 (> 6 months post-HT). A positive ddcfDNA score was defined as >0.12%. A positive biopsy was defined as grade ≥1B/1RResults246 HT patients were enrolled and followed for a minimum of 3 months. Mean age was 56±14, 71.5% were male, and median time from transplant was 2.7 years. The average distance of patients from the hospital was 25.6 miles. 359 blood tests were drawn for detection of GEP and ddcfDNA and 102 biopsies performed (Figure). Among 32 patients who had negative results on both tests and had a biopsy, 0 had a positive biopsy. Of 25 patients who had positive results on both tests and had a biopsy, 3 (12%) had a positive biopsy. The biopsy positivity rate in patients who were GEP+/ddcfDNA- was 6% and in patients who were GEP-/ddcfDNA+ was 8%. None of the positive biopsies were associated with hemodynamic compromise. 15 (6%) of patients were admitted due to allograft rejection during the study period. There were no deaths.ConclusionUsing a remote monitoring protocol with home phlebotomy and noninvasive rejection surveillance was feasible and safe in HT recipients. In this cohort, the combination of negative GEP and ddcfDNA scores was accurate at predicting a lack of allograft rejection.

Plasma Donor-Derived Cell-Free DNA Levels are Not Affected by Prednisone Dose nor Time after Heart Transplant: Pilot Data from DOAR and SHORE

<h3>Purpose</h3> Gene expression profiling (GEP; AlloMap®) scores can be impacted in the presence of >20 mg/day of prednisone due to effects on steroid-responsive genes (IL-1R2, ITGAM, FLT3) and is not validated for use prior to 55 days post-heart transplant. Donor-derived cell-free DNA (dd-cfDNA) can be used as early as 14 days after heart transplant (HT), but the impact of early use and the impact of steroid dosing has not yet been demonstrated. <h3>Methods</h3> The dd-cfDNA (AlloSure®) from the <i>Donor-Derived Cell-Free DNA Outcomes AlloMap Registry</i> (D-OAR) and <i>Surveillance Heart Care Outcome Registry</i> (SHORE) registries had 17 visits < 55 days and 106 visits 55 - 90 days post-HT. dd-cfDNA was also as analyzed by Kruskal-Wallis for prednisone dose ranges: 0-5, >5-10, >10-15, >15-20, and >20 mg/day (n=1113, 542, 203, 82, and 30). <h3>Results</h3> For those < 55 and >= 55 days post-HT the median dd-cfDNA result was 0.15% (IQR: 0.15-0.15%) and 0.15% (0.15-0.19%), respectively (p=NS). When stratified by prednisone doses (0-5], (5-10], (10-15], (15-20], and (>20, there was also no difference in median dd-cfDNA: 0.12%, 0.12%, 0.15%, 0.15%, and 0.15%, respectively, p=0.7. <h3>Conclusion</h3> This preliminary data suggests that dd-cfDNA levels are not significantly affected when assessed earlier than 55 days post-HT or by varying prednisone dosage. Implementation of dd-cfDNA as an early biomarker of allograft injury and rejection may represent a valuable clinical tool during the early post-HT period.

A Comprehensive Analysis of Baseline Clinical Characteristics and Biomarkers Associated with Outcome in Advanced Melanoma Patients Treated with Pembrolizumab

Simple SummaryPembrolizumab, a monoclonal antibody targeting programmed cell death 1, improves the survival of patients with advanced melanoma. This study aimed to investigate the association of baseline clinical characteristics, laboratory and imaging variables, and gene expression profiling scores on tumor tissue analysis of advanced melanoma patients who were treated with pembrolizumab, with survival using univariate and multivariate analysis. Baseline organ function (reflected by the presence of active brain metastases, number of metastatically affected organs, albumin) and systemic inflammatory/immunologic status (reflected by albumin, C-reactive protein, absolute lymphocyte count, neutrophil-to-lymphocyte ratio) are the most important clinical and/or laboratory parameters predictive of survival. Novel biomarkers include the baseline presence of BRAFV600 or NRASQ61/G12/G13 mutant circulating tumor DNA and baseline total metabolic tumor volume assessed by whole-body 18F-FDG-PET/CT. Gene expression profiling scores by the NanoString PanCancer IO360 panel were not conclusive in our patient population.Background: Pembrolizumab improves the survival of patients with advanced melanoma. A comprehensive analysis of baseline variables that predict the benefit of pembrolizumab monotherapy has not been conducted. Methods: Survival data of patients with advanced melanoma who were treated with pembrolizumab in a single university hospital were collected. A multivariate Cox regression analysis was performed to correlate baseline clinical, laboratory, and radiologic characteristics and NanoString IO360 gene expression profiling (GEP) with survival. Results: 183 patients were included (stage IV 85.2%, WHO performance status ≥1 31.1%; pembrolizumab first-line 25.7%), of whom 112 underwent baseline 18F-FDG-PET/CT imaging, 58 had circulating tumor DNA (ctDNA) assessments, and GEP was available in 27 patients. Active brain metastases, a higher number of metastatic sites, lower albumin and absolute lymphocyte count (ALC), higher C-reactive protein (CRP) and neutrophil-to-lymphocyte ratio, higher total metabolic tumor volume (TMTV), and higher ctDNA levels were associated with worse survival. Elevated lactate dehydrogenase (LDH) ≥ 2ULN (upper limit of normal), CRP ≥ 10ULN, or ALC < 750/mm3 delineate a subpopulation where treatment with pembrolizumab is futile. A TMTV ≥ 80 mL encompassed 17/21 patients with LDH ≥ 2ULN, CRP ≥ 10ULN, or ALC < 750/mm3. No significant associations were observed between baseline GEP scores and survival. Conclusion: Multiple baseline variables correlate with survival on pembrolizumab. TMTV is a more comprehensive baseline biomarker than CRP, LDH, or ALC in predicting the futility of pembrolizumab.

