Mass spectrometry-based assessment of M protein in peripheral blood during maintenance therapy in multiple myeloma.

ABSTRACT: Mass spectrometry (MS) can detect multiple myeloma-derived monoclonal proteins in the peripheral blood (PB) with high sensitivity, potentially serving as a PB assay for measurable residual disease (MRD). This study evaluated the significance of PB MS MRD negativity during posttransplant therapy in patients with newly diagnosed multiple myeloma. Serum samples from 138 patients treated in the phase 3 ATLAS trial of posttransplant maintenance with either carfilzomib, lenalidomide, and dexamethasone, or with lenalidomide alone were analyzed using EXENT MS methodology. We established feasibility of measuring MRD by MS in the PB in the posttransplant setting, despite unavailability of pretreatment calibration samples. There was high agreement between MRD by MS in the PB and paired bone marrow (BM) MRD results at the 10-5 threshold, assessed by either next-generation sequencing (NGS) or multiparameter flow cytometry (MFC) (70% and 67%, respectively). Agreement between PB MS and both BM MRD methods was lowest early after transplant and increased with time. MS negativity was associated with improved progression-free survival (PFS), which, in landmark analysis, reached statistical significance after 18 cycles after transplant. Combined PB/BM MRD negativity by MFC or NGS was associated with superior PFS compared with MRD negativity by only 1 modality. Sustained MS negativity carried similar prognostic performance to sustained BM MRD negativity at the 10-5 threshold. Overall, posttransplant MS assessment was feasible and provided additional prognostic information to BM MRD negativity. Further studies are needed to confirm the role and optimal timing of MS in disease evaluation algorithms. The ATLAS trial is registered at www.clinicaltrials.gov as #NCT02659293.

A Review of Resistance Mechanisms to Bruton's Kinase Inhibitors in Chronic Lymphocytic Leukemia.

Bruton's Tyrosine Kinase (BTK) inhibitors have become one of the most vital drugs in the therapy of chronic lymphocytic leukemia (CLL). Inactivation of BTK disrupts the B-cell antigen receptor (BCR) signaling pathway, which leads to the inhibition of the proliferation and survival of CLL cells. BTK inhibitors (BTKi) are established as leading drugs in the treatment of both treatment-naïve (TN) and relapsed or refractory (R/R) CLL. Furthermore, BTKi demonstrate outstanding efficacy in high-risk CLL, including patients with chromosome 17p deletion, TP53 mutations, and unmutated status of the immunoglobulin heavy-chain variable region (IGHV) gene. Ibrutinib is the first-in-class BTKi which has changed the treatment landscape of CLL. Over the last few years, novel, covalent (acalabrutinib, zanubrutinib), and non-covalent (pirtobrutinib) BTKi have been approved for the treatment of CLL. Unfortunately, continuous therapy with BTKi contributes to the acquisition of secondary resistance leading to clinical relapse. In recent years, it has been demonstrated that the predominant mechanisms of resistance to BTKi are mutations in BTK or phospholipase Cγ2 (PLCG2). Some differences in the mechanisms of resistance to covalent BTKi have been identified despite their similar mechanism of action. Moreover, novel mutations resulting in resistance to non-covalent BTKi have been recently suggested. This article summarizes the clinical efficacy and the latest data regarding resistance to all of the registered BTKi.

Treatment of Double-Refractory Chronic Lymphocytic Leukemia-An Unmet Clinical Need.

Recent years have seen significant improvement in chronic lymphocytic leukemia (CLL) management. Targeting B-cell lymphoma (BCL-2) and Bruton's kinase (BTK) have become the main strategies to restrain CLL activity. These agents are generally well tolerated, but the discontinuation of these therapies happens due to resistance, adverse effects, and Richter's transformation. A growing population of patients who have previously used both BTK inhibitors and BCL2 suffer from the constriction of the following regimens. This review explores the resistance mechanisms for both ibrutinib and venetoclax. Moreover, we present innovative approaches evaluated for treating double-refractory CLL.

Characteristics and outcomes of patients with lymphoma who developed therapy-related acute myeloid leukemia or myelodysplastic syndrome - a retrospective analysis of the Polish Adult Leukemia Group.

