Outcomes after intensive chemotherapy for secondary and myeloid-related changes acute myeloid leukemia patients aged 60 to 75 years old: a retrospective analysis from the PETHEMA registry.

Treatment options for patients with secondary acute myeloid leukemia (sAML) and AML with myeloid-related changes (AMLMRC) aged 60 to 75 years are scarce and unsuitable. A pivotal trial showed that CPX-351 improved complete remission with/without incomplete recovery (CR/CRi) and overall survival (OS) as compared with standard "3+7" regimens. We retrospectively analyze outcomes of 765 patients with sAML and AML-MRC aged 60 to 75 years treated with intensive chemotherapy, reported to the PETHEMA registry before CPX-351 became available. The CR/CRi rate was 48%, median OS was 7.6 months (95% confidence interval [CI]: 6.7-8.5) and event-free survival (EFS) 2.7 months (95% CI: 2-3.3), without differences between intensive chemotherapy regimens and AML type. Multivariate analyses identified age ≥70 years, Eastern Cooperative Oncology Group performance status ≥1 as independent adverse prognostic factors for CR/CRi and OS, while favorable/intermediate cytogenetic risk and NPM1 were favorable prognostic factors. Patients receiving allogeneic stem cell transplant (HSCT), autologous HSCT, and those who completed more consolidation cycles showed improved OS. This large study suggests that classical intensive chemotherapy could lead to similar CR/CRi rates with slightly shorter median OS than CPX-351.

Comprehensive evaluation of genetic and acquired thrombophilia markers for an individualized prediction of clinical thrombosis in patients with lymphoma and multiple myeloma.

Patients diagnosed with lymphoma or multiple myeloma are at elevated risk of venous thromboembolism (VTE). Optimum risk stratification and effective thromboprophylaxis can only be achieved through the development of a multiple-specific risk score that successfully captures all aspects of the heterogeneous prothrombotic environment existing in these patients. Our aim was to identify risk factors for thrombosis and suggest an improved tool combining clinical data, thrombo-inflammatory biomarkers and genetic (Thrombo inCode® test) variables for predicting thrombotic risk in patients with lymphoma and multiple myeloma. A prospective longitudinal study was conducted on newly-diagnosed lymphoma and multiple myeloma patients who presented at our institution between February 2020 and January 2021. The study included 47 patients with lymphoma and 16 patients with multiple myeloma. We performed a follow-up of 1 year or until September 2021. The incidence of venous thrombosis and associated risk factors were analysed, including the genetic Thrombo inCode® test. Khorana and ThroLy scores for lymphoma patients and IMPEDE VTE score for myeloma patients were calculated. At a median follow-up of 9.1 months, VTE incidence was 9.5% (6/63), with 4 and 2 patients with lymphoma and myeloma who developed the events, respectively. Univariate analysis showed that the incidence of thrombosis was significantly higher in patients with ECOG ≥ 2 and prior immobility. Median factor VIII levels were significantly higher in patients with thrombosis (with increased values in all of them). Moreover, there was a trend in genetic variant rs5985 (factor XIII) as a protective factor, and a trend to higher thrombotic risk in patients with factor V Leiden, rs2232698 variant (serpinA10), low total protein S activity, elevated D-dimer, aggressive lymphoma and treatment with dexamethasone. The results of our study demonstrate promise for the potential use of widely accessible markers to increase precision in risk prediction for VTE in patients with lymphoma and multiple myeloma, particularly ECOG ≥ 2, immobility and higher factor VIII levels, as well as lymphoma aggressiveness, treatment with dexamethasone and the haemostatic biomarkers D-dimer and total protein S activity. Additionally, genetic variants factor V Leiden, serpinA10 rs2232698 and factor XIII-A Val34Leu warrant further investigation for use in the research setting.

Impact of early palliative care intervention in autologous bone marrow transplantation: feasibility of a multicentric study.

INTRODUCTION: This prospective multicentre study evaluates the impact of Palliative Care Unit (PCU) intervention (Experimental Group, EG), during autologous hematopoietic stem cell transplantation (AHSCT) on quality of life (QoL), symptom control and healthcare resource use compared to standard practice (Control Group, CG). We used validated scales on Days 0 (stem cell infusion), + 7 (bone marrow aplasia, acute symptoms) and + 21 (aplasia recovery). RESULTS: In 40 patients (20 EG/ 20 CG: 45%/25% female, median age 57.5/59), QoL differed significantly at Day + 7 (EG: median 0.50; CG: -63.00; p < 0.001) and Day + 21 (EG: -2.00; CG: -129.00; p < 0.001). On Day 0, mean FACT-BMT scores were CG/EG: 131/ 89.35, reflecting the pre-transplant intervention of the PCU in EG patients. For pain (EG median 0.00, CG median 2.50; p = 0.01), 45% EG patients used opioids on day 0 (mean 38.5 mg morphine/day/patient). Reduced pain control impacted nutritional support (parenteral nutrition 45% CG, 5% EG; p = 0.08). Hospitalisation duration was longer in CG (median 18.5; EG median 13.00; p < 0.001). Despite the short follow-up and small sample size, PCU and HD collaboration improves QoL and symptom management during acute AHSCT, evident through pain control, analgesia management, reduced parenteral nutrition need and shorter hospital stays.

