Cerebrospinal Fluid Immunophenotyping Research Articles

DETECTION OF HEMATOPOIETIC NEOPLASMS IN CEREBROSPINAL FLUID BY IMMUNOPHENOTYPING: SINGLE-CENTER DATA ANALYSIS

Cerebrospinal fluid (CSF) infiltration can be identified at diagnosis and during the course of many hematopoietic malignancies, mainly acute leukemia and aggressive non-Hodgking lymphomas (NHL). The accurate diagnosis of central nervous system (CNS) infiltration allows patients with infiltrated CSF to be properly treated, avoiding the use of unnecessary CNS therapy for patients who do not have compromised CSF. Cytomorphology is the classic method for diagnosing CSF neoplastic infiltration, but it presents variability in sensitivity and specificity according to the experience of the morphologist and cell abnormalities resulting from sample preparation. Currently, multiparametric flow cytometry (MFC) has great sensitivity and specificity to detect cells of hematopoietic neoplasms in the CSF. This is a retrospective study to assess the frequency of CSF infiltration in a cohort of patients with hematologic malignancies treated at our institution. to evaluate the casuistry and frequency of CSF infiltration by hematologic neoplasms diagnosed by immunophenotyping in our institution. the results of CSF immunophenotyping were collected using the flow cytometry laboratory database. Assess period: from March 2008 to June 2021. Patients were grouped by type of disease and the frequency with which each one was represented in the cohort was verified. a total of 1572 CSF samples were analyzed by the laboratory during the study period. The frequency of diagnoses of patients whose samples were sent for evaluation were, in decreasing order: 956(60.8%), B-cell precursor acute lymphoblastic leukemia (BCP ALL) samples 238(15.1%) T-cell acute lymphoblastic leukemia (T-ALL) samples, 168(10.6%) AML samples, 120 (7.6%) B and T NHL, 42 (2.6%) mixed lineage acute leukemia (MPAL), 34(2.1%) blastic crisis of chronic myeloid leukemia (CML), 14(0.8%) other mature lymphoid neoplasms. Immunophenotyping showed 311(19.78%) positive results for CSF infiltration. The diagnoses of these patients and their respective frequency were: 156 (50.1%) BCP-ALL, 49(15.7%) AML, 41(31.1%) T-ALL, 37(11.8%) NHL, 13 (4.1%) blastic crisis of CML, 8 (2.5%) MPAL, 7(2.2%) other mature lymphoid neoplasms. The frequencies of NHL subtypes observed in this cohort were: 16 (1%) Large B Cell Lymphoma, 7(0.44%) Burkitt Lymphoma, 6 (0.39%) Mantle Cell Lymphoma, 5(0.31%) Adult T-Cell Leukemia-Lymphoma, 1 (0.06%) Marginal Zone Lymphoma, 1 (0.06%) Folicular Lymphoma and 1 (0.06%) Sezary Syndrome. It should be emphasized that among the samples from patients with other mature lymphoid neoplasms, 4 (0.25%) were from patients with Chronic Lymphocytic Leukemia (CLL), which is an unusual finding. this study observed that 91% of the samples with CSF infiltration were from patients diagnosed with acute leukemia and blastic crisis of CML. Mature lymphoid neoplasms have a lower frequency of CNS infiltration than acute leukemia, as also reflected in our series. These findings show the importance of an accurate assessment of the CSF for these diseases, because the results influence the choice of the therapeutic modality and intensity to be adopted in the different treatment protocols. we demonstrate the ability of immunophenotyping as a robust method for evaluating CSF samples from patients diagnosed with hematologic malignancies.

Rapid cognitive decline in a patient with chronic lymphocytic leukaemia: a case report

