The Role of Methylation in Chronic Lymphocytic Leukemia and Its Prognostic and Therapeutic Impacts in the Disease: A Systematic Review.

Epigenetic regulation has been thoroughly investigated in recent years and has emerged as an important aspect of chronic lymphocytic leukemia (CLL) biology. Characteristic aberrant features such as methylation patterns and global DNA hypomethylation were the early findings of the research during the last decades. The investigation in this field led to the identification of a large number of genes where methylation features correlated with important clinical and laboratory parameters. Gene-specific analyses investigated methylation in the gene body enhancer regions as well as promoter regions. The findings included genes and proteins involved in key pathways that play central roles in the pathophysiology of the disease. Τhe application of these findings beyond the theoretical understanding can not only lead to the creation of prognostic and predictive models and scores but also to the design of novel therapeutic agents. The following is a review focusing on the present knowledge about single gene/gene promoter methylation or mRNA expression in CLL cases as well as records of older data that have been published in past papers.

A Patient with Synchronous Gallbladder and Bone Plasmacytoma.

Multiple myeloma (MM) is the most common primary bone-originating tumor, whereas extramedullary plasmacytoma (EMP) is a plasma cell tumor that arises outside the bone and is most commonly found in the head and neck area. Gastrointestinal and particularly gallbladder involvement is exceedingly rare, and symptoms, if any are present, are usually similar to those seen with cholelithiasis. Treatment options usually include surgical resection and/or chemotherapy. In this report, we present a rare case of a clinically unexpected plasmablastic extramedullary plasmacytoma that was found on abdominal ultrasound (US) and magnetic resonance imaging (MRI) in a 61-year-old asymptomatic patient and led him to undergo cholecystectomy. A fluorodeoxyglucose positron emission computed tomography (FDG PET-CT) that was performed due to the onset of left thigh pain also demonstrated concurrent bone plasmacytoma. The patient is currently receiving chemotherapy and is also being prepared for autologous stem cell transplantation. In this context, we further present the diagnostic, therapeutic and prognostic challenges of EMPs. Lastly, we point out the distinct features of the plasmablastic subtype and analyze its differences compared to other histologic subtypes in achieving a successful diagnosis and management.

The Clinical and Prognostic Significance of Ribonucleotide Reductase Subunits RRM1 and RRM2 mRNA Levels in Patients with Chronic Lymphocytic Leukemia.

Ribonucleotide Reductase (RNR) converts ribonucleotides to deoxyribonucleotides required for DNA replication and repair. RNR consists of subunits M1 and M2. It has been studied as a prognostic factor in several solid tumors and in chronic hematological malignancies, but not in chronic lymphocytic leukemia (CLL). Peripheral blood samples were collected from 135 CLL patients. M1/M2 gene mRNA levels were measured and expressed as a RRM1-2/GAPDH ratio. M1 gene promoter methylation was studied in a patients' subgroup. M1 mRNA expression was higher in patients without anemia (p = 0.026), without lymphadenopathy (p = 0.005) and 17p gene deletion (p = 0.031). Abnormal LDH (p = 0.022) and higher Rai stage (p = 0.019) were associated with lower M1 mRNA levels. Higher M2 mRNA levels were found in patients without lymphadenopathy (p = .048), Rai stage 0 (p = 0.025) and Trisomy 12 (p = 0.025). The correlation between RNR subunits and clinic-biological characteristics in CLL patients demonstrate RNR's potential role as a prognostic factor.

Immunogenicity of a third dose of the BNT162b2 COVID-19 vaccine in patients with CLL: effects on treatment selection.

