Abstract

Key words : Fertility, Gonadal damage, Ovarian insufficiency

Introduction

Sickle cell disease (SCD) is among the world’s most common genetic disease.¹ Recurrent vaso-occlusive crises and chronic inflammation can lead to tissue and organ damage, including damage to gonads.^2-4 For patients with SCD, both life expectancy and quality of life of patients are affected.⁵ According to the National Center for Health Statistics multiple cause of death databases, the median life expectancy is 42 years for women and 38 years for men with adult SCD.⁶ In a mortality study from the Eastern Mediterranean Region, the mean age of mortality was around 40 years.⁷

For patients with SCD, the only curative option is allogeneic hematopoietic stem cell transplant (allo-HSCT). Today, overall survival of >90% and event-free survival between 84% and 97% can be achieved in patients with adult SCD by treatment with nonmyeloablative allo-HSCT from an HLA-matched family donor.^8-13 The rate of acute and chronic graft-versus host disease (GVHD) may be reduced by <10%, and graft rejection may be overcome.^9-11 Transplant seems to improve organ function and organ damage in children.^14,15

Patients with adult SCD are susceptible to complications after transplant and to the toxic effects of drugs. Therefore, transplants are mostly done in patients at preschool age.^12,16,17 It has been reported that allo-HSCT can damage the gonads as a result of medications, radiotherapy, and complications such as GVHD.^18-20

The Baskent Organ Damage Mitigation and Medical Care Development Program for SCD (BASCARE) has worked to significantly improve age of mortality and the management of SCD, including development of electronic health recording systems in accordance with international accreditation rules and electronic vaccination schedule. The Program has also implemented the use of low citrate for erythrocyte exchange, has improved maternity care, and has introduced use of magnetic resonance imaging methods for management of transfusion-related hemosiderosis. In 2014, allo-HSCT was added to the Program.¹⁶

It is unclear how transplant procedures performed with appropriate dose-reduction regimens in adult SCD patients may affect the gonads and how the gonads are affected late after cure by allo-HSCT in adults. We speculated that gonadal damage can be prevented with dose reductions of posttransplant agents and with a medical care development program, such as BASCARE.

In this study, we aimed to determine the late effects of anti-T-lymphocyte globulin (ATLG) and cyclophosphamide-containing nonmyeloablative allo-HSCT regimen on gonads of adult patients with SCD.

Materials and Methods

Study design and patients
This was a prospective, single-center, observational study that included data from September 2013 to July 2019. All consecutive adult patients with SCD who were older than 17 years and who underwent allo-HSCT using peripheral blood stem cells from their HLA-full matched siblings were invited to participate. Because chronic GVHD may develop up to 2 years after transplant, patients who were GVHD free and not immunosuppressed at 2 years after transplant were included in the study.²¹ The study was conducted in a Joint Accreditation Committee: International Society for Cellular Therapy and European Blood and Marrow Transplantation accredited center.

To evaluate gonadal status, pre- and postprocedure menstrual cycle patterns were compared, anti-Mullerian hormone (AMH) and follicle stimulating hormone (FSH) levels were measured for ovarian reserve, and if married, the number of antral follicles were evaluated. In male participants, the quality of ejaculate was evaluated and compared with pretransplant values and testosterone was measured. According to the agreement between the transplant center and in vitro fertilization center (KIT-KY-33), participants were required to have routine preventive fertilization methods. To assess fertilization, concep-tions that occurred spontaneously or by in vitro fertilization method before and after transplant were noted.

Sexual function by transplant was measured by using the International Index of Erectile Function (IIEF) questionnaire and the Female Sex Function Index (FSFI) questionnaire. The results were compared with those that had been routinely collected before transplant.^22,23

Inclusion criteria
All consecutive adult patients (≥17 years) with homozygous or a heterozygous combination of SCD who underwent allo-HSCT from September 2013 to July 2019 and were GVHD-free for 2 years and not immunosuppressed were included in the study.

