Follicular lymphoma (FL) is the second most common non-Hodgkin lymphoma in the Western world, generally characterized by a disseminated disease at diagnosis, an indolent clinical course, and recurrent, increasingly chemoresistant relapses. Overt FL is preceded by an insidious phase of asymptomatic growth and might emerge from common precursor clones, evolving over decades, and which might participate in subsequent relapses. Consequently, the cell of origin remains ambiguous. Although FL results from the malignant transformation of germinal center B cells, FL precursors emerge much earlier in B-cell ontogeny with the hallmark t(14;18) translocation and ensuing constitutive expression of BCL2 in bone marrow pre-B cells. While t(14;18) is considered the necessary early first hit to transformation, BCL2 as such is a very weak driver, as evidenced by the detection of t(14;18) in peripheral blood from a large fraction of the adult healthy population and the long latency and low penetrance observed in various BCL2 mouse models. Recent data of clonal dynamics in human and mouse models strongly suggest that additional illegitimate events likely accumulate over iterative GC passages, generating a highly mutagenic dynamics linked with slow oncogenic progression. Combined with the power of next-generation sequencing, molecular interrogations of t(14;18)+ clones in healthy individuals and FL patients recently provided the extent of FL genetic diversity in space (distinct tissues) and time (diagnosis vs. relapse). Thus, the path to transformation appears as a complex multi-hit process occurring along B-cell ontogeny, escalating along successive derailments of B-cell receptor diversification mechanisms, and subversion of specific immunologic properties of B cells. This protracted clonal evolution generates a massive accumulation of subclones and an associated complex overall genetic heterogeneity. This raises several questions. While FL originate from germinal center B cells, it is unclear to what extent overt lymphoma B cells retain GC B-cell functional dynamics or are blocked in a particular stage of the GC reaction. Furthermore, although one might intuitively assume that distinct subclones likely have a different gene expression profile associated to specific functions (and potentially driving distinct capacity to respond to therapy), it is still unclear to which extent subclonal genetic heterogeneity imprints functional heterogeneity. Because subclonal heterogeneity is hidden in bulk, where all distinct subclones are mixed and averaged, we recently used integrative single-cell analysis of phenotype, gene expression, and IGH sequence to track the characteristic human GC B-cell program in FL B cells. We used pseudo-time approaches to model the cyclic continuum of GC B-cell transitional states and identified characteristic patterns of synchronously expressed gene clusters in GC B cells. Strikingly this GC-specific gene expression synchrony was lost in single lymphoma B cells. Yet, distinct and conserved FL-specific cell states co-existed within single patient biopsies. Our data show that lymphoma B cells are not blocked in a GC B-cell state but may adopt new dynamic modes of functional diversity, opening novel definitions of lymphoma identity.

Citation Format: Bertrand Nadel. Follicular lymphoma dynamics through single-cell analysis [abstract]. In: Proceedings of the AACR Virtual Meeting: Advances in Malignant Lymphoma; 2020 Aug 17-19. Philadelphia (PA): AACR; Blood Cancer Discov 2020;1(3_Suppl):Abstract nr IA35.