Validation of Digital-PCR Analysis through Programmed imatinib Interruption in Q-RT-PCR Negative Chronic Myeloid Leukemia Patients
Authors: Mori Silvia , Vagge Elisabetta , le Coutre Philipp , Abruzzese Elisabetta , Martino Bruno , Pungolino Ester , Elena Chiara , Pierri Ivana , Assouline Sarit , D'Emilio Anna , Gozzini Antonella , Giraldo Pilar , Stagno Fabio , Nagler Arnon , Iurlo Alessandra , Rizzo Christian , Fossati Alessia , Pogliani Enrico Maria , Kim Dong-Wook , Pirola Alessandra , Piazza Rocco , Gambacorti-Passerini Carlo 
Published: October 2019

Abstract

Introduction
Imatinib induces complete cytogenetic response (CCyR) in up to 80% of chronic myeloid leukemia (CML) patients (pts) and major molecular response (MMR) in 33-60% of them. These patients enjoy life expectancy similar to general population. However even undetectable BCR-ABL may not equate to eradication of the disease because the sensitivity of the standard diagnostic method, the Q-RT-PCR, is limited. A new diagnostic method, the digital-PCR (dPCR), able to detect 1 BCR-ABL+ cell out of 107 cells, corresponding to a 100 times increased sensitivity as compared to conventional Q-RT-PCR, was developed (Goh HG et al., Leuk Lymphoma 52(5): 896-904. 2011). Therefore dPCR, assessing with more sensitivity the presence of minimal residual disease, could potentially identify pts in whom CML is eradicated. The Imatinib Suspension And Validation (ISAV) study is aimed at assessing the capability of dPCR to predict relapses after imatinib discontinuation in CML pts with negative Q-RT-PCR results.

Methods
This study involves 15 sites, 10 in Italy and 5 in each of the following countries: Germany, Spain, The Netherlands, Canada and Israel. In this study CML patients (Chronic Phase or Accelerated Phase) under imatinib therapy since more than 2 years and in complete molecular remission (CMR) were eligible. Patients had to be in CMR for at least 18 months, with a minimum of 3 Q-RT-PCR performed in their own centers. After signing the informed consent, pts were tested for dPCR and discontinued imatinib therapy. They are being monitored by standard Q-RT-PCR for 36 months to assess the maintenance of the molecular remission. At the end of this period, a peripheral blood sample for dPCR analysis will be obtained from those pts who will still have undetectable BCR-ABL transcripts by Q-RT-PCR, to verify CML eradication. The loss of molecular remission is defined as two consecutive positive Q-RT-PCR tests with at least one BCR-ABL/ABL value above 0.1%. Patients losing molecular remission resumed imatinib treatment at the same dosage used before interruption. Patients’ quality of life during imatinib discontinuation/resumption is being evaluated trough the EORTC – C30 Quality of Life questionnaire.

Results
The enrollment in the ISAV study began in November 2011 and ended in June 2013. The study enrolled 112 pts: Italy 69.6%, Berlin 21.4%, Montreal 5.3%, Zaragoza 2.6% and Tel Hashomer 0.9%. Sixty-one percent of the pts were male and 38% were aged 65 or older; median duration of imatinib treatment is 102 months with median duration of CMR of 32 months before imatinib discontinuation. To date, the median follow-up (FUP) time is 4.6 months [95% CI: 4.1-5.8] and 92 pts out of 112 (82%) had at least one Q-RT-PCR performed after imatinib discontinuation. The following analysis is restricted to 48 pts with a minimum of 6 months of FUP. Of these 48 pts, 20 remained Q-RT-PCR negative (42%, 95% CI:29-56%, median duration of negativity after imatinib discontinuation: 10.3 months). Nineteen pts (40%, 95% CI:27-53%) relapsed and resumed imatinib. All relapses occurred in the first 10 months and all but 3 of them in the first 6 months. A loss of CCyR happened in 5 pts out of 19 (26%): 1 pt regained CCyR after 3 months of re-treatment and is now in CMR, 1 pt died shortly after the diagnosis of relapse because of lung adenocarcinoma and 3 pts are now being monitored after imatinib resumption. No case of progression of CML was observed. After the resumption of imatinib the median time to either MMR or CMR, whichever came first, was 2.1 [95% CI: 0.9-5.8] months. Finally, nine pts (18%, 95% CI:10-31%) regained Q-RT-PCR positivity but never lost MMR. The median time to Q-RT-PCR positivity in this group of pts was 2.92 months (range 1-5 months), and the range of duration of Q-RT-PCR positivity (below 0.1%) is between 2 and 14 months. No significant correlation between relapse and previous duration of imatinib treatment, time to CCyR or duration of CMR was present. Patients previously treated with interferon showed a trend toward lower risk of relapse which is not significant so far. Finally, 19% of pts complained of musculoskeletal/articular pain after imatinib discontinuation.

Conclusions
After 21 months from the beginning of the study with a median follow-up of 4.6 months, 40% of pts relapsed; the majority of relapses happened in the first 6 months after imatinib discontinuation. The correlation of dPCR results with clinical outcomes will be presented at the meeting.

Disclosures:
le Coutre: Novartis: Honoraria; BMS: Honoraria; Pfizer: Honoraria; Novartis: Research Funding. Gozzini:Novartis: Consultancy; Bristol Myers Squibb: Consultancy.