Exclusion criteria
Patients who received transplant from an unrelated donor, were not 2 years posttransplant, were using contraceptive methods, required transfusion, devel-oped chronic GVHD, were using immunosup-pressive drugs, had diabetes, or had kidney and liver failure were excluded.

Ethical approval
Local ethics committee approval was obtained from the Baskent University Clinical Research Ethics Committee (KA20/457). Written informed consent was obtained from all patients.

Measurements
Hormone analyses were performed with chemilu-minescence immunoassay method (RIQAS for AMH, Bayer Corp for FSH, Siemens Atellica reagent for testosterone). Blood samples for AMH were collected regardless of menstrual status. Antral follicle count was done by vaginal ultrasonography.²⁴

For cryopreservation, thawing, and reanalysis of ejaculate material, semen parameters (volume, concentration, motility, and morphology) were evaluated according to the World Health Organization laboratory guidelines.^25-27 The cryopreserved semen samples were carefully transferred to the liquid nitrogen tank and stored at -196 °C.

For cryopreservation and recovery of embryos, the vitrification method was used for cryopreser-vation in accordance with the European Society of Human Reproduction and Embryology guidelines.^28,29

Survey components
The IIEF form consisted of 5 questions, with scoring between 1 and 5 points. The FSFI questionnaire consisted of 19 questions and 6 subscales that included (1) sexual desire, (2) arousal, (3) lubrication, (4) orgasm, (5) satisfaction, and (6) sexual pain.²³

Within the scope of the BASCARE program, patients were followed regularly and trained about IIEF and FSFI practices.19 The survey was administered by the same family physician before and 2 years after transplant in order to prevent individual differences between practitioners.

Definitions
All patients had been diagnosed with homozygous sickle cell hemoglobin (HbS) or heterozygous combinations of HbS and thalassemia (HbS/β-thalassemia, HbS/α-thalassemia) by HbS elec-trophoresis or genetic screening. A painful crisis was defined as the need for hospital admission because of pain not related to any cause other than SCD or for the use of parenteral nonsteroidal anti-inflammatory drugs, metamizole, or narcotics. Diminished ovarian reserve was defined as an AMH level <1.07 ng/mL or an FSH >10 IU/L on day 3 of the menstrual cycle. Ovarian insufficiency was defined as amenorrhea or irregular cycles with an FSH >40 IU/L and antral follicle count <5 by transvaginal ultrasonography (if applicable).^24,30,31 Male patients with score of <26 per IIEF were defined to have erectile dysfunction, and female patients with FSFI score <26.55 were defined to have sexual dysfunction.²²

Study endpoints
Primary endpoints included incidence of amenorrhea, diminished ovarian reserve, ovarian insufficiency, and azoospermia; risk of ovarian insufficiency; risk of azoospermia; and number of spontaneous or assisted conceptions. Secondary endpoints included rate of erectile dysfunction rate in men and rate of sexual dysfunction in women.

Statistical analyses
Data were analyzed with the Statistical Package for the Social Sciences (SPSS) software, version 17. Baseline categorical variables were compared using the Fisher exact test, and continuous variables were compared using the Mann-Whitney U test or the 2-sample t test. Incidence was estimated by calculating the events per 100 persons. Comparisons between estimates of proportions were made with the chi-square test. Estimates for risk assessment (odds ratios [OR]) and corresponding 95% confidence intervals (CIs) were obtained for each significant prognostic factor. Cox regression models were constructed for use of hydroxyurea, age, or painful crises as factors influencing gonadal damage. Variables considered in the model were those significant at P = .20 from the univariable models. Variables remaining in the final models were significant at P = .05.

Results

Among the 61 patients with SCD who received allo-HSCT transplant, 49 had completed 2-yearsof follow-up and met the inclusion criteria. Of these, 43 patients (22 women, 21 men) agreed to participate.