NOTCH3 is a Prognostic Factor and Is Correlated With Immune Tolerance in Gastric Cancer.

IntroductionAlthough traditional treatments confer survival benefits to patients with gastric cancer (GC), many patients experience relapse soon after postoperative adjuvant therapy. Immune-related mechanisms play an important role in GC, and immunotherapeutic strategies are considered to be a promising direction for the treatment of GC. Thus, our study aimed to investigate the expression and prognostic significance of immune-related genes in GC.MethodsFormalin-fixed, paraffin-embedded samples were collected from 48 resectable GC patients. The transcriptome data of the tumor immune microenvironment were assessed using an immuno-oncology 395-gene panel RNA sequencing platform. The prognostic value of the 395 genes was analyzed and validated in the KM plotter and GEPIA databases. The data from The Cancer Genome Atlas (TCGA, downloaded from UCSC Xena repository) and Tumor IMmune Estimation Resource (TIMER) were used to evaluate the correlations between prognostic factors and immune signatures.ResultsAmong the 395 genes, NOTCH3 was identified as a good prognostic factor for GC patients. Its prognostic value was also suggested in both our GC cohort from Zhongshan Hospital and the public databases (KM plotter and GEPIA database). Mechanistically, high NOTCH3 expression correlated with a lower infiltration of activated CD8+ T cells and a higher infiltration of immunosuppressive cells including Tregs and M2 macrophages in the tumor microenvironment. Moreover, high NOTCH3 expression was accompanied by the increased expression of a series of immune checkpoint inhibitors, resulting in a dampened immune response. Interestingly, NOTCH3 expression had a negative association with well-documented predictive biomarkers of immune checkpoint blockade (ICB) immunotherapy, including tumor mutation burden (TMB), gene expression profiling (GEP) score and innate anti-PD-1 resistance (IPRES) signature.ConclusionThese findings uncovered a new mechanism by which NOTCH3 participates in the immune tolerance of GC, implying the potential of NOTCH3 as a therapeutic target or predictive marker for GC patients.

Impact of cytomegalovirus infection on gene expression profile in heart transplant recipients

Cytomegalovirus (CMV) infection has been implicated in the pathogenesis of allograft rejection in heart transplant (HT) recipients. The effect of a CMV infection on the gene expression profiling (GEP, AlloMap) scores in the absence of acute rejection is not known. Data from 14,985 samples collected from 2,288 adult HT recipients enrolled in Outcomes AlloMap Registry were analyzed. Patients with known CMV serology at the time of HT who had at least 1 AlloMap score reported during follow-up were included. AlloMap scores for those patients with CMV (but no ongoing rejection) were compared with those who were never infected. An exploratory analysis on the impact of CMV on available donor-derived cell-free DNA (AlloSure) was also performed. A total of 218 patients (10%) were reported to have CMV infection after transplantation. AlloMap score in those samples with CMV infection (n = 311) had a GEP score (34; range: 29-36) significantly higher than the GEP score from samples (n = 14,674) obtained in the absence of CMV infection (30; range: 26-34; p < 0.0001). Both asymptomatic viremia and CMV disease demonstrated significantly higher AlloMap scores than no CMV infection samples (median scores: 33, 35, and 30, respectively; p < 0.0001). AlloSure levels, available for 776 samples, were not significantly different (median: 0.23% in 18 samples with CMV infection vs 0.15% in 776 samples without CMV infection; p = 0.66). CMV infection in HT recipients is associated with an increase in AlloMap score, whereas AlloSure results do not appear to be impacted. This information should be considered when clinically interpreting abnormal/high AlloMap scores in HT recipients.

