Pakistan Journal of Pharmaceutical Sciences

Abstract: Background: Triple-negative breast cancer (TNBC) is predominant among younger Pakistani women with advanced and biologically aggressive disease. Ethnic, genetic and environmental factors, as well as neoadjuvant chemotherapy (NACT), may affect the tumor immune microenvironment, including stromal tumor-infiltrating lymphocytes (TILs), immune checkpoint proteins expression like lymphocyte activation gene-3 (LAG-3), programmed cell death-1 (PD-1) and programmed cell death ligand-1 (PD-L1). Objectives: To investigate the expression of LAG-3, PD-1 and PD-L1 immune checkpoint proteins and evaluate their association with TILs in patients with TNBC receiving NACT with or without carboplatin. Methods: This quasi-experimental study included 54 patients with TNBC aged 24 to 45 years treated at two tertiary care hospitals in Karachi, Pakistan. Thirty-nine patients received anthracycline-taxane-carboplatin therapy (A+T+C group), whereas fifteen received anthracycline-taxane therapy (A+T group). Clinical response was assessed using Response Evaluation Criteria in Solid Tumor (RECIST) based on mammography and breast ultrasonography before and after NACT. TILs were assessed on hematoxylin and eosin-stained (H&E) post-NACT resection sections and categorized as low (0-10%), intermediate (11-40%), or high (>40%) according to international recommendations. Immunohistochemistry was performed for PD-1, PD-L1 and LAG-3 expression in tumor cells (TC) and TILs using compartment-specific proportional scoring with a 1% positivity cutoff. Associations were analyzed using chi-square/Fisher’s exact tests. Results: The A+T+C group attained a higher RECIST-based clinical response rate than the A+T group (92.31% vs. 60.0%) with a reduction in tumor size (2.67 cm, 95% CI: 2.14–3.23; p=0.001). High TILs density was significantly associated with PD-L1 expression on TC (χ² = 18.72; p < 0.001), LAG-3 expression on immune cells (IC) (χ² = 13.67; p < 0.001) and PD-1 expression on TC (χ² =  12.38; p = 0.002). No significant associations were observed with LAG-3 expression on TC, PD-L1 and PD-1 expressions on IC. Conclusion: Post-NACT TILs density was significantly associated with PD-L1 expression on TC and LAG-3 expression on IC in TNBC. Carboplatin improved the clinical response without significantly affecting the TIL density or immune checkpoint expression.

Abstract: Background: Few studies examine combined shockwave and core training for chronic low back pain (CLBP) analgesic dependence. Objectives: This study compared multimodal rehabilitation (shock wave therapy plus core muscle training) with pharmacotherapy alone for analgesic dependence in CLBP patients. Methods: Clinical data were retrospectively collected from 187 consecutive CLBP patients admitted to Jiangbin Hospital of Guangxi Zhuang Autonomous Region between January 2024 and June 2025. After exclusion, 184 patients were enrolled and divided into a multimodal rehabilitation group (shock wave therapy, core muscle exercises and routine medications; n=94) and a monotherapy group (pharmacotherapy only; n=90). 1:1 propensity score matching (PSM) produced 86 matched cases per group. The primary endpoints were analgesic dosage and the incidence of analgesic dependence. Secondary outcomes included the Visual Analogue Scale (VAS), Oswestry Disability Index (ODI), back extensor endurance, side-bridge holding time, lumbar range of motion, Hamilton Anxiety Rating Scale (HAMA) and Hamilton Depression Rating Scale (HAMD) scores and adverse events. Multivariate logistic regression was performed to identify independent predictors of analgesic dependence in CLBP patients. Results: After PSM, baseline characteristics were balanced between groups (all P > 0.05). At 12-week follow-up, the multimodal rehabilitation group showed lower analgesic dosage and analgesic dependence rate than the medication-only group (both P < 0.05). Both groups showed improvements in VAS, ODI, HAMA, and HAMD scores (all P < 0.05), with the multimodal group showing better outcomes (all P < 0.001). The multimodal group also exhibited greater enhancements in Sorensen test time, side-bridge duration and lumbar range of motion (all P < 0.001). No significant between-group difference in adverse events was observed (P = 0.109). Therapeutic strategy and baseline analgesic dosage were independent predictors of analgesic dependence (all P < 0.05). Conclusion: Multimodal rehabilitation combining shock wave therapy and core muscle training is more effective in reducing analgesic dependence in CLBP patients.

