Survival after allogeneic transplantation according to pretransplant minimal residual disease and conditioning intensity in patients with acute myeloid leukemia.

Background: The measurement of minimal residual disease (MRD) by multiparametric flow cytometry (MFC) before hematopoietic stem cell transplantation (HSCT) in patients with acute myeloid leukemia (AML) is a powerful prognostic factor. The interaction of pretransplant MRD and the conditioning intensity has not yet been clarified. Objective: The aim of this study is to analyze the transplant outcomes of patients with AML who underwent HSCT in complete remission (CR), comparing patients with positive MRD (MRD+) and negative MRD (MRD-) before HSCT, and the interaction between conditioning intensity and pre-HSCT MRD. Study design: We retrospectively analyzed the transplant outcomes of 118 patients with AML who underwent HSCT in CR in a single institution, comparing patients with MRD+ and MRD- before HSCT using a cutoff of 0.1% on MFC, and the interaction between conditioning intensity and pre-HSCT MRD. Results: Patients with MRD+ before HSCT had a significantly worse 2-year (2y) event-free survival (EFS) (56.5% vs. 32.0%, p = 0.018) than MRD- patients, due to a higher cumulative incidence of relapse (CIR) at 2 years (49.0% vs. 18.0%, p = 0.002), with no differences in transplant-related mortality (TRM) (2y-TRM, 19.0% and 25.0%, respectively, p = 0.588). In the analysis stratified by conditioning intensity, in patients who received MAC, those with MRD- before HSCT had better EFS (p = 0.009) and overall survival (OS) (p = 0.070) due to lower CIR (p = 0.004) than MRD+ patients. On the other hand, the survival was similar in reduced intensity conditioning (RIC) patients regardless of the MRD status. Conclusions: Patients with MRD+ before HSCT have worse outcomes than MRD- patients. In patients who received MAC, MRD- patients have better EFS and OS due to lower CIR than MRD+ patients, probably because they represent a more chemo-sensitive group. However, among RIC patients, results were similar regardless of the MRD status.

Generation and external validation of a histological transformation risk model for patients with follicular lymphoma.

Follicular lymphoma (FL) is the most frequent indolent lymphoma. 10-15% of patients suffer histological transformation (HT) to a more aggressive lymphoma, usually diffuse large B cell lymphoma (DLBCL). This study aimed to validate and improve a genetic risk model to predict HT at diagnosis. We collected mutational data from diagnosis biopsies of 64 FL patients. We combined them with the data from a previously published cohort (total n = 104, 62 from non-transformed, and 42 from patients who did transform to DLBCL). This combined cohort was used to develop a nomogram to estimate the risk of HT. Prognostic mutated genes and clinical variables were assessed using Cox regression analysis to generate a risk model. The model was internally validated by bootstrapping and externally validated in an independent cohort. Its performance was evaluated using a concordance index and a calibration curve. The clinicogenetic nomogram included the mutational status of three genes (HIST1HE1, KMT2D, and TNFSR14) and high-risk FLIPI and predicted HT with a concordance index of 0.746. Patients were classified as being at low or high risk of transformation. The probability HT function at 24 months was 0.90 in the low-risk group vs. 0.51 in the high-risk group and, at 60 months, 0.69 vs. 0.15, respectively. In the external validation cohort, the probability HT function in the low-risk group was 0.86 vs. 0.54 in the high-risk group at 24 months, and 0.71 vs. 0.32 at 60 months. The concordance index in the external cohort was 0.552. In conclusion, we propose a clinicogenetic risk model to predict FL HT to DLBLC, combining genetic alterations in HIST1H1E, KMT2D, and TNFRSF14 genes and clinical features (FLIPI) at diagnosis. This model could improve the management of FL patients and allow treatment strategies that would prevent or delay transformation.

Patients with secondary acute myeloid leukemia undergoing allogeneic stem-cell transplant have inferior outcomes than de novo acute myeloid leukemia regardless minimal residual disease level by flow cytometry.