The age range at the time of transplant was 18 to 45 years in women and 18 to 43 years in men. Six women (27%) were 40 years of age and older. Eleven women (50%) and 7 men (33%) were married. Except for divorce of 1 male and 1 female patient, the marital status of the other patients did not change after transplant. None of the patients needed hydroxyurea after transplant. Posttransplant comorbid conditions were similar to those before transplant. Three patients (7%) improved their educational status after treatment compared with pretransplant. One patient interrupted his education temporarily due to transplant (Table 1).

Menstrual cycle patterns and gonadal status in women
No incidences of amenorrhea were detected in any of the female patients undergoing allo-HSCT. None had received hormone replacement therapy during the 1 year before transplant. After transplant, amenorrhea was detected in 15 women (68%). Seven patients (32%) with no amenorrhea received regular hormonal support. In all women, AMH levels (<1 ng/mL) indicated diminished ovarian reserve at 2 years posttransplant. In 10 patients (45%), FSH levels (>40 IU/mL) revealed ovarian insufficiency. The antral follicle count values in married women supported these findings (Table 2 and Table 3).

Gonadal status in men
The incidence of male patients with normal semen analysis was significantly reduced after transplant compared with before allo-HSCT transplant (76% vs 10%; P < .001). The incidence of men with azoospermia significantly increased after transplant versus before transplant (17% vs 74%; P < .001). The incidence of male patients with cryptozoospermia before transplant was similar to that after transplant (6% vs 16%; P = .370) (Table 3 and Table 4). Testosterone level was found to be within normal limits in all male patients after transplant.

Risk of gonadal damage after transplant
Allogeneic stem cell transplant was associated with an increased risk of secondary amenorrhea (OR = 93; 95% CI, 4.94-17.50; P = .002) and ovarian insufficiency (OR = 37.8; 95% CI, 2.03 to -700.94; P = .014). Transplant was associated with a significant risk of azoospermia (OR = 4.35; 95% CI, 1.02-18.45; P = .017).

Factors impeding gonadal insufficiency
A Cox regression model that included the 22 female patients revealed that, although age was associated with increased risk of gonadal damage beyond 2 years (OR = 1.31; 95% CI, 1.04-1.64; P = .02), gonadal damage was not influenced by use of hydroxyurea (OR = 0.66; 95% CI, 0.02-18.12; P = .80) or number of painful crisis (OR = 0.19; 95% CI, 0.001-3.63; P = .14) (Table 5).

Fertility
Seven of 22 female patients (32%) underwent pretransplant fertility preservation procedures. Embryos were saved for 1 married couple, ovarian tissue was preserved in 1 patient (5%), and ovum was preserved in 3 patients (14%). Fourteen of 21 male patients (67%) had sperm preservation.

Before transplant, 5 (23%) of the married female patients had children. After transplant, spontaneous conception occurred in 1 female patient (9%); however, it resulted in a miscarriage. In another female patient, the embryo was successfully recovered after embryo preservation. She gave birth to a healthy baby girl. The other patients did not want to become pregnant (Table 2 and Table 3).

Four male patients (19%) had children. Among them, 1 male patient (14%) who developed azoospermia was able to achieve a successful in vitro fertilization 3 years posttransplant. One male patient (14%) who developed azoospermia experienced a spontaneous conception. Clinical status and gonadal reserve are presented in Table 3 and Table 4.

Sexual function by transplant
Sexual function could be evaluated in the 8 married female patients (36%). Total FSFI score after transplant was similar to that before transplant (mean [SD] score = 24 [9] vs 25 [3]; P = .20). The FSFI scores obtained for desire, arousal, lubrication, orgasm, satisfaction, and pain subscales were not different after transplant versus before transplant. The incidence of sexual dysfunction in female patients was not changed after transplant compared with before transplant (75% vs 37%; P = .13) (Table 2 and Table 3).