Introduction: Enhancing lymphoma outcomes increases the risk of therapy-related neoplasms such as acute myeloid leukemia (t-AML) and myelodysplastic syndrome (t-MDS). Material and methods: Our study, conducted at seven Polish hematology centers between 2011 and 2018, explores clinical features, outcomes, and prognostic factors of t-AML and t-MDS arising after initial lymphoid neoplasms. Results: The analysis included 57 patients of median age 65 with t-MDS (n = 38) and t-AML (n = 19). The median time to the onset of t-MDS/AML was 58.7 months. The median overall survival (OS) was 16.1 months. The presence of unfavorable cytogenetics and molecular risk factors (HR 2.88, 95% CI: 1.29-6.42, p = 0.009), hemoglobin level (HR 0.79, 95% CI: 0.65-0.95, p = 0.01) and platelets (HR 0.99, 95% CI: 0.99-0.9996, p = 0.03) were independent prognostic factors influencing OS. Therapy- related myelodysplastic syndrome/acute myeloid leukemia after lymphoma treatment is associated with a dismal prognosis mainly due to poor cytogenetic risk. Conclusions: Anemia and thrombocytopenia may indicate more severe impairment of bone marrow function, resulting in further inferior treatment outcomes.

Carfilzomib, lenalidomide, and dexamethasone or lenalidomide alone as maintenance therapy after autologous stem-cell transplantation in patients with multiple myeloma (ATLAS): interim analysis of a randomised, open-label, phase 3 trial.

BACKGROUND: Lenalidomide is a cornerstone of maintenance therapy in patients with newly diagnosed multiple myeloma after autologous stem-cell transplantation. We aimed to compare the efficacy and safety of maintenance therapy with carfilzomib, lenalidomide, and dexamethasone versus lenalidomide alone in this patient population. METHODS: This study is an interim analysis of ATLAS, which is an investigator-initiated, multicentre, open-label, randomised, phase 3 trial in 12 academic and clinical centres in the USA and Poland. Participants were aged 18 years or older with newly diagnosed multiple myeloma, completed any type of induction and had stable disease or better, autologous stem-cell transplantation within 100 days, initiated induction 12 months before enrolment, and an Eastern Cooperative Oncology Group performance status of 0 or 1. Patients were randomly assigned (1:1) using permuted blocks of sizes 4 and 6 and a web-based system to receive up to 36 cycles of carfilzomib, lenalidomide, and dexamethasone (28-day cycles of carfilzomib 20 mg/m2 administered intravenously in cycle one on days 1 and 2 then 36 mg/m2 on days 1, 2, 8, 9, 15, and 16 in cycles one to four and 36 mg/m2 on days 1, 2, 15, and 16 from cycle five up to 36 [per protocol]; lenalidomide 25 mg administered orally on days 1-21; and dexamethasone 20 mg administered orally on days 1, 8, 15, and 22) or lenalidomide alone (10 mg administered orally for the first three cycles and then at the best tolerated dose [≤15 mg for 28 days in 28-day cycles]) until disease progression or unacceptable toxicity as maintenance therapy. After 36 cycles, patients in both treatment groups received lenalidomide maintenance. Randomisation was stratified by response to previous treatment, cytogenetic risk factors, and country. Investigators and patients were not masked to treatment allocation. Patients in the carfilzomib, lenalidomide, and dexamethasone group with no detectable minimal residual disease after cycle six (as per International Myeloma Working Group criteria) and standard-risk cytogenetics were switched to lenalidomide maintenance as of cycle nine. The primary endpoint was progression-free survival in the intention-to-treat population (defined as all randomly assigned patients). Safety was analysed in all randomly assigned patients who received at least one dose of study treatment. This unplanned interim analysis was triggered by the occurrence of 59 (61%) of the expected 96 events for the primary analysis and the results are considered preliminary. This trial is registered with ClinicalTrials.gov, NCT02659293 (active, not recruiting) and EudraCT, 2015-002380-42. FINDINGS: Between June 10, 2016, and Oct 21, 2020, 180 patients were randomly assigned to receive either carfilzomib, lenalidomide, and dexamethasone (n=93) or lenalidomide alone (n=87; intention-to-treat population). The median age of patients was 59·0 years (IQR 49·0-63·0); 84 (47%) patients were female and 96 (53%) were male. With a median follow-up of 33·8 months (IQR 20·9-42·9), median progression-free survival was 59·1 months (95% CI 54·8-not estimable) in the carfilzomib, lenalidomide, and dexamethasone group versus 41·4 months (33·2-65·4) in the lenalidomide group (hazard ratio 0·51 [95% CI 0·31-0·86]; p=0·012). The most common grade 3 and 4 adverse events were neutropenia (44 [48%] in the carfilzomib, lenalidomide, and dexamethasone group vs 52 [60%] in the lenalidomide group), thrombocytopenia (12 [13%] vs six [7%]), and lower respiratory tract infections (seven [8%] vs one [1%]). Serious adverse events were reported in 28 (30%) patients in the carfilzomib, lenalidomide, and dexamethasone group and 19 (22%) in the lenalidomide group. One treatment-related adverse event led to death (respiratory failure due to severe pneumonia) in the carfilzomib, lenalidomide, and dexamethasone group. INTERPRETATION: This interim analysis provides support for considering carfilzomib, lenalidomide, and dexamethasone therapy in patients with newly diagnosed multiple myeloma who completed any induction regimen followed by autologous stem-cell transplantation, which requires confirmation after longer follow-up of this ongoing phase 3 trial. FUNDING: Amgen and Celgene (Bristol Myers Squibb).