Patients with chronic lymphocytic leukemia (CLL) show suboptimal responses to the vaccines against SARS-CoV-2; it has been shown though that a booster dose of the BNT162b2 vaccine may lead to a significant increase in the seroconversion rates of immunocompromised patients. We conducted a prospective, non-interventional study to evaluate the immunogenicity of a third dose of the BNT162b2 vaccine in adult patients with CLL. Sera were tested before the first, after the second, and before and after the third dose for anti-SARS-CoV-2 receptor binding domain (RBD) spike protein IgG (anti-RBD). Thirty-nine patients with CLL were included in the study. The seroconversion rate increased from 28.2% before the third dose to 64.1% after the third dose and was higher in treatment-naïve patients (72.7% versus 47.1% in actively treated patients, p = 0.042). All but one patient achieving a seroconversion after the second dose retained after the third, while eight patients not achieving a seroconversion after the second dose (38.1%), did so after the third. Moreover, patients actively treated with venetoclax had a higher seroconversion rate than those treated with ibrutinib (87.5% versus 14.3%, p = 0.001). This study confirms the beneficial effect of a third dose of the BNT162b2 vaccine on the seroconversion rate in patients with CLL. Our results also strongly suggest that the use of venetoclax is correlated with higher immunogenicity/seroconversion rates than that of ibrutinib, a finding that has been reported by another study. A treatment strategy change during the pandemic favoring the use of venetoclax may be suggested based on our results, although these results should be validated in larger studies.

Safety and immunogenicity of the BNT162b2 mRNA Covid-19 vaccine in patients with chronic lymphocytic leukemia: a prospective study.

Introduction: Immunization of patients with chronic lymphocytic leukemia (CLL) with vaccines against several infectious diseases has proven insufficient. Data on seroconversion of patients with CLL after vaccination against severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) are still young, but accumulating evidence shows low seroconversion rates. Methods: We conducted a prospective, noninterventional study evaluating the safety and immunogenicity of two doses of the BNT162b2 mRNA Covid-19 vaccine, administered 21 days apart in consecutive adult patients with CLL. Patients vaccinated with other vaccines against SARS-CoV-2, with a history of confirmed Coronavirus Disease 19 (COVID-19), with known human immunodeficiency virus infection, or with an inability to provide written informed consent were excluded. Sera were tested before the first and after the second dose of the vaccine for anti-SARS-CoV-2 receptor binding domain (RBD) spike protein IgG (anti-RBD), using the Abbott SARS-CoV-2 IgG II Quant assay (Abbott Laboratories, Abbott Park, IL, USA), with a cutoff value for seroconversion at 50 AU/ml. Results: Sixty-one patients (28 males/33 females) with CLL, with a median age of 61 years, were included in the study. The majority of the patients (82.0%) were lower (0-2) stage per the RAI staging system. The seroconversion rate at 14 days after the second dose was 45% and was correlated with RAI stage (0-2 versus 3-4; 51.0% versus 18.3%, p = 0.047), the treatment status (treatment naïve, previously treated, or actively treated patients; 63.0% versus 40.0% versus 26.1%, respectively, p = 0.031), the number of previous treatment lines (0-2 versus >2; 55.3% versus 8.3%, p = 0.004), and the platelet count of the patients (over or under 100 × 109/L; 52.9% versus 10.0%, p = 0.015). Moreover, there was a positive linear relationship between the antibody titers and the gamma-globulin levels (r = 0.182, p = 0.046) and platelet count (r = 0.277, p = 0.002). Finally, patients actively treated with venetoclax had higher antibody titers than those treated with ibrutinib (15.8 AU/ml versus 0.0 AU/ml, p = 0.047). No safety issues were identified while the emergence of adverse events was not correlated with immunogenicity. Discussion: This study confirms results from previous studies on the low seroconversion rates in patients with CLL vaccinated with the BNT162b2 mRNA Covid-19 vaccine and on the detrimental effect of advanced disease and multiple treatment lines on seroconversion, while it is suggested that treatment with venetoclax may offer a chance for higher antibody titers, suggesting a treatment strategy change during the pandemic provided that this result is confirmed by larger studies specifically designed to address this issue.

