The Molecular Microscope Diagnostic System: Assessment of Rejection and Injury in Heart Transplant Biopsies

Abstract

This review describes the development of the Molecular Microscope Diagnostic System (MMDx) for heart transplant endomyocardial biopsies (EMBs). MMDx-Heart uses microarrays to measure biopsy-based gene expression and ensembles of machine learning algorithms to interpret the results and compare each new biopsy to a large reference set of earlier biopsies. MMDx assesses T cell–mediated rejection (TCMR), antibody-mediated rejection (AMR), recent parenchymal injury, and atrophy-fibrosis, continually “learning” from new biopsies. Rejection-associated transcripts mapped in kidney transplants and experimental systems were used to identify TCMR, AMR, and recent injury-induced inflammation. Rejection and injury emerged as gradients of intensity, rather than binary classes. AMR was one-third donor-specific antibody (DSA)-negative, and many EMBs first considered to have no rejection displayed minor AMR-like changes, with increased probability of DSA positivity and subtle inflammation. Rejection-associated transcript–based algorithms now classify EMBs as “Normal,” “Minor AMR changes,” “AMR,” “possible AMR,” “TCMR,” “possible TCMR,” and “recent injury.” Additionally, MMDx uses injury-associated transcript sets to assess the degree of parenchymal injury and atrophy-fibrosis in every biopsy and study the effect of rejection on the parenchyma. TCMR directly injures the parenchyma whereas AMR usually induces microcirculation stress but relatively little initial parenchymal damage, although slowly inducing parenchymal atrophy-fibrosis. Function (left ventricular ejection fraction) and short-term risk of failure are strongly determined by parenchymal injury. These discoveries can guide molecular diagnostic applications, either as a central MMDx system or adapted to other platforms. MMDx can also help calibrate noninvasive blood-based biomarkers to avoid unnecessary biopsies and monitor response to therapy.

The Molecular Phenotype of Kidney Transplants: Insights From the MMDx Project

“…58 The technology is detailed in a previous review. 12 The MMDx project approaches the disease states as latent variables-unknowable truths that can only ever be estimated by various approaches (Figure 2). We consider the phenotype labels assigned by clinical experience and histology as guides to the discovery of the underlying truth (the real disease).…”

“…Some aspects have previously been described, [7][8][9][10][11] and the technology and workflow are outlined in a recent MMDx-Heart review. 12 The assignment of the probability that a disease is present is typically a 2-step approach: step 1, measure designated features; and step 2, interpret feature measurements using predefined algorithms. In MMDx step 1, microarrays assess messenger RNA (mRNA) expression in the biopsy, currently measuring expression of 49 495 probe sets reflecting the expression of 19 462 unique genes.…”

Anything But a Biopsy: The Quest for Noninvasive Alternatives in Heart Transplantation

“…13 Other approaches have included application of newer technologies to the EMB specimen itself, including artificial intelligence techniques [14][15][16] and molecular analysis of specimens with iterative improvement in analysis (segmenting biopsy results into archetypes). 17,18 It remains difficult to know what to do with discrepant information where cell damage/injury is seen in the absence of clear rejection or allograft dysfunction. [18][19][20] Given the cost, complexity, and uncertainties that exist coupled with the push to optimize noninvasive surveillance pathways, there remains keen interest in novel biomarker approaches.…”

“…17,18 It remains difficult to know what to do with discrepant information where cell damage/injury is seen in the absence of clear rejection or allograft dysfunction. [18][19][20] Given the cost, complexity, and uncertainties that exist coupled with the push to optimize noninvasive surveillance pathways, there remains keen interest in novel biomarker approaches.…”

“…17,18 It remains difficult to know what to do with discrepant information where cell damage/injury is seen in the absence of clear rejection or allograft dysfunction. 18-20…”