BackgroundThe use of monoclonal antibodies in various settings has been linked to the development of progressive multifocal leukoencephalopathy (PML). Whilst this association is well-described with agents such as rituximab and natalizumab, the literature describing the occurrence of PML with ofatumumab therapy (especially in a haematology setting) is sparse. This case aims to draw attention to the above association with a particular focus on the mechanisms by which B-cell-depleting therapy can precipitate PML during the treatment of haematological malignancy.Case presentationA 68-year-old Caucasian man presented with acute-on-subacute confusion and reduced mobility. He had a history of chronic lymphocytic leukaemia for which he had completed six cycles of ofatumumab and chlorambucil 2 months prior to presentation. Biochemistry, physical examination and imaging were unremarkable on admission. Subsequent neurological examination demonstrated diminished reflexes and an extensor right plantar, while magnetic resonance imaging (MRI) assessment revealed white matter hyperintensities in the frontal lobes with restricted diffusion surrounding these areas. Cerebrospinal fluid (CSF) analysis demonstrated normal cell counts and chemistry but detected John Cunningham virus (JCV) via polymerase chain reaction (PCR), with a quantitative value of 41,850 gEg/ml. CSF immunophenotyping excluded malignant processes. A diagnosis of PML was confirmed, and with the support of palliative care, the patient was discharged to a hospice for ongoing care with the family’s agreement.ConclusionPML remains a rare complication of ofatumumab treatment. Nevertheless, clinicians should maintain a certain level of suspicion for this risk, especially in the context of patients presenting with clinical syndromes of encephalopathy and focal neurologic deficits. Furthermore, research to better our understanding of the manifold links between B-cell function and JCV regulation could provide valuable information for use in the future prevention and treatment of PML.

Immunophenotyping of the cerebrospinal fluid as a prognostic factor at diagnosis of acute lymphoblastic leukemia in children and adolescents

ABSTRACTThis study aimed at evaluating the use of immunophenotyping (IMP) in the identification of blast cells in the cerebrospinal fluid (CSF) of children and adolescents with acute lymphoblastic leukemia (ALL). Sixty-seven patients aged 18 years or younger were included. Fifty-five CSF samples were analyzed at initial diagnosis and 17 at the time of relapse. A cytological analysis (CA) was performed in all 72 samples, while IMP was done in 63. Blasts were identified in only three samples by CA, whereas all three samples were found negative by IMP, one of which had no isolation of nucleated cells after centrifugation. Among the samples analyzed by IMP, 11 showed a positive blast count, two of which had been inconclusive using CA. No equivalence was found between CA and IMP results (p = 0.55). CSF IMP positivity was not associated with other risk factors for ALL relapse. Among the 55 patients included at the time of diagnosis of ALL, eight relapsed during follow-up. Considering the cases of central nervous system (CNS) relapse, one of the patients belonged to the CSF IMP-positive group (11%) at diagnosis, and the other two cases, to the IMP-negative (5%) group. Detection of CSF blast cells using IMP was associated with a worse overall (p < 0.0001) and event-free survival (p < 0.0001). These results show that CSF IMP may be a useful additional method to conventional CA in the diagnosis of CNS involvement in ALL, and for the identification of high-risk subgroups that would benefit from an intensified therapy.

Comprehensive Immunophenotyping of Cerebrospinal Fluid Cells in Patients with Neuroimmunological Diseases

We performed unbiased, comprehensive immunophenotyping of cerebrospinal fluid (CSF) and blood leukocytes in 221 subjects referred for the diagnostic work-up of neuroimmunological disorders to obtain insight about disease-specific phenotypes of intrathecal immune responses. Quantification of 14 different immune cell subsets, coupled with the assessment of their activation status, revealed physiological differences between intrathecal and systemic immunity, irrespective of final diagnosis. Our data are consistent with a model where the CNS shapes intrathecal immune responses to provide effective protection against persistent viral infections, especially by memory T cells, plasmacytoid dendritic cells, and CD56(bright) NK cells. Our data also argue that CSF immune cells do not simply reflect cells recruited from the periphery. Instead, they represent a mixture of cells that are recruited from the blood, have been activated intrathecally and leave the CNS after performing effector functions. Diagnosis-specific differences provide mechanistic insight into the disease process in the defined subtypes of multiple sclerosis (MS), neonatal onset multisystem inflammatory disease, and Aicardi-Goutières syndrome. This analysis also determined that secondary-progressive MS patients are immunologically closer to relapsing-remitting patients as compared with patients with primary-progressive MS. Because CSF immunophenotyping captures the biology of the intrathecal inflammatory processes, it has the potential to guide optimal selection of immunomodulatory therapies in individual patients and monitor their efficacy. Our study adds to the increasing number of publications that demonstrate poor correlation between systemic and intrathecal inflammatory biomarkers in patients with neuroimmunological diseases and stresses the importance of studying immune responses directly in the intrathecal compartment.