A case report of a fulminant Aeromonas hydrophila soft tissue infection in a patient with acute lymphoblastic leukemia harboring a rare translocation.

INTRODUCTION: Aeromonads are gram-negative opportunistic bacteria, mainly found in aquatic environments. Hematologic patients are particularly at risk of Aeromonas soft tissue infections and septicemia, especially during chemotherapy-induced neutropenia. CASE DESCRIPTION: A 46-year-old man was diagnosed with acute lymphoblastic leukemia characterized by the rare t(12;17)(p13;q21)/TAF15-ZNF384 aberration. On day 22 of chemotherapy, he developed febrile neutropenia followed by necrotizing fasciitis in his upper right extremity. Despite appropriate antibiotic therapy and prompt surgical intervention, he died within 36 h after the appearance of a fever. A multi-sensitive Aeromonas hydrophila was isolated from all cultural sites. DISCUSSION AND CONCLUSIONS: In a previous paper we characterized the patient's aberration with cytogenetic and FISH analysis. Here, we provide details regarding the patient's rapidly progressing infection and underline the importance of maintaining high clinical suspicion of Aeromonas infections in acute leukemia. Given the unusually rapid progression of an infection caused by a rare non-resistant pathogen, and after considering data on the implication of metalloproteinase function in immune system regulation, a correlation between risk of severe infection and TAF15-ZNF384 aberrated acute lymphoblastic leukemia cannot be ruled out.

Serum ferritin levels in previously untreated classical Hodgkin lymphoma: correlations and prognostic significance.

Serum ferritin (SF) is frequently elevated in classical Hodgkin lymphoma (cHL). We report on its prognostic significance in an unselected series of 529 cHL patients treated with state-of-the-art therapy. Higher baseline levels correlated with markers of advanced/aggressive disease. SF levels were significantly higher in male and older patients, those with high body mass index and mixed cellularity histology. The strongest correlation was recorded between SF and complement reactive protein (CRP) levels. Gender-specific SF cutoffs which provided the best discrimination in terms of freedom from progression (FFP) were identified. In multivariate analysis elevated SF levels, advanced stage and high lactate dehydrogenase (LDH) were independent prognostic factors of inferior FFP. SF also appears to retain independent prognostic significance for progression-free survival (PFS) but not for overall survival (OS). In conclusion, SF levels in cHL reflect disease activity and are associated with adverse patient outcomes.

Progressive multifocal leukoencephalopathy in the context of newer therapies in hematology and review of new treatment strategies.

Progressive multifocal leukoencephalopathy (PML) is a rare, often fatal demyelinating disease of the central nervous system (CNS) caused by the reactivation of JC polyomavirus in the CNS. We present a case of a 54-year-old man with follicular lymphoma diagnosed with PML after being treated with anti-CD20 monoclonal antibody-based regimens for several years. Due to the lack of effective treatment choices for PML, the patient was treated with nivolumab, based on recent reports, but succumbed to his disease a few months after diagnosis. In this paper, we focus on reviewing the literature of PML cases correlated with newer agents used in hematology, possible factors affecting disease prognosis, as well as the available data on upcoming therapeutic options for patients with PML. Though newer promising treatments such as anti-PD1 monoclonal antibodies arise, a definitive treatment option is yet to be found. Vigilance, early detection, and prompt intervention play a crucial role in the prognosis of PML in patients with hematological malignancies.

Bone marrow ribonucleotide reductase mRNA levels and methylation status as prognostic factors in patients with myelodysplastic syndrome treated with 5-Azacytidine.