Molnupiravir is effective in patients with haematological malignancies.

Patients with hematologic malignancies are particularly vulnerable to severe infectious complications. SARS-CoV-2 infection is associated with a high risk of severe course and death in this patient population. In addition, immune deficits associated with both the blood cancer and the treatment used make vaccination against SARS-CoV-2 less effective than in immunocompetent individuals. Molnupiravir is one of the first oral antiviral drugs to demonstrate a significant benefit in reducing hospitalisation and death in COVID-19 in the general population. In this context, 175 haematology patients with diagnosed COVID-19, and treated with MOL between January and April 2022, came under our scrutiny with a view to defining their clinical characteristics and outcomes. The most common underlying conditions were lymphomas (45%), multiple myelomas (21%) and acute leukaemias or myelodysplastic syndrome (35%). Of all, 77% of the patients were vaccinated, and half of them received a booster. At 28 days after the breakthrough COVID-19 diagnosis, 35 (20%) subjects required hospital admission. Out of those patients, seven (4%) died during the follow-up due to the progression of COVID. Our results corroborate what has been established to date with regard to the positive clinical and safety outcomes of MOL in haematology patients with mild or moderate COVID-19.

Response to anti-SARS-CoV-2 mRNA vaccines in multiple myeloma and chronic lymphocytic leukemia patients.

Multiple myeloma (MM) and chronic lymphocytic leukemia (CLL) patients have increased morbidity and mortality rates of COVID-19 due to immunosuppression associated with the disease and ongoing therapy. The same immune impairment accompanying CLL and MM also affects suboptimal vaccine response. The study assessed the effectiveness of the humoral and T cell-mediated immunity following mRNA COVID-19 vaccination (using either BNT162b2 or mRNA-1273) in short-term (2-5 weeks after second dose) and long-term follow-up (12 weeks after vaccination). Between March and August 2021, blood samples were obtained from 62 CLL and 60 MM patients from eight different hematology departments in Poland. Total anti-RBD antibodies were detected in 37% MM patients before vaccination, increased to 91% and 94% in short- and long-term follow-up, respectively. In CLL, serological responses were detectable in 21% of patients before vaccination and increased to 45% in the short-term and 71% in long-term observation. We detected a tendency to higher frequencies of specific CD8+ T cells against SARS-CoV-2 after vaccination compared to samples before vaccination in MM patients and no changes in frequencies of specific T cells in CLL patients. Our study provides novel insights into mRNA vaccination efficacy in immunocompromised MM and CLL patients, and our findings highlight that specific CD8+ T cells against SARS-CoV-2 might be induced by vaccination but do not correlate positively with serological responses.

