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The clinical benefit of pharmaceutical cares in improving the quality-of-care outcomes is well demonstrated. Clinical pharmacy services are not systematically deployed in cancer units in the absence of economic data. The aim of this prospective, observational 1-year study was to evaluate the clinical, economic and organisational impacts of pharmaceutical care into a multidisciplinary day hospital for patients treated with oral cancer drugs.All pharmacists' interventions (PI) were documented and their impact and the probability of adverse drug events were assessed using the clinical, economic and organisational tool.Among 360 admissions, an average of 1.81 PI per admission was accepted. Among 452 PI leading to a clinical benefit on the patient, 16.9% had a major impact, and 1.9% had an impact on survival. The large majority of PIs (87%) increased the quality-of-care organisation. The budget impact model showed a total cost savings and cost avoidance of €539,047 per year and a cost-benefit ratio of 7.07:1. The direct cost-benefit was €201,741, and the cost avoidance was €337,306.Multidisciplinary care and pharmaceutical care are key elements to improve cancer patients' outcomes and avoid evitable healthcare costs.

The risk of immune thrombocytopenia (ITP) worsening during pregnancy and neonatal ITP (NITP) have never been prospectively studied. We included 180 pregnant and 168 nonpregnant women with ITP in a prospective, multicenter, observational cohort study. A total of 131 pregnant women with ITP were matched to 131 nonpregnant women with ITP by history of splenectomy, ITP status (no response, response, complete response), and duration. Groups were followed for 15 months. The primary outcome was the first occurrence of ITP worsening defined by a composite end point including bleeding events and/or severe thrombocytopenia (

Since the development of tyrosine kinase inhibitors (TKIs) targeting BCR/ABL1 and significantly improved outcomes on long-term disease-free survival in Philadelphia chromosome–positive (Ph+) acute ...

International audience

Immunocompromised individuals such as patients with chronic lymphocytic leukemia (CLL) are at risk of impaired immune responses to vaccination. The objective of our study was to evaluate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–specific antibody responses in patients with CLL after the first, second, and third doses of the BNT162b2 or mRNA-1273 vaccines and after a single dose for patients with confirmed previous COVID-19. In all, 530 patients were included in the study. Patients received 2 doses at a 4-week interval and a third dose if they were seronegative after the second dose. Response rate was 27% after dose 1 and 52% after dose 2. Post-dose 2 treatment-naïve patients had the highest response rate (72%) followed by patients previously treated by chemoimmunotherapy (60%). Among patients receiving therapy, those receiving Bruton tyrosine kinase inhibitor alone (22%) or in combination with anti-CD20 monoclonal antibodies or venetoclax (0%) had the poorer response rate whereas patients who received venetoclax monotherapy achieved a significantly higher response rate (52%). A multivariable analysis identified age older than 65 years, ongoing CLL treatment, and gamma globulin ≤6 g/L as independent predictors of the absence of seroconversion. Post-dose 2 seronegative patients had a global response rate of 35% after dose 3. This study provides an argument for the use of a third dose and for prophylactic SARS-CoV-2 neutralizing monoclonal antibodies.

The treatment of relapsed/refractory (R/R) acute myeloid leukaemia (AML) remains a challenge. Among salvage chemotherapy regimens, the clofarabine and cytarabine (CLARA) combination has been widely evaluated and has a favourable safety/efficacy balance. Predictive factors of efficacy in patients with R/R AML are unclear, particularly the impact of AML-related gene mutations. We report our single-centre experience on 34 R/R AML patients treated with CLARA, with a focus on the genetic characterization of our cohort. CLARA yielded a 47% response rate among this poor-prognosis AML population, while two patients (5·8%) died due to treatment-related toxicity. The two-year progression-free survival and overall survival rates were 29·4% and 35·3%, respectively. Nine patients (26%) had long-term response with a median follow-up of 39·5 months among the responders, of whom six underwent haematopoietic stem cell transplantation. Adverse karyotype did not correlate with response or survival, and secondary AML were more frequent among responders to CLARA, suggesting that this combination may successfully salvage R/R AML patients regardless of adverse prognostic markers. We also observed that a low mutational burden and absence of splice mutations correlated with prolonged survival after CLARA, suggesting that extensive genotyping may have prognostic implications in R/R AML.

B-cell acute lymphoblastic leukemia (B-ALL) can be cured in most children and in the majority of young adults with sufficiently intensive – and myelosuppressive – combination chemotherapy regimens ...

TO THE EDITOR : Patients with chronic lymphocytic leukemia (CLL) exhibit a variety of skin lesions including mostly non-specific cutaneous manifestations (such as cutaneous infections or exaggerated reactions to insect bite) and secondary cutaneous malignancies, as patients are at high risk of developing basal cell carcinoma, squamous cell carcinoma, melanoma, and Merkel cell carcinoma. Specific cutaneous infiltration by neoplastic B lymphocytes with clinically identifiable skin lesions, also called leukemia cutis (LC), is more uncommon and has seldom been reported in chronic lymphocytic leukemia. [...]

Recent years have witnessed a dramatic improvement in multiple myeloma (MM) patient care with a current median life expectancy of 6 to 10 years dependent on age. High-dose melphalan followed by aut...

