This retrospective single-center observational study was conducted at the Bone Marrow Transplantation Unit of Dokuz Eylül University, a tertiary referral center in İzmir, Türkiye. Adult patients (≥18 years) with a confirmed diagnosis of MM who underwent ASCT using freshly collected, non-cryopreserved (DMSO-free) PBSC between 2023 and 2025 were consecutively included. Transplant eligibility and response assessment were based on International Myeloma Working Group (IMWG) criteria, and only patients achieving at least a partial response prior to transplantation were eligible for inclusion [16].

Mobilization was performed using granulocyte colony-stimulating factor (G-CSF) at 10 µg/kg/day administered subcutaneously for five consecutive days, either alone or combined with plerixafor (0.24 mg/kg) in patients with predicted poor mobilization according to institutional practice [17]. Peripheral blood CD34⁺ counts were monitored daily starting on Day 4, and leukapheresis was initiated when circulating CD34⁺ cells exceeded 20 × 10⁶ cells/L using continuous-flow apheresis systems. The minimum target yield was ≥2 × 10⁶ CD34⁺ cells/kg. Collected grafts were stored at 4 °C in sterile transfusion bags and infused within 72 h without cryopreservation or exposure to DMSO in accordance with previously described fresh PBSC transplantation protocols [18].

High-dose melphalan was administered as the conditioning regimen at 200 mg/m² on Day −1. In patients with renal impairment (creatinine clearance <50 mL/min), the dose was reduced to 140 mg/m² according to established transplant practice guidelines [19]. PBSC, without cryopreservation or DMSO exposure, were infused on Day 0 via a central venous catheter over 15–30 min under continuous vital sign monitoring.

All patients received institutional standard prophylaxis, including antibacterial (levofloxacin), antifungal (fluconazole), and antiviral (valaciclovir) therapy consistent with international supportive care recommendations for ASCT recipients [20]. G-CSF at 5 µg/kg/day was initiated when leukocyte counts declined below 1.0 × 10⁹/L and continued until neutrophil engraftment. Patients were monitored daily for infectious complications, mucositis, gastrointestinal toxicity, and infusion-related adverse events.

Neutrophil engraftment was defined as the first of three consecutive days with an absolute neutrophil count (ANC) ≥0.5 × 10⁹/L, and platelet engraftment as the first of three consecutive days with platelet counts ≥20 × 10⁹/L without transfusion support [21]. Adverse events, including mucositis, nausea, vomiting, and infusion-related reactions, were graded using the National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE), version 5.0 [22]. Length of hospitalization was defined as the interval from transplant admission to discharge following hematopoietic recovery and resolution of major complications.

Demographic, clinical, and laboratory variables were retrospectively extracted from institutional electronic health records and the bone marrow transplantation registry. Collected variables included age, sex, disease stage, induction regimen, mobilization strategy, CD34⁺ cell yield, engraftment kinetics, toxicity incidence, and length of hospital stay.

Descriptive statistics were used to summarize baseline characteristics and outcomes. Categorical variables are expressed as frequencies and percentages, whereas continuous variables are summarized as medians with ranges. Given the limited follow-up and absence of relapse or mortality events during the observation period, time-to-event analyses such as overall survival or event-free survival could not be performed. The primary objective was to evaluate procedural safety, tolerability, engraftment kinetics, toxicity profile, and length of hospital stay associated with fresh, DMSO-free ASCT rather than long-term therapeutic efficacy.

Patient characteristics are summarized in Table 1. A total of 40 consecutive MM patients underwent ASCT using freshly collected, non‑cryopreserved PBSC. Induction therapy consisted primarily of VCD (velcade, cyclophosphamide, dexamethasone) in 27 patients (67.5%) and VRD (velcade, revlimid, dexamethasone) in 11 patients (27.5%), while two patients received alternative regimens. Prior radiotherapy exposure was rare (2.5%). Stem cell mobilization was successful in all cases, including patients previously treated with lenalidomide-based induction. Notably, none of the patients had received anti‑CD38 monoclonal antibodies therapy prior to mobilization.

At the time of transplantation, disease responses included complete response in 12 patients (30%), very good partial response in 15 (37.5%), partial response in 12 (30%), and stable disease in one patient (2.5%). According to the Revised International Staging System, 25% of patients were Stage I, 45% in Stage II, and 30% in Stage III. Heavy‑chain myeloma predominated (IgG: n = 26, IgA: n = 6, IgM: n = 1), whereas seven patients presented with light‑chain disease.