Polyclonal immunoglobulin recovery in patients with newly diagnosed myeloma receiving maintenance therapy after autologous haematopoietic stem cell transplantation with either carfilzomib, lenalidomide and dexamethasone or lenalidomide alone: Subanalysis of the randomized phase 3 ATLAS trial.

Previous studies suggest that postautologous stem cell transplant (ASCT) recovery of polyclonal immunoglobulin from immunoparesis in patients with multiple myeloma is a positive prognostic marker. We performed a longitudinal analysis of polyclonal immunoglobulin concentrations and unique B-cell sequences in patients enrolled in the phase 3 ATLAS trial that randomized 180 subjects to either carfilzomib, lenalidomide, dexamethasone (KRd) or lenalidomide (R) maintenance. In the KRd arm, standard-risk patients with minimal residual disease negativity after six cycles de-escalated to R alone after cycle 8. One year from the initiation of maintenance at least partial recovery of polyclonal immunoglobulin was observed in more patients on the R arm (58/66, p < 0.001) and in those who de-escalated from KRd to R (27/38, p < 0.001) compared to the KRd arm (9/36). In patients who switched from KRd to R, the concentrations of uninvolved immunoglobulin and the number of B-cell unique sequences increased over time, approaching values observed in the R arm. There were no differences in progression-free survival between the patients with at least partial immunoglobulin recovery and the remaining population. Our analysis indicates that patients receiving continuous therapy after ASCT experience prolonged immunoparesis, limiting prognostic significance of polyclonal immunoglobulin recovery in this setting.

Clinical efficacy and tolerability of venetoclax plus rituximab in patients with relapsed or refractory chronic lymphocytic leukemia-a real-world analysis of the Polish Adult Leukemia Study Group.

The results of the MURANO trial showed encouraging progression-free survival (PFS) and overall survival (OS) in relapsed/refractory chronic lymphocytic leukemia (RR-CLL) patients treated with venetoclax-rituximab (VEN-R). A retrospective analysis was performed to evaluate the efficacy and safety of VEN-R within the Polish Adult Leukemia Study Group (PALG) centers. The study group included 117 patients with RR-CLL (with early relapse after immunochemotherapy or bearing TP53 aberrations) treated with VEN-R in 2019-2023 outside clinical trials. Patients were treated with a median of 2 (range 1-9) previous lines of therapy. Twenty-two participants were previously treated with BTKi (18.8% out of 117). The median follow-up was 20.3 months (range 0.27-39.1). The overall response rate (ORR) was 95.3% in the group of patients in whom a response to treatment was assessed and 86.3% for all patients. Twenty patients (17.1% out of 117) achieved a complete response (CR), 81 (69.2%) achieved a partial response (PR), and in 5 patients (4.3%), disease progression was noted (assessed as the best response during treatment). The median PFS in the whole cohort was 36.97 (95% CI 24.5, not reached) months, and the median OS was not reached (95% CI 27.03, not reached). Thirty-six patients died during the follow-up, 10 (8.5%; 27.8% of deaths) due to COVID-19 infection. All grade neutropenia (n = 87/117, 74.4%; grade 3 or higher n = 67/117, 57.3%) was the most common treatment adverse event. Forty-five patients (38.5%) remained on treatment, and twenty-two (18.8%) completed 24 months of therapy, while it was discontinued in fifty cases (42.7%). In this real-world setting of early access in very high-risk RR-CLL patients, the VEN-R regimen was associated with shorter median PFS compared with the results of the MURANO trial. This outcome, however, could be attributed to patients' exposure to SARS-CoV-2 infection and the aggressive course of the disease as very high-risk patients, after multiple lines of prior therapies, were included in the Polish Ministry of Health reimbursement program.

Predictive significance of selected gene mutations in relapsed and refractory chronic lymphocytic leukemia patients treated with ibrutinib.