The total IIEF scores in male patients were similar before and after transplant (mean [SD] score = 25 [5] vs 23 [6]; P = .29). The IIEF scores for desire, organic function, sexual satisfaction, and general satisfaction items were similar to those before transplant. The incidence of IIEF score ≥26 (normal) before versus after transplant was also similar (55% vs 42%, respectively; P = .34). Although no patients had severe or moderate erectile dysfunction before transplant, severe erectile dysfunction developed in 1 male patient (5%) and moderate erectile dysfunction developed in 1 male patient (5%) after transplant (Table 3 and Table 4).

Discussion

Allogeneic stem cell transplant with low-intensity conditioning regimens are increasingly used to treat patients with SCD.^32-34 Most adult SCD patients undergoing transplant receive reduced-intensity conditioning regimens with chemotherapy or chemotherapy-free (alemtuzumab-based) protocols.^9,32-34 The ATLG and posttransplant cyclophosphamide-containing protocol (modified Johns Hopkins protocol) has been provided to adult patients with SCD after allo-HSCT as it provides a higher survival advantage (>95% overall survival and disease-free survival) and absence of graft failure or disease recurrence. This protocol also allows for successful transplants in the presence of major blood group incompatibility or erythrocyte alloimmunization.^9,10,16,34 Although the stem cell source was peripheral blood, acute GVHD and chronic GVHD have been observed in only a few cases, and permanent complete chimerism has been achieved in almost all adult patients with SCD. Therefore, more than half of patients can recover from immunosuppressive therapy within 1 year.^10,11,16

Experimental and clinical studies have demon-strated the deleterious effects of ablative peripheral blood stem cell transplant on gonadal reserve in both female and male patients.^24,33-35 In the European Society for Medical Oncology guideline, cyclophos-phamide, busulfan, and fludarabine are stated as high-risk chemotherapy agents in terms of azoospermia and male infertility risk.²⁸ A limited number of studies are available in the literature investigating fertility in pediatric SCD patients after transplant.^9,24,36 It is unknown whether reduc-tion of chemotherapy can overcome the harmful effects to gonads in patients with adult SCD after transplant.

Adult patients with SCD are prone to gonadal damage as a result of inflammation, microvascular ischemia, and, potentially, treatment medications.^5,24,36 Studies have found that 24% of male patients develop hypogonadism characterized by erectile dysfunction, sperm abnormalities, and low libido.^3,4 Although these deleterious effects are less pronounced in women, female patients are considered to be at high risk for transplant-related amenorrhea.²⁴

To prevent toxicity, busulfan has been reduced at up to 25% of the ablative dose, the total body irradiation has been administered at a low dose (200 cGy), and posttransplant cyclophosphamide has been reduced by 30%.^10,11,16,35 Our study is the first to evaluate the effects of a reduced dose of posttransplant cyclophosphamide with regard to gonadal toxicity.

In this study, although our premenopausal female patients did not have amenorrhea before transplant, approximately two-thirds seemed to be affected by the transplant protocol. In some patients who did not have amenorrhea after transplant, amenorrhea appeared later and the ovarian reserve decreased. In the literature, there is evidence supporting that menstruation can be restored with hormone replacement therapy in patients who underwent nonmyeloablative allo-HSCT.^24,37 However, despite restoration of menstruation, it was found that all women in our study had diminished ovarian reserve in the late posttransplant period, and nearly half of our female patients had ovarian insufficiency. That is, even with improved patient care after transplant and dose reduction, ovarian insufficiency could not be prevented. We also found that age seems to be important in the emergence of this effect. In 1 study, 110 of 708 postpubertal women without SCD recovered normal ovarian function and 32 became pregnant.³⁸ However, our data demonstrated that adult female patients with SCD who underwent transplant and received ATLG and a cyclophos-phamide containing regimen after transplant were likely to have an increased risk of ovarian insufficiency.