Abstract: Background: Ligustrazine is a pivotal active component of Ligusticum wallichii Franch; it is used clinically for treating cardiovascular diseases. Derivatization of ligustrazine might promote its biological activities and bioavailability. Objectives: A series of ligustrazine-acylhydrazone derivatives (TMPAD-1–TMPAD-13) were synthesized and their inhibitory activity against A549 cells was investigated. Methods: These derivatives were synthesized by condensing aromatic aldehydes with ligustrazine-based hydrazides to form acylhydrazone bonds and their chemical structures were characterized through a combination of spectral methods, including Fourier transform infrared (FT-IR) spectroscopy, electrospray ionization high resolution mass spectrometry (ESI-HRMS), ¹H and ¹³C nuclear magnetic resonance (NMR) spectra. Results: In-vitro experimental results displayed that TMPAD-2 exhibited marked inhibitory activity against A549 cells as shown by impeding cell proliferation (IC₅₀ = 48.08 μmol/L), colony formation (colony forming efficiency ≤ 1.5 ± 0.02%), invasion and migration, enhancing tumor necrosis factor (TNF)-α and reactive oxygen species (ROS) levels, decreasing interleukin (IL)-6 and mitochondrial membrane potential (MMP) levels, hindering cell cycle progression and inducing apoptosis. RNA-sequencing (RNA-seq) analysis disclosed 17 lung cancer-related differentially expressed genes (DEGs). These results were statistically supported with significant p-value. Conclusion: These results suggest that dysregulation of the cyclin–cyclin-dependent kinase (CDK)–cyclin-dependent kinase inhibitor (CKI) regulatory network is primarily responsible for the inhibitory action of TMPAD-2 on A549 cells. Current findings might provide valuable insights for further development of ligustrazine derivatives as novel anticancer agents.

Abstract: Background: Diabetic wounds are challenging due to delayed healing, inflammation, oxidative stress and bacterial infections, which conventional dressings often inadequately address. Objectives: To develop and characterize Aloe vera-mediated ZnO-loaded chitosan-pectin composite hydrogel films and evaluate their physicochemical, antibacterial, antioxidant, and in-vivo wound-healing efficacy in streptozotocin-induced diabetic male Wistar rat. Methods: AV-ZnO was synthesized using Aloe vera gel extract and incorporated into CS/PT hydrogels via solution casting to formulate three groups: PC (without AV-ZnO), PCG-1 (10 mg AV-ZnO), and PCG-2 (20 mg AV-ZnO). Physicochemical characterization included FTIR, SEM, AFM, TGA, gel fraction, contact angle, and tensile testing. Antibacterial activity was assessed against Staphylococcus aureus and Escherichia coli via disk diffusion, and antioxidant activity via DPPH radical-scavenging assay. In-vivo wound-healing efficacy was evaluated in streptozotocin-induced diabetic male Wistar rats (N = 24, n = 6/group) across four groups: untreated control, gentamicin-treated control, topical PCG-2, and exploratory oral PCG-2 hydrogel. Results: AV-ZnO incorporation improved tensile strength (10.1 MPa for PCG-2), hydrophilicity, gel fraction, and thermal stability. PCG-2 exhibited the strongest antibacterial activity, with inhibition zones of 28 mm against S. aureus and 20 mm against E. coli, attributed to AV-ZnO (Zn²⁺ release kinetics not evaluated). Antioxidant activity rose concentration-dependently, with PCG-2 showing 60.33% radical scavenging at 0.6 mg/mL. In-vivo, topical PCG-2 showed the highest wound contraction (89.09% ± 0.61 %, p < 0.05 vs. control) by day 15, surpassing exploratory oral treatment (72.84% ± 0.65%) and gentamicin (topical treated group) (83.77% ± 0.25%). Histology indicate improved re-epithelialization and collagen organization with topical treatment versus controls.