Sickle cell disease (SCD) is a hematological disorder caused by a single-gene mutation in which valine replaces glutamic acid at the sixth position of the β-globin chain, leading to sickle hemoglobin (HbS) formation. Under low-oxygen conditions, red blood cells (RBCs) become rigid and sickle-shaped and obstruct the microvasculature, resulting in vaso-occlusive crises (VOCs) and increased cellular adhesiveness. The clinical course of SCD varies significantly, with complications, including chronic hemolysis and end-organ damage affecting the lungs, kidneys, liver, and brain. Homozygosity for the HbS mutation (HbSS) causes sickle cell anemia (SCA), the most severe form of SCD, while compound heterozygous states (HbSC, HbSE, HbSβ⁰) can produce similar symptoms [1]. SCA occurs when an individual inherits two copies of the mutated β-globin gene (HBB), one from each parent, leading to the exclusive production of HbS instead of normal hemoglobin (Hb) A. The resulting sickle-shaped RBCs cause vaso-occlusion, hemolysis, and various complications associated with SCD.

Globally, SCD has a high incidence, particularly in sub-Saharan Africa, where prevalence ranges from 1 to 2% [2]. India, the second most affected country, reported an estimated 42,016 SCA births in 2010, reflecting a rising prevalence of up to 4.3% in various populations, including indigenous tribes [3].

Available treatment options for HbSS include disease-modifying drugs targeting several pathophysiological pathways and definitive therapies, such as gene therapy and hematopoietic stem cell transplantation [4]. Hydroxyurea (HU), a fetal hemoglobin (HbF) inducer, has been a cornerstone of SCD management since its Food and Drug Administration approval in 1998, demonstrating its efficacy in reducing pain episodes, acute chest syndrome, and mortality [5, 6]. Recently, novel therapeutic agents, such as voxelotor, L-glutamine, and crizanlizumab, have expanded the treatment landscape by targeting different aspects of SCA pathobiology [7].

Thalidomide and its analogs are emerging as promising agents for treating HbSS because of their diverse mechanisms of action, including induction of HbF, anti-inflammatory properties, and immunomodulatory effects. It inhibits tumor necrosis factor-alpha and activates the p38 mitogen-activated protein kinase pathway, stimulating HbF production [8, 9].

We evaluated the efficacy and safety of thalidomide in combination with HU compared with HU alone in patients with SCA. The frequency of VOCs, need for transfusions, variations in hematological parameters, and side effects between treatment groups were assessed to determine whether combination therapy provides superior clinical benefit and safety profile in managing SCA (HbSS).

An open-label quasi-experimental clinical trial was conducted at the Department of Hematology and Hemato-Oncology between January 2022 and May 2023. Male patients of 12 years and above and postmenopausal females of 45 years and older were enrolled in the trial. Hemoglobin electrophoresis (MINICAP SEBIA FLEX-PIERCING capillary electrophoresis system) was performed to confirm the diagnosis of SCA. The study included two groups of participants: Group A received HU (20 mg/kg/day), thalidomide (50 mg/day), and folic acid (5 mg/day), while Group B received HU (20 mg/kg/day) and folic acid (5 mg/day). The exclusion criteria included females of childbearing age (15–45 years); significant liver, renal, cardiac, pulmonary, or neurological conditions; a history of thrombotic episodes; active human immunodeficiency virus infection; or hepatitis B or C infections.

All patients were administered a fixed low dose of HU at 20 mg/kg/day, along with supportive care. Since HU is available in India only in a 500 mg capsule formulation, the daily dosage was adjusted as necessary to maintain an equivalent dose of 20 mg/kg/day for each patient. The patients were also advised to consume folic acid (5 mg/day) and ensure adequate hydration.

The clinical responses and laboratory parameters were monitored from the initiation of HU therapy to each follow-up visit over 16 months. Monthly assessments were performed using a standardized clinical protocol. Laboratory evaluations at baseline and follow-up included Hb, mean corpuscular volume (MCV), HbF, reticulocyte count, and serum creatinine. Additional data collected included adverse effects, consanguinity, patient age, and sex. The frequency of VOCs and blood transfusions in the previous year was compared with post-treatment values. Baseline clinical and laboratory parameters (Hb, HbS, HbF, and MCV) were analyzed after 16 months of low-fixed-dose HU therapy. Patient adherence was assessed by counting the remaining HU capsules during follow-up visits.

