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In these two short reports, the authors approach the issue of whether hydroxyurea (HU) use in young males has major irreversible effects on sperm production. Joseph et al analyzed and compared sperm parameters in male patients with sickle cell disease (SCD) who were exposed or not exposed to HU before puberty. They report semen abnormalities in all patients but no differences between groups. Independently, Gille et al provide evidence for the lack of in vivo HU-related decreases in the spermatogonial pool in biopsy specimens from young males with SCD but evidence for a negative effect of SCD itself. Together, these reports suggest that the use of HU in young males does not adversely affect fertility.

To the editor: The prognosis of sickle cell anemia (SCA), initially disastrous in the severe forms of the disease, has greatly improved, leading to increased life expectancy. More than 95% to 99% of the children diagnosed with SCA in developed countries will become young adults, reaching

Background: Sperm parameters alteration is documented in untreated men with sickle cell disease (SCD). An aggravating effect of hydroxyurea (HU) on sperm parameters is also well established: HU, at current doses, causes significant, rapid, and unpredictable impairment of spermatogenesis. Reversal of its effect when treatment is stopped has also been documented, albeit in patients given very low doses of treatment (10 mg/kg/d). In France, sperm banking is free of charge and recommended whenever possible before initiation of HU treatment. While recent guidelines have broadened the indication of HU for asymptomatic infantssuch banking is impossible in younger boys before puberty. In addition, little is known on the effect of HU on sperm parameters when given at this specific period. Objectives and Methods: The main objective of this study was to compare sperm parameters after treatment resolution in young males treated by HU prior to puberty with sperm parameters of untreated males. Secondary objective was to analyze longitudinally sperm parameters during HU washout in those treated prior to puberty.Data regarding indication of HU, dosage, date of initiation and stop, clinical profile including vaso occlusive events (VOE) transfusion episodes and date of puberty was collected. Alternative treatment before or during semen analysis was also documented. A period of 3 months of HU wash out was required prior to semen analysis in treated patients. Results: A total of 26 patients (43 semen samples) were studied, with 16 patients treated with HU prior to puberty (HU-PP) and 10 untreated (HU-naive). Characteristics of patients are presented in Table 1. Indication of HU was cerebral vasculopathy (n=3), VOE (n=5), severe anemia (n=1) or combined (n= 7). The median stopping of the HU before CECOS is 4.5 years [0.5-11.0]. An alternative treatment based on a transfusion program was initiated in 14 patients (87.5%) at the time of sperm analysis in the HU-PP group versus 5 patients (50%) in the HU-naive group. Duration of transfusion program was 126 months [3-188] in HU-PP versus 13 [7-100] in the HU naive group. We compared the fraction of abnormal values in semen samples in both groups (25 samples in the HU-PP group and 18 in the HU-naive). No significant difference was observed regarding volume of ejaculate, spermatozoid concentration, total sperm count, spermatozoid motility, morphology and vitality, and sexual abstinence before sampling in both groups (Table 1). In the HU-PP group, there was a trend in improvement of sperm parameters with the duration of transfusion program. In addition, a kinetic analysis of sperm parameters during the period of HU wash out was performed in 3 patients exposed to HU prior to puberty, demonstrating reversibility of HU toxicity in all. All 26 patients were offered semen cryopreservation. For 22 of them, semen parameters after thawing indicated a possible use for assisted reproductive technologies, mostly in vitro fertilization with intracytoplasmic sperm injection. In the remaining 4 (3 in HU-PP group and 1 in HU naïve), there was serious concern about the possible use because of a very low initial sperm concentration and the absence of motile spermatozoids after thawing. Discussion/Conclusion: Toxicity of HU upon spermatogenesis has been well documented in animal studies and in adult males. In many settings, such issues may be a drawback for parents and/or caregivers to treat young boys before semen banking can be performed, given the lack of robust information on reversibility after treatment resolution. Abnormalities of sperm count are also common in non-treated males so that demonstrating the specific effect of HU may be complex. Oseggbe et al. and Berthaud et al. showed that 91% of untreated SCD males have at least one abnormal sperm parameter. Here, we show that after treatment resolution, there are no differences in sperm parameters in patients exposed to HU before puberty compared to untreated males. Because a majority of patients benefitted from transfusion therapy at the time of analysis, we were however unable to demonstrate whether reversal of toxicity occurs spontaneously or requires transfusion. Notwithstanding possible limitations due to small sample size, this study shows that in boys with severe disease requiring HU treatment before puberty, toxicity of HU on sperm parameters should not be a major drawback. Disclosures Bernaudin: AddMedica: Honoraria, Other: Help for travel; GBT: Membership on an entity's Board of Directors or advisory committees; BlueBirdBio: Consultancy. De Montalembert:Novartis: Consultancy, Honoraria; Addmedica: Consultancy, Honoraria; Bluebird Bio: Membership on an entity's Board of Directors or advisory committees. Brousse:bluebird bio: Consultancy; Add medica: Consultancy.