Ribonucleotide Reductase (RNR) is a two-subunit (RRM1, RRM2) enzyme, responsible for the conversion of ribonucleotides to deoxyribonucleotides required for DNA replication. To evaluate RNR as a biomarker of response to 5-azacytidine, we measured RNR mRNA levels by a quantitative real-time PCR in bone marrow samples of 98 patients with myelodysplastic syndrome (MDS) treated with 5-azacytidine with parallel quantification of the gene promoter's methylation. Patients with low RRM1 levels had a high RRM1 methylation status (p = 0.005) and a better response to treatment with 5-azacytidine (p = 0.019). A next-generation sequencing for genes of interest in MDS was also carried out in a subset of 61 samples. Splicing factor mutations were correlated with lower RRM1 mRNA levels (p = 0.044). Our results suggest that the expression of RNR is correlated with clinical outcomes, thus its expression could be used as a prognostic factor for response to 5-azacytidine and a possible therapeutic target in MDS.

PARP1 as a therapeutic target in acute myeloid leukemia and myelodysplastic syndrome.

Poly(ADP-ribose) polymerase 1 (PARP1) is a key mediator of various forms of DNA damage repair and plays an important role in the progression of several cancer types. The enzyme is activated by binding to DNA single-strand and double-strand breaks. Its contribution to chromatin remodeling makes PARP1 crucial for gene expression regulation. Inhibition of its activity with small molecules leads to the synthetic lethal effect by impeding DNA repair in the treatment of cancer cells. At first, PARP1 inhibitors (PARPis) were developed to target breast cancer mutated cancer cells. Currently, PARPis are being studied to be used in a broader variety of patients either as single agents or in combination with chemotherapy, antiangiogenic agents, ionizing radiation, and immune checkpoint inhibitors. Ongoing clinical trials on olaparib, rucaparib, niraparib, veliparib, and the recent talazoparib show the advantage of these agents in overcoming PARPi resistance and underline their efficacy in targeted treatment of several hematologic malignancies. In this review, focusing on the crucial role of PARP1 in physiological and pathological effects in myelodysplastic syndrome and acute myeloid leukemia, we give an outline of the enzyme's mechanisms of action and its role in the pathophysiology and prognosis of myelodysplastic syndrome/acute myeloid leukemia and we analyze the available data on the use of PARPis, highlighting their promising advances in clinical application.

JAK2V617F positive polycythemia vera with paroxysmal nocturnal hemoglobinuria and visceral thromboses: a case report and review of the literature.

BACKGROUND: Polycythemia vera (PV) is characterized by red cell mass expansion in the peripheral blood and can be complicated with thrombosis, bleeding, evolution to acute myeloid leukemia (AML) or a fibrotic phase. Paroxysmal nocturnal hemoglobinuria (PNH) in an acquired clonal haematopoietic stem cell disorder associated with chronic intravascular hemolysis, venous thrombosis, defective hematopoiesis, frequent episodes of infection and, rarely, leukemic transformation. Herein, we report an interesting case of a patient with coexistence of PNH clones and a JAK2V617F positive PV, with unusual thromboses without hemolysis. CASE PRESENTATION: A 51-year-old woman presented with increased levels of hematocrit, multiple liver, spleen, and left kidney infarctions and ascites; further investigation revealed a JAK2V617F-positive polycythemia vera and the presence of a significant PNH population (more than 90% CD55- CD59- cells among both granulocytes and red blood cells). Interestingly, the patient has experienced severe thrombotic events without any signs or symptoms of hemolysis. CONCLUSIONS: This case raises questions over uncharted aspects of the PNH etiopathogenesis and its potential association with myeloproliferative neoplasms (MPN) and highlights the difficulty of diagnosing and managing patients with more than one potentially thrombophilic conditions, especially with established and severe thromboses.

A case report of Hodgkin lymphoma in a patient treated with ustekinumab for psoriasis.