Secondary acute myeloid leukemia (s-AML) patients have a poor prognosis and currently the only curative therapy is allogeneic stem-cell transplant (HSCT). However, we do not yet know whether transplantation is sufficient to reverse the poor prognosis compared to de novo AML patients. We analyzed survival after HSCT comparing a cohort of 58 patients with s-AML versus 52 de novo patients who were transplanted between 2012 and 2020. Patients with s-AML had worse event-free survival (EFS) (p = 0.001) and overall survival (OS) (p < 0.001) compared to de novo AML due to an increased risk of relapse (p = 0.06) and non-relapse mortality (p = 0.03). The main difference in survival was observed in patients who achieved complete remission (CR) before HSCT (EFS p = 0.002 OS and <0.001), regardless minimal residual disease (MRD) by |multiparametric flow cytometry cohorts. In patients transplanted with active disease (AD), the prognosis was adverse in both s-AML and de novo AML groups (EFS p = 0.869 and OS p = 0.930). After excluding patients with AD, we stratified the cohort according to conditioning intensity, noticing that s-AML who received MAC had comparable outcomes to de novo AML, but the survival differences remained among reduce intensity conditioning group. In conclusion, transplanted s-AML patients have worse survival among patients in CR before HSCT, regardless of MRD level by flow cytometry compared to de novo AML. MAC patients had similar outcomes irrespective of leukemia ontogeny.

Lymphoplasmacytic lymphoma and marginal zone lymphoma involving bone marrow: A diagnostic dilemma. Useful clinicopathological features to accurate the diagnosis.

Lymphoplasmacytic lymphoma (LPL) and marginal zone lymphoma (MZL) frequently infiltrate the bone marrow with similar histologic and immunohistochemical characteristics posing diagnostic problems. Bone marrow biopsy specimens from 25 LPL and 16 MZL have been studied, correlating with clinical, laboratory parameters and the MYD88_p.L265P mutation. Paratrabecular and interstitial infiltration pattern, serum IgM paraprotein levels, and MYD88_p.L265P mutation were significantly more frequent in LPL. Nodular or intrasinusoidal pattern with lymphocytosis and splenomegaly were associated with MZL diagnosis. Different clinical and histological parameters should be collected when LPL or MZL is suspected in bone marrow biopsy specimens.

Neuropathological findings in fatal COVID-19 and their associated neurological clinical manifestations.

Severe cases of Coronavirus Disease 2019 (COVID-19) can present with multiple neurological symptoms. The available neuropathological studies have described different lesions; the most frequent was the presence of neuroinflammation and vascular-related lesions. The objective of this study was to report the neuropathological studies performed in a medical institution, with abundant long intensive care unit stays, and their associated clinical manifestations. This is a retrospective monocentric case series study based on the neuropathological reports of 13 autopsies with a wide range of illness duration (13-108 days). A neuroinflammatory score was calculated based on the quantification of CD8- and CD68-positive cells in representative areas of the central nervous system. This score was correlated afterwards with illness duration and parameters related to systemic inflammation. Widespread microglial and cytotoxic T-cell activation was found in all patients. There was no correlation between the neuroinflammatory score and the duration of the illness; nor with parameters of systemic inflammation such as the peak of IL-6 or the HScore (a parameter of systemic macrophage activation syndrome). Two patients had global hypoxic ischaemic damage and five patients had subacute infarcts. One patient had many more brain vascular microthrombi compared to the others and multiple subacute pituitary infarcts. SARS-CoV-2 RNA was not detected with qRT-PCR. The proportion of brain lesions in severe COVID-19 patients could be related to illness duration. In our series, with abundant long hospitalisation stays, neuroinflammation was present in all patients and was more prominent between day 34 and day 45 after onset of symptoms. Clinical correlation showed that two patients with the highest neuroinflammatory scores had severe encephalopathies that were not attributable to any other cause. The second most frequent lesions were related to vascular pathology.

Secondary haemophagocytic lymphohistiocytosis in COVID-19: correlation of the autopsy findings of bone marrow haemophagocytosis with HScore.

BACKGROUND: Secondary haemophagocytic lymphohistiocytosis (sHLH) is characterised by a hyper activation of immune system that leads to multiorgan failure. It is suggested that excessive immune response in patients with COVID-19 could mimic this syndrome. Some COVID-19 autopsy studies have revealed the presence of haemophagocytosis images in bone marrow, raising the possibility, along with HScore parameters, of sHLH. AIM: Our objective is to ascertain the existence of sHLH in some patients with severe COVID-19. METHODS: We report the autopsy histological findings of 16 patients with COVID-19, focusing on the presence of haemophagocytosis in bone marrow, obtained from rib squeeze and integrating these findings with HScore parameters. CD68 immunohistochemical stains were used to highlight histiocytes and haemophagocytic cells. Clinical evolution and laboratory parameters of patients were collected from electronic clinical records. RESULTS: Eleven patients (68.7%) displayed moderate histiocytic hyperplasia with haemophagocytosis (HHH) in bone marrow, three patients (18.7%) displayed severe HHH and the remainder were mild. All HScore parameters were collected in 10 patients (62.5%). Among the patients in which all parameters were evaluable, eight patients (80%) had an HScore >169. sHLH was not clinically suspected in any case. CONCLUSIONS: Our results support the recommendation of some authors to use the HScore in patients with severe COVID-19 in order to identify those who could benefit from immunosuppressive therapies. The presence of haemophagocytosis in bone marrow tissue, despite not being a specific finding, has proved to be a very useful tool in our study to identify these patients.