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for an ongoing global pandemic. Phase III trials have demonstrated excellent efficacies of mRNA vaccines against SARS-CoV-2 in large population studies (Baden LR, NEJM, 2021; Polack FP, NEJM, 2020). Immunosuppressed individuals such as chronic lymphocytic leukemia (CLL) patients are at risk for a suboptimal response to 2 vaccine doses (Herishanu Y, Blood, 2021). The French National Authority for Health recommends the use of a third dose in immunosuppressed patients. However, seroconversion rate after the triple-dose vaccine is not yet known. The objective of our study was to evaluate SARS-CoV-2 antibody responses after the first, second and third doses of the BNT162b2 and mRNA-1273 vaccines. Data were collected from 17 French Innovative Leukemia Organization (FILO) investigating centers and the French CLL patients' association (SILLC). SARS-CoV-2 IgG anti-Spike levels were measured at 4-6 weeks after each vaccine dose. A total of 530 patients and 14 controls were included in the study. Vaccine response was evaluated post-dose 1 for 158 CLL patients, post-dose 2, for 506 patients and post-dose 3 for 66 patients. Peripheral blood lymphocyte subsets were studied post-dose 2 by flow cytometry in 80 CLL patients and 14 controls. The median age of the patients was 71 years (range 37-93), 218 (40%) were treatment-naïve (TN), 136 (26%) had a prior CLL treatment and 176 (34%) were on therapy. Post-dose 1, the global response rate was 27% (43/158). TN patients had a response rate of 34% (23/67), similar to those who had a prior CLL treatment (33%,12/36), and higher compared to on-therapy patients (15%, 8/55, P=0.02). Post-dose 2, the global response rate was 52% (265/506). TN patients had the highest response rate of 72% (151/210) compared to previously treated patients, mostly by immunochemotherapy (60%, 78/130, P=0.02) and on-therapy patients (22%, 36/166, P65 years (OR 0.55, 95% CI 0.33-0.92, P=0.02), ongoing CLL treatment (OR 0.13, 95% CI 0.07-0.23, P Flow cytometry results suggest a differential balance of the T CD4+ cell subpopulations in Binet stage A and in patients on targeted therapy compared to healthy controls. Post-dose 2 seronegative patients were proposed a third dose and to date, 66 have been tested for the antibody response 4-6 weeks post-dose 3. The post-dose 3 response rate was 42% (28/66). TN patients and previously treated patients had a significantly higher response rate (57%, 16/28) compared to on-therapy patients (32%, 12/38, P=0.03). We further analyzed patients tested post-dose 2 with the Abbott Architect SARS-CoV-2 IgG anti-Spike assay (n=24). Those who achieved seroconversion after the third dose (n=10) had significantly higher titers post-dose 2 (median 12, IQR 3.0-40.8) compared to those who remained seronegative (n=14) (median 2.2, IQR 0.5-5.1, p An additional cohort of 40 CLL patients who presented a SARS-CoV-2 infection prior to vaccination participated to the study and was analyzed independently. All patients achieved seroconversion after infection and a single dose of vaccine, even though 30% (n=12) had an ongoing CLL treatment. In conclusion, double-dose mRNA vaccination generated a humoral response in 52% of our CLL cohort and a third dose induced seroconversion in 42% of the patients who remained seronegative after the second dose. The major independent predictor of negative antibody response was ongoing treatment with BTKi. The strongest boost to immune response against the virus seems to be SARS-CoV-2 infection, as a substantial increase in anti-Spike antibodies was observed in all CLL patients with prior infection, after a single dose vaccination. Figure 1 Figure 1. Disclosures Letestu: Janssen: Research Funding, Speakers Bureau; Roche: Speakers Bureau; AbbVie: Research Funding, Speakers Bureau. Dartigeas: Astra-Zeneca: Membership on an entity's Board of Directors or advisory committees, Other: travel grants/Congress; Abbvie: Membership on an entity's Board of Directors or advisory committees, Other: travel grants/Congress; Janssen: Membership on an entity's Board of Directors or advisory committees, Other: travel grants/Congress. Laribi: Astellas Phama, Inc.: Other: Personal Fees; BeiGene: Other: Personal Fees; AstraZeneca: Other: Personal Fees; IQONE: Other: Personal Fees; AbbVie: Other: Personal Fees, Research Funding; Takeda: Other: Personal Fees, Research Funding; Novartis: Other: Personal Fees, Research Funding; Le Mans Hospital: Research Funding; Jansen: Research Funding. Cymbalista: Lilly-LOXO: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; ASTRA ZENECA: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Introduction: TP53 gene mutations (m TP53), found in up to 20% of MDS or AML pts and 30-40% of therapy-related (TR) MDS/AML cases, represent a distinct molecular cohort with poor outcomes. Hypomethylating agents (HMA) are the frontline standard of care, with CR rates of ~20% and median OS of < 12 months. APR-246 is a novel, first-in-class small molecule that reactivates the mutant p53 protein and targets cellular redox balance, ultimately inducing apoptosis and ferroptosis in m TP53 cancer cells. We previously reported the Phase 2 results of 2 parallel trials of APR-246+AZA (Sallman et al and Cluzeau et al., JCO 2021). We now report analyses of the combined Phase 2 cohorts and long-term follow-up. Methods: This is a multicenter, international collaboration of the US MDS clinical research consortium and the GFM of HMA-naïve m TP53 higher-risk MDS, MDS/MPN and oligoblastic AML (≤ 30% blasts) pts (NCT03072043/NCT03588078). Pts received APR-246 4500mg IV (days 1-4) + AZA 75 mg/m 2 SC/IV x 7 days (days 4-10 or 4-5 and 8-12) in 28-day cycles. Primary objective was CR rate by International Working Group (IWG) 2006 criteria. Secondary objectives included ORR, OS, outcome following allogeneic hematopoietic stem cell transplant (allo-HSCT), with serial high depth next generation sequencing (NGS, 0.1% cutoff) for evaluation of measurable residual disease (MRD). Results: As of July 15, 2021, 100 pts had been enrolled with a median age of 68 years (range, 34-87; 47% male). By WHO, 74 pts had MDS, 22 AML-MRC and 4 CMML/MDS-MPN; 83% complex karyotype (CK) and 88% were CK and/or biallelic for TP53 mutations; 92% had a TP53 missense mutation in the DNA binding domain. In 63 pts, TP53 was the only mutation detected (i.e. isolated m TP53). Median time on treatment was 6 cycles (1-25+) with 5 pts ongoing and 23 pts who proceeded to allo-HSCT. Non-hematologic treatment (Tx)-related adverse