Single cell analysis of clonal architecture in acute myeloid leukaemia

Nicola Potter,Peter Campbell,John Gribben,Mel Greaves,David Taussig,Farideh Miraki-Moud,Gowri Vijayaraghavan,Elli Papaemmanuil,Ian Titley,Luca Ermini

doi:10.1038/s41375-018-0319-2

Abstract

We used single cell Q-PCR on a micro-fluidic platform (Fluidigm) to analyse clonal, genetic architecture and phylogeny in acute myeloid leukaemia (AML) using selected mutations. Ten cases of NPM1c mutant AML were screened for 111 mutations that are recurrent in AML and cancer. Clonal architectures were relatively simple with one to six sub-clones and were branching in some, but not all, patients. NPM1 mutations were secondary or sub-clonal to other driver mutations (DNM3TA, TET2, WT1 and IDH2) in all cases. In three of the ten cases, single cell analysis of enriched CD34+/CD33− cells revealed a putative pre-leukaemic sub-clone, undetectable in the bulk CD33+ population that had one or more driver mutations but lacked NPM1c. Cells from all cases were transplanted into NSG mice and in most (8/10), more than one sub-clone (#2-5 sub-clones) transplanted. However, the dominant regenerating sub-clone in 9/10 cases was NPM1+ and this sub-clone was either dominant or minor in the diagnostic sample from which it was derived. This study provides further evidence, at the single cell level, for genetic variegation in sub-clones and stem cells in acute leukaemia and demonstrates both a preferential order of mutation accrual and parallel evolution of sub-clones.

Highlights

Almost all cancers originate in a single cell, the sequential acquisition of necessary additional mutations fuels sub-clonal diversity which is a substrate for positive or negative selection within the tissue ecosystems and with therapy [1, 2]
Individual cells sorted as CD34+/CD33− or CD33+/CD3− were assayed by multiplex Q-PCR for each driver mutation identified in that patient’s sample
Different driver mutations have epistatic or synergistic functional impacts in acute myeloid leukaemia (AML) [7, 22, 27] and the order of mutation accrual may impact on stem/progenitor cell function and clinical features [28]

Summary

Introduction

Almost all cancers originate in a single cell, the sequential acquisition of necessary additional mutations fuels sub-clonal diversity which is a substrate for positive or negative selection within the tissue ecosystems and with therapy [1, 2]. This process frequently results in complex cell population structures and highly variegated genetics [3,4,5]. The genomics of AML have been described in considerable detail, revealing multiple sub-types [6, 7] and sequential transition between clinically silent pre-leukaemia

Methods

Results

Conclusion