Proposal and validation of a method to classify genetic subtypes of diffuse large B cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease whose prognosis is associated with clinical features, cell-of-origin and genetic aberrations. Recent integrative, multi-omic analyses had led to identifying overlapping genetic DLBCL subtypes. We used targeted massive sequencing to analyze 84 diagnostic samples from a multicenter cohort of patients with DLBCL treated with rituximab-containing therapies and a median follow-up of 6 years. The most frequently mutated genes were IGLL5 (43%), KMT2D (33.3%), CREBBP (28.6%), PIM1 (26.2%), and CARD11 (22.6%). Mutations in CD79B were associated with a higher risk of relapse after treatment, whereas patients with mutations in CD79B, ETS1, and CD58 had a significantly shorter survival. Based on the new genetic DLBCL classifications, we tested and validated a simplified method to classify samples in five genetic subtypes analyzing the mutational status of 26 genes and BCL2 and BCL6 translocations. We propose a two-step genetic DLBCL classifier (2-S), integrating the most significant features from previous algorithms, to classify the samples as N12-S, EZB2-S, MCD2-S, BN22-S, and ST22-S groups. We determined its sensitivity and specificity, compared with the other established algorithms, and evaluated its clinical impact. The results showed that ST22-S is the group with the best clinical outcome and N12-S, the more aggressive one. EZB2-S identified a subgroup with a worse prognosis among GCB-DLBLC cases.

Development and validation of a sequential two-step algorithm for the screening of individuals with potential polycythaemia vera.

In 2016, the WHO included haemoglobin values within normal ranges as a diagnostic criterion for Polycythaemia Vera (PV). Since then, concerns have arisen that a large number of patients are undergoing unnecessary screening for PV. To address this issue, we estimated the prevalence of JAK2 p.V617F in individuals with elevated haemoglobin or haematocrit and developed and validated a screening algorithm for PV. A total of 15,366 blood counts performed in seven non-consecutive days were reviewed, of which 1001 were selected for subsequent JAK2 p.V617F mutation screening. Eight (0.8%) new JAK2 p.V617F-mutated cases were detected. From ROC curves, a two-step algorithm was developed based on the optimal cut-off for the detection of the JAK2 p.V617F mutation. The algorithm was prospectively validated in an independent cohort of 15,298 blood counts. A total of 1595 (10.4%) cases met the criterion for haemoglobin or haematocrit, of whom 581 passed to step 2 (3.8% of the total). The JAK2 p.V617F mutation was detected in 7 of the 501 patients tested, which accounts for 0.04% of the total cohort and 0.4% of patients with erythrocytosis. In conclusion, this data show that our two-step algorithm improves the selection of candidates for JAK2 p.V617F testing.

Identification of Immunological Parameters as Predictive Biomarkers of Relapse in Patients with Chronic Myeloid Leukemia on Treatment-Free Remission.

BCR-ABL is an aberrant tyrosine kinase responsible for chronic myeloid leukemia (CML). Tyrosine kinase inhibitors (TKIs) induce a potent antileukemic response mostly based on the inhibition of BCR-ABL, but they also increase the activity of Natural Killer (NK) and CD8+ T cells. After several years, patients may interrupt treatment due to sustained, deep molecular response. By unknown reasons, half of the patients relapse during treatment interruption, whereas others maintain a potent control of the residual leukemic cells for several years. In this study, several immunological parameters related to sustained antileukemic control were analyzed. According to our results, the features more related to poor antileukemic control were as follows: low levels of cytotoxic cells such as NK, (Natural Killer T) NKT and CD8±TCRγß+ T cells; low expression of activating receptors on the surface of NK and NKT cells; impaired synthesis of proinflammatory cytokines or proteases from NK cells; and HLA-E*0103 homozygosis and KIR haplotype BX. A Random Forest algorithm predicted 90% of the accuracy for the classification of CML patients in groups of relapse or non-relapse according to these parameters. Consequently, these features may be useful as biomarkers predictive of CML relapse in patients that are candidates to initiate treatment discontinuation.

