On the Robustness of Bayesian Neural Networks to Adversarial Attacks.

Vulnerability to adversarial attacks is one of the principal hurdles to the adoption of deep learning in safety-critical applications. Despite significant efforts, both practical and theoretical, training deep learning models robust to adversarial attacks is still an open problem. In this article, we analyse the geometry of adversarial attacks in the over-parameterized limit for Bayesian neural networks (BNNs). We show that, in the limit, vulnerability to gradient-based attacks arises as a result of degeneracy in the data distribution, i.e., when the data lie on a lower dimensional submanifold of the ambient space. As a direct consequence, we demonstrate that in this limit, BNN posteriors are robust to gradient-based adversarial attacks. Crucially, by relying on the convergence of infinitely-wide BNNs to Gaussian processes (GPs), we prove that, under certain relatively mild assumptions, the expected gradient of the loss with respect to the BNN posterior distribution is vanishing, even when each NN sampled from the BNN posterior does not have vanishing gradients. The experimental results on the MNIST, Fashion MNIST, and a synthetic dataset with BNNs trained with Hamiltonian Monte Carlo and variational inference support this line of arguments, empirically showing that BNNs can display both high accuracy on clean data and robustness to both gradient-based and gradient-free adversarial attacks.

NFKBIE mutations are selected by the tumor microenvironment and contribute to immune escape in chronic lymphocytic leukemia.

Loss-of-function mutations in NFKBIE, which encodes for the NF-κB inhibitor IκBε, are frequent in chronic lymphocytic leukemia (CLL) and certain other B-cell malignancies and have been associated with accelerated disease progression and inferior responses to chemotherapy. Using in vitro and in vivo murine models and primary patient samples, we now show that NFKBIE-mutated CLL cells are selected by microenvironmental signals that activate the NF-κB pathway and induce alterations within the tumor microenvironment that can allow for immune escape, including expansion of CD8+ T-cells with an exhausted phenotype and increased PD-L1 expression on the malignant B-cells. Consistent with the latter observations, we find increased expression of exhaustion markers on T-cells from patients with NFKBIE-mutated CLL. In addition, we show that NFKBIE-mutated murine CLL cells display selective resistance to ibrutinib and report inferior outcomes to ibrutinib treatment in NFKBIE-mutated CLL patients. These findings suggest that NFKBIE mutations can contribute to CLL progression through multiple mechanisms, including a bidirectional crosstalk with the microenvironment and reduced sensitivity to BTK inhibitor treatment.

Real-World Outcome of Treatment with Single-Agent Ibrutinib in Italian Patients with Chronic Lymphocytic Leukemia: Final Results of the EVIdeNCE Study.

Real-world data in clinical practice are needed to confirm the efficacy and safety that ibrutinib has demonstrated in clinical trials of patients with chronic lymphocytic leukemia (CLL). We described the real-world persistence rate, patterns of use, and clinical outcomes in 309 patients with CLL receiving single-agent ibrutinib in first line (1L, n = 118), 2L (n = 127) and ≥3L (n = 64) in the prospective, real-world, Italian EVIdeNCE study. After a median follow-up of 23.9 months, 29.8% of patients discontinued ibrutinib (1L: 24.6%, 2L: 29.9%, ≥3L: 39.1%), mainly owing to adverse events (AEs)/toxicity (14.2%). The most common AEs leading to discontinuation were infections (1L, ≥3L) and cardiac events (2L). The 2-year retention rate was 70.2% in the whole cohort (1L: 75.4%, 2L: 70.1%, ≥3L: 60.9%). The 2-year PFS and OS were, respectively, 85.4% and 91.7% in 1L, 80.0% and 86.2% in 2L, and 70.1% and 80.0% in ≥3L. Cardiovascular conditions did not impact patients' clinical outcomes. The most common AEs were infections (30.7%), bleeding (12.9%), fatigue (10.0%), and neutropenia (9.7%), while grade 3-4 atrial fibrillation occurred in 3.9% of patients. No new safety signals were detected. These results strongly support ibrutinib as a valuable treatment option for CLL.

PlentiPlex™ MYD88 Waldenström lymphoma qPCR assay: A highly sensitive method for detection of MYD88 L265P mutation.