As per the enrollment, allocation, follow-up, and analysis of participants in the clinical trial, 91 patients were initially approached, of whom 8 declined to participate, leaving 83 assessed for eligibility. Among them, 8 were excluded because they did not meet the eligibility criteria, resulting in 75 enrolled patients. These participants were randomly assigned to two groups: Group A (n = 36) received the intervention, and Group B (n = 39) served as the control group. During the follow-up period, 3 patients from Group A and 6 from Group B were lost to follow-up, leading to the final analyzed samples of 33 patients in each group (Fig. 1).

Based on the order in which they were presented, patients were enrolled in either group based on their willingness to participate. This study did not employ blinding or treatment concealment. Using the G Power 3.1.9.4 program, the sample size calculation indicated that 66 patients, or at least 30 in each group, were required to attain adequate statistical power.

At the start of the trial, each participant underwent comprehensive clinical and laboratory evaluations, followed by periodic follow-up assessments at 3, 6, 9, and 12 months. Parameters, including hemoglobin electrophoresis, renal and liver function tests, complete blood counts, frequency of VOCs, need for blood transfusions, and safety profile, were monitored and compared over time.

Pain intensity was assessed using the Numeric Rating Scale, while adverse events were monitored following the Common Terminology Criteria for Adverse Events Version 5.0 [10, 11]. The five primary outcome variables were Hb increment, reduction in VOCs, decreased transfusion requirement, decreased HbS%, and increased HbF%. Transfusion independence, defined as a reduction in the need for transfusion, was used as the response criterion. Patients were categorized as major responders if their hemoglobin increased by ≥ 2 g/dL, minor responders if it increased by 1–2 g/dL, and non-responders if there was no significant change [12].

The choice of a quasi-experimental study design was driven by several practical and ethical considerations. First, we included patients with SCA (HbSS), a vulnerable population with significant disease-related challenges. Randomization was deemed impractical as it could limit patient autonomy and adherence to treatment regimens. Instead, the participants were enrolled in treatment groups based on their willingness, reflecting real-world clinical scenarios and optimizing compliance. Second, distinct therapeutic agents (HU and thalidomide) made blinding and treatment concealment infeasible, reinforcing the suitability of a quasi-experimental design. Additionally, recruiting a sufficient number of participants for a fully randomized controlled trial in a single-center setting posed logistical constraints, particularly given the rarity of SCA (HbSS) and its complex clinical management. This approach allowed us to evaluate the comparative effectiveness of the interventions in routine clinical practice while ensuring ethical standards, feasibility, and resource optimization. These factors collectively justified the adoption of a quasi-experimental design for this investigation. Ethical clearance for the study was obtained from the institutional ethics committee (ref. no. IEC/IMS.SH/SOA/2021/270 dated December 27, 2021).

IBM SPSS Statistical Software v27.0 was employed for the data analysis. Continuous variables are expressed as mean and standard deviation. Frequency distribution and Chi-square tests were applied to analyze categorical variables, and paired/independent sample t-tests were used to evaluate continuous variables. Repeated-measures analysis of variance was used to assess the changes across the observation period of 12 months, with the F-value calculated using Wilks’ Lambda test. Mauchly’s Test of Sphericity was used to test the assumption of sphericity, and when violated, a Greenhouse–Geisser correction was used. A p-value of < 0.05 was used to signify statistical significance.

This study included 66 patients with SCA (HbSS). Figure 1 shows a schematic representation of the patient enrollment. The average age was 32.9 ± 11.5 years. Most (56.1%) patients were in the 21–40 age group, with males constituting 57.6%. Table 1 shows the baseline characteristics of the study participants, including their clinical and biochemical parameters. The baseline packed red blood cell (PRBC) requirement/year (5.27 vs. 5.03) and VOC episodes/year (11.18 vs. 10.79) between the groups before enrollment were statistically comparable (p = 0.467 and 0.501, respectively), although hemoglobin, percentage of reticulocytes, and HbF, among other hematological parameters, were not comparable.

Figures 2 and 3 show comparisons between the two intervention arms. The 12-month treatment regimen with thalidomide in Group A patients had a statistically significant effect on all five outcome measures compared with their counterparts in Group B.