A team-based multidisciplinary approach to managing peripherally inserted central catheter complications in high-risk haematological patients: a prospective study.

PURPOSE: Use of peripherally inserted central catheters (PICCs) has markedly increased during the last decade. However, there are few studies on use of PICCs in patients with haematological malignancies (HM) receiving intensive chemotherapy. Preliminary data suggest a higher rate of PICC-related complications in these high-risk patients. This prospective observational single-centre study aimed to investigate PICC-related complications after implementation of a multidisciplinary approach to PICC care and compared it with previous literature. METHODS: A total of 44 PICCs were inserted in 36 patients (27.3%, thrombocytopenia <50 × 10(9)/L at insertion) over 5045 PICC days (median duration, 114.5 days). RESULTS: No major insertion-related complications were observed. Major late complications were obstruction in 13.6% (1.19/1000 PICC days) of patients, catheter-related bloodstream infection in 6.8% (0.59/1000 PICC days), and catheter-related thrombosis in 4.5% (0.39/1000 PICC days). Premature PICC removal occurred in 34% (2.97/1000 PICC days) of patients. The overall rate of potentially major dangerous complications was particularly low (11.36%, 0.99/1000 PICC days) compared with previous studies. CONCLUSIONS: This study highlights the utility of a multidisciplinary approach for PICC care in adults with HM receiving intensive chemotherapy. We provide further data to support use of PICCs in such patient populations.

COVID-19 Outcomes in Patients with Hematologic Malignancies in the Era of COVID-19 Vaccination and the Omicron Variant.

A greater understanding of clinical trends in COVID-19 outcomes among patients with hematologic malignancies (HM) over the course of the pandemic, particularly the Omicron era, is needed. This ongoing, observational, and registry-based study with prospective data collection evaluated COVID-19 clinical severity and mortality in 1818 adult HM patients diagnosed with COVID-19 between 27 February 2020 and 1 October 2022, at 31 centers in the Madrid region of Spain. Of these, 1281 (70.5%) and 537 (29.5%) were reported in the pre-Omicron and Omicron periods, respectively. Overall, patients aged ≥70 years (odds ratio 2.16, 95% CI 1.64-2.87), with >1 comorbidity (2.44, 1.85-3.21), or with an underlying HM of chronic lymphocytic leukemia (1.64, 1.19-2.27), had greater odds of severe/critical COVID-19; odds were lower during the Omicron BA.1/BA.2 (0.28, 0.2-0.37) or BA.4/BA.5 (0.13, 0.08-0.19) periods and among patients vaccinated with one or two (0.51, 0.34-0.75) or three or four (0.22, 0.16-0.29) doses. The hospitalization rate (75.3% [963/1279], 35.7% [191/535]), rate of intensive care admission (30.0% [289/963], 14.7% [28/191]), and mortality rate overall (31.9% [409/1281], 9.9% [53/536]) and in hospitalized patients (41.3% [398/963], 22.0% [42/191]) decreased from the pre-Omicron to Omicron period. Age ≥70 years was the only factor associated with higher mortality risk in both the pre-Omicron (hazard ratio 2.57, 95% CI 2.03-3.25) and Omicron (3.19, 95% CI 1.59-6.42) periods. Receipt of prior stem cell transplantation, COVID-19 vaccination(s), and treatment with nirmatrelvir/ritonavir or remdesivir were associated with greater survival rates. In conclusion, COVID-19 mortality in HM patients has decreased considerably in the Omicron period; however, mortality in hospitalized HM patients remains high. Specific studies should be undertaken to test new treatments and preventive interventions in HM patients.

Two-weekly dose-adjusted (DA)-EPOCH-like chemotherapy with high-dose dexamethasone plus rituximab (DA-EDOCH14-R) in poor-prognostic untreated diffuse large B-cell lymphoma.

The activity and safety of two-weekly dose-adjusted (DA)-EPOCH (etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin)-like chemotherapy with high-dose dexamethasone plus rituximab (DA-EDOCH14-R) was explored in 20 patients with previously untreated poor prognosis diffuse large B-cell lymphoma (DLBCL). The main outcomes were compared with those of 27 poor-prognosis patients enrolled into a previous trial of 3-weekly DA-EPOCH-R. Toxicity was manageable and there were no therapy-related deaths. Three-year progression-free survival (PFS) was superior in the DA-EDOCH14-R group (95% vs. 74%, P = 0·08). Importantly, this improvement in PFS with the two-weekly DA-EDOCH14-R was particularly notable in patients with an age-adjusted International Prognostic Index of 3 (100% vs. 30%, P < 0·001).

Severe infections in patients with lymphoproliferative diseases treated with new targeted drugs: A multicentric real-world study.

BACKGROUND: Lymphoid neoplasms treatment has recently been renewed to increase antitumor efficacy and conventional chemotherapies toxicities. Limited data have been published about the infection risk associated with these new drugs, therefore this study analyzes the infectious complications in patients with lymphoproliferative diseases (LPD) treated with monoclonal antibodies (obinutuzumab, ofatumumab, brentuximab, nivolumab, or pembrolizumab), BTK inhibitors (ibrutinib and acalabrutinib), PI3K inhibitors (idelalisib