Researchers share molecular insights from second phase of AMP

The Accelerating Medicines Partnership (AMP) is beginning to outline molecular features of rheumatoid arthritis that could have direct clinical impact.

AMP is a consortium linking academia, industry, and nonprofit groups, overseen by the National Institutes of Health, designed to accelerate basic research in RA and fast track findings into clinical development.

“Phase 1 showed the key cytokines that drive RA emerge from a few key cell states,” said Soumya Raychaudhuri, MD, PhD, Professor of Medicine and Biomedical Informatics at Harvard Medical School and the JS Coblyn and MB Brenner Chair of Rheumatology/Immunology and Director of the Center for Data Sciences at Brigham & Women’s Hospital. “Now we want to know what those cell states are and whether they are consistently expanded in all types of RA to develop new targets. Immune suppression doesn’t consistently lead to remission. We need novel therapeutics.”

Dr. Raychaudhuri and three other researchers provided the first overview of AMP Phase 2 during the Saturday session Molecular Insights from the Accelerating Medicines Partnership (AMP) 2021: RA. The session, which was originally presented Nov. 6, can be viewed by registered meeting participants through March 11, 2022.

Improved single-cell sequencing and imaging technologies have expanded the AMP sample base from roughly 21 samples and 5,000 cells to 80 samples and more than 300,000 cells. Initial insights from AMP Phase 2 include improved definitions of cell-type populations in RA, the identification of unique RA cell-type abundance phenotypes (CTAPs), and potential clinical implications of different cell-type populations. The AMP data set is public.

AMP Phase 2 focused on synovial tissue samples from 70 RA and nine osteoarthritis patients who were either DMARD-naïve (28), or had inadequate response to methotrexate (27) or inadequate response to TNF inhibitors (15). Seven major cell types were identified in synovial samples: NK, myoid, T, stromal, endothelial, plasma, and B cells. The RA synovial samples clustered into six CTAPs: endothelial + fibroblast + myeloid (E+F+M), fibroblast (F), T cells + fibroblasts (T+F), T + B cells (T+B), T cells + myeloid cells (T+M), and myeloid cells (M).

The CTAPs track with histology and serology, Dr. Raychaudhuri said, but are independent of disease activity. What does track with disease activity is the spatial associations between different CTAPs, or neighborhoods of phenotypically associated cell types.

“This may have therapeutic implications,” he said. “The IL-1β population is missing in the E+F+M CTAP, and anti-IL-1 therapy with agents such as anakinra may not be effective. The T+F CTAP is dominated by IFNγ+ CD16+ NK cells, so targeting IFNγ with JAK/STAT inhibitors may be appropriate.”

T, B cell populations

Lymphocytes in RA synovial tissue cluster in two CTAPs, T+F and T+B, said Deepak Rao, MD, PhD, Assistant Professor of Medicine and Co-Director of the Center for Cellular Profiling at Brigham and Women’s Hospital and Harvard Medical School. Researchers have identified two distinct T cell population clusters, CD4 and CD8 T cells.

The CD4 clusters are composed primarily of Tph and Tfh cell clusters and two distinct populations of Treg cells. There are also two distinct types of CD8 cells — a cluster of cytotoxic cells and a cluster of pro-inflammatory GZMK (granzyme K)-expressing cells.

B cells also cluster into two distinct populations. One cluster has plasmablasts and plasma cells. The second cluster has a variety of other B cell types, including memory, naïve, age-associated, GC-like, and others.

“We see a remarkable diversity of lymphocytes in the RA synovium,” Dr. Rao said. “We are trying to tease out some of the clinically relevant associations.”

Synovial myeloid cells

Myeloid cells in RA synovium segregate into four cell types and 15 distinct subsets, said Laura Donlin, PhD, Co-Director of the Derfner Foundation Precision Laboratory at the Hospital for Special Surgery and Assistant Professor of Immunology at Weill Cornell Medical College.

Five macrophage subsets are phagocytic and mostly non-inflamed. Four monocyte types fall into different inflammatory subsets, IL-1β and CXCL 10. Five classical dendritic cell (cDC) types are antigen-presenting and one plasmacytoid dendritic cell (pDC) type has IFNα. These myeloid cells are seen in three CTAPs — E+F+M, T+B, and M.

“Many of the cell types we thought were found universally in RA we now think may be specific to a subset of tissues,” Dr. Donlin said. “As an example, dendritic cells are mostly found only in CTAP tissues where there are large quantities of T cells.”

The M CTAP has only rare dendritic cells and an abundance of inflammatory-infiltrating monocytes that create high-density inflammation but few lymphocyte aggregations. The E+F+M CTAP is enriched for macrophages, but has few inflammatory monocytes or dendritic cells.

“Some clinical data suggest that these tissues may be less responsive to treatment, possibly because there is little inflammation going on,” Dr. Donlin said. “M CTAP tissues may be more responsive because there are more inflammatory monocytes present. Understanding the different crosstalk pathways between specific combinations of cell types may lead to better-tailored and novel treatments for RA.”

High-dimensional histology and clinical outcomes

CTAPs map to RA synovium histology by density, aggregates, and Krenn inflammation score, and immunofluorescence imaging is beginning to identify cell-type neighborhoods and interactions that are consistently associated, explained Andrew Filer, MD, PhD, FRCP, Senior Lecturer at the University of Birmingham, United Kingdom.

An interactome — a neighborhood analysis of consistent cell-type interactions — suggests that CTAPs do not map to RA disease activity, though they may map to treatment response. The M CTAP is enriched in inflammatory infiltrating monocytes and RA patients who are DMARD treatment-naïve respond very well. But that response is lost early and patients become unresponsive to methotrexate and TNF inhibitors.

“CTAPs may suggest stratification and therapy targets,” Dr. Filer said. “Spatial data coming in the next few months will tell us a lot more about the underlying mechanisms.”