INTRODUCTION: Agarose gel-based conventional and real-time allele-specific polymerase chain reaction (AS-PCR) assays are currently used for sensitive detection and quantification of MYD88 L265P mutation. Visual inspection of an agarose gel can often be ambiguous. We propose a new allele-specific quantification PCR (AS-qPCR) assay, PlentiPlex™ MYD88 Waldenström lymphoma qPCR assay, that uses Intercalating Nucleic Acid (INA®) technology for increased affinity and specificity. METHODS: This study compares PlentiPlex™ MYD88 Waldenström lymphoma qPCR assay with conventional AS-PCR. We included a total of 102 peripheral and bone marrow blood samples from 94 patients with a lymphoproliferative disorder. Droplet digital PCR (ddPCR) was used as a third method in case of discrepancy. RESULTS: A positive percent agreement of 100% (95% CI 0.92-1.0) and a negative percent agreement of 98% (95% CI 0.90-1.0) were found between the conventional AS-PCR and the AS-qPCR methods. Including the ddPCR results to validate the discrepant cases, the sensitivity and specificity of PlentiPlex™ MYD88 Waldenström lymphoma qPCR Assay were 1.0 (95% CI 0.97-1.0) and 1.0 (95% CI 0.96-1.0), respectively. CONCLUSION: Our data demonstrate that PlentiPlex™ MYD88 Waldenström lymphoma qPCR assay is a fast, highly sensitive, and specific method for the detection of MYD88 L265P compared with conventional AS-PCR.

Ibrutinib as first line therapy in chronic lymphocytic leukemia patients over 80 years old: A retrospective real-life multicenter Italian cohort.

Although chronic lymphocytic leukemia (CLL) predominantly affects the elderly, limited data exists about the outcomes of over 80-year-old patients, usually underrepresented in clinical trials. We conducted a multicenter study enrolling 79 consecutive CLL patients ≥80 years at the time of frontline therapy, all treated with ibrutinib. Nearly 48% of cases exhibited unmutated IGHV genes, 32% 17p deletion, and 39.2% TP53 mutations; 63.3% displayed a cumulative illness rating scale (CIRS) > 6. The overall response rate on ibrutinib, computed in 74/79 patients (5 patients excluded for early withdrawal), was 89.9%. After a median follow-up of 28.9 months, the median progression-free survival (PFS) and overall survival (OS) were 42.5 and 51.8 months, respectively. CIRS>6 and temporary discontinuation of ibrutinib lasting for 7-30 days were the only parameters associated with a significantly shorter PFS and were both relevant in predicting a shorter PFS compared to patients with CIRS≤6 and therapy discontinuation ≤7 days. The most common grade≥3 adverse events were infections (25.5%), neutropenia (10.1%), and anemia (2.5%). Eighteen patients (22.8%) experienced a cardiovascular event, including grade-2 atrial fibrillation (n = 9; 11%), grade-2 hypertension (n = 5; 6%), heart failure (n = 3; 3%), and acute coronary syndrome (n = 1; 1%). Mild bleeding events were observed in 27 patients (34.2%). Ibrutinib was permanently discontinued in 26 patients due to progressive disease (n = 11, including 5 Richter's syndromes), secondary malignancies (n = 6), infections (n = 3), cardiac failure (n = 3), severe bleeding (n = 2), and sudden death (n = 1). In conclusion, our analyses confirmed the overall effectiveness and favorable safety profile of the ibrutinib-single agent therapeutic approach in CLL patients ≥80 years.

Unselected donor-derived hematopoietic stem cells boost for Chimeric Antigen Receptor T-cell associated hematotoxicity.

Hematological toxicity following Chimeric Antigen Receptor-T therapy in a patient with a prior allogeneic stem cell transplantation was resolved by the infusion of unselected donor-derived stem cell boost. Due to the donor's lymphocytes, the patient experienced a well-controlled flare-up of acute graft versus host disease.

What Does Atypical Chronic Lymphocytic Leukemia Really Mean? A Retrospective Morphological and Immunophenotypic Study.

Atypical chronic lymphocytic leukemia (CLL) is still defined according to morphological criteria. However, deviance from the typical surface immunological profile suggests an atypical immunological-based CLL. A large cohort of patients with CLL was retrospectively evaluated aiming at assessing morphological (FAB criteria), immunophenotypical (two or more discordances from the typical profile), and clinical-biological features of atypical CLL. Compared to typical cases, morphologically atypical CLL showed a greater percentage of unmutated IgVH and CD38 positivity, and a higher expression of CD20. Immunophenotypically atypical CLL was characterized by more advanced clinical stages, higher expression of CD20, higher rate of FMC7, CD79b and CD49d positivity, and by an intermediate-high expression of membrane surface immunoglobulin, compared to typical cases. When patients were categorized based on immunophenotypic and morphologic concordance or discordance, no difference emerged. Finally, morphological features better discriminated patients' prognosis in terms of time-to-first treatment, while concordant atypical cases showed overall a worse prognosis. Discordant cases by immunophenotype and/or morphology did not identify specific prognostic groups. Whether-in the era of molecular markers used as prognostic indicators-it does make sense to focus on morphology and immunophenotype features in CLL is still matter of debate needing further research.