Sperm abnormalities, including low sperm count, morphology defects, poor motility, and low sperm density, are commonly shown in patients with SCD (up to 91%).^33-37 Alkylating agents may damage germinal epithelial cells and weaken spermatogenesis, and sperm abnormalities may develop as a result of testicular vaso-occlusion.³¹ Hydroxyurea and sirolimus are other possible factors.^31,33,39 Discontinuation of these agents has been shown to improve sperm parameters in rats and humans.³⁹ In our study, pretransplant sperm analysis was normal in approximately two-thirds of our male patients. Although patients had absence of SCD and GVHD, sirolimus had been discontinued for at least 1 year, hydroxyurea was not used, and patients received improved standard medical care, only one-tenth of men in our study had normal spermiogram values after transplant. Dose reduction and using a standard two-sided clam shell scrotal shield did not prevent testicular injury. Interestingly, testosterone production does not seem to be affected after transplant. This situation has been interpreted in the literature as the continued activity of Leydig cells after transplant.³¹

Before transplant, female patients with SCD can become pregnant; although the pregnancy process is risky, these patients can have a healthy birth with adequate medical support.³⁴ For patients who achieve cure from SCD after transplant, to maintain these chances, pretransplant embryo or sperm cryopreservation procedures should be performed. In married female patients, transplant can be delayed until conception occurs. In our present study, we reported that these methods were successfully applied after transplant. On the other hand, the occurrence of spontaneous conceptions in both female and male patients strengthened the claim that fertility can be possible after reduced-dose transplants.

About 21% to 35% of male patients with SCD may have erectile dysfunction probably due to recurrent priapism.^24,40 In cases where extensive penile fibrosis has not developed, the course of this defect after transplant is not clear.^24,41 In the present study, some male patients had erectile dysfunction before transplant. Eradication of SCD did not change the rate of erectile dysfunction. This can be explained by nonorganic (mostly psychogenic) factors such as concern that the disease may return, in addition to organic problems. In women, sexual dysfunction was not influenced by transplant. However, although there was no significant difference before and after transplant, there was an increasing trend in female recipients with sexual dysfunction after transplant. We suggest that ovarian insufficiency may contribute to the development of sexual dysfunction after transplant.

The present study makes 2 significant contri-butions to the literature: reporting gonadal status (including functionality) from a vulnerable group of SCD patients with a relatively high mean age and detailing results obtained in the late period after transplant. Strengths of this study are that our SCD population represented a homogeneous group of the same ethnicity, reproductive age, conditioning regimen, and GVHD prophylaxis. Because chronic GVHD can develop within 2 years after treatment, pregnancy was allowed 2 years after transplant. In addition, only patients without transplant complications were included in the study. Another important factor is that the patients were under standard and advanced medical care (BASCARE protocol). We suggest that the number of patients in the study was also notable, as the number of allo-HSCT procedures performed in adult patients with SCD is limited (around 400 in the literature).

Several limitations of our study should be discussed. First, the small number of married women created difficulties in evaluation of sexual function according to their own culture, and the results may not reflect the overall results. Second, the lack of an interim evaluation during the early posttransplant period may be a limitation in the follow-up of gonadal reserves. Finally, detailed hormonal data were not collected during the pretransplant period; these data may be helpful to explain the mechanisms.

Conclusions

Despite improved medical care and a rational dose reduction of drugs, including cyclophosphamide, our current nonmyeloablative allo-HSCT and ATLG and cyclophosphamide-containing regimen posttransplant created a risk of gonadal insufficiency in patients with adult SCD whose disease was eradicated after transplant. Nevertheless, spontaneous conception was possible in both men and women. Risk of gonadal damage was associated with age in female patients. Our findings emphasized the need for fertility preservation methods before transplant. Decision-making at the time of transplant in adult patients with SCD is difficult due to disease-specific risks, and transplant should be postponed until the embryo forms and is preserved in married female patients.

