Maurizio Cavallari

Chromosome aberrations detected by conventional karyotyping using novel mitogens in chronic lymphocytic leukemia: Clinical and biologic correlations

To clarify whether karyotype aberrations (KA) involving regions not covered by the standard fluorescence in situ hybridization (FISH) panel have independent prognostic relevance, we evaluated KA by conventional cytogenetics in a learning cohort (LC; n = 166) and a validation cohort (VC; n = 250) of untreated chronic lymphocytic leukemia (CLL) patients. In the VC, novel mitogens were used to improve metaphase generation and TP53, NOTCH1, and SF3B1 mutations were assessed. KA undetected by FISH were found in 35 and 35% of the cases in the LC and VC, respectively. In addition to FISH, KA allowed reclassification of 23 and 26% of cases in the LC and VC, respectively, into a higher cytogenetic risk group. By multivariate analysis, both in the LC and VC, KA other than isolated 13q deletion correlated with a shorter time to first treatment (TFT; P < 0.001 and 0.003, respectively), while a complex karyotype predicted a worse overall survival (OS, P = 0.015 and 0.010, respectively). In the VC, where a comprehensive biologic assessment was performed, a shorter TFT was also predicted by stage (P < 0.001), IGHV mutational status (P = 0.05), and del(17p)/TP53 mutations (P = 0.033) while stage (P = 0.023) and del(17p)/TP53 mutations (P = 0.024) independently predicted a shorter OS. FISH results did not independently impact on TFT and OS, in the LC and VC cohorts; this was also the case for NOTCH1 and SF3B1 mutations in the VC. We suggest that in CLL, conventional karyotyping with novel mitogens could be more effective than FISH for the detection of KA allowing for a more precise refinement of prognosis.

In CLL, comorbidities and the complex karyotype are associated with an inferior outcome independently of CLL-IPI

To the editor: Chronic lymphocytic leukemia (CLL) represents the most common form of leukemia in Western countries.[1][1] The clinical course of the disease is quite heterogeneous with some patients living for years with asymptomatic disease and others experiencing early progression and requiring

Response to ibrutinib of refractory life-threatening autoimmune hemolytic anemia occurring in a relapsed chronic lymphocytic leukemia patient with 17p deletion

Autoimmune hemolytic anemia (AIHA) is one of the most common disease-specific complications of chronic lymphocytic leukemia (CLL), occurring in 3–10% of patients, with a higher incidence in advanced stages or in cases with an unmutated configuration of the variable portion of the immunoglobulin heavy chain gene (IGHV).[1] AIHA is mediated most frequently by IgG auto-antibodies produced by nonmalignant B cells, or by IgM antibodies in <10% of cases.[2] Activated CLL cells may bind and present the erythrocyte protein band 3 and activate CD4þ cells in a HLA-DR-dependent way, playing a role as possible initiators of the hemolytic process.[3] Furthermore, CLL cells have been shown capable of causing regulatory T-cell dysfunction which may contribute to the development of this complication.[4] Even though AIHA had no independent effect on survival in two studies,[5,6] this complication has been shown to be more frequent in CLL subsets with unfavorable prognostic features, i.e. 11q-, 17p-, CD38þ [6] and patients with severe anemia refractory to steroids and alkylating agents have been described.[5] In steroid-refractory patients, CLL-directed treatment including rituximab, with or without cytotoxic agents, or alemtuzumab has been shown to induce response in some cases.[1] Chemorefractory disease and/or inability to tolerate chemoimmunotherapy represent a major problem in patients not responding to steroids who may succumb to infections, AIHA-related complications [5] or disease progression. Hence, effective and tolerable treatment represents an unmet need in refractory CLL-associated AIHA. In March 2011, a 62-year-old male was referred to our institution with a diagnosis of Rai stage II asymptomatic CLL and abnormal bilirubin excretion (Gilbert syndrome) with indirect bilirubin levels ranging between 2 and 3 mg/dl. Hematologic work-up was performed as previously reported,[7] documenting a typical morphology and immunophenotype and negativity for CD38 and ZAP-70. The karyotype was normal and no aberration was detected by fluorescence in situ hybridization (FISH). The Ig heavy chain variable (IGHV) region was mutated; no mutations of exons 4–9 of the TP53 gene were detected. In October 2012, the patient presented disease progression with fatigue, peripheral edema, hyperleukocytosis (lymphocytes 195,000/ml), adenopathies and symptomatic splenomegaly (8 cm below the left costal margin). Clonal evolution with 17p deletion in 22% of the cells was documented. Fludarabine, cyclophosphamide and rituximab (FCR regimen) treatment was started and soon after the second cycle, the patient presented with dyspnea and jaundice. Hemoglobin (Hb) levels were low (Hb 6.5 g/ dl) with high reticulocyte count. Elevated serum lactate dehydrogenase (LDH) levels (694 U/l normal values 240–480 U/l) and indirect bilirubin levels (5.08 mg/dl; normal values 0.10–1.00 mg/dl), low haptoglobulin levels (5 mg/dl; normal values 30–200 mg/dl) and strong positivity for IgG (5þ) on direct antiglobulin test (DAT) were consistent with a diagnosis of AIHA. High-dose methylprednisolone and rituximab were administered along with intravenous (iv) Ig (1 g/kg for two consecutive days). Due to persistent hemolysis after 4 weeks, the patient was treated with bendamustine and rituximab for a total of six cycles. Normalization of Hb levels and hemolysis-associated laboratory parameters was observed over a 3-month period and a partial response (PR) according to NCI criteria [8] was documented at the end of treatment, with a normalization of blood

