Preexisting and treatment-emergent autoimmune cytopenias in patients with CLL treated with targeted drugs.

Autoimmune cytopenias (AICs) affect 5% to 9% of patients with chronic lymphocytic leukemia (CLL). Targeted drugs-ibrutinib, idelalisib, and venetoclax-have a prominent role in the treatment of CLL, but their impact on CLL-associated AICs is largely unknown. In this study, we evaluated the characteristics and outcome of preexisting AICs and described the incidence, quality, and management of treatment-emergent AICs during therapy with targeted drugs in patients with CLL. We collected data from 572 patients treated with ibrutinib (9% in combination with an anti-CD20 monoclonal antibody), 143 treated with idelalisib-rituximab, and 100 treated with venetoclax (12% in combination with an anti-CD20 monoclonal antibody). A history of preexisting AICs was reported in 104 (13%) of 815 patients. Interestingly, 80% of patients whose AICs had not resolved when treatment with a targeted drug was started experienced an improvement or a resolution during therapy. Treatment-emergent AICs occurred in 1% of patients during ibrutinib therapy, in 0.9% during idelalisib therapy, and in 7% during venetoclax therapy, with an estimated incidence rate of 5, 6, and 69 episodes per 1000 patients per year of exposure in the 3 treatment groups, respectively. The vast majority of patients who developed treatment-emergent AICs had unfavorable biological features such as an unmutated IGHV and a del(17p) and/or TP53 mutation. Notably, despite AICs, 83% of patients were able to continue the targeted drug, in some cases in combination with additional immunosuppressive agents. Overall, treatment with ibrutinib, idelalisib, or venetoclax seems to have a beneficial impact on CLL-associated AICs, inducing an improvement or even a resolution of preexisting AICs in most cases and eliciting treatment-emergent AICs in a negligible portion of patients.

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.

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.

Beyond ibrutinib: novel BTK inhibitors for the treatment of chronic lymphocytic leukemia.

PURPOSE OF THE REVIEW: Ibrutinib was the first Bruton tyrosine kinase inhibitor (BTKi) approved for clinical use, contributing to a dramatic change in the treatment landscape of chronic lymphocytic leukemia (CLL). This review provides an overview of next-generation BTKi that have been recently approved or are being investigated for the treatment of CLL, specifically highlighting differences and similarities compared to ibrutinib. RECENT FINDINGS: Acalabrutinib presented comparable response rates to ibrutinib with lower rates of adverse events and is currently approved for the treatment of CLL. Zanubrutinib displayed excellent response rates with a lower incidence of BTKi-related adverse events, but major rates of neutropenia, and its approval is awaited. With the aim of overcoming drug resistance, noncovalent BTKi have been developed. Of all the explored agents to date, pirtobrutinib has shown promising results with manageable toxicities. SUMMARY: For the treatment of CLL, several effective therapeutic strategies to target BTK are or will soon be available: these drugs present different safety profiles, thus making it possible to tailor the treatment choice according to patient's characteristics. Importantly, noncovalent BTKi will provide a therapeutic chance also for those relapsed/refractory CLL patients who are BTKi-resistant and are considered an unmet clinical need.

Clinical implications of cancer gene mutations in patients with chronic lymphocytic leukemia treated with lenalidomide.

Lenalidomide is clinically active in chronic lymphocytic leukemia (CLL), but its effectiveness in the context of the CLL mutational landscape is unknown. We performed targeted capture sequencing of 295 cancer genes in specimens from 102 CLL patients with treatment-naïve disease (TN patients) and 186 CLL patients with relapsed/refractory disease (R/R patients) who received lenalidomide-based therapy at our institution. The most frequently mutated gene was SF3B1 (15%), followed by NOTCH1 (14%) and TP53 (14%), with R/R patients having significantly more TP53 mutations than did TN patients. Among all lenalidomide-treated patients, del(17p) (P ≤ .001), del(11q) (P = .032), and complex karyotype (P = .022), along with mutations in TP53 (P ≤ .001), KRAS (P = .034), and DDX3X (P ≤ .001), were associated with worse overall response (OR). R/R patients with SF3B1 and MGA mutations had significantly worse OR (P = .025 and .035, respectively). TN and R/R patients with del(17p) and TP53 mutations had worse overall survival (OS) and progression-free survival (PFS). In R/R patients, complex karyotype and SF3B1 mutations were associated with worse OS and PFS; DDX3X mutations were associated with worse PFS only. Weibull regression multivariate analysis revealed that TP53 aberrations (del(17p), TP53 mutation, or both), along with complex karyotype and SF3B1 mutations, were associated with worse OS in the R/R cohort. Taken together, cancer gene mutations in CLL contribute to the already comprehensive risk stratification and add to prognosis and response to treatment. The related trials were registered at www.clinicaltrials.gov as #NCT00267059, #NCT00535873, #NCT00759603, #NCT01446133, and #NCT01002755.