The last three authors contributed equally to this work. Introduction: Allogeneic hematopoietic stem cell transplantation (HSCT) may improve long-term survival of patients with acute lymphoblastic leukemia (ALL) at high risk of relapse. However, ovarian failure is observed in 70 to 90% of patients who received myeloablative HSCT. Autotransplantation of cryopreserved ovarian cortex harvested before gonadotoxic treatments has been shown to re-establish the menstrual cycles and lead to the birth of healthy children (Donnez J. N Engl J Med, 2018). In patients treated for acute leukemia, concern has been expressed about the risk of leukemia recurrence after the ovarian autotransplantation due to the potential presence of residual leukemic cells in the ovarian tissue (Dolmans MM. Blood, 2010). To date, there were only few studies evaluating the presence of leukemic cells in cryopreserved ovarian tissue, and in most cases ovarian tissues were harvested at diagnosis of ALL or early after onset of chemotherapy. The present study prospectively investigated, in the context of fertility preservation, the presence of leukemic cells in cryopreserved ovarian samples harvested before allogeneic HSCT in patients in complete remission (CR) of ALL. Patients and Methods: From October 2015 to May 2018, all female patients with ALL who had a leukemia specific marker and underwent ovarian cryopreservation before allogeneic HSCT as part of preservation fertility program in 3 centers were included in the study. Consents were obtained from the guardians and/or age appropriate patients. The specific local ethical committees approved the study. Ovariectomy was performed after patients achieved CR, mostly in the weeks preceding HSCT. Ovarian cortex was separated into several fragments and cryopreserved as previously described (Poirot C. Human Reprod, 2002). When more than 14 cortical fragments were obtained, one was dedicated to molecular analysis, as well as the ovarian medulla. MRD quantification was performed by quantitative PCR of clonal rearrangements of immunoglobulin/T-cell receptor genes (Ig/TCR) or oncogenic fusion genes, according to EuroMRD and European Against Cancer (EAC) guidelines. MRD in ovarian samples were compared to MRD in bone marrow (BM) sample obtained at the same time point. Results: Fourteen patients were included in the study: 12 with B-cell precursor ALL, 1 with T-cell ALL and 1 with mixed phenotype acute leukemia. MRD marker was Ig/TCR rearrangement in 12 cases, M-BCR-ABL transcript in 1 case and genomic MLL-AF4 in 1 case. Median age at transplant was 18.2 years (range 1.1 - 35.2 years). Criteria for HSCT were poor early response to treatment (N=8), Philadelphia positive ALL (N=1), or previous relapse (N=5). With a median follow-up after HSCT of 8.1 months (range 0.8 - 29.8 months), all patients are disease-free. At the time of ovariectomy, 7/14 (50%) patients had undetectable MRD in ovarian samples, 6 had low positivity below 10-4 and one had positive MRD higher than 10-4. Unexpectedly of the 7 patients with positive ovarian MRD, 4 were undetectable in BM. One of the 7 patients with undetectable MRD in ovarian samples was positive in bone marrow. Concordant results were observed between the cortex and medulla samples in 10 of the 11 cases where both could be tested. Conclusions: To our knowledge, this series is the largest cohort evaluating MRD in ovarian samples from patients with ALL in complete remission after full chemotherapy regimen. We detected low levels of residual leukemic cells in ovarian tissues in half of the patients, some of them having no detectable MRD in BM at the time of ovariectomy, suggesting that leukemic cells could preferentially persist in ovary. Our results warrant further analyses of an extended cohort and longer post-transplant follow-up to better assess the potential presence of leukemic cells in ovarian samples and evaluate the impact of ovarian MRD status on post HSCT relapse. Still, autotransplantation of cryopreserved ovarian cortex could be discussed after allogeneic HSCT in patients with undetectable ovarian MRD. Table 1. MRD evaluation in bone marrow and ovarian samples. BM, bone marrow; BCP-ALL, B-cell precursor ALL; Ph-positive ALL, ALL with Philadelphia chromosome; NA: not available. Undetectable MRD means negative result obtained with a sensitivity of 10-4 or 10-5. BCR-ABL1 MRD results are expressed as the BCR-ABL1/ABL1 transcripts ratio. Disclosures No relevant conflicts of interest to declare.

Abstract 242 Introduction Hydroxyurea (HU) has proved its efficacy in reducing vaso-occlusive events in patients with SCD and increasing life expectancy. However, effects on fertility in adult males represent a major issue for acceptance and adherence, as HU has been reported to impair spermatogenesis through direct cytotoxic effects and hypogonadism. There are only limited data in the literature in this field and mainly retrospective. Patients and methods 49 adult patients aged 20–52 years with homozygous SCD eligible for first line HU treatment were enrolled after informed consent between June 2010 and April 2012 in this prospective study: HYDREP.The main objective was to compare at Day 0 and 180 of HU treatment the semen parameters according to the WHO (1999) criteria(volume, sperm concentration, viability, forward motility and morphology ). Vaso-occlusive events and other complications, number of transfusions during the 6 months preceding and following HU initiation were recorded. Hematological, biochemical, hormonal parameters, HU dosage were recorded at Day 0, Day 90 and 180. HU was begun at 15mg/Kg dose (Platt *) and modulated following haematological tolerance and renal function. Results and discussion Preliminary results concern the 24/49 patients, who complete the 6 months protocol. A statistically significant impairment in all sperm parameters was observed, after a 6 months HU therapy, with great variations between individuals, which are to be explained by complementary tests. These preliminary deleterious findings must be interpreted with caution as in our experience 1) spermatogenesis may recover after drug withdrawal 2) Repeated vaso-occlusive events are also potentially deleterious on fertility (Berthaut **)and 3)this study must be completed with a survey on effective fertility seen in men treated with HU. Thus, regarding to the great benefit offered by HU on prognosis allowing a real “life project” in young symptomatic adults with SCD, it is worth providing each patient of the most accurate information on fertility as an essential prerequisite for their adhesion to HU treatment. Conclusion Sperm cryopreservation is advised before beginning a treatment with HU. Prospective follow up studies of HU male cohort regarding fertility are warranted. Health education has a major role to play to increase HU compliance especially in view of these results. Disclosures: No relevant conflicts of interest to declare.