Patients' preferences for chronic lymphocytic leukemia treatment: The CHOICE study.

Chronic lymphocytic leukemia (CLL) therapies differ in efficacy, side effects, route, frequency, and duration of administration. We assessed patient preferences for treatment attributes and evaluated associations with disease stage, treatment line, and socio-demographic characteristics in a cross sectional, observational study conducted at 16 Italian hematology centers. Study visits occurred between February and July 2020; 401 adult patients with CLL (201 Watch and Wait (W&W), 200 treated) participated in a discrete choice experiment (DCE), composed of 8 choices between pairs of treatment profiles with different levels of 5 attributes of currently available CLL treatments (length of response, route and duration of administration, risk of side effects including diarrhea, infections, or organ damage). Health-related quality of life was assessed with the EQ-5D-5L, EORTC QLQ-C30 and QLQ CLL-16. Previously treated patients had longer disease duration (7 vs. 5 years), higher prevalence of serious comorbidities (45.5% vs. 36.2%) and high-risk molecular markers (unmutated IGHV 55.6% vs. 17.1%; TP53 mutation 15.2% vs. 4.0%). Health-related quality of life scores were similar between groups. In the DCE, W&W patients rated "possible occurrence of infections" highest (relative importance [RI] = 36.2%), followed by "treatment and relevant duration" (RI = 28.0%) and "progression-free survival (PFS)" (RI = 16.9%). Previously treated patients rated "treatment and relevant duration" highest (RI = 33.3%), followed by "possible occurrence of infections" (RI = 28.8%), "possible occurrence of organ damage" (RI = 19.4%), and "PFS" (RI = 9.8%). Concern over infection was rated highest overall; unexpectedly PFS was not among the most important criteria in either group, suggesting that the first COVID-19 pandemic wave may have influenced patient preferences and concerns about CLL therapy options.

What is the best treatment strategy before autologous peripheral blood stem cell transplantation in POEMS syndrome?

Autologous peripheral blood stem cell transplantation (aPBSCT) provides optimal outcomes in POEMS syndrome but the definition of the best treatment before aPBSCT remains to be defined because of the rarity of the disease and the heterogeneity of published case series. We collected clinical and laboratory data of patients with POEMS syndrome undergoing aPBSCT from 1998 to 2020 in ten Italian centers. The primary endpoint of the study was to evaluate the impact of prior therapies and mobilization regimen on outcome. We divided the patients into three groups: patients who did not receive any treatment before transplant (15 patients, group A: front-line), patients pre-treated with other agents (14 patients, group B) and patients treated with cyclophosphamide as their mobilizing regimen (16 patients, group C). The three groups did not show differences in terms of demographic and clinical characteristics. All 45 patients underwent aPBSCT after a high-dose melphalan conditioning regimen, with a median follow-up of 77 months (range, 37-169 months). The responses were not statistically different between the three groups (P=0.38). Progression-free and overall survival rates at 6 years were: 70% (95% confidence interval: 55-85%) and 91% (95% confidence interval: 82-99) 65%, respectively, and did not differ between the three groups. The cumulative incidence of transplant-related mortality and relapse was 4% and 36%, respectively. In conclusion, in a relatively large number of patients with POEMS syndrome, undergoing an autologous transplant, pre-treatment and disease status at transplant did not appear to have an impact on major transplant outcomes.

Donor specific anti-HLA antibodies in hematopoietic stem cell transplantation. Single Center prospective evaluation and desensitization strategies employed.