Diffuse follicular lymphoma variant with a typical diagnostic pattern and an unusually aggressive clinical presentation.

Diffuse follicular lymphoma (FL) variant is a rare condition that shows distinctive clinical, morphological, immunophenotypic, and molecular features that distinguish it from classical FL. Diffuse FL variant is characterized by a predominantly diffuse growth pattern, absence of the t (14;18) IGH/BCL2 translocation, CD23 expression, and presence of 1p36 deletion. Gene mutations involving STAT6 have been reported, with nuclear expression of STAT6 and phosphorylated STAT6 detected by immunohistochemistry. Patients frequently present with inguinal node involvement and low clinical stage. We describe the case of an 80-year-old female diagnosed with diffuse FL variant, presented with a classic diagnostic pattern and an unusual aggressive clinical onset.

Systemic Mastocytosis: Following the Tyrosine Kinase Inhibition Roadmap.

Systemic mastocytosis is a rare and heterogeneous disease characterized by mast cell proliferation and activation. KIT is a transmembrane tyrosine kinase which plays a key role in mast cell growth, differentiation and survival. After interaction with its ligand, the stem cell factor, KIT dimerizes activating downstream pathways involving multiple tyrosine kinases (PI3K, JAK/STAT, RAS/ERK). Activating mutations in KIT are detected in most cases of systemic mastocytosis, being the most common KIT D816V. Therefore, since the emergence of tyrosine kinase inhibitors, KIT inhibition has been an attractive approach when facing mastocytosis treatment. Initial reports showed that only the rare KIT D816V negative cases were responsive to tyrosine kinase inhibitors. However, the development of new tyrosine kinase inhibitors such as midostaurin or avapritinib with activity against mast cells carrying the D816V KIT mutation, has changed the landscape of this disease.

Early detection of lysosomal diseases by screening of cases of idiopathic splenomegaly and/or thrombocytopenia with a next-generation sequencing gene panel.

Lysosomal diseases (LD) are a group of about 70 rare hereditary disorders (combined incidence 1:5000) in which diverse lysosomal functions are impaired, impacting multiple organs and systems. The first clinical signs and symptoms are usually unspecific and shared by hundreds of other disorders. Diagnosis of LD traditionally relies on performing specific enzymatic assays, if available, upon clinical suspicion of the disorder. However, the combination of the insidious onset of LD and the lack of awareness on these rare diseases among medical personnel results in undesirable diagnostic delays, with unchecked disease progression, appearance of complications and a worsened prognosis. We tested the usefulness of a next-generation sequencing-based gene panel for quick, early detection of LD among cases of idiopathic splenomegaly and/or thrombocytopenia, two of the earliest clinical signs observed in most LD. Our 73-gene panel interrogated 28 genes for LD, 1 biomarker and 44 genes underlying non-LD differential diagnoses. Among 38 unrelated patients, we elucidated eight cases (21%), five with LD (GM1 gangliosidosis, Sanfilippo disease A and B, Niemann-Pick disease B, Gaucher disease) and three with non-LD conditions. Interestingly, we identified three LD patients harboring pathogenic mutations in two LD genes each, which may result in unusual disease presentations and impact treatment. Turnaround time for panel screening and genetic validation was 1 month. Our results underline the usefulness of resequencing gene panels for quick and cost-effective screening of LDs and disorders sharing with them early clinical signs.

Bone marrow infiltration by flow cytometry at diffuse large B-cell lymphoma NOS diagnosis implies worse prognosis without considering bone marrow histology.