Correlation of Longitudinal Gene-Expression Profiling (GEP) Score to Cytomegalovirus (CMV) Infection: Results from the Outcomes Allomap® Registry

Cytomegalovirus (CMV) infection has been implicated in the pathogenesis of allograft rejection, which is reflected by high gene-expression profiling (GEP) scores. The effect of CMV infection on serial GEP scores in the absence of acute cellular rejection (ACR) is unknown. Data from 14,985 samples collected from 2288 heart transplant (HT) recipients enrolled in Outcomes AlloMap® Registry (OAR) were analyzed. The empirical distribution of GEP scores at the visit closest to CMV infection were compared to GEP scores obtained at other visits via Kolmogorov-Smirnov tests. AlloSure measurements from those who underwent concurrent dd-cfDNA testing were also reviewed. 311 samples were collected from 213 patients with CMV infection at the time of GEP testing. The mean time to CMV infection from transplant was 346 days. Median GEP score at the time of diagnosis of CMV infection was significantly higher when compared to GEP score obtained at other visits (34 v 30, p<; 0.0001) (Figure 1). For patients with samples assessed via AlloMap and AlloSure, there was no significant difference in the distribution of AlloSure measurements at the time of CMV infection (Figure 2). CMV infections in HT recipients are associated with an increase in GEP score independent of associated ACR. The likely mechanism is immune activation/modulation of one or more of the 11 genes in the GEP signature. Hence, CMV status should be considered when clinically interpreting high GEP scores. AlloSure testing in conjunction with AlloMap demonstrates AlloSure is invariant to CMV infection.

Gene Expression Profiling Scores (Allomap) Can Successfully Predict the Absence of Serious Rejection in a Pediatric Heart Transplant Population; Single Center Experience

Allomap has been used successfully in adult cardiac transplantation as a screening tool to predict the absence of serious rejection. In adult studies, an Allomap score less than 34 has been highly correlated (Negative Predictive Value [NPV] of 98%) with biopsy grade 0-1R rejection. However, there are minimal studies using Allomap in pediatric patients. We sought to determine if using an Allomap score in pediatric patients ages 10-21 years old could also predict absence of serious rejection in our population. Heart transplant recipients >more than 3 months after transplant, who were between 10-21 years of age at the time of testing, who were taking a prednisone dose less than 15 mg/day and who had cardiac catheterization with biopsy performed had blood drawn for Allomap testing the same day. The blood was handled by a certified lab according to CareDx protocol and then sent to CareDx for testing. Allomap scores were then compared to cellular biopsy grading (serious rejection = 2R or greater). The data points were assessed for sensitivity and specificity using the Youden index and distance to (0,1), as was done in the CARGO studies. Eighty-one patients (mean age 14 +/- 2.9 years, 44% female) had 265 total Allomap tests and biopsy pairs evaluated between 2013 and 2019. Sixty-three (63) patients had more than one Allomap sample and biopsy pair. Race mix was Caucasian 44%, Hispanic 38%, African American 12%, Asian 2.5%, others 3.5%. Biopsy grade 0-1R represented 94% of the sample population. The NPVs for Allomap scores were 100% at 32 and 98.8% at 34, similar to what was found for adults in the CARGO studies. ROC curve demonstrates a c-statistic of 0.7566. Allomap scores in children and adolescents can be used to successfully predict absence of rejection with NPVs between 98%-100%; scores less than or equal to 32 correlated 100% with biopsy grades of 0-1R in this population. Given these results, we speculate that Allomap use will lead to a reduced need for cardiac catheterization and biopsy in this population but further studies in a larger pediatric transplant population is warranted.

Correlation of gene expression profiling score, cardiac hemodynamics and echocardiographic parameters in asymptomatic, rejection-free pediatric heart transplant recipients.

To correlate gene expression profiling scores obtained by AlloMap® with cardiac hemodynamics, cardiac allograft vasculopathy (CAV), and echocardiographic parameters in asymptomatic, rejection-free pediatric heart transplant (HT) recipients. Single-institution retrospective study of 210 AlloMap scores obtained concomitantly with cardiac catheterization and echocardiogram from 55 children during follow-up after cardiac transplantation. The median age at HT was 5.1years (range, 0.9-14.1), with 29 males and 26 females. AlloMap scores were high in <2years vs ≥2years of age at the time of HT (P=.001), and trending higher with time after HT (R2 =.04, P=.004). There was no significant difference in scores between ACR grades 0 and 1R or CAV. There was mild to modest correlation of AlloMap scores with the mean right atrial pressure (P=.002), and pulmonary capillary wedge pressure (P=.02), but no correlation was found with LV SF% (P=.3), LV EF% (P=.5), or RV FAC % (P=.8). Our study provides preliminary data that the AlloMap score must be studied carefully before it can be used in children, particularly in those under 2years of age. Monitoring of serial scores for each patient could potentially reflect changes in allograft performance that may determine indications for catheterization and biopsy which needs to be validated in future studies.