BACKGROUND: In the setting of mismatched-hematopoietic stem cells transplantation, the detection of antibodies directed against donor-specific HLA allele(s) or antigen(s) (DSA) represents a barrier for engraftment. It is thus necessary to plan an immunosuppressive strategy, or to select an alternative donor. This prospective study aimed at evaluating the efficacy of our strategy for testing DSAs and the efficacy of the desensitization strategy (DS) employed between November 2017 and November 2020. MATERIALS AND METHODS: The anti-HLA antibody search was performed using the Luminex bead assays (Lifecode ID and LSA I/II-Immucor) and expressed as mean fluorescence intensity (MFI >1,000 positive). If the patient had DSAs and no alternative donors, a DS was employed with rituximab (day -15), 2 single volume plasmaphereses (PP; days -9 and -8), intravenous immunoglobulins (day -7) and infusion of HLA selected platelets, if persistent DSAs were directed against class I HLA. DS was scheduled with or without PP, according to the DSA MFI (>1,000 or <5,000) and FCXM (flow cytometry crossmatch). RESULTS: Twenty-two out of 126 patients (17.46%) showed anti-HLA antibodies, 5 of them DSAs (3.97% of total); 3 patients underwent DS obtaining engraftment. Female gender (p=0.033) and a history of previous pregnancies or miscarriages (p=0.009) showed a statistically significant impact on alloimmunization. Factors associated with a delayed neutrophil engraftment were patient's female gender (p=0.039), stem cell source (p=0.025), and a high HSCT-specific comorbidity index (p=0.028). None of the analyzed variables, including the DSA detection, influenced engraftment. CONCLUSIONS: Our study confirms the importance to test DSAs in mismatched-hematopoietic stem cells transplantation The DS used proved successful in removing DSAs. Prospective multicenter studies are needed to better define and validate consensus strategies on DSA management in HSCT.

Treatment Sequencing and Outcome of Chronic Lymphocytic Leukemia Patients Treated at Fondazione Policlinico Universitario Agostino Gemelli IRCCS: A Thirty-Year Single-Center Experience.

BACKGROUND: This monocentric retrospective study describes the treatment patterns and outcomes of chronic lymphocytic leukemia (CLL) patients. METHODS: Adult CLL patients treated between 1992 and 2022 were included. The time to next treatment (TTNT) was defined as the time from the treatment's start to the start of a subsequent therapy or death. The time to next treatment failure or death (TTNTF) was defined as the time from treatment discontinuation to the discontinuation of a subsequent therapy or death. RESULTS: Of 637 registered patients, 318 (49.9%) received treatment. We evaluated 157 cBTKi-exposed, 34 BCL2i-exposed cBTKi-naïve, and 26 double-exposed patients. The five-year TTNT values in the cBTKi-exposed patients were 80% (median NR), 40% (median 40 months), and 21% (median 24 months) months in the first line (1L), second line (2L), and beyond the second line (>2L), respectively (p < 0.0001). The five-year TTNT values in the BCL2i-exposed patients were 83% (median NR), 72% (median NR), 12% (median 28 months) in the 1L, 2L, and >2L, respectively (p = 0.185). The median TTNTF was 9 months (range 1-87) after cBTKi and 17 months (range 8-49) after both a cBTKi and BCL2i. CONCLUSIONS: This study suggests that, in CLL patients, the earlier we used targeted therapies, the better was the outcome obtained. Nonetheless, the poor outcomes in the advanced lines of therapy highlight the need for more effective treatments.

Role of microRNAs in Chronic Lymphocytic Leukemia.

Chronic Lymphocytic Leukemia (CLL) is the most common form of leukemia in adults, with a highly variable clinical course. Improvement in the knowledge of the molecular pathways behind this disease has led to the development of increasingly specific therapies, such as BCR signaling inhibitors and BCL-2 inhibitors. In this context, the emerging role of microRNAs (miRNAs) in CLL pathophysiology and their possible application in therapy is worth noting. MiRNAs are one of the most important regulatory molecules of gene expression. In CLL, they can act both as oncogenes and tumor suppressor genes, and the deregulation of specific miRNAs has been associated with prognosis, progression, and drug resistance. In this review, we describe the role of the miRNAs that primarily impact the disease, and how these miRNAs could be used as therapeutic tools. Certainly, the use of miRNAs in clinical practice is still limited in CLL. Many issues still need to be solved, particularly regarding their biological and safety profile, even if several studies have suggested their efficacy on the disease, alone or in combination with other drugs.

Lymphadenopathy as a predictor of progression during venetoclax treatment in chronic lymphocytic leukemia. A campus chronic lymphocytic leukemia study.

Clinical or biological parameters useful to predict progression during treatment in real-life setting with ibrutinib, idelalisib and venetoclax in relapsed/refractory chronic lymphocytic leukemia (CLL) are still debated. We conducted a multi-center retrospective study on CLL patients treated with ibrutinib and/or idelalisib who were switched to venetoclax for progression or due to adverse events to identify any clinical and/or biological parameters useful to predict progression during treatment with venetoclax. Of all the 128 evaluable patients, 81 had received ibrutinib prior to switching to venetoclax, 35 had received idelalisib and 12 both. When comparing the three subgroups, we did not notice any statistical difference in terms of clinical or biological features. No variable at baseline and at different time points during the follow-up (at 6, 12, 18 and 24 months) was found to predict progression nor to have significance for Progression Free Survival (PFS) in the ibrutinib group and in the idelalisib group and in subgroups according to the line of treatment. Analyzing the data of the venetoclax treatment, after a median follow up of 14.3 months, median PFS was not reached and estimated 3-year PFS was 54%. Of the 128 patients treated with venetoclax, 28 (22%) experienced progressive disease. At multivariate analysis for predictive factors for progression, lymph node diameter >56.5 mm before starting treatment emerged as an independent risk factor for progression. The lymph node predictive role for progression during venetoclax treatment could be a new parameter that deserves to be investigate in future studies.

The immunomodulatory molecule TIGIT is expressed by chronic lymphocytic leukemia cells and contributes to anergy.

T-cell immunoreceptor with Ig and ITIM domains (TIGIT) is an inhibitory checkpoint receptor that negatively regulates Tcell responses. CD226 competes with TIGIT for binding to the CD155 ligand, delivering a positive signal to the T cell. Here we studied the expression of TIGIT and CD226 in a cohort of 115 patients with chronic lymphocytic leukemia (CLL) and report expression of TIGIT and CD226 by leukemic cells. By devising a TIGIT/CD226 ratio, we showed that CLL cells favoring TIGIT over CD226 are typical of a more indolent disease, while those favoring CD226 are characterized by a shorter time to first treatment and shorter progression-free survival after first treatment. TIGIT expression was inversely correlated to the B-cell receptor (BCR) signaling capacity, as determined by studying BTK phosphorylation, cell proliferation and interleukin- 10 production. In CLL cells treated with ibrutinib, in which surface IgM and BCR signaling capacity are temporarily increased, TIGIT expression was downmodulated, in line with data indicating transient recovery from anergy. Lastly, cells from patients with Richter syndrome were characterized by high levels of CD226, with low to undetectable TIGIT, in keeping with their high proliferative drive. Together, these data suggest that TIGIT contributes to CLL anergy by downregulating BCR signaling, identifying novel and actionable molecular circuits regulating anergy and modulating CLL cell functions.

High rate of durable responses with undetectable minimal residual disease with front-line venetoclax and rituximab in young, fit patients with chronic lymphocytic leukemia and an adverse biological profile: results of the GIMEMA phase II LLC1518 - VERITAS study.

The GIMEMA phase II LLC1518 VERITAS trial investigated the efficacy and safety of front-line, fixed-duration venetoclax and rituximab (VenR) in combination in young (≤65 years), fit patients with chronic lymphocytic leukemia and unmutated IGHV and/or TP53 disruption. Treatment consisted of the venetoclax ramp-up, six monthly courses of the VenR combination, followed by six monthly courses of venetoclax as a single agent. A centralized assessment of minimal residual disease (MRD) was performed by allele-specific oligonucleotide polymerase chain reaction assay on the peripheral blood and bone marrow at the end of treatment (EOT) and during the follow-up. The primary endpoint was the complete remission rate at the EOT. Seventy-five patients were enrolled; the median age was 54 years (range, 38-65), 96% had unmutated IGHV, 12% had TP53 disruption, and 4% had mutated IGHV with TP53 disruption. The overall response rate at the EOT was 94.7%, with a complete remission rate of 76%. MRD was undetectable in the peripheral blood of 69.3% of patients and in the bone marrow of 58.7% of patients. The 12-month MRD-free survival in the 52 patients with undetectable MRD in the peripheral blood at the EOT was 73.1%. After a median follow-up of 20.8 months, no cases of disease progression were observed. Three patients had died, two due to COVID-19 and one due to tumor lysis syndrome. The first report of the VERITAS study shows that front-line VenR was associated with a high rate of complete remissions and durable response with undetectable MRD in young patients with chronic lymphocytic leukemia and unfavorable genetic characteristics. ClinicalTrials.gov identifier: NCT03455517.