References:

1. Ware WE, de Montalembert M, Tshilolo L, Abboud MR. Sickle cell disease. Lancet. 2017;390(10091):311-323. doi:10.1016/S0140-6736(17) 30193-9
   CrossRef - PubMed
   
2. Sachdev V, Tian X, Gu Y, et al. A phenotypic risk score for predicting mortality in sickle cell disease. Br J Haematol. 2021;192(5):932-941. doi:10.1111/bjh.17342
   CrossRef - PubMed
   
3. Taddase A, Woldie IL, Khana P, et al. Hypogonadism in patients with sickle cell disease; central or peripheral? Acta Haematol. 2012;128(2):65-68. doi:10.1159/000337344
   CrossRef - PubMed
   
4. Smith-Whitley K. Reproductive issues in sickle cell disease. Blood. 2014;124(24):3538-3543. doi:10.1182/blood-2014-07-577619
   CrossRef - PubMed
   
5. Shafrin J, Thom HHZ, Keeney E, et al. The impact of vaso-occlusive crises and disease severity on quality of life and productivity among patients with sickle cell disease in the US. Curr Med Res Opin. 2021;37(5):761-768. doi:10.1080/03007995.2021.1897556
   CrossRef - PubMed
   
6. Lanzkron S, Carroll CP, Haywood C Jr. Mortality rates and age at death from sickle cell disease: US, 1979-2005. Public Health Rep. 2013;128(2):110-116. doi:10.1177/003335491312800206
   CrossRef - PubMed
   
7. Karacaoglu PK, Asma S, Korur A, et al. East Mediterranean region sickle cell disease mortality trial: retrospective multicenter cohort analysis of 735 patients. Ann Hematol. 2016;95(6):993-1000. doi:10.1007/s00277-016-2655-5
   CrossRef - PubMed
   
8. Alzahrani M, Damlaj M, Jeffries N, et al. Non-myeloablative human leukocyte antigen-matched related donor transplantation in sickle cell disease: outcomes from three independent centres. Br J Haematol. 2021;192(4):761-768. doi:10.1111/bjh.17311
   CrossRef - PubMed
   
9. Saraf SL, Oh AL, Patel PR, et al. Nonmyeloablative stem cell transplantation with alemtuzumab/ low-dose irradiation to cure and improve the quality of life of adults with sickle cell disease. Biol Blood Marrow Transplant. 2016;22:441-448. doi:10.1016/j.bbmt.2015.08.036
   CrossRef - PubMed
   
10. Ozdogu H, Boga C, Yeral M, et al. Allogenic peripheral stem cell transplantation from HLA-matched related donors for adult sickle cell disease: remarkable outcomes from a single-center trial. Bone Marrow Transplant. 2018;53(7):880-890. doi:10.1038/s41409-018-0111-y
    CrossRef - PubMed
    
11. Ozdogu H, Boga C, Yeral M, et al. Excellent outcomes of allogeneic transplantation from peripheral blood of HLA-matched related donors for adult sickle cell disease with ATLG and posttransplant cyclophosphamide-containing regimen: an update work. Bone Marrow Transplant. 2020 55(8):1647-1651. doi:10.1038/s41409-020-0805-9
    CrossRef - PubMed
    
12. Gluckman E, Cappelli B, Bernaudin F, et al. Sickle cell disease: an international survey of results of HLA-identical sibling hematopoietic stem cell transplantation. Blood. 2017;129:1548-1556. doi:10.1182/blood-2016-10-745711
    CrossRef - PubMed
    
13. Eapen M, Brazauskas R, Walters MC, et al. Impact of donor type and conditioning regimen intensity on allogeneic transplantation outcomes in patients with sickle cell disease: a retrospective, cohort study. Lancet Haematol. 2019;6(11):e585-e596. doi:10.1016/S2352-3026(19)30154-1
    CrossRef - PubMed
    
14. Friedman D, Dozor AJ, Milner J, et al. Stable to improved cardiac and pulmonary function in children with high-risk sickle cell disease following haploidentical stem cell transplantation. Bone Marrow Transplant. 2021;56(9):2221-2230. doi:10.1038/s41409-021-01298-7
    CrossRef - PubMed
    