events (AEs) in ≥20% of pts included nausea/vomiting (58%), ataxia (26%), and dizziness (23%). Neurologic AEs were reversible in 100% of cases. Febrile neutropenia occurred in 37% of pts. Thirty and 60-day mortality was 1% (n=1) and 7% (n=7), respectively. Dose reductions of APR-246 and AZA occurred in 16% and 1% of pts, respectively, with only 1 treatment discontinuation due to a treatment-related AE. By intention-to-treat (ITT) analysis, ORR by IWG was 69% with 43 CR, 1 PR, 10 marrow CR (mCR)+HI, 9 HI alone, and 6 with mCR. Of non-responders, 6 had stable disease and 7 pts had progressive disease. The median duration of CR/PR was 10.6 months (95% CI 8.8-12.3, 23+ months ongoing). CR/PR rate for MDS was 49% (36/74), 36% for AML (8/22) and 0% for MDS/MPN (0/4) with an ORR rate of 70%/64%/75%, respectively. Isolated m TP53 was predictive for a higher CR rate (52% vs 30%; P=.04). Patients who had biallelic TP53 or CK had a significantly higher CR rate vs pts without (49% vs 8%; P=0.01). On serial TP53 NGS using a VAF cutoff of 5%, 40 pts achieved NGS negativity with 6 pts MRD negative (VAF < 0.1%). Of NGS negative pts (TP53 VAF At data-cutoff by ITT analysis with a median follow up of 27.8 months, median OS was 11.8 months (95% CI 9.4-14.3). Pts undergoing allo-HSCT had a median OS of 16.1 months (95% CI 14-18.1). Impact of response and NGS clearance was evaluated by landmark analysis at 6 months. Pts achieving CR/PR or NGS negativity had improved OS (15.8 vs 10.1 months; P=0.002; Fig 1A). Additionally, pts who became MRD negative had a 2-year OS of 50% vs 23% (P=0.21). Although allo-HSCT was not predictive of OS in the overall cohort by landmark analysis (14.7 vs 14.4 months; P=0.15; Fig 1B), significant OS improvement was noted in allo-HSCT pts based on CR/PR or NGS negativity (P=0.002; Fig 1C). Notably, pts who achieved CR/PR/NGS negativity and were bridged to allo-HSCT had a median OS that was not reached (95% CI 10.4-NR) vs 9.1 months (95% CI 7.4-NR) in allo-HSCT pts who did not achieve this response (P=0.01). Conclusions: In this international, combined analysis of P2 APR-246+AZA pts, the combination was well-tolerated with high response rates in m TP53 MDS/AML. Quality of response and NGS negativity strongly predicted OS, particularly in the setting of allo-HSCT, validating NGS clearance as a critical biomarker of allo-HSCT outcomes in m TP53 pts. Figure 1 Figure 1. Disclosures Sallman: Intellia: Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite: Membership on an entity's Board of Directors or advisory committees; Aprea: Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees; Magenta: Consultancy; Shattuck Labs: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Syndax: Membership on an entity's Board of Directors or advisory committees; Incyte: Speakers Bureau; Agios: Membership on an entity's Board of Directors or advisory committees. Komrokji: BMS: Honoraria, Speakers Bureau; JAZZ: Honoraria, Speakers Bureau; Abbvie: Honoraria, Speakers Bureau; Novartis: Honoraria; Geron: Honoraria; Acceleron: Honoraria; Agios: Honoraria, Speakers Bureau. DeZern: Taiho: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees. Sebert: BMS: Consultancy; Abbvie: Consultancy. Steensma: Novartis: Current Employment. Roboz: Astellas: Consultancy; Otsuka: Consultancy; Blueprint Medicines: Consultancy; AbbVie: Consultancy; Bayer: Consultancy; Glaxo SmithKline: Consultancy; Celgene: Consultancy; Bristol Myers Squibb: Consultancy; Helsinn: Consultancy; Janssen: Consultancy; Novartis: Consultancy; Jasper Therapeutics: Consultancy; MEI Pharma - IDMC Chair: Consultancy; Janssen: Research Funding; AstraZeneca: Consultancy; Jazz: Consultancy; Amgen: Consultancy; Daiichi Sankyo: Consultancy; Mesoblast: Consultancy; Agios: Consultancy; Pfizer: Consultancy; Astex: Consultancy; Actinium: Consultancy; Roche/Genentech: Consultancy. Sekeres: BMS: Membership on an entity's Board of Directors or advisory committees; Takeda/Millenium: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Sweet: Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; AROG: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Bristol Meyers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees. Kuykendall: Incyte: Consultancy; Novartis: Honoraria, Speakers Bureau; Prelude: Research Funding; PharmaEssentia: Honoraria; CTI Biopharma: Honoraria; Celgene/BMS: Honoraria, Speakers Bureau; BluePrint Medicines: Honoraria, Speakers Bureau; Abbvie: Honoraria; Protagonist: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Recher: Agios: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS/Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Honoraria; Janssen: Honoraria; Jazz: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; MaatPharma: Research Funding; Macrogenics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees. Raffoux: ASTELLAS: Consultancy; ABBVIE: Consultancy; PFIZER: Consultancy; CELGENE/BMS: Consultancy. Padron: Incyte: Research Funding; BMS: Research Funding; Blueprint: Honoraria; Kura: Research Funding; Taiho: Honoraria; Stemline: Honoraria. Attar: Aprea Therapeutics: Current Employment, Current equity holder in publicly-traded company. Kantarjian: Amgen: Honoraria, Research Funding; KAHR Medical Ltd: Honoraria; BMS: Research Funding; AbbVie: Honoraria, Research Funding; Ascentage: Research Funding; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Astellas Health: Honoraria; Jazz: Research Funding; Aptitude Health: Honoraria; Astra Zeneca: Honoraria; Ipsen Pharmaceuticals: Honoraria; NOVA Research: Honoraria; Precision Biosciences: Honoraria; Immunogen: Research Funding; Daiichi-Sankyo: Research Funding; Taiho Pharmaceutical Canada: Honoraria. List: Precision BioSciences: Current Employment, Current equity holder in publicly-traded company; Aileron Therapeutics: Consultancy; CTI Biosciences: Consultancy; Halia Therapeutics: Consultancy, Current holder of individual stocks in a privately-held company. Ades: Celgene: Honoraria, Research Funding; Abbvie: Honoraria; Takeda: Honoraria; Novartis: Honoraria; JAZZ: Honoraria. Lancet: BerGenBio: Consultancy; Celgene/BMS: Consultancy; Millenium Pharma/Takeda: Consultancy; Jazz: Consultancy; Astellas: Consultancy; Daiichi Sankyo: Consultancy; ElevateBio Management: Consultancy; Agios: Consultancy; AbbVie: Consultancy. Fenaux: Janssen: Honoraria, Research Funding; Takeda: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; JAZZ: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Celgene/BMS: Honoraria, Research Funding; Syros Pharmaceuticals: Honoraria.