BACKGROUND: The significance of discrepant findings between histology (BMB) and flow cytometry (FC) in bone marrow (BM) examination at diffuse large B-cell lymphoma (DLBCL) diagnosis is uncertain. METHODS: We performed a 5-year retrospective single-center study of patients diagnosed by DLBCL not otherwise specified (n = 82), divided into three groups according to BM infiltration at diagnosis: BMB-/FC- (75.6%), BMB+/FC+ (13.4%), and BMB-/FC+ (11%). RESULTS: Median infiltration by FC analysis of the BMB-/FC+ group was 0.8% and if we considered BM infiltration as positive in all cases, 4/9 would be upstaged. Median follow was 33 months. Event-free survival (EFS) after 18 months was 82, 23, and 27% for BMB-/FC-, BMB-/FC+, and BMB+/FC+, respectively (p < .001). After 18 months of observation, OS was 87, 46, and 55% for BMB-/FC-, BMB-/FC+, and BMB+/FC+, respectively (p = .001). In multivariate analysis (BM infiltration vs. cell-of-origin according to Hans algorithm and standard IPI), BM infiltration was independently associated with EFS (HR: 1.94, 95% CI: 1.3-2.9) and overall survival (HR: 1.69, 95% CI: 1.1-2.7). CONCLUSION: In summary, minimal BM infiltration, detected by FC but not by BMB, has same prognostic implications than overt BM infiltration and should be considered as extranodal involvement regardless the infiltration quantity.

Safety and efficacy of bosutinib in fourth-line therapy of chronic myeloid leukemia patients.

Bosutinib is a second-generation tyrosine kinase inhibitor (2GTKI) approved at 400 mg once daily (QD) as first-line therapy in patients with chronic myeloid leukemia (CML) patients and at 500 mg QD in patients who are resistant to or intolerant of prior therapy. In clinical practice, bosutinib is often given to patients who have failed imatinib, nilotinib, and dasatinib (i.e., as fourth-line treatment), despite the limited data on its clinical benefit in this setting. We have retrospectively evaluated the results of bosutinib in a series of 62 CML patients who have failed to prior treatment with all three, imatinib, nilotinib, and dasatinib. Median time on TKI treatment before bosutinib start was 105 (9-163) months, and median duration on bosutinib was 9 months (1-30). Overall, probabilities to achieve complete cytogenetic response (CCyR) and major molecular response (MMR) were 25% and 24% respectively. After a median follow-up period of 14 months, the event-free survival and progression-free survival were 68 and 85%, respectively. Sixty-four percent of patients in CCyR at the time of bosutinib start were able to achieve MMR. In contrast, patients without CCyR, probabilities to obtain CCyR and MMR were 25% and 14%. Bosutinib was well tolerated in this heavily pretreated patients' cohort. Pleural effusions and diarrhea were the most frequent grade II-IV side effects, leading to treatment discontinuation in 16% of patients. Bosutinib is an effective treatment option for patients who have failed previous 2GTKIs due to intolerance. However, efficacy seems to be related to the molecular response that the patient achieved prior to bosutinib.

Prostatic-Like Syndrome in a Woman with Chronic Lymphocytic Leukemia: Sequential Kinase Inhibitor Therapy.

Chronic lymphocytic leukemia (CLL) is an incurable lymphoproliferative disorder with a heterogeneous genetic and clinical course. Two kinase inhibitors, ibrutinib and idelalisib, have demonstrated achievement of complete and durable remissions in relapse/refractory genetically unselected CLL patients. We present a case of relapsed CLL with extensive disease and hourglass deformity of urinary bladder as a result of the compression of two extraperitoneal paravesical soft tissue bulky masses, with excellent response to sequential kinase inhibitor therapy.

Results of a phase II study of lenalidomide and rituximab for refractory/relapsed chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is an incurable disease in need of new therapeutic strategies. The immunomodulatory agent, lenalidomide, has shown activity as salvage therapy for CLL. In this phase II trial, we combined lenalidomide with rituximab in 25 patients (range, 41-79) with refractory/relapsed CLL. Lenalidomide was administered orally on escalating doses, with cycle 1 doses of 2.5mg daily on days 1-7, 5mg on days 8-14, and 10mg on days 15-21 followed by 7days off. On cycle 2 and beyond, lenalidomide was administered at 20mg daily on days 1-21. Rituximab was administered at 375mg/m(2) intravenously on a weekly basis for the first cycle starting on day 15 for 4 doses, with each cycle being 28days. Treatment was continued until disease progression or toxicity. Overall response rate was 45.8% on intent-to-treat and 61.1% in evaluable patients (all partial responses). Median time to treatment failure was 14.3 months for evaluable patients, and median overall survival was not reached. The most common grade 3/4 toxicity was neutropenia (72% of patients). The most common nonhematologic toxicity was infection (29% of patients). Lenalidomide combined with rituximab showed activity in heavily treated refractory CLL with an acceptable toxicity profile.