15. Pittman C, Hsieh MM, Coles W, Tisdale JF, Weir NA, Fitzhugh CD. Reversal of pre-capillary pulmonary hypertension in a patient with sickle cell anemia who underwent haploidentical peripheral blood stem cell transplantation. Bone Marrow Transplant. 2017;52(4):641-642. doi:10.1038/bmt.2016.335
    CrossRef - PubMed
    
16. Ozdogu H, Boga C, Asma S, et al. Organ damage mitigation with the Baskent Sickle Cell Medical Care Development Program (BASCARE). Medicine (Baltimore). 2018;97(6):e9844. doi:10.1097/MD.0000000000009844
    CrossRef - PubMed
    
17. Angelucci E, Matthes-Martin S, Baronciani D, et al. Hematopoietic stem cell transplantation in thalassemia major and sickle cell disease: indications and management recommendations from an International Expert Panel. Haematologica. 2014;99:811-820. doi:10.3324/haematol.2013.099747
    CrossRef - PubMed
    
18. Zavattaro M, Felicetti F, Faraci D, et al. Impact of allogeneic stem cell transplantation on testicular and sexual function. Transplant Cell Ther. 2021;27:182.e1-182.e8. doi:10.1016/j.jtct.2020.10.020
    CrossRef - PubMed
    
19. Guida M, Castaldi AM, Rosamillio M, Giudice V, Orio F, Selleri C. Reproductive issues in patients undergoing hematopoietic stem cell transplantation: an update. J Ovarian Res. 2016;9:72. doi:10.1186/s13048-016-0279-y
    CrossRef - PubMed
    
20. Tauchmanovà L, Selleri C, De Rosa G, et al. Gonadal status in reproductive age women after haematopoietic stem cell transplantation for haematological malignancies. Hum Reprod. 2003;18(7):1410-1416. doi:10.1093/humrep/deg295
    CrossRef - PubMed
    
21. Tirri BF, Häusermann P, Bertz H, et al. Clinical guidelines for gynecologic care after hematopoietic SCT. Report from the international consensus project on clinical practice in chronic GVHD. Bone Marrow Transplant. 2015;50(1):3-9. doi:10.1038/bmt.2014.242
    CrossRef - PubMed
    
22. Turunç T, Deveci S, Güvel S, Pe?kircioglu L. The assessment of Turkish validation with 5 question version of International Index of Erectile Function (IIEF-5). Türk Üroloji Dergisi. 2007:33(1):45-49.
    CrossRef - PubMed
    
23. Öksüz E, Malhan S. The assessment of Turkish validation with 5 question version of International Index of Erectile Function (IIEF-5). Sendrom. 2005(1);54-59. https://app.trdizin.gov.tr/
    CrossRef - PubMed
    
24. Elchuri SV, Lewis RV, Quarmyne M, Haight AE, Cottrell HN, Meacham LR. Longitudinal description of gonadal function in sickle-cell patients treated with hematopoietic stem cell transplant using alkylator-based conditioning regimens. J Pediatr Hematol Oncol. 2020;42:e575-e582. doi:10.1097/MPH.0000000000001782
    CrossRef - PubMed
    
25. World Health Organization. WHO Laboratory Manual for the Examination and Processing of Human Semen. Geneva: World Health Organization; 2010. https://apps.who.int/
    CrossRef - PubMed
    
26. Chung K, Donnez J, Ginsburg E, Meirow D. Emergency IVF versus ovarian tissue cryopreservation: decision making in fertility preservation for female cancer patients. Fertil Steril. 2013; 99(6):1534-1542. doi:10.1016/j.fertnstert.2012.11.057
    CrossRef - PubMed
    
27. Roness H, Kalich-Philosoph L, Meirow D. Prevention of chemotherapy-induced ovarian damage: possible roles for hormonal and non-hormonal attenuating agents. Hum Reprod Update. 2014; 20(5):759-74. doi:10.1093/humupd/dmu019
    CrossRef - PubMed
    