Mutational analyses performed following acquired ibrutinib resistance have suggested that chronic lymphocytic leukemia (CLL) progression on ibrutinib is linked to mutations in Bruton tyrosine kinase (BTK) and/or phospholipase Cγ2 (PLCG2) genes. Mutational information for patients still on ibrutinib is limited. We report a study aimed to provide a “snapshot” of the prevalence of mutations in a real-life CLL cohort still on ibrutinib after at least 3 years of treatment. Of 204 patients who initiated ibrutinib via an early-access program at 29 French Innovative Leukemia Organization (FILO) centers, 63 (31%) were still on ibrutinib after 3 years and 57 provided a fresh blood sample. Thirty patients had a CLL clone ≥0.5 × 109/L, enabling next-generation sequencing (NGS); BTK and PLCG2 mutations were detected in 57% and 13% of the NGS samples, respectively. After median follow-up of 8.5 months from sample collection, the presence of a BTK mutation was significantly associated with subsequent CLL progression (P = .0005 vs no BTK mutation). Our findings support that mutational analysis should be considered in patients receiving ibrutinib who have residual clonal lymphocytosis, and that clinical trials are needed to evaluate whether patients with a BTK mutation may benefit from an early switch to another treatment.

We would like to report a study on solitary plasmacytoma of the bone (SPB) treatment.SPB is a rare plasma cell dyscrasia characterized by a single bone tumor with no evidence of other lesions or mu...

Introduction : TP53 mutated (TP53m) MDS and AML have very poor outcome irrespective of the treatment received, including 40% responses (20% CR) with azacitidine (AZA) with short response duration and a median overall survival (OS) of about 8 months (Bejar, Blood 2014). APR-246 is a prodrug spontaneously converted to methylene quinuclidinone (MQ), a Michael acceptor that binds covalently to cysteines in mutant p53, leading to protein reconformation that reactivates its pro apoptotic and cell cycle arrest functions. The combination of AZA and APR 246 showed promising results in a phase Ib study in TP53m MDS (Sallman, ASH 2018). We report interim results of a GFM phase 2 study of AZA+ APR-246 in TP53m MDS and AML, conducted in parallel with a similar US MDS consortium study. Patients and Methods : The study included hypomethylating agent (HMA) naïve and not previously allografted intermediate, high or very high IPSS-R TP53m MDS and AML adult patients. Patients received APR-246 4500 mg IV /d (6 hour infusions) (days 1-4) followed by AZA 75 mg/m²/d (days 4-10) in 28 day cycles. Response (primary endpoint, assessed by IWG 2006 for MDS and ELN criteria for AML) was evaluated after 3 and 6 cycles in the intent to treat (ITT) population, ie all patients who had received any protocol treatment, and in patients who had at least a blood and bone marrow evaluation after cycle 3 (evaluable population). Allo-SCT, when possible, was proposed after 3 to 6 cycles, and treatment with reduced APR 246 and AZA doses could be continued after allo-SCT. Results : 53 patients were enrolled between Sept 2018 and July 2019 in 7 GFM centers, with a median age of 73 years (range 44-87), and M/F: 28/25. 34 patients had MDS (including 74% very high IPSS-R) and 19 had AML. IPSS-R cytogenetic risk was very poor in 30/34 MDS, and unfavorable in 18/19 AML, complex in 89% of the patients. Median baseline mutated TP53 VAF was 21% (range 3-76). Nineteen of the 53 patients had been included at least 7 months before date of analysis (25 July 2019), had received protocol treatment and were thus potentially evaluable for response after 6 treatment cycles (ITT population). One of them died after only one cycle from an unrelated cause (cerebral ischemic stroke), and 2 during the third cycle (from bleeding and sepsis, respectively). In the remaining 16 patients (evaluable population per protocol), the response rate was 75% including 9 (56%) CR, 3 (19%) marrow CR or stable disease with hematological improvement (HI), and 4 treatment resistance. In the ITT population, the response rate was 63%, including 47% CR, and 16% stable or marrow CR+ HI. Among CR patients, complete cytogenetic CR and negative NGS for TP53 mutation (VAF cutoff of 2%) were achieved in 7/9 (78%) and 8/8 (100%), respectively. So far, 1 patient has undergone allo-SCT. All 53 patients had received at least one treatment cycle, and no increased myelosuppression, compared with AZA alone, was apparent. Treatment related AEs observed in ≥ 20% of patients were febrile neutropenia in 19 (36%) and neurological AEs in 21 (40%) of the patients. The latter, reviewed with a neurological team, were mainly grade 1 or 2 and consisted of ataxia (n=13), sometimes associated with cognitive impairment (n=4), suggesting a cerebellar origin. Other patients experienced acute confusion (n=4), isolated dizziness (n=3) and facial paresthesia (n=1). Neurological AEs reached grade III in 3 cases (1 acute confusion, 2 ataxia). Occurrence of neurological AEs was correlated with lower glomerular filtration rate at treatment onset (p Conclusion : In this very high-risk elderly population of TP53m MDS and AML, generally with complex karyotype, a promising 56% CR rate at 6 cycles was reached in the evaluable population with AZA+ APR 246 combination, with deep molecular remission in all CR patients. We observed manageable neurologic AEs, mainly in elderly patients with reduced renal function, who therefore require close monitoring and dose reduction if necessary. An update regarding safety and efficacy in the 53 patients, including survival data, will be available at the meeting. A phase III international trial comparing AZA alone and AZA+ APR 246 in TP53m MDS is ongoing. Disclosures Cluzeau: Abbvie: Consultancy; Jazz Pharma: Consultancy; Menarini: Consultancy. Peterlin:AbbVie Inc: Consultancy; Astellas: Consultancy; Jazz Pharma: Consultancy; Daiichi-Sankyo: Consultancy. Recher:Daiichi-Sankyo: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; chugai: Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Agios: Research Funding; Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Astellas Pharma: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Stamatoullas:Celgene: Honoraria; Takeda: Consultancy. Berthon:JAZZPHARMACEUTICAL: Other: DISCLOSURE BOARD; CELGEN: Other: DISCLOSURE BOARD; PFIZER: Other: DISCLOSURE BOARD. Sallman:Celyad: Membership on an entity's Board of Directors or advisory committees. Ades:Amgen: Research Funding; Astellas: Membership on an entity's Board of Directors or advisory committees; Silence Therapeutics: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Jazz: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Helsinn Healthcare: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Fenaux:Celgene Corporation: Honoraria, Research Funding; Astex: Honoraria, Research Funding; Jazz: Honoraria, Research Funding; Aprea: Research Funding.

l-asparaginase (L-ase) is a major drug used to treat acute lymphoblastic leukemia (ALL) [1]. L-ase kills leukemic cells by depleting circulating asparagine pools related to its asparaginase activit...

