The two patients, a girl aged 3 years and 5 months and a 3-year-old boy, had both experienced repeated and prolonged infections despite normal findings on standard immunological workup. The girl had suffered multiple respiratory tract infections, including nasopharyngitis, bronchiolitis, angina, and a confirmed respiratory syncytial virus infection that required hospitalization, along with several extended courses of antibiotics. Her background also included suspected adenoid hypertrophy and signs of non-specific respiratory allergy. Laboratory assessment showed normal levels of serum immunoglobulins IgG, IgM, and IgA, as well as normal complement components C3 and C4. The boy presented with recurrent respiratory and gastrointestinal infections, including bronchitis, otitis media, pneumonia requiring hospitalization, and an episode of acute appendicitis treated surgically. He too had received multiple antibiotic treatments, and laboratory evaluation confirmed normal humoral immune function, with no comorbidities identified.

Importantly, both children had previously received or were concurrently receiving other immunomodulatory interventions, specifically oral bacterial lysates and beta-glucans, as part of their preventive care. The authors note that these therapies could have influenced baseline immune status and must be considered when interpreting any immunophenotypic changes observed during the study. The girl’s broader regimen included bacterial lysates and beta-glucans administered for 10 and 20 consecutive days every 3 months, along with IAD at a dose of 10 mL per day for 5 days followed by a 2-day break, repeated over a 3-week cycle. During one acute infection episode, she was additionally treated with IAD at 1 mL/kg per day for 7 days, and her follow-up period lasted 10 months. The boy followed the same preventive schedule with bacterial lysates and beta-glucans, but received IAD only once during an acute infection episode, at 1 mL/kg daily for 7 days, with a 6-month follow-up.

IAD itself is a synthetic immunomodulating agent composed of a 3:1 molar complex of p-acetamidobenzoate dimethylaminopropanol and inosine. It has been used clinically since 1971 in more than 70 countries for managing viral infections, including those caused by herpes simplex virus, varicella-zoster virus, Epstein-Barr virus, and cytomegalovirus, as well as human papillomavirus-related conditions, subacute sclerosing panencephalitis, acute respiratory infections, and immunocompromised states. Five decades of clinical experience support a favorable safety profile, with transient hyperuricemia attributable to inosine metabolism as the most frequently observed adverse effect, alongside rare instances of dose-dependent nausea. The compound exhibits dual antiviral and immunomodulatory activities, although its precise mechanism remains incompletely characterized. Three key immunomodulatory effects have been described: augmented T-cell proliferation, upregulated cytokine secretion particularly of interleukin-2, and altered lymphocyte activation and memory profiles.

For the immunophenotyping, peripheral venous blood samples were collected in K2EDTA vacutainers and leukocytes were stained with fluorochrome-conjugated monoclonal antibodies targeting specific surface markers. Samples underwent either a lysis-and-wash protocol or direct lysis without washing when absolute cell counts were needed, using BD TruCount tubes. Analysis was performed on a BD FACSLyric system with data acquired and interpreted using BD FACSuite software. A four-tube antibody panel allowed the team to identify the main lymphocyte subpopulations as percentages and absolute counts, to separate naïve, central memory, effector memory, and terminal effector T-cell subsets based on CD45RA, CCR7, and CD27 expression, to distinguish naïve from memory B-cells using CD19, CD27, IgD, and IgM, and to measure activation markers including CD38, CD25, and HLA-DR on T- and B-lymphocytes.

The baseline analysis revealed distinct immune profiles in the two cases, reflecting individual variability and prior immunomodulatory therapy. Both children exhibited absolute lymphocytosis, but with different lineage predominance: in the girl, the increase was mainly due to B-cells, whereas in the boy it was primarily attributable to T-cells. The girl presented with a markedly elevated percentage of CD19+ B-cells at 37% at admission, while her T-cell percentages were low, with 49% CD3+, 30% CD4+, and 16% CD8+ cells. The boy showed the opposite pattern, with T-cells and their CD4+ and CD8+ subsets increasing further after therapy to 74%, 44%, and 23% of lymphocytes respectively, along with a decrease in CD19+ B-cells to 20%. His absolute counts after therapy reached 4496 cells/µL for CD3+ T-cells, 2700 cells/µL for CD4+ cells, and 1381 cells/µL for CD8+ cells, while his initially elevated B-cell counts decreased following IAD therapy toward the reference value of 1211 cells/µL.