RATIONALE: Ustekinumab is a biological agent that inhibits interleukin 12 and 23 and has been approved for the treatment of moderate and severe plaque psoriasis. There have been case reports that raise concerns about its oncogenic potential. We are the first authors to report a case of Hodgkin lymphoma in a psoriatic patient receiving ustekinumab. PATIENT CONCERNS: A 22-year-old asymptomatic female patient presented to our department to investigate an enlarged cervical lymph node. Her past history was unremarkable, except for psoriasis since age 13. Two months before presentation the decision to administer Ustekinumab was taken and the patient had already received 3 doses. DIAGNOSES: During workup a Stage IV Hodgkin lymphoma was discovered. INTERVENTIONS: Ustekinumab administration was discontinued. The patient received treatment with the ABVD regimen. OUTCOMES: The patient's disease was refractory to the above-mentioned treatment. Therefore, a more aggressive regimen (BEACOPP escalated) was administered. LESSONS: Growing postmarketing surveillance data and case reports indicate that further research is warranted in order to elucidate a potential association between Ustekinumab and malignancy.

Post-Myelofibrosis Acute Myeloid Leukemia Effectively Treated with a Combination of Ruxolitinib and 5-Azacytidine.

BACKGROUND: Leukemic transformation is an unfavorable event emerging in a minority of patients with myelofibrosis (MF) and carrying a poor prognosis. Patients with post-MF acute myeloid leukemia (AML) may be treated with curative or noncurative intent, but there is no standard of care. The use of hypomethylating agents has been correlated with limited activity and low overall survival rates, while ruxolitinib has been proven to have modest antileukemic activity. CASE PRESENTATION: In this case study, we present the case of a 67-year-old female patient with post-polycythemia vera MF who developed AML and was successfully treated with a combination of 5-azacytidine and ruxolitinib, without any significant toxicity. SUMMARY: To our knowledge, this is the first report of a patient with post-MF AML achieving a complete remission with a combination of ruxolitinib and a hypomethylating agent. This combination is actively studied in phase I/II clinical trials and may be proven effective in this hard-to-treat group of patients.

An Adult Patient with Early Pre-B Acute Lymphoblastic Leukemia with t(12;17)(p13;q21)/ZNF384-TAF15.

This is a case report of a 46-year-old man diagnosed with early pre-B acute lymphoblastic leukemia (ALL), bearing the translocation t(12;17)(p13;q21) as the sole chromosomal abnormality. This is a rare chromosomal abnormality that has been reported in approximately 25 cases worldwide. FISH analysis revealed a rearrangement of ZNF384 (12p13) and TAF15 (17q12) genes, which is usually associated with a pre-B ALL phenotype with co-expression of the myeloid markers CD13 and/or CD33. ZNF384 encodes a zinc finger protein, which acts as a transcription factor, regulating the expression of several matrix metalloproteinases and TAF15 belongs to the FET (FUS, EWS, and TAF15) family, consisting of RNA and DNA-binding proteins. Unlike most of the cases where CD10 expression was absent or weak, in our case CD10 was highly expressed. The prognostic significance of ZNF384/TAF15 fusion is not very clear since several reports support a generally good prognosis, while others support a poor clinical outcome. Our patient was treated with the German multicenter ALL (GMALL) protocol for B-ALL, but experienced a fulminant gram-negative sepsis and eventually died during induction therapy.

A case report of a very late response to 5-azacytidine in a patient with lower risk myelodysplastic syndrome: Time to update treatment guidelines for lower risk patients.

RATIONALE: The hypomethylating agent 5-azacytidine has been approved in Europe for patients with intermediate 2 and high (i.e., higher) risk myelodysplastic syndrome according to the International Prognostic Scoring System (IPSS). A total of 91% of all first responses in higher risk patients occur within 6 cycles of treatment; however, data regarding the time to first response in clinical trials with lower risk patients are not available. PATIENT CONCERNS: Our case describes the late response of a lower risk (intermediate 1 according to the IPSS and intermediate according to the IPSS-R) patient to 5-azacytidine treatment.Diagnosis and interventions: Once diagnosed, the patient started supportive treatment due to persistent pancytopenia and recurrent infections. The use of a hypomethylating agent was decided because the patient was transfusion dependent, and suffering from recurrent severe febrile infections due to neutropenia. Other possible causes of fever except infections in the context of his neutropenia were excluded. OUTCOMES: After the 12th cycle of 5-azacytidine the patient showed a hematologic response, with transfusion independency and with no recurrent febrile episodes. LESSONS: This case report probably indicates that a subset of patients who belong to the lower risk category according to the previous prognostic systems and to the intermediate one according to the IPSS-R, may benefit from prolonged treatment with the drug. The indication of 5-azacytidine in Europe for patients with higher risk myelodysplastic syndrome (MDS) (according to the IPSS) could possibly include a wider range of patients if updated according to the IPSS-R.