Sustained Leukemia-Free State and Molecular Response to Sorafenib in a Patient With Chronic Myelomonocytic Leukemia in Transformation Driven by Homozygous FLT3-ITD Malignant Hematopoiesis Olivier Kosmider, Nicolas Chapuis, Sophie Kaltenbach, Romain Coriat, Pascaline Boudou Rouquette, Lise Willems, Virginie Chesnais, Isabelle Radford-Weiss, Valerie Bardet, Patrick Mayeux, Jerome Tamburini, Michaela Fontenay, Didier Bouscary

International audience; Primary bone diffuse large B-cell lymphoma (PB-DLBCL) is a rare DLBCL location variant. We treated 76 PB-DLBCL patients by immuno-chemotherapy, resulting in an 84% sustained complete remission rate and a 78.9% survival over a 4.7-year median follow-up period. Ann Arbor stage IV and high age-adjusted international prognostic index were predictive of adverse outcome in univariate analysis. In multivariate analysis using a Cox model, only aa-IPI predicted long-term survival. While based on a limited number of cases, we suggested that radiotherapy may be useful as a consolidation modality in PB-DLBCL. We also suggested that positron emission tomography/CT scan should be interpreted with caution due to a persistent [18F]fluorodeoxyglucose [18FDG] uptake of bone lesions even after remission in some in PB-DLBCL patients. Our study based on a homogeneous cohort of PB-DLBCL patients confirmed the favorable outcome of this DLBCL variant and support the implementation of prospective clinical trials in this disease.

Myelodysplastic syndromes (MDSs) are hematopoietic stem cell disorders in which recurrent mutations define clonal hematopoiesis. The origin of the phenotypic diversity of non-del(5q) MDS remains unclear. Here, we investigated the clonal architecture of the CD34+CD38- hematopoietic stem/progenitor cell (HSPC) compartment and interrogated dominant clones for MDS-initiating cells. We found that clones mainly accumulate mutations in a linear succession with retention of a dominant subclone. The clone detected in the long-term culture-initiating cell compartment that reconstitutes short-term human hematopoiesis in xenotransplantation models is usually the dominant clone, which gives rise to the myeloid and to a lesser extent to the lymphoid lineage. The pattern of mutations may differ between common myeloid progenitors (CMPs), granulomonocytic progenitors (GMPs), and megakaryocytic-erythroid progenitors (MEPs). Rare STAG2 mutations can amplify at the level of GMPs, from which it may drive the transformation to acute myeloid leukemia. We report that major truncating BCOR gene mutation affecting HSPC and CMP was beneath the threshold of detection in GMP or MEP. Consistently, BCOR knock-down (KD) in normal CD34+ progenitors modifies their granulocytic and erythroid differentiation. Clonal architecture of the HSPC compartment and mutations selected during differentiation contribute to the phenotypic heterogeneity of MDS. Defining the hierarchy of driver mutations provides insights into the process of transformation and may guide the search for novel therapeutic strategies.

Clostridium difficile (C. difficile) infection was observed in 13% of recipients after hematopoietic stem cell transplantation (HSCT), mainly in the first month posttransplantation. Risk factors were cord blood as the source of stem cells, acute graft-versus-host disease (GVHD), and total body irradiation (TBI). No association was found with an increased risk of mortality. The purpose of this study was to evaluate the incidence, risk factors, and outcome of C. difficile infection (CDI) after HSCT. We conducted a single-center, retrospective, cohort study on all patients who received an allogeneic HSCT from January 2004 to December 2007. All patients with diarrhea in the first year after HSCT were tested for the presence of C. difficile in stools. Among the 407 assessable patients, 53 presented at least 1 CDI in the first year post-HSCT. The total incidence rate was 5.6 cases of CDI per 10,000 patient-days. Fifty percent of cases were diagnosed in the first month after HSCT, and 95% occurred during the first 6 months. Fewer than 5% of patients with CDI had severe diarrhea and severe complications were never observed. TBI in the conditioning regimen, cord blood as the source of stem cells, and acute graft-versus-host disease (aGVHD) were independently associated with CDI. Six patients (11%) had a recurrence of CDI. Four patients required second-line treatment with vancomycin. With a median follow-up of 22 months, the 2-year overall survival rates were similar between patients who presented a CDI and those who did not. CDI was observed in approximately 13% of recipients after HSCT, mainly in the first month posttransplantation and was associated with CB, aGVHD, and TBI. CDI was not associated either with severe complications or with an increased risk of mortality in this large cohort of patients.

Mammalian target of rapamycin (mTOR) is a protein kinase implicated in the regulation of various cellular processes, including those required for tumor development, such as the initiation of mRNA translation, cell-cycle progression and cellular proliferation. In a wide range of hematological malignancies, the mTORC1 signaling pathway has been found to be deregulated and has been designed as a major target for tumor therapy. Given that pre-clinical studies have clearly established the therapeutic value of mTORC1 inhibition, numerous clinical trials of rapamycin and its derivates (rapalogs) are ongoing for treatment of these diseases. At this time, although disease stabilization and tumor regression have been observed, objective responses in some tumor types have been modest. Nevertheless, some of the mechanisms underlying cancer-cell resistance to rapamycin have now been described, thereby leading to the development of new strategy to efficiently target mTOR signaling in these diseases. In this review, we discuss the rationale for using mTOR inhibitors as novel therapies for a variety of hematological, malignancies with a focus on promising new perspectives for these approaches.

Cancer cells require glutamine to adapt to increased biosynthetic activity. The limiting step in intracellular glutamine catabolism involves its conversion to glutamate by glutaminase (GA). Different GA isoforms are encoded by the genes GLS1 and GLS2 in humans. Herein, we show that glutamine levels control mitochondrial oxidative phosphorylation (OXPHOS) in acute myeloid leukemia (AML) cells. Glutaminase C (GAC) is the GA isoform that is most abundantly expressed in AML. Both knockdown of GLS1 expression and pharmacologic GLS1 inhibition by the drug CB-839 can reduce OXPHOS, leading to leukemic cell proliferation arrest and apoptosis without causing cytotoxic activity against normal human CD34(+) progenitors. Strikingly, GLS1 knockdown dramatically inhibited AML development in NSG mice. The antileukemic activity of CB-839 was abrogated by both the expression of a hyperactive GAC(K320A) allele and the addition of the tricarboxyclic acid cycle product α-ketoglutarate, indicating the critical function of GLS1 in AML cell survival. Finally, glutaminolysis inhibition activated mitochondrial apoptosis and synergistically sensitized leukemic cells to priming with the BCL-2 inhibitor ABT-199. These findings show that targeting glutamine addiction via GLS1 inhibition offers a potential novel therapeutic strategy for AML.