Flow Cytometric Immunophenotyping of the Blood and CSF of Patients with Multiple Sclerosis (P02.081)

Objective: To monitor intrathecal immune responses and to investigate their role in pathogenesis of multiple sclerosis (MS). Background Immune cells in the cerebrospinal fluid (CSF) better reflect intrathecal inflammation than peripheral blood. However, low cellularity and susceptibility to apoptosis makes immunophenotyping of CSF challenging. Design/Methods: CSF and concomitant peripheral blood was collected in prospective manner in untreated patients (N=36) who presented for the diagnostic work-up of neuroimmunological disorder. Cells were isolated from 10-15cc of CSF within 15 min of collection. An optimized 12 color flow cytometric immunophenotyping (FCI) protocol was used to identify immune subpopulations (CD4, CD8 T cells, B cells, CD56bright- and CD56dim NK cells, monocytes, myeloid and plasmacytoid dendritic cells (DCs)) and their activation markers. FCI results were analyzed in an unbiased manner with the absolute cell count in blood and CSF and with the diagnosis established by the clinicians. Results: Upon unblinding, our cohort consisted of clinically isolated syndrome (CIS, N=2), relapsing-remitting MS (RR-MS, N=18), primary progressive MS (PP-MS, N=4), secondary progressive MS (SP-MS, N=4), non-inflammatory neurological diseases (NIND, N=4) and other inflammatory neurological diseases (OIND, N=5) controls. CSF was enriched for T cells, HLA-DR+ (i.e. activated) T cells and CD56bright NK cells in comparison to blood. We observed positive correlations between blood and CSF B cells, and blood and CSF HLA-DR+ T cells and terminally-differentiated double negative T cells, suggesting a link between systemic and intrathecal immune activation. Intrathecal differences between MS patients and controls consisted of elevated B cell to monocyte ratio (p=0.0004) and elevated CD4/CD8 T cell ratio (p=0.0007). Conclusions: The CSF immunophenotyping has been adapted to all NIB clinical protocols, including interventional trials, so updated results will be presented. We expect that with expanded cohort and longitudinal data, CSF immunophenotyping will provide essential information about intrathecal immune responses in neuroimmunological diseases. Supported by: NIH Intramural Research Program. Disclosure: Dr. Han has nothing to disclose. Dr. Salgado has nothing to disclose. Dr. Alfahad has nothing to disclose. Dr. Bielekova has received royalty payments from NIH.

Cerebrospinal fluid B‐cell expansion in longitudinally extensive transverse myelitis associated with neuromyelitis optica immunoglobulin G

A first episode of central nervous system (CNS) demyelination may represent heterogeneous entities such as acute disseminated encephalomyelitis, clinically isolated syndrome, neuromyelitis optica (NMO), or multiple sclerosis. As new immune therapies become available, it is increasingly important to make an early diagnosis. Autoantibodies such as NMO immunoglobulin G (IgG) and myelin oligodendrocyte glycoprotein IgG are increasingly being employed to define subgroups of CNS demyelination or guide treatment. Similarly, cerebrospinal fluid (CSF) immunophenotyping can demonstrate B-lymphocyte subpopulation expansion, which has been used to guide therapy in other autoimmune CNS disorders. We present a report on a 15-year-old male with longitudinally extensive transverse myelitis with magnetic resonance imaging findings of oedema, cavitation, and gadolinium enhancement. NMO-IgG and aquaporin 4 IgG were positive; thus, we diagnosed a limited form of NMO. Acute CSF immunophenotyping revealed a 3.6% expansion of CD19 B-cell populations, whereas a comparison group of five children (4 males, age range 2-15y; mean age 7y) with other neurological disorders showed only a 0.51% expansion (SD 0.25%). In view of the diagnosis of a 'limited form of neuromyelitis optica', we therefore elected to treat him aggressively from the outset with a prolonged steroid regimen and mycophenylate mofetil. This case demonstrates a correlation between autoantibody production and CSF B lymphocyte expansion in an individual with CNS demyelination. These approaches could be used in individuals with a first episode of CNS demyelination to help delineate immunological subgroups and guide treatment.

Enterovirus infection of neuronal cells post‐Rituximab

Enterovirus infection of neuronal cells post‐Rituximab