BACKGROUND: Ibrutinib, an inhibitor of the Bruton's kinase (BTK), is characterized by high efficacy in the therapy of patients with relapsed and refractory chronic lymphocytic leukemia (RR-CLL). AIMS: To analyze the potential significance of the mutational status of selected 30 genes on the disease outcome in 45 patients with RR-CLL using custom-made gene panel and sequencing on Illumina MiSeq FGx platform. RESULTS: The highest rate of mutations was observed in TP53 (n = 18; 40.0%), NOTCH1 (n = 13; 28.8%), SF3B1 (n = 11; 24.4%), ATM (n = 7; 15.6%), MED12 (n = 6, 13.3%), CHD2 (n = 5; 11.1%), XPO1 (n = 5; 11.1%), NFKBIE (n = 5; 11.1%), BIRC3 (n = 4; 8.9%), SPEN (n = 4; 8.9%), POT1 (n = 4; 8.9%), EGR2 (n = 3; 6.7%), and RPS15 (n = 3; 6.7%). With a median observation time of 45.9 months, the median progression-free survival (PFS) and overall survival (OS) were not reached. The 36-month estimated rate of PFS and OS were 64% and 68.2%, respectively. The overall response rate was noted in 23 patients (51.1%), while twenty (44.4%) patients achieved stability. Progression was noted in 2 (4.5%) cases. Analyzed molecular factors had no impact on PFS and OS. CONCLUSION: Despite accumulation of several poor prognostic factors in our real-life cohort of heavily pretreated patients with CLL, ibrutinib treatment showed long-term clinical benefit.

Outcomes of treatment with dose-adjusted EPOCH-R or R-CHOP in primary mediastinal large B-cell lymphoma.

BACKGROUND: The standard first-line treatment for primary mediastinal B-cell lymphoma (PMBCL) patients is rituximab-based immunochemotherapy; however, this is not due to the result of randomized clinical trials. AIMS: We retrospectively investigated 53 PMBCL patient outcomes treated either with R-CHOP-21 or DA-EPOCH-R-28. The primary endpoint was overall survival (OS). Secondary endpoints were complete remission (CR), overall response rate (ORR), progression-free survival (PFS), and treatment-related complications. RESULTS: Treatment with R-CHOP-21 resulted in a 92.0% ORR (60% CR), while DA-EPOCH-R yielded a 92.6% ORR (70.4% CR). There were no differences in the occurrence of grade 3-4 hematological adverse events, but grade 1-2 cardiologic complications (P = .003) were observed more frequently in the DA-EPOCH-R arm. Median PFS and OS were not achieved. The differences in estimated 12-month PFS in R-CHOP and DA-EPOCH-R group (87% vs 73.9%) and OS (100% vs 92%) were insignificant. Patients treated with R-CHOP-21 and autologous hematopoietic stem cell transplantation (auto-HSCT) had an improved OS (P = .03) but not PFS (P = .43) compared to those treated solely with R-CHOP-21. No differences in PFS or OS were observed between patients treated with R-CHOP-21/auto-HSCT and DA-EPOCH-R. CONCLUSION: The results of this study suggest that R-CHOP-21 may be an alternative to DA-EPOCH-R treatment for PMBCL patients.

Microenvironment-induced PIM kinases promote CXCR4-triggered mTOR pathway required for chronic lymphocytic leukaemia cell migration.

Lymph node microenvironment provides chronic lymphocytic leukaemia (CLL) cells with signals promoting their survival and granting resistance to chemotherapeutics. CLL cells overexpress PIM kinases, which regulate apoptosis, cell cycle and migration. We demonstrate that BCR crosslinking, CD40 stimulation, and coculture with stromal cells increases PIMs expression in CLL cells, indicating microenvironment-dependent PIMs regulation. PIM1 and PIM2 expression at diagnosis was higher in patients with advanced disease (Binet C vs. Binet A/B) and in those, who progressed after first-line treatment. In primary CLL cells, inhibition of PIM kinases with a pan-PIM inhibitor, SEL24-B489, decreased PIM-specific substrate phosphorylation and induced dose-dependent apoptosis in leukaemic, but not in normal B cells. Cytotoxicity of SEL24-B489 was similar in TP53-mutant and TP53 wild-type cells. Finally, inhibition of PIM kinases decreased CXCR4-mediated cell chemotaxis in two related mechanisms-by decreasing CXCR4 phosphorylation and surface expression, and by limiting CXCR4-triggered mTOR pathway activity. Importantly, PIM and mTOR inhibitors similarly impaired migration, indicating that CXCL12-triggered mTOR is required for CLL cell chemotaxis. Given the microenvironment-modulated PIM expression, their pro-survival function and a role of PIMs in CXCR4-induced migration, inhibition of these kinases might override microenvironmental protection and be an attractive therapeutic strategy in this disease.