Isolated skeletal muscle recurrence of an originally nodal diffuse large B cell lymphoma: A case report and review of the literature.

RATIONALE: Diffuse large B cell lymphoma (DLBCL) is a malignancy of the B cells with extranodal primary involvement being estimated at 30% to 40% of cases. Primary skeletal muscle presentation of DLBCL is extremely rare, with an estimated incidence of about 0.5% of extranodal lymphomas, presenting mostly in the lower extremities. The possible mechanisms of muscle involvement of DLBCL include primary extranodal disease, extension from adjacent organs (such as lymph nodes) or disseminated disease. PATIENT CONCERNS: We report a case of a 70-year-old woman with an advanced initially nodal DLBCL, treated with R-CHOP, that presented with an enlargement of her left thigh and restricted mobility 3 months after completion of chemotherapy. Imaging studies were performed, which showed possible infiltration of the muscles of the left thigh, without any nodal disease present. DIAGNOSES: Muscle biopsy documented the recurrence of the lymphoma at the left thigh. INTERVENTIONS: The patient started second-line treatment with gemcitabine and vinorelbine. OUTCOMES: A partial response was achieved after the first cycle. LESSONS: The remarkable element lies in the reappearance of the lymphoma at the left thigh muscles, with no radiographic or clinical evidence of involvement of lymph nodes, despite the extensive lymph node disease at initial presentation. The further management of such recurrences remains to be clarified, as the odd biological behavior of the malignant cells dictates a special handling of the disease.

Tumor Protein 53 Gene Mutations Without 17p13 Deletion Have No Significant Clinical Implications in Chronic Lymphocytic Leukemia. Detection of a New Mutation.

BACKGROUND: The tumor protein p53 (TP53) gene may be inactivated through 17p13 deletion, somatic mutations, or both. In chronic lymphocytic leukemia (CLL) although 17p13 deletion is correlated with poor prognosis, the role of sole TP53 mutations remains controversial. MATERIALS AND METHODS: We carried out a mutation analysis of TP53 gene in 72 patients with CLL. RESULTS: Seventy-one (98.6%) patients carried the polymorphic site c.215C>G, p.Pro72Arg, but its presence was not correlated with overall survival (OS). Moreover, 19 (26.4%) patients carried a mutation of TP53. Among the eight detected mutations, to our knowledge, one (c.587G>A) has never been reported in the past. There was a correlation of the mutation burden with the stage of the disease (p=0.022), but not with OS. None of the detected mutations was individually correlated with OS. CONCLUSION: The clinical significance of TP53 mutations is still a matter of debate and larger studies and meta-analyses are required to reach an unequivocal conclusion.

Hypereosinophilic Syndrome as a Rare Cause of Reversible Biventricular Heart Failure.

Hypereosinophilic syndrome is a rare entity that can develop secondary to overproduction of eosinophilopoietic cytokines or as idiopathic disease. Cardiac involvement, which occurs often, is divided into 3 stages, the latter of which is nonreversible and leads to severe heart failure. Early detection and treatment of the syndrome is essential. For this reason, genetic testing for the FIP1L1-PDGFRA fusion gene has recently been added to the diagnostic algorithm. Patients with this mutation are at increased risk for the development of cardiac involvement and typically respond to treatment with the tyrosine kinase inhibitor imatinib mesylate.