Cancer cells require nutrients and energy to adapt to increased biosynthetic activity, and protein synthesis inhibition downstream of mammalian target of rapamycin complex 1 (mTORC1) has shown promise as a possible therapy for acute myeloid leukemia (AML). Glutamine contributes to leucine import into cells, which controls the amino acid/Rag/mTORC1 signaling pathway. We show in our current study that glutamine removal inhibits mTORC1 and induces apoptosis in AML cells. The knockdown of the SLC1A5 high-affinity transporter for glutamine induces apoptosis and inhibits tumor formation in a mouse AML xenotransplantation model. l-asparaginase (l-ase) is an anticancer agent also harboring glutaminase activity. We show that l-ases from both Escherichia coli and Erwinia chrysanthemi profoundly inhibit mTORC1 and protein synthesis and that this inhibition correlates with their glutaminase activity levels and produces a strong apoptotic response in primary AML cells. We further show that l-ases upregulate glutamine synthase (GS) expression in leukemic cells and that a GS knockdown enhances l-ase–induced apoptosis in some AML cells. Finally, we observe a strong autophagic process upon l-ase treatment. These results suggest that l-ase anticancer activity and glutamine uptake inhibition are promising new therapeutic strategies for AML.

Introduction Myelodysplastic syndromes (MDS) are clinically diverse malignant disorders of aging with a propensity to evolve to acute myeloid leukemia (AML) or bone marrow failure. In early MDS, whole genome sequencing identified mutations distributed in few clones. Evidence have been provided for the existence of a MDS-initiating cell in cases harboring a 5q deletion. However, the clonal heterogeneity and its consequences on the phenotypic diversity of non-del(5q) MDS is little documented. Here we focused on studying the hierarchical organization and the functionality of clones defined by molecular profiling. The clonal architecture of the CD34+CD38- hematopoietic stem/progenitor cell (HSPC) compartment was investigated and dominant clones were examined as MDS-initiating cells. Material and methods Bone marrow (BM) samples were obtained from 20 patients with non-del(5q) MDS enrolled in the national Programme Hospitalier de Recherche Clinique MDS-04 after informed consent in accordance with ethics committee guidelines. BM samples, cytaphereses from age-matched healthy individuals and cord bloods were used as controls. Targeted NGS of a selected panel of 39 genes was used to define the mutational landscape on BM mononuclear cells (MNC). To study the clonal architecture at the HSPC level, single CD34+CD38- cells were seeded in 96-well plates coated with MS-5 stromal cells and cultured in H5100 MyeloCult medium (StemCell Technologies, Vancouver, Canada) with cytokines for six weeks. For long-term culture-initiating cell (LTC-IC) assays, CD34+ progenitors were cultured for six weeks on MS-5-coated plates without cytokines and then tested for colony-forming cells. For clonogenic assays, CD34+CD38- cells were seeded in methylcellulose for two weeks. All animal experimentations were performed in NSG mice. Results In the 20 cases of non-del(5q) MDS, genomic lesions were traced down to single CD34+CD38- HSPC-derived colonies. Clonal organization was mostly linear in 13/17 patients and branched in 4 cases with retention of a dominant subclone. The clone detected in LTC-IC compartment and that reconstituted short-term human hematopoiesis in xenotransplantation models was usually the dominant clone, which gave rise to the myeloid and to a lesser extent to the lymphoid lineage. Other mutations not detected in LTC-IC can appear in CD34+CD38- compartment or at the level of lineage-committed progenitors. The pattern of mutations may differ between common myeloid (CMP), granulo-monocytic (GMP) and megakaryocytic-erythroid (MEP) progenitors. For instance, a major truncating BCOR gene mutation affecting HSPC and CMP was beneath the threshold of detection in GMP or MEP. Consistently, BCOR knockdown by shRNA in normal CD34+ progenitors impaired their granulocytic and erythroid differentiation. By contrast, a STAG2 gene mutation, not detected in CMP or MEP, amplified in a GMP, which drove the transformation to AML. Conclusion In the present study, we characterized the first genetic hits that initiate disease in a dominant clone of the CD34+CD38- HSPC compartment, which exhibits LTC-IC activity and reconstitutes human short-term hematopoiesis in NSG mice. The genetic heterogeneity in non-del(5q) MDS arises within the HSPC compartment and in lineage-committed progenitors which ultimately support the transformation into AML. The clonal architecture of HSPC compartment and mutations selection along differentiation contribute to the phenotype of MDS. Defining the hierarchy of driver mutations provides insights into the process of transformation, and may guide the search for novel therapeutic strategies. Disclosures No relevant conflicts of interest to declare.

Background: Myelofibrosis (MF) is the less frequent Philadelphia-negative myeloproliferative neoplasms (MPN). We have included in a nationwide database primary (P), post-polycythemia (PPV) and post-essential thrombocythemia (PET) MF diagnosed in France since 2005. Methods: Inclusion criteria were: diagnostic of MF after 2005; according strictly to WHO (bone marrow biopsy mandatory); informed consent. The registry was launched in Oct. 2013, and 26 hematology centers included patients (pts). Summary statistics were reported, namely median [Interquartile range, IQR] or percentage. Baseline characteristics were compared across IPSS groups using chi-square test or Mann and Whitney test. Kaplan-Meier survival curves were plotted and compared by the log-rank test. Results: At time of analysis (June 2016) a total of 527 pts were included in the registry, complete baseline data were available in 499 (95%), and follow-up (FU) data in 433 (87%). Median [IQR] age and M/F sex ratio were 71 [63-78] years and 315/184 (1.7), respectively (resp). 301 (60%) pts had PMF, 182 (36%) had secondary MF (including 64 PPVMF and 118 PETMF) and 16 had pre-fibrotic MF. Five percent of pts had a familial history of hematologic malignancy, and 22% a history of thrombosis or hemorrhage. Splenomegaly was present in 386 (77%) with a median [IQR] spleen size of 4 cm [1-8] below costal margin, and was symptomatic in 11% of pts. Constitutional symptoms were present in 107 (21%) (weight loss in 56, night sweats in 61, fever in 14), and ECOG score was 0, 1, 2, and 3 in 53%, 36%, 11% and 0.3% of pts, resp. Median [IQR] Hemoglobin, WBC and platelet counts were 109 g/L [94-122], 9.3 G/L [5.7-16.0] and 257 G/L [138-430], resp. Circulating blast cells were present in 41%, LDH was above normal value in 95% of pts, median EPO level was 54 [11 - 57] U/L. Grade of fibrosis (WHO) was 1, 2, and 3 in 2%, 66% and 32% of pts, resp. Karyotype was done in 321 pts, normal in 173 (54%), abnormal with favorable prognostic value in 89 (30%) and unfavorable in 30 (10%) (29 failures). A total of 461 (92%) pts had molecular testing: 60% were JAK2V617F positive, 4% had MPL and 7% had CALR mutations, and 99 (28%) over the 352 pts with triple testing were triple negative. IPSS risk categories were low, int-1, int-2 and high in 68 (14%), 168 (34%), 158 (32%), and 105 (21%) pts, resp. In addition to constitutional symptoms, there was a significant increase in the prevalence of clinical signs across IPSS categories (from low to high risk): symptomatic spleen (p=0.016) -though no difference in spleen size was found (p=0.18)-, early satiety (p=0.013), ECOG score (p= 0.0001), bone pain (p=0.002). Moreover, among biological parameters, there was an increase across IPSS groups in WBC (p=0.029), LDH (p=0.0007), ferritin (p=0.003), circulating CD34+ cells (p=0.020), EPO level (p=0.035). In contrast, a decrease was seen for hemoglobin and platelets (p=0.0001 for both). Lastly, frequency of grade 3 fibrosis increased with IPSS (p=0.043), while no evidence of difference was found regarding abnormal karyotype and mutational pattern. Median FU was 33 months [9-63 months]. Among those 433 pts with FU data, median FU was 38 months [19-68], and 124 (29 %) pts had died at the time of analysis, including 13%, 17%, 31%, and 40% of pts from the Low, Int-1, Int-2, and High risk groups, resp (p= 0.0001, figure 1). In the 450 pts with treatment data, treatments received during FU included cytoreductive drug (41%), Jak-inhibitors (35%), Interferon alpha (18%), IMIDs (4%). Splenectomy was performed in only 14 (3%) pts. Forty percent of pts received packed red blood cells, and 12% platelets transfusions. 49 (11%) patients participated in a clinical trial, and 27 (6%) were allografted. Conclusion: This is the first analysis of the French MF observatory after inclusion of more than 500 pts diagnosed and treated during the past 10 years. Complete baseline data and follow-up information available for the majority of pts should allow for new studies of outcomes and influence of clinical and biological parameters, as well as reassessment of prognostic models in the era of new targeted therapies. Figure 1 Comparison of overall survival according to IPPS Figure 1. Comparison of overall survival according to IPPS Figure 2 Figure 2. Disclosures Etienne: Pfizer: Speakers Bureau; BMS: Speakers Bureau; ARIAD: Speakers Bureau; novartis: Consultancy, Speakers Bureau. Tavitian:Novartis: Membership on an entity's Board of Directors or advisory committees. Ugo:Novartis: Membership on an entity's Board of Directors or advisory committees. Kiladjian:Novartis: Honoraria, Research Funding; AOP Orphan: Membership on an entity's Board of Directors or advisory committees, Research Funding.