28. Lambertini M, Peccatori A, Demeestere I, Meirow D. Fertility preservation and posttreatment pregnancies in post-pubertal cancer patients: ESMO Clinical Practice Guidelines. Ann Oncol. 2020;31(12):1664-1678. doi:10.1016/j.annonc.2020.09.006
    CrossRef - PubMed
    
29. Anderson RA, Amant F, Braat D, et al. ESHRE guideline: female fertility preservation. Hum Reprod Open. 2020;2020(4):hoaa052. doi:10.1093/hropen/hoaa052
    CrossRef - PubMed
    
30. Lambertini M, Del Mastro L, Pescio MC, et al. Cancer and fertility preservation: international recommendations from an expert meeting. BMC Med. 2016;14:1. doi:10.1186/s12916-015-0545-7
    CrossRef - PubMed
    
31. Randolph J, Sowers M, Gold EB, et al. Reproductive hormones in the early menopausal transition: relationship to ethnicity, body size, and menopausal status. J Clin Endocrinol Metabol. 2003;88(4):1516-1522. doi:10.1210/jc.2002-020777
    CrossRef - PubMed
    
32. Hsieh MM, Fitzhugh CD, Weitzel RP, et al. Nonmyeloablative HLA-matched sibling allogeneic hematopoietic stem cell transplantation for severe sickle cell phenotype. JAMA. 2014;312:48-56. doi:10.1001/jama.2014.7192
    CrossRef - PubMed
    
33. Hsieh MM. A standard nonmyeloablative transplantation regimen for adults with sickle cell disease: are we there yet? Biol Blood Marrow Transplant. 2016;22:397-398. doi:10.1016/j.bbmt.2015.12.002
    CrossRef - PubMed
    
34. Guilcher GMT, Truong TH, Saraf SL, Joseph JJ, Rondelli D, Hsieh MM. Curative therapies: allogeneic hematopoietic cell transplantation from matched related donors using myeloablative, reduced intensity, and nonmyeloablative conditioning in sickle cell disease. Semin Hematol. 2018;55(2):87-93. doi:10.1053/j.seminhematol.2018.04.011.3
    CrossRef - PubMed
    
35. Ozdogu H, Boga C. Hematopoietic stem cell transplantation in adult sickle cell disease: problems and solutions. Turk J Haematol. 2015;32(3):195-205. doi:10.4274/tjh.2014.0311
    CrossRef - PubMed
    
36. Berhaut I, Guignedoux G, Kirch-Noir F, et al. Influence of sickle cell disease and treatment with hydroxyurea on sperm parameters and fertility of human males. Haematologica. 2008;93(7):988-993. doi:10.3324/haematol.11515
    CrossRef - PubMed
    
37. Gharvan H, Neary NM, Link M, et al. Successful fertility restoration after allogeneic haematopoietic stem cell transplantation. Endocr Pract. 2014;20(9):e157-e161. doi:10.4158/EP13474.CR
    CrossRef - PubMed
    
38. Sanders JE, Hawley J, Levy W, et al. Pregnancies following high-dose cyclophosphamide with or without high-dose busulfan or total-body irradiation and bone marrow transplantation. Blood. 1996;87(7):3045-3052.
    CrossRef - PubMed
    
39. Zuber J, Anglicheau D, Elie C, et al. Sirolimus may reduce fertility in male renal transplant recipients. Am J Transplant. 2008;8:1471-1479. doi:10.1111/j.1600-6143.2008.02267.x
    CrossRef - PubMed
    
40. Lukusa AK, Vermylen C, Vanabelle B, et al. Bone marrow transplantation or hydroxyurea for sickle cell anemia: long-term effects on semen variables and hormone profiles. Pediatr Hematol Oncol. 2009;26(4):186-194. doi:10.1080/07357900902892780
    CrossRef - PubMed
    
41. Jacob A, Barker H, Goodman A, Holmes J. Recovery of spermatogenesis following bone marrow transplantation. Bone Marrow Transplant. 1998;22(3):277-279. doi:10.1038/sj.bmt.1701332
    CrossRef - PubMed