An extensive molecular cytogenetic characterization in high-risk chronic lymphocytic leukemia identifies karyotype aberrations and TP53 disruption as predictors of outcome and chemorefractoriness

We investigated whether karyotype analysis and mutational screening by next generation sequencing could predict outcome in 101 newly diagnosed chronic lymphocytic leukemia patients with high-risk features, as defined by the presence of unmutated IGHV gene and/or 11q22/17p13 deletion by FISH and/or TP53 mutations. Cytogenetic analysis showed favorable findings (normal karyotype and isolated 13q14 deletion) in 30 patients, unfavorable (complex karyotype and/or 17p13/11q22 deletion) in 34 cases and intermediate (all other abnormalities) in 36 cases. A complex karyotype was present in 21 patients. Mutations were detected in 56 cases and were associated with unmutated IGHV status (p = 0.040) and complex karyotype (p = 0.047). TP53 disruption (i.e. TP53 mutations and/or 17p13 deletion by FISH) correlated with the presence of ≥ 2 mutations (p = 0.001) and a complex karyotype (p = 0.012). By multivariate analysis, an advanced Binet stage (p < 0.001) and an unfavorable karyotype (p = 0.001) predicted a shorter time to first treatment. TP53 disruption (p = 0.019) and the unfavorable karyotype (p = 0.028) predicted a worse overall survival. A shorter time to chemorefractoriness was associated with TP53 disruption (p = 0.001) and unfavorable karyotype (p = 0.025). Patients with both unfavorable karyotype and TP53 disruption presented a dismal outcome (median overall survival and time to chemorefractoriness of 28.7 and 15.0 months, respectively). In conclusion, karyotype analysis refines risk stratification in high-risk CLL patients and could identify a subset of patients with highly unfavorable outcome requiring alternative treatments.

Biological significance and prognostic/predictive impact of complex karyotype in chronic lymphocytic leukemia

The complex karyotype (CK) is an established negative prognostic marker in a number of haematological malignancies. After the introduction of effective mitogens, a growing body of evidence has suggested that the presence of 3 or more aberrations by conventional banding analysis (CBA) is associated with an unfavorable outcome in chronic lymphocytic leukemia (CLL). Thus, the importance of CBA was recognized by the 2018 guidelines of the International Workshop on CLL, which proposed the introduction of CBA in clinical trials to validate the value of karyotype aberrations. Indeed, a number of observational studies showed that cytogenetic aberrations and, particularly, the CK may have a negative independent impact on objective outcome measures (i.e. time to first treatment, progression free survival, time to chemorefractoriness and overall survival) both in patients treated with chemoimmunotherapy and, possibly, in patients receiving novel mechanism-based treatment. Here, we set out to present the scientific evidence supporting the significance of CK as a prognostic marker in CLL and to discuss the biological basis showing that the CK is a consequence of genomic instability.

In chronic lymphocytic leukaemia with complex karyotype, major structural abnormalities identify a subset of patients with inferior outcome and distinct biological characteristics

Complex karyotype (CK) is a negative prognostic factor in chronic lymphocytic leukaemia (CLL). However, CK is a heterogeneous cytogenetic category. Unbalanced rearrangements were present in 73·3% of 90 CLL patients with CK (i.e. ≥3 chromosome aberrations in the same clone), and were associated with a shorter overall survival (P = 0·025) and a shorter time to first treatment (P = 0·043) by multivariate analysis. Patients with unbalanced rearrangements presented a distinct mRNA expression profile. In conclusion, CLL patients with unbalanced rearrangements might represent a subset of very high‐risk CLL patients with distinct clinical and biological characteristics.

“Hemophagocytic Lymphohistiocytosis after EBV reactivation and ibrutinib treatment in relapsed/refractory Chronic Lymphocytic Leukemia”

Hemophagocytic Lymphohistiocytosis (HLH) is a rare syndrome characterized by ineffective T-cell and NK response. We report the clinical course of a patient with relapsed CLL who developed acute symptoms soon after starting ibrutinib. Hyperpyrexia, splenomegaly, hyperferritinemia, hypertriglyceridemia, cytopenias, and a typical cytokine pattern, i.e. high interleukin (IL)−6, IL10 and IL18, were consistent with a diagnosis of HLH. Coexistent Epstein Barr virus reactivation was documented. Ibrutinib-induced impairment of NK degranulation, associated with EBV reactivation and CLL-related immunodeficiency may have contributed to the development of HLH in our patient.