Current Treatment of Chronic Lymphocytic Leukemia.

OPINION STATEMENT: A number of new treatment options have recently emerged for chronic lymphocytic leukemia (CLL) patients, including the Bruton's tyrosine kinase (BTK) inhibitor ibrutinib, phosphatidylinositol-3-kinase (PI3K) delta isoform inhibitor idelalisib combined with rituximab, the Bcl-2 antagonist venetoclax, and the new anti-CD20 antibodies obinutuzumab and ofatumumab. Most of these agents are already included into treatment algorithms defined by international practice guidelines, but more clinical investigations are needed to answer still remaining questions. Ibrutinib was proven as a primary choice for patients with the TP53 gene deletion/mutation, who otherwise have no active treatment available. Idelalisib with rituximab is also an active therapy, but due to increased risk of serious infections, its use in first-line treatment is limited to patients for whom ibrutinib is not an option. A new indication for ibrutinib was recently approved for older patients with comorbidities, as an alternative to the already existing indication for chlorambucil with obinutuzumab. The use of kinase inhibitors is already well established in recurrent/refractory disease. Immunochemotherapy with fludarabine, cyclophosphamide, rituximab (FCR) remains a major first-line option for many CLL patients without the TP53 gene deletion/mutation, and who have no significant comorbidities or history of infections, and is particularly effective in patients with favorable features including mutated IGHV status. There are a number of issues regarding novel therapies for CLL that need further investigation such as optimum duration of treatment with kinase inhibitors, appropriate sequencing of novel agents, mechanisms of resistance to inhibitors and response to class switching after treatment failure, along with the potential role of combinations of targeted agents.

The Molecular Effect of Diagnostic Absorbed Doses from 131I on Papillary Thyroid Cancer Cells In Vitro.

Diagnostic whole-body scan is a standard procedure in patients with thyroid cancer prior to the application of a therapeutic dose of 131I. Unfortunately, administration of the radioisotope in a diagnostic dose may decrease further radioiodine uptake-the phenomenon called "thyroid stunning". We estimated radiation absorbed dose-dependent changes in genetic material, in particular in the sodium iodide symporter (NIS) gene promoter, and the NIS protein level in a K1 cell line derived from the metastasis of a human papillary thyroid carcinoma exposed to 131I in culture. The different activities applied were calculated to result in absorbed doses of 5, 10 and 20 Gy. Radioiodine did not affect the expression of the NIS gene at the mRNA level, however, we observed significant changes in the NIS protein level in K1 cells. The decrease of the NIS protein level observed in the cells subjected to the lowest absorbed dose was paralleled by a significant increase in 8-oxo-dG concentrations (p < 0.01) and followed by late activation of the DNA repair pathways. Our findings suggest that the impact of 131I radiation on thyroid cells, in the range compared to doses absorbed during diagnostic procedures, is not linear and depends on various factors including the cellular components of thyroid pathology.

Metallothioneins in Normal and Cancer Cells.

Metallothioneins (MTs) are low molecular weight proteins, which are present in almost all types of organisms. In mammals, four main MT isoforms designated from MT-1 to MT-4 were identified. Their biological role, according to their characteristic structure, was shown to be mostly associated with cellular metabolism of metal ions, especially zinc. Moreover, the available evidence suggests broad regulatory properties of MTs in the control of cell senescence and various pathological processes including neurodegeneration, cardiovascular pathology, metabolic disorders, and various malignancies. This extensive review provides general in formation on the structure of MT family members and the cellular functions of MT-1, MT-2, and MT-4 isoforms as well as insights into divergent biological roles of MT-3. Due to the involvement of MT molecules in various processes related to carcinogenesis, an organ-specific presentation of current data concerning their potential impact on the progression of various tumors is given. The regulatory role of MT family members in the function of the immune system is also discussed in depth.