At 12 months, the mean Hb level in Group A patients increased significantly from 8.2 ± 1.8 g/dL to 11.8 ± 1.2 g/dL (p < 0.0001), while no significant improvement was observed in Group B (p = 0.726) (Fig. 2). A significant reduction in mean HbS% was noted in Group A, decreasing from 72.5 ± 5.5% to 64.5 ± 5.4% (p < 0.0001). In contrast, Group B showed a smaller yet significant decline from 73.3 ± 7.7% to 69.9 ± 6.2% (Fig. 4). HbF levels in Group A increased progressively from 18.9 ± 5.1% to 28.4 ± 5.6% (p < 0.0001), while no statistically significant increase was observed in Group B (p = 0.115) (Fig. 5). Additionally, patients in Group A required significantly fewer PRBC transfusions over 12 months, with a mean of 3.61 ± 2.19 transfusions (Fig. 6).

Table 2 describes the changes in blood parameters across the timeline at 3, 6, 9, and 12 months of treatment compared with those at baseline. At 12 months, the mean Hb level in patients in Group A increased steadily from 8.2 ± 1.8 g/dL to 11.8 ± 1.2 g/dL (p < 0.0001), while in Group B, no significant improvement was observed (p = 0.726). A steady decrease in mean HbS% from the baseline values of 72.5 ± 5.5 to 64.5 ± 5.4 in 12 months in Group A (p < 0.0001) was seen. Similarly, in patients of Group B, a significant reduction in HbS% was observed, albeit to a lesser extent (73.3 ± 7.7 to 69.9 ± 6.2). After treatment, Group A patients’ HbF values increased gradually, from 18.9 ± 5.1 to 28.4 ± 5.6 (p < 0.0001) in 12 months. Group B patients did not show a statistically significant increase in HbF values (p < 0.115).

Patients in Group A had significantly fewer episodes of VOCs (3.48 ± 2.81) and PRBC transfusions (3.61 ± 2.19) over 12 months than VOCs (11.36 ± 4.20) and PRBC transfusions (13.27 ± 3.70) in Group B (p = 0.0001 and 0.0001, respectively).

Furthermore, we observed that approximately 85% of the patients in Group A were major responders, exhibiting an increase in hemoglobin of ≥ 2 g/dL, compared with only 15% in Group B, as shown in Table 3. The mean white blood cell (WBC) count was 8.5 ± 4.1 × 103/µL at baseline, which increased to 12.1 ± 15.4 × 103/µL at 3 months and subsequently declined to 7.0 ± 2.5 × 103/µL at 12 months. The mean WBC counts at 6, 9, and 12 months were significantly lower than the baseline values (p < 0.05). The mean difference in WBC count between baseline and 12 months was -1.5 ± 3.6 in Group A and -2.1 ± 3.8 in Group B (p = 0.561).

The safety profile of the drugs was assessed over the treatment period by serial monitoring of renal and liver function tests, and there were no significant differences between the groups (Table 1). Constipation (grade 1) (n = 19, 57.6%) and somnolence (grade 1) (n = 6, 18.2%) were the considerable side effects in Group A, while nausea (grade 1) (n = 11, 33%) was the predominant adverse effect in Group B. Only two patients in Group A (n = 2, 6%) had grade 1 peripheral neuropathy.

Management of SCA poses a significant challenge because of its chronic and often debilitating nature. HU has been established as a cornerstone in SCA treatment by increasing HbF levels, thereby reducing HbS polymerization and associated complications. However, not all patients respond adequately to HU, necessitating the exploration of adjunctive therapies to optimize outcomes [13]. Several studies have demonstrated the effectiveness of HU as an HbF inducer [14, 15–16].

After a 6-month trial on mean therapeutic dose, HU failure is defined as the inability to reduce the frequency and severity of painful episodes or acute chest syndrome [17]. The ability to tolerate moderate HU doses with little myelotoxicity depends on the “bone marrow reserve,” which, if reduced, leads to HU intolerance even at moderate doses [18]. Recently, thalidomide, which is known to induce immunomodulatory and anti-inflammatory effects in combination with HU, has been studied in vitro for SCA with good outcomes [9].

However, to our knowledge, few studies on patients with SCA have compared the effectiveness and safety of this combination therapy (HU + thalidomide). In this quasi-experimental clinical trial, we investigated the combination of thalidomide with HU compared with HU alone in patients with SCA to evaluate whether adding thalidomide to standard HU therapy could offer better clinical benefits than HU alone.

This study provides preliminary evidence supporting the efficacy and safety of combining thalidomide with HU for SCA treatment. Combination therapy is a promising therapeutic option for patients who do not achieve optimal outcomes with HU alone, highlighting the need for further research to refine and optimize treatment strategies for SCA management.