Key transplant-related variables are summarized in Table 2. The median Hematopoietic Cell Transplantation Comorbidity Index was 0 (range: 0–2), and the median Revised International Staging System Stage was I. Mobilization was performed using G‑CSF alone in 26 patients (65%) and G‑CSF plus plerixafor in 14 patients (35%). All patients received high‑dose melphalan conditioning. The median infused CD34⁺ cell dose was 6.3 × 10⁶/kg (range: 3.1–11), thereby exceeding standard minimum thresholds for autologous transplantation. Baseline hematologic parameters prior to conditioning were within clinically acceptable limits across the cohort.

Successful hematopoietic engraftment was achieved in all patients (100%). Median neutrophil recovery occurred at a median of Day 9 (range: 6–14), while platelet engraftment was observed at a median of Day 11 (range: 7–16). No cases of graft failure, delayed engraftment, or engraftment syndrome were documented. Overall engraftment kinetics appeared numerically favorable compared with historical benchmarks reported in cryopreserved ASCT series.

The DMSO‑free transplantation approach demonstrated an excellent safety profile. No infusion‑related adverse events (nausea, vomiting, bradycardia, hypotension, or acute infusion reactions) were observed during stem cell administration. Early transplant-related toxicity was limited, and no patients required intensive care unit admission. Importantly, there were no early transplant‑related deaths within the first 100 days. The median length of hospital stay was 13 days (range: 9–18), reflecting efficient hematopoietic recovery and favorable clinical tolerability.

The median time from diagnosis to ASCT was five months (range: 4–7). Patients required a median of zero red blood cell units (range: 0–3) and three platelet units (range: 2–7). Additionally, systemic antibiotics were administered to 27 patients (67.5%), with a median of one episode per patient (range: 0–2).

The use of freshly collected PBSCs eliminated the need for cryopreservation, including freezing consumables, liquid nitrogen storage, and a controlled‑rate freezing infrastructure. This simplified the transplantation workflow and reduced preparation time within the transplant unit. Operationally, fresh ASCT was associated with improved logistical efficiency while maintaining consistent clinical outcomes, supporting its potential as a streamlined and resource‑efficient transplantation strategy.

Conventional cryopreserved ASCT series typically report median neutrophil engraftment at approximately 10–12 days and platelet engraftment at 12–16 days, with inpatient stays frequently ranging from 15–20 days [14,23,24]. Moreover, infusion of DMSO-containing grafts is associated with adverse events, including nausea, vomiting, hypotension, bradycardia, and occasional cardiopulmonary complications, with infusion-related toxicity rates reaching 20%−40% in some cohorts [25,26]. In contrast, the current cohort demonstrated median neutrophil recovery at nine days, platelet engraftment at 11 days, and a median hospitalization of 13 days, suggesting numerically earlier hematopoietic recovery and streamlined post-transplant care.

Interest in fresh, non-cryopreserved ASCT has re-emerged internationally as advances in mobilization strategies and transplant coordination have enabled reliable same-cycle graft infusion [27–29]. Prior observational studies have reported comparable, or in some cases accelerated, engraftment kinetics with fresh transplantations, alongside reduced infusion-related adverse events and improved patient comfort. The findings of this study are consistent with these reports.

Data on DMSO-free ASCT from Türkiye remain limited, and most centers continue to rely on cryopreservation as standard practice [15,30]. The present cohort represents one of the most homogeneous MM populations reported nationally using a fresh transplantation strategy.

As MM increasingly resembles a chronic disease, treatment decisions are shaped not only by efficacy but also by patient experience and quality of life [2,3]. Eliminating DMSO exposure removes a major contributor to infusion-related discomfort, particularly nausea and vomiting. The rapid engraftment and shortened hospitalization observed in the present cohort suggest that, under appropriate institutional conditions, fresh ASCT may facilitate early discharge or even outpatient-based transplant models.

Cryopreservation requires controlled-rate freezing systems, liquid nitrogen storage, cryogenic consumables, and additional laboratory handling, all of which increase procedural complexity and cost [12,31]. Previous reports have suggested that fresh ASCT may reduce transplantation expenses through elimination of these requirements, with estimated cost reductions of up to 25%−30% in some analyses [32].

Several limitations should be acknowledged. The retrospective, single-center design and relatively small sample size may limit generalizability, and the absence of a contemporaneous cryopreserved control group precludes definitive comparative conclusions. Long-term outcomes, including progression-free and overall survival, were not assessed.

Taken together, these results contribute to a growing body of evidence suggesting that fresh, DMSO-free ASCT may represent a safe and efficient alternative to conventional cryopreserved transplantation. Future studies incorporating prospective comparisons, patient-reported outcomes, and health-economic analyses are essential.