Subset-level analysis added further detail. In the girl, the proportion of naïve CD4+ T-cells, defined as CD197+CD45RA+CD27+, increased following IAD treatment to 71%, while memory CD45RA− T-cell populations remained stable and within normal limits. In the boy, naïve CD4+ cells remained unchanged at the upper reference limit of 83%, and his memory T-cell values, already low, decreased further to 9%, below the lower reference limit. B-cell subsets also diverged: the girl showed very high naïve B-cell levels of up to 92% and very low memory B-cell levels at 8%, with both non-switched and switched memory subtypes below the lower limit. In the boy, naïve CD19+CD27− B-cells were 88% and decreased after therapy, while memory B-cells and both subsets increased, especially the non-switched CD27+IgD+IgM+ subset, which rose from 5.4% to 10.8%. Both children maintained a slight persistent decrease in NK-cell counts during the observation period.

Activation marker analysis showed that before treatment, both children exhibited evidence of chronic immune activation, reflected in elevated levels of CD3+CD38+ and CD19+CD38+ cells, pointing toward ongoing antigenic stimulation. The authors caution, however, that these elevations could also represent normal antigen-specific immune responses during or shortly after recent infections, and that spontaneous normalization of activation markers can occur naturally over time independent of therapy. After IAD, most activation marker values moved closer to the reference range: CD3+HLA-DR+ cells decreased slightly, CD3+CD38+ cells showed a slight reduction in the boy, and a difference was found in CD3+CD4+CD25+ cells, which were above the upper reference limit in the girl and normalized after treatment to 4%, while in the boy they rose from 3% to 6%, exceeding the upper reference limit.

The authors interpret the two cases as distinct models of immune response, one driven predominantly by T-cell dynamics and the other by B-cell alterations, each influencing the effects of immunomodulation differently. They suggest that in the girl, normalization of naïve CD4+ T-cell frequencies with stable memory compartments may indicate an enhancement of T-cell renewal without disrupting established immunological memory, while in the boy the upward trend in memory B-cells, particularly the non-switched subset, may indicate a favorable modulation of the memory B-cell profile, though this compartment remained functionally limited. They also note that residual NK-cell and double-negative T-cell dysregulation persisted in both cases, potentially contributing to susceptibility to recurrent infections, and that physiological maturation of the pediatric immune system, in which naïve T-cell proportions naturally decline with age while memory compartments expand, must be considered in longitudinal follow-up.

The study’s limitations are substantial. Beyond the minimum number of patients, the case-report nature of the analysis limits the generalizability of the findings, and the authors state explicitly that it is not possible to attribute the immune alterations solely to IAD therapy, as they may represent transient or combined effects of multiple interventions or the natural dynamics of immune recovery after infection. The authors argue that the observations nonetheless demonstrate the clinical utility of subset-specific flow cytometry monitoring for detecting subtle immune abnormalities not apparent in standard lymphocyte count evaluation, and for tracking therapeutic responses that can guide personalized treatment strategies. They call for future prospective controlled studies with larger cohorts and standardized monitoring of immune activation markers to define optimal dosing, duration, and patient selection criteria for IAD therapy in children with recurrent infections.

Subject of Research: Short-term immunophenotypic changes after inosine acedoben dimepranol therapy in two children with recurrent infections

Article Title: Immunophenotypic changes in pediatric patients with recurrent infections following inosine acedoben dimepranol therapy: a case-based analysis

Article References: Velikova, T., Mihova, A., Ali, H., Angelova, A., Ivanova, V., Pentchev, E., Mihaylov, R., & Pencheva, B. (2026). Immunophenotypic changes in pediatric patients with recurrent infections following inosine acedoben dimepranol therapy: a case-based analysis. Pharmacia, 73, Article e168282. https://doi.org/10.3897/pharmacia.73.e168282

Image Credits: AI Generated

DOI: 10.3897/pharmacia.73.e168282

Keywords: inosine acedoben dimepranol, immunophenotyping, flow cytometry, recurrent infections, pediatric immunology, T-cells, B-cells, NK cells, activation markers, case report, immunomodulation, CD38