Effect of Collateral Sprouting on Donor Nerve Function After Nerve Coaptation: A Study of the Brachial Plexus.

BACKGROUND The aim of the present study was to evaluate the donor nerve from the C7 spinal nerve of the rabbit brachial plexus after a coaptation procedure. Assessment was performed of avulsion of the C5 and C6 spinal nerves treated by coaptation of these nerves to the C7 spinal nerve. MATERIAL AND METHODS After nerve injury, fourteen rabbits were treated by end-to-side coaptation (ETS), and fourteen animals were treated by side-to-side coaptation (STS) on the right brachial plexus. Electrophysiological and histomorphometric analyses and the skin pinch test were used to evaluate the outcomes. RESULTS There was no statistically significant difference in the G-ratio proximal and distal to the coaptation in the ETS group, but the differences in the axon, myelin sheath and fiber diameters were statistically significant. The comparison of the ETS and STS groups distal to the coaptation with the controls demonstrated statistically significant differences in the fiber, axon, and myelin sheath diameters. With respect to the G-ratio, the ETS group exhibited no significant differences relative to the control, whereas the G-ratio in the STS group and the controls differed significantly. In the electrophysiological study, the ETS and STS groups exhibited major changes in the biceps and subscapularis muscles. CONCLUSIONS The coaptation procedure affects the histological structure of the nerve donor, but it does not translate into changes in nerve conduction or the sensory function of the limb. The donor nerve lesion in the ETS group is transient and has minimal clinical relevance.

Divergent expression patterns of SATB1 mRNA and SATB1 protein in colorectal cancer and normal tissues.

Special AT-rich sequence-binding protein 1 (SATB1) is a 'genome organizer,' and it has been proposed as a factor that affects the development and progression of various human neoplasms, including colorectal cancer (CRC). This study aimed to compare SATB1 expression in a group of CRC patients and healthy subjects at the mRNA and protein levels. We collected paired tumor tissue and unchanged mucosa of the large intestine from 102 CRC patients as well as 53 biopsies of normal colon mucosa obtained from healthy patients during screening colonoscopy. Tissue samples were quantified for SATB1 mRNA by quantitative PCR, while SATB1 protein expression was determined by Western blotting and immunohistochemistry. SATB1 mRNA level in tumor tissues was over twofolds lower than in samples of corresponding unchanged tissues and fourfolds lower than in biopsies of healthy colon mucosa. Western blotting analysis revealed that SATB1 protein content in tumor and unchanged tissues of CRC patients was over sixfold and fivefolds higher than in biopsies of healthy colon mucosa, respectively. Immunohistochemical staining demonstrated higher nuclear and cytoplasmic SATB1 reactivity in the tumor tissue compared to unchanged mucosa of CRC patients. Despite these differences, SATB1 mRNA, protein, and immunoreactivity levels did not correlate with patients' clinicopathological data and their overall survival, but the latter analysis was limited by a relatively short period of follow-up. In conclusion, we suggest that some as yet unidentified posttranscriptional mechanisms that regulate SATB1 expression may be altered in the CRC tissue.

Interferon-beta affects mitochondrial activity in CD4+ lymphocytes: Implications for mechanism of action in multiple sclerosis.