The serine/threonine kinase mammalian target of rapamycin (mTOR) is crucial for cell growth and proliferation, and is constitutively activated in primary acute myeloid leukemia (AML) cells, therefore representing a major target for drug development in this disease. We show here that the specific mTOR kinase inhibitor AZD8055 blocked mTORC1 and mTORC2 signaling in AML. Particularly, AZD8055 fully inhibited multisite eIF4E-binding protein 1 phosphorylation, subsequently blocking protein translation, which was in contrast to the effects of rapamycin. In addition, the mTORC1-dependent PI3K/Akt feedback activation was fully abrogated in AZD8055-treated AML cells. Significantly, AZD8055 decreased AML blast cell proliferation and cell cycle progression, reduced the clonogenic growth of leukemic progenitors and induced caspase-dependent apoptosis in leukemic cells but not in normal immature CD34+ cells. Interestingly, AZD8055 strongly induced autophagy, which may be either protective or cell death inducing, depending on concentration. Finally, AZD8055 markedly increased the survival of AML transplanted mice through a significant reduction of tumor growth, without apparent toxicity. Our current results strongly suggest that AZD8055 should be tested in AML patients in clinical trials.

The FOXO transcription factors are involved in multiple signaling pathways and have tumor-suppressor functions. In acute myeloid leukemia (AML), deregulation of oncogenic kinases, including Akt, extra-signal–regulated kinase, or IκB kinase, is frequently observed, which may potentially inactivate FOXO activity. We therefore investigated the mechanism underlying the regulation of FOXO3a, the only FOXO protein constantly expressed in AML blast cells. We show that in both primary AML samples and in a MV4-11/FOXO3a-GFP cell line, FOXO3a is in a constant inactive state due to its cytoplasmic localization, and that neither PI3K/Akt nor extra-signal–regulated kinase–specific inhibition resulted in its nuclear translocation. In contrast, the anti-Nemo peptide that specifically inhibits IKK activity was found to induce FOXO3a nuclear localization in leukemic cells. Furthermore, an IKK-insensitive FOXO3a protein mutated at S644 translocated into the nucleus and activated the transcription of the Fas-L and p21Cip1 genes. This, in turn, inhibited leukemic cell proliferation and induced apoptosis. These results thus indicate that IKK activity maintains FOXO3a in the cytoplasm and establishes an important role of FOXO3a inactivation in the proliferation and survival of AML cells. The restoration of FOXO3a activity by interacting with its subcellular distribution may thus represent a new attractive therapeutic strategy for AML.

Finding an effective treatment for acute myeloid leukemia (AML) remains a challenge, and all cellular processes that are deregulated in AML cells should be considered in the design of targeted therapies. We show in our current study that the LKB1/AMPK/TSC tumor suppressor axis is functional in AML and can be activated by the biguanide molecule metformin, resulting in a specific inhibition of mammalian target of rapamycin (mTOR) catalytic activity. This induces a multisite dephosphorylation of the key translation regulator, 4E-BP1, which markedly inhibits the initiation step of mRNA translation. Consequently, metformin reduces the recruitment of mRNA molecules encoding oncogenic proteins to the polysomes, resulting in a strong antileukemic activity against primary AML cells while sparing normal hematopoiesis ex vivo and significantly reducing the growth of AML cells in nude mice. The induction of the LKB1/AMPK tumor-suppressor pathway thus represents a promising new strategy for AML therapy.

