STAG2/LMO2 Gamma-Delta (γδ) T-ALL: Identification and Characterization of an Extremely High Risk Group of T-ALL in the Very Young
Authors: Kimura Shunsuke , Pölönen Petri , Montefiori Lindsey , Caldwell Kenneth , Iacobucci Ilaria , Chen Chelsey , Brown Anthony , Han Katie , Liu Yen-Chun , Chang Yunchao , Cheng Zhongshan , Yinmei Zhou , Park Chun Shik , Mitchell Sharnise , Reyes Noemi , Yeoh Allen , Attarbaschi Andishe , Moore Andrew , Manabe Atsushi , Buldini Barbara , Chiew Kean Hui , Li Chi Kong , Pui Ching-Hon , Qu Chunxu , Tomizawa Daisuke , Paietta Elisabeth , Locatelli Franco , Escherich Gabriele , Qingsong Gao , Muhle Hannah Elisa , Marquart Hanne Vibeke , de Groot-Kruseman Hester A. , Rowe Jacob M. , Stary Jan , Trka Jan , Choi John Kim , Meijerink Jules P.P. , Takita Junko , Pawinska-Wasikowska Katarzyna , Schmiegelow Kjeld , Eguchi Mariko , Schrappe Martin , Zimmermann Martin , Takagi Masatoshi , Maybury Melissa , Svaton Michael , Reiterova Michaela , Kicinski Michal , Kato Motohiro , Spinelli Orietta , Mazilier Pauline , Thomas Paul G. , Masetti Riccardo , Kotecha Rishi Sury , Pieters Rob , Elitzur Sarah , Luger Selina M , Shen Shuhong , Jeha Sima , Kornblau Steven M. , Skoczen Szymon , Miyamura Takako , Vincent Tiffaney , Imamura Toshihiko , Conter Valentino , Tang Yanjing , Gu Zhaohui , Roberts Kathryn G. , Teachey David T. , Crews Kristine R , Cheng Cheng , Yang Jun J. , Inaba Hiroto , Mullighan Charles G. 
Published: December 2023

Background
The prognosis of pediatric T-cell acute lymphoblastic leukemia (T-ALL) has improved with minimal residual disease (MRD)-stratified therapy, however, gamma delta T cell receptor positive (γδ) T-ALL remains a high-risk (HR) group. Limited studies have explored the clinical and genomic characteristics of γδ T-ALL, prompting us to conduct a comprehensive analysis of this entity and to identify determinants of outcome.
Methods
Through a consortium of 13 groups, we assembled a cohort of 200 patients up to 25 years of age with γδ T-ALL enrolled in clinical trials between 2000 and 2018. Clinical data of patients with non-γδ T-ALL enrolled on the same clinical trials were collected (n = 1,067). Complete remission (CR) was defined when bone marrow (BM) showed M1 cytomorphology and/or MRD &lt;1% without evidence of extramedullary disease at end of induction/consolidation (EOI/EOC) and failure to achieve CR was considered treatment failure. A total of 76 γδ T-ALL samples were analyzed by whole genome (WGS) and/or transcriptome (RNAseq) sequencing.
Results
The frequency of γδ T-ALL was 8.0% of T-ALL cases. Patients with γδ T-ALL exhibited a higher rate of poor prednisone response ( P&lt;0.01), MRD &gt;1% at day 15 ( P&lt;0.01), at EOI ( P&lt;0.01) and EOC ( P&lt;0.01), compared to non-γδ T-ALL cases. Furthermore, patients with γδ T-ALL showed significantly worse 5-year event free survival (EFS, 65% v. 78%, P&lt;0.01) and overall survival (OS, 77% vs 83%, P=0.048). Almost all relapses of γδ T-ALL were isolated BM, while the central nervous system was the main site of relapse in non-γδ T-ALL, suggesting slow treatment response and chemo-resistance to the current treatment in γδ T-ALL. However, γδ T-ALL showed a higher rate of toxic death during treatment (7.6% vs 4.0%, P&lt;0.01), suggesting the need for different therapeutic strategies and risk-classification, rather than treatment intensification.
Strikingly, patients less than 3 years of age with γδ T-ALL exhibited significantly poor EFS (33% v. 70% [3-10 years] and 73% [&gt;10], P&lt;0.01) and OS (49% v. 78% [3-10] and 82% [&gt;10], P&lt; 0.01) ( Fig. A), a difference not observed in non-γδ T-ALL. MRD &gt;1% at EOI showed poor EFS (51% v. 96% [MRD&lt;0.01%] and 91% [1%&gt;MRD&gt;0.01%], P&lt;0.01) and OS (66%).
Integrated analysis of WGS and RNAseq identified enrichment of several genomic subtypes in γδ T-ALL, including STAG2/LMO2, hyperdiploidy with recurrent gains of chromosomes 8, 10, 11, 13q and 19, a recently identified “LMO2 γδ-like” subtype with distinct gene expression and LMO2/MYC/MYCN alterations, TLX3-rearranged (-R), and PICALM::MLLT10. No TAL1 nor TLX1-R were detected. STAG2/LMO2 was associated with age at diagnosis before 3 years, and extremely poor outcome, with 4 out of 5 cases dying within three years of diagnosis ( Fig. B).
Of 24 STAG2/LMO2 T-ALL (additional 5 non-γδ, 13 TCR unknown cases), 22 of which were diagnosed by age three. All STAG2/LMO2 cases had alterations resulting in LMO2 activation and STAG2 inactivation, most commonly a single rearrangement between these two genes, and upregulation of HBE1, the LIN28-let7 pathway and stem cell proliferation pathways, suggesting a fetal hematopoietic origin.
STAG2 has a critical role in the maintenance of enhancer-promoter looping mediated by the cohesin complex. To examine the consequences of STAG2 alterations, we performed integrated genomic/epigenomic analysis of the STAG2/LMO2 (MOLT-14 and PER-117) and STAG2 knockout (KO)/addback T-ALL lines. Chromatin loop sizes defined by H3K27ac HiChIP was highest in STAG2/LMO2 lines compared to other T-ALL. Following restoration of STAG2 expression in MOLT-14, CD34 and ID1/2 were down-regulated and H3K27ac was enriched in pathways related to T-cell differentiation. STAG2 KO in the non- STAG2/LMO2, LMO2-activated line PF382 identified genes also upregulated in STAG2/LMO2 primary samples, including CDK4 and STAG1. STAG2 KO lines exhibited partial compensation of STAG2 binding sites by STAG1 and upregulation of γδ-related genes, RORC and ID1/3. High throughput screening of 2,050 small molecules identified efficacy of HDAC, CDK and PARP inhibitors in STAG2/LMO2 lines.
Conclusion
Very young onset γδ T-ALL, but not non-γδ T-ALL, is enriched for the STAG2/LMO2 subtype and is a very high risk form of T-ALL. STAG2 loss perturbs chromatin organization and hematopoietic differentiation. Moreover, we demonstrate efficacy of novel therapeutic approaches that are needed to cure this form of leukemia.