BACKGROUND: Whereas cellular immune function depends on energy supply and mitochondrial function, little is known on the impact of immunotherapies on cellular energy metabolism. OBJECTIVE: The objective of this paper is to assess the effects of interferon-beta (IFN-ß) on mitochondrial function of CD4(+) T cells. METHODS: Intracellular adenosine triphosphate (iATP) in phytohemagglutinin (PHA)-stimulated CD4(+) cells of multiple sclerosis (MS) patients treated with IFN-ß and controls were analyzed in a luciferase-based assay. Mitochondrial-transmembrane potential (ΔΨm) in IFN-ß-treated peripheral blood mononuclear cells (PBMCs) was investigated by flow cytometry. Expression of genes involved in mitochondrial oxidative phosphorylation (OXPHOS) in CD4(+) cells of IFN-ß-treated individuals and correlations between genetic variants in the key metabolism regulator PGC-1α and IFN-ß response in MS were analyzed. RESULTS: IFN-ß-treated MS patients exhibited a dose-dependent reduction of iATP levels in CD4(+) T cells compared to controls (p < 0.001). Mitochondrial effects were reflected by depolarization of ΔΨm. Expression data revealed changes in the transcription of OXPHOS-genes. iATP levels in IFN-ß-responders were reduced compared to non-responders (p < 0.05), and the major T allele of the SNP rs7665116 of PGC-1α correlated with iATP-levels. CONCLUSION: Reduced iATP-synthesis ex vivo and differential expression of OXPHOS-genes in CD4(+) T cells point to unknown IFN-ß effects on mitochondrial energy metabolism, adding to potential pleiotropic mechanisms of action.

The Effect of Diagnostic Absorbed Doses from 131I on Human Thyrocytes in Vitro.

BACKGROUND: Administration of diagnostic activities of 131I, performed in order to detect thyroid remnants after surgery and/or thyroid cancer recurrence/metastases, may lead to reduction of iodine uptake. This phenomenon is called "thyroid stunning". We estimated radiation absorbed dose-dependent changes in genetic material, in particular in sodium iodide symporter (NIS) gene promoter, and NIS protein level in human thyrocytes (HT). MATERIALS AND METHODS: We used unmodified HT isolated from patients subjected to thyroidectomy exposed to 131I in culture. The different 131I activities applied were calculated to result in absorbed doses of 5, 10, and 20 Gy. RESULTS: According to flow cytometry analysis and comet assay, 131I did not influence the HT viability in culture. Temporary increase of 8-oxo-dG concentration in HT directly after 24 h (p < 0.05) and increase in the number of AP-sites 72 h after termination of exposition to 20 Gy dose (p < 0.0001) were observed. The signs of dose-dependent DNA damage were not associated with essential changes in the NIS expression on mRNA and protein levels. CONCLUSIONS: Our observation constitutes a first attempt to evaluate the effect of the absorbed dose of 131I on HT. The results have not confirmed the theory that the "thyroid stunning" reduces the NIS protein synthesis.

Expression of matrix metalloproteinase 2 (MMP-2), E-cadherin and Ki-67 in metastatic and non-metastatic canine mammary carcinomas.

BACKGROUND: The aim of the study was to demonstrate the immunohistochemical expression of proteins that affect the metastatic potential of a tumour, including matrix metalloproteinase 2 (MMP-2) and E-cadherin. Another objective was to determine their correlation with the expression of the Ki-67 antigen in metastasizing and non-metastasizing mammary carcinomas in female dogs. The study was conducted on 32 canine mammary carcinomas (12 metastatic and 20 non-metastatic), classified as simple tubular and tubulopapillary carcinomas. Immunohistochemistry was performed to evaluate the expression of MMP-2, E-cadherin and Ki-67 antigen. RESULTS: MMP-2 was expressed in 85 % of the non-metastatic tumours and in all the metastatic tumours, while E-cadherin was expressed in 85 % of the non-metastatic tumours and in 66 % of the metastatic tumours. The Ki-67 antigen was expressed in 65 % of the non-metastatic tumours and in 91 % of the metastatic tumours. The mean Ki-67 expression was slightly higher in tumours that had metastasized (1.5 ± 0.90 vs 1.1 ± 0.94; p = 0.22). A similar relationship was found in terms of the intensity of the MMP-2 expression (2.9 ± 1.9 vs 2.7 ± 2.4; p = 0.50). A decrease in the expression of E-cadherin (2.8 ± 2.5) was found in metastatic tumours compared to the expression in non-metastatic tumours (3.2 ± 2.3). However, these differences were not statistically significant (p = 0.63). CONCLUSION: We did not show significant differences in MMP-2, E-cadherin and Ki-67 expression between metastatic and non-metastatic tumours due to low number of cases studied, however further experiments are necessary to assess the role of these antigens in the process of canine mammary tumours metastasis.