Background Alterations in the PI3K/Akt pathway are found in a wide range of cancers and the development of PI3K inhibitors represents a promising approach to cancer therapy. Constitutive PI3K activation, reflecting an intrinsic oncogenic deregulation of primary blast cells, is detected in 50% of patients with acute myeloid leukemia. However, the mechanisms leading to this activation are currently unknown. As we previously reported IGF-1 autocriny in acute myeloid leukemia cells, we investigated whether IGF-1 signaling was involved in the constitutive activation of PI3K. Design and Methods We analyzed the IGF-1/IGF-1R signaling pathway and PI3K activity in 40 acute myeloid leukemia bone marrow samples. Specific inhibition of IGF-1/IGF-1R signaling was investigated using neutralizing anti-IGF-1R, anti-IGF-1 antibodies or IGF-1 short interfering RNA. The anti-leukemic activity of the neutralizing anti-IGF-1R was tested by analyzing its effects on leukemic progenitor clonogenicity, blast cell proliferation and survival. Results In all samples tested, we found that functional IGF-1R was constantly expressed in leukemic cells. In the acute myeloid leukemia samples with PI3K activation, we found that the IGF-1R was constitutively phosphorylated, although no IGF-1R activating mutation was detected. Specific inhibition of IGF-1R signaling with neutralizing anti-IGF-1R strongly inhibited the constitutive phosphorylation of both IGF-1R and Akt in 70% of the PI3K activated samples. Moreover, both incubation with anti-IGF-1 antibody and IGF-1 short interfering RNA inhibited Akt phosphorylation in leukemic cells. Finally, neutralizing anti-IGF-1R treatment decreased the clonogenicity of leukemic progenitors and the proliferation of PI3K activated acute myeloid leukemia cells. Conclusions Our current data indicate a critical role for IGF-1 autocriny in constitutive PI3K/Akt activation in primary acute myeloid leukemia cells and provide a strong rationale for targeting IGF-1R as a potential new therapy for this disease.

The PI3K/AKT and mTOR signaling pathways are activated in acute myeloid leukemia, including in the more immature leukemic populations. Constitutive PI3K activation is detectable in 50% of acute myeloid leukemia samples whereas mTORC1 is activated in all cases of this disease. In leukemic cells, the PI3K activity relates to the expression of the p110delta isoform of class IA PI3K. Constitutive PI3K activation is the result of autocrine IGF-1/IGF-1R signaling in 70% of acute myeloid leukemia samples but specific inhibition of this pathway does not induce apoptosis. Specific inhibition of PI3K/AKT or mTORC1 alone in vitro has anti-leukemic effects which are essentially exerted via the suppression of proliferation. However, as mTORC1 activation is independent of PI3K/AKT in acute myeloid leukemia, dual PI3K and mTOR inhibitors may induce apoptosis in blast cells. Moreover, mTORC1 inhibition using sirolimus overactivates PI3K/AKT via the upregulation of IRS2 expression and by favoring IGF-1/IGF-1R autocrine signaling. Recent data also indicate that mTORC1 does not control protein translation in acute myeloid leukemia. These results open the way for the design of direct inhibitors of protein synthesis as novel acute myeloid leukemia therapies and also for the development of second generation mTOR inhibitors (the TORKinhibs).

The deregulation of translation markedly contributes to the malignant phenotype in cancers, and the assembly of the translation initiating complex eIF4F is the limiting step of this process. The mammalian Target of Rapamycin Complex 1 (mTORC1) is thought to positively regulate eIF4F assembly and subsequent oncogenic protein synthesis through 4E-BP1 phosphorylation. We showed here that the translation inhibitor 4EGI-1 decreased the clonogenic growth of leukemic progenitors and induced apoptosis of blast cells, with limited toxicity against normal hematopoiesis, which emphasize the importance of translation deregulation in acute myeloid leukemia (AML) biology. However, the mTORC1 inhibitor RAD001 (a rapamycin derivate) did not induce AML blast cell apoptosis. We herein demonstrated that mTORC1 disruption using raptor siRNA or RAD001 failed to inhibit 4E-BP1 phosphorylation in AML. Moreover, RAD001 failed to inhibit eIF4F assembly, to decrease the proportion of polysome-bound c-Myc mRNA, and to reduce the translation-dependent accumulation of oncogenic proteins. We identified the Pim-2 serine/threonine kinase as mainly responsible for 4E-BP1 phosphorylation on the S65 residue and subsequent translation control in AML. Our results strongly implicate an mTORC1-independent deregulation of oncogenic proteins synthesis in human myeloid leukemogenesis. Direct inhibition of the translation initiating complex thus represents an attractive option for the development of new therapies in AML.

The phosphatidylinositol 3-kinase (PI3K)/Akt and mammalian target of rapamycin complex 1 (mTORC1) signaling pathways are frequently activated in acute myelogenous leukemia (AML). mTORC1 inhibition with RAD001 induces PI3K/Akt activation and both pathways are activated independently, providing a rationale for dual inhibition of both pathways. PI-103 is a new potent PI3K/Akt and mTOR inhibitor. In human leukemic cell lines and in primary blast cells from AML patients, PI-103 inhibited constitutive and growth factor-induced PI3K/Akt and mTORC1 activation. PI-103 was essentially cytostatic for cell lines and induced cell cycle arrest in the G1 phase. In blast cells, PI-103 inhibited leukemic proliferation, the clonogenicity of leukemic progenitors and induced mitochondrial apoptosis, especially in the compartment containing leukemic stem cells. In contrast, apoptosis was not induced with RAD001 and IC87114 association, which specifically inhibits mTORC1 and p110delta activity, respectively. PI-103 had additive proapoptotic effects with etoposide in blast cells and in immature leukemic cells. Interestingly, PI-103 did not induce apoptosis in normal CD34(+) cells and had moderate effects on their clonogenic and proliferative properties. Here, we demonstrate that multitargeted therapy against PI3K/Akt and mTOR with PI-103 may be of therapeutic value in AML.

The phosphatidylinositol 3-kinase (PI3K)/Akt and mTORC1 pathways are frequently activated, representing potential therapeutic targets in acute myeloid leukemia (AML). In 19 AML samples with constitutive PI3K/Akt activation, the rapamycin derivative inhibitor everolimus (RAD001) increased Akt phosphorylation. This mTOR C1-mediated Akt up-regulation was explained by an insulin-like growth factor-1 (IGF-1)/IGF-1 receptor autocrine loop: (1) blast cells expressed functional IGF-1 receptors, and IGF-1-induced Akt activation was increased by RAD001, (2) a neutralizing anti-IGF-1R α-IR3 monoclonal antibody reversed the RAD001-induced Akt phosphorylation, and (3) autocrine production of IGF-1 was detected in purified blast cells by quantitative reverse transcription-polymerase chain reaction and immunofluorescence. This RAD001-induced PI3K/Akt up-regulation was due to an up-regulated expression of the IRS2 adaptor. Finally, we observed that concomitant inhibition of mTORC1 and PI3K/Akt by RAD001 and IC87114 induced additive antiproliferative effects. Our results suggest that dual inhibition of the mTORC1 complex and the IGF-1/IGF-1R/PI3K/Akt pathway in AML may enhance the efficacy of mTOR inhibitors in treatment of this disease.

Successful outcome of a disseminated adenovirus infection 6 years after allogeneic bone marrow transplantation