Abstract 5022: The selection of TP53 sub-clonal variants over time identifies MM patients with adverse clinical outcome
Authors: Martello Marina , Remondini Daniel , Borsi Enrica , Santacroce Barbara , Procacci Mauro , Dico Angela Flores , Pezzi Annalisa , Zamagni Elena , Tacchetti Paola , Pantani Lucia , Marzocchi Giulia , Zannetti Beatrice Anna , Mancuso Katia , Rocchi Serena , Martinelli Giovanni , Cavo Michele , Terragna Carolina 
Published: July 2016

Abstract
Introduction In most human cancer p53 impairment is a driver event, which confers a survival advantage to affected cells. In Multiple Myeloma (MM), the role of p53 clonal aberrations (mainly 17p del) is well recognised, whereas the prognostic relevance of TP53 mutations is less clear, due to the very limited frequency of clonal lesions. Here we aim at characterizing by ultra-deep sequencing (UDS) the TP53 mutational state in both newly diagnosed and relapsed MM pts, to assess the prognostic role and evolution over time of small TP53 mutated sub-clones.
Pts and methods A cohort of 99 newly diagnosed MM pts treated up-front with bortezomib-based regimens and ASCT was included in this study. In 29 cases, samples were collected also at relapse(s). DNA was obtained from CD138+ highly purified plasma cells and SNPs array profiled (Affymetrix). TP53 gene was analysed by amplicon-targeted UDS approach (GSJ, 454 Roche). A specific bioinformatics pipeline was set up to discriminate between low frequency TP53 variants and sequencing errors.
Results With a median coverage of 1386X, 129 correctly called TP53 variants were detected. Most newly diagnosed MM pts (55%) carried at least one TP53 sub-clonal variant (on average 1.08 variants per pts).. According to TP53 sub-clonal mutational load, pts were stratified in two sub-groups, including 28 pts with ≥2 (high load) and 71 with &lt;2 variants (low load), respectively. The clinical impact of the TP53 sub-clonal mutational load was evaluated in 90/99 MM (median follow up = 70 months). Pts carrying high TP53 sub-clonal mutational load had significantly shorter OS and OS after relapse (% at 5 years: 38 vs 74 and 30 vs 73, respectively), as compared to the others, while no difference between these two groups was seen regarding PFS and TTP. Multivariate analysis showed that high TP53 mutational load, resulted independent factors adversely affecting OS and OS after relapse (HR = 2.38, CI: 1.31-5.98; HR = 2.6, CI: 1.01-6.42). None of the detected genomic aberrations significantly influenced the response to front-line induction therapy. The distribution of both TP53 sub-clonal variants and genomic CNAs was overall modified in longitudinally collected samples: overall 90% of relapsed pts carried at least one sub-clonal variant (on average 1.63 variants per pts). Moreover, 5 different sub-clonal lesions proved a linear increment of both TP53 VAFs (from 29.4% to 54.6%; from 7.8% to 12.4%; from 0.5% to 4.3%) and TP53 CN loss smooth signal (from 7% to 89% and from 50% to 100%).
Conclusion The TP53 UDS analysis in newly diagnosed MM highlighted for the first time a high rate of variants, recurring with a wide range of frequencies among samples. The increased number of TP53 sub-clonal variants per pts in samples collected at relapse(s), compared to that seen at the onset of the disease, suggests a sub-clonal dynamics over time.
Acknowledgements: Roche Diagnostics, FP7 NGS-PTL project, Fondazione Berlucchi.
Citation Format: Marina Martello, Daniel Remondini, Enrica Borsi, Barbara Santacroce, Mauro Procacci, Angela Flores Dico, Annalisa Pezzi, Elena Zamagni, Paola Tacchetti, Lucia Pantani, Giulia Marzocchi, Beatrice Anna Zannetti, Katia Mancuso, Serena Rocchi, Giovanni Martinelli, Michele Cavo, Carolina Terragna. The selection of TP53 sub-clonal variants over time identifies MM patients with adverse clinical outcome. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5022.