HIF-1α is over-expressed in leukemic cells from TP53-disrupted patients and is a promising therapeutic target in chronic lymphocytic leukemia.

In chronic lymphocytic leukemia (CLL), the hypoxia-inducible factor 1 (HIF-1) regulates the response of tumor cells to hypoxia and their protective interactions with the leukemic microenvironment. In this study, we demonstrate that CLL cells from TP53-disrupted (TP53 dis) patients have constitutively higher expression levels of the α-subunit of HIF-1 (HIF-1α) and increased HIF-1 transcriptional activity compared to the wild-type counterpart. In the TP53 dis subset, HIF-1α upregulation is due to reduced expression of the HIF-1α ubiquitin ligase von Hippel-Lindau protein (pVHL). Hypoxia and stromal cells further enhance HIF-1α accumulation, independently of TP53 status. Hypoxia acts through the downmodulation of pVHL and the activation of the PI3K/AKT and RAS/ERK1-2 pathways, whereas stromal cells induce an increased activity of the RAS/ERK1-2, RHOA/RHOA kinase and PI3K/AKT pathways, without affecting pVHL expression. Interestingly, we observed that higher levels of HIF-1A mRNA correlate with a lower susceptibility of leukemic cells to spontaneous apoptosis, and associate with the fludarabine resistance that mainly characterizes TP53 dis tumor cells. The HIF-1α inhibitor BAY87-2243 exerts cytotoxic effects toward leukemic cells, regardless of the TP53 status, and has anti-tumor activity in Em-TCL1 mice. BAY87-2243 also overcomes the constitutive fludarabine resistance of TP53 dis leukemic cells and elicits a strongly synergistic cytotoxic effect in combination with ibrutinib, thus providing preclinical evidence to stimulate further investigation into use as a potential new drug in CLL.

CD200 included in a 4-marker modified Matutes score provides optimal sensitivity and specificity for the diagnosis of chronic lymphocytic leukaemia.

CD200, a transmembrane type Ia glycoprotein belonging to the immunoglobulin superfamily, has been shown to have a differential expression in B-cell neoplasms. Here, we retrospectively assessed the diagnostic relevance of CD200 on 427 patients with B-cell chronic neoplasms in leukemic phase (median age, 69 y; range, 35-97 y). The final diagnosis based on the investigator's assessment was chronic lymphocytic leukaemia (CLL) in 75% of cases and non-CLL in 25% of cases. Sensitivity and specificity for the diagnosis of CLL (vs non-CLL) were calculated for the following markers: CD200, CD5, CD22, CD23, CD79b, FMC7, and SmIg. CD23 was the only marker without a statistically significant difference between the investigator assessment and the flowcytometric analysis. The other markers were unable-when individually evaluated-to discriminate between CLL and non-CLL, requiring the integration into a scoring system. The modified score no. 1 (addition of CD200) showed superimposable sensitivity and specificity compared with the Matutes score. The substitution of CD79b (modified score no. 2), surface membrane immunoglobulins (SmIg) (modified score no. 3), and CD79b and FMC7 (modified score no. 4) with CD200 showed that only the modified score no. 4 had both higher sensitivity and higher specificity compared with standard Matutes score. In conclusion, this work defines a simplified score, compared with the classical Matutes score, for the differential diagnosis of chronic B-cell leukaemia-which only requires 4 markers instead of 5 (CD5, CD23, CD200, and SmIg).

Prognostic relevance of oxidative stress measurement in chronic lymphocytic leukaemia.

OBJECTIVE: To evaluate the prognostic significance of oxidative stress (OS) and antioxidant defence status measurement in patients with chronic lymphocytic leukaemia (CLL). METHODS: d-ROMs test and BAP test were evaluated at diagnosis of 165 patients with CLL and correlated with clinical-biological features and prognosis. RESULTS: An increased oxidative damage (d-ROMs test) and a reduced antioxidant potential (BAP test) were found in CLL patients than normal controls (P<.0001). CLL patients with higher d-ROMs values had higher number of circulating white blood cells and lymphocytes, and higher values of ß2 -microglobulin. Higher d-ROMs values were found in female (P=.0003), in patients with unmutated IgVH (P=.04), unfavourable cytogenetics (P=.002) and more advanced clinical stage (P=.002). Higher BAP test values were found in patients expressing CD49d (P=.01) and with more advanced clinical stage (P=.004). At a median follow-up of 48 months, CLL patients with d-ROMs ≥418 CARR U were found to have a shorter time to first treatment (TFT) (P=.0002), and a reduced survival (P=.006) than others. CLL patients with BAP test values ≥2155 µmol/L had a shorter TFT (P=.008) and a shorter survival (P=.003). CONCLUSIONS: OS can affect CLL patients by concomitantly increasing reactive oxygen metabolites production and decreasing antioxidant defences.

Dysfunctional Vγ9Vδ2 T cells are negative prognosticators and markers of dysregulated mevalonate pathway activity in chronic lymphocytic leukemia cells.

The role of Vγ9Vδ2 T cells in chronic lymphocytic leukemia (CLL) is unexplored, although these cells have a natural inclination to react against B-cell malignancies. Proliferation induced by zoledronic acid was used as a surrogate of γδ TCR-dependent stimulation to functionally interrogate Vγ9Vδ2 T cells in 106 untreated CLL patients. This assay permitted the identification of responder and low-responder (LR) patients. The LR status was associated with greater baseline counts of Vγ9Vδ2 T cells and to the expansion of the effector memory and terminally differentiated effector memory subsets. The tumor immunoglobulin heavy chain variable region was more frequently unmutated in CLL cells of LR patients, and the mevalonate pathway, which generates Vγ9Vδ2 TCR ligands, was more active in unmutated CLL cells. In addition, greater numbers of circulating regulatory T cells were detected in LR patients. In multivariate analysis, the LR condition was an independent predictor of shorter time-to-first treatment. Accordingly, the time-to-first treatment was significantly shorter in patients with greater baseline numbers of total Vγ9Vδ2 T cells and effector memory and terminally differentiated effector memory subpopulations. These results unveil a clinically relevant in vivo relationship between the mevalonate pathway activity of CLL cells and dys-functional Vγ9Vδ2 T cells.

Prevention and management of infectious complications in patients with chronic lymphocytic leukemia (CLL) treated with BTK and BCL-2 inhibitors, focus on current guidelines.

CLL is associated with an increased risk of infectious complications. Treatment with BTK or BCL-2 inhibitors does not seem to increase significantly the risk of opportunistic infections, but the role of combination therapies including BTK and/or BCL-2 inhibitors remains to be established. Various infectious complications can be successfully prevented with appropriate risk management strategies. In this paper we reviewed the international guidelines on prevention and management of infectious complications in patients with CLL treated with BTK or BCL-2 inhibitors. Universal pharmacological anti-herpes, antibacterial or antifungal prophylaxis is not warranted. Reactivation of HBV should be prevented in HBsAg-positive subjects. For HBsAg-negative/HBcAb-positive patients recommendations differ, but in case of combination treatment should follow those for other, particularly anti-CD20, agent. Immunization should be provided preferably before the onset of treatment. Immunoglobulin therapy has favourable impact on morbidity but not mortality in patients with hypogammaglobulinemia and severe or recurrent infections. Lack of high-quality data and heterogeneity of patients or protocols included in the studies might explain differences among the main guidelines. Better data collection is warranted.

Severe infections in patients with chronic lymphocytic leukemia included in trials investigating BTK and BCL2 inhibitors.

Bruton tyrosine kinase inhibitors (BTKi) and the BCL-2 inhibitor venetoclax have significantly improved the prognosis of patients with chronic lymphocytic leukemia (CLL). However, the incidence of severe infections in patients receiving these agents needs to be better understood. Our review aimed to provide an overview of grade ≥3 infections in patients with CLL who received BTKi and venetoclax-based therapy in prospective trials. Infection rates were influenced by the age of patients and the duration of follow-up. For treatment-naive (TN) patients receiving BTKi, infection rates ranged between 11.4 % and 27.4 % and were close to 30 % in relapsed/refractory (R/R) patients. TN and R/R patients receiving fixed-duration venetoclax-based treatments showed variable rates, with maximum values around 20 %. Opportunistic and fatal infections were uncommon. In conclusion, infections remain a concern in patients with CLL receiving targeted agents. A better definition of factors increasing infection vulnerability could help identify those patients who require infection prophylaxis.

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.

CAR-modified Cellular Therapies in Chronic Lymphocytic Leukemia: Is the Uphill Road Getting Less Steep?

The clinical development of chimeric antigen receptor (CAR) T-cell therapy has been more challenging for chronic lymphocytic leukemia (CLL) compared to other settings. One of the main reasons is the CLL-associated state of immune dysfunction that specifically involves patient-derived T cells. Here, we provide an overview of the clinical results obtained with CAR T-cell therapy in CLL, describing the identified immunologic reasons for the inferior efficacy. Novel CAR T-cell formulations, such as lisocabtagene maraleucel, administered alone or in combination with the Bruton tyrosine kinase inhibitor ibrutinib, are currently under investigation. These approaches are based on the rationale that improving the quality of the T-cell source and of the CAR T-cell product may deliver a more functional therapeutic weapon. Further strategies to boost the efficacy of CAR T cells should rely not only on the production of CAR T cells with an improved cellular composition but also on additional changes. Such alterations could include (1) the coadministration of immunomodulatory agents capable of counteracting CLL-related immunological alterations, (2) the design of improved CAR constructs (such as third- and fourth-generation CARs), (3) the incorporation into the manufacturing process of immunomodulatory compounds overcoming the T-cell defects, and (4) the use of allogeneic CAR T cells or alternative CAR-modified cellular vectors. These strategies may allow to develop more effective CAR-modified cellular therapies capable of counteracting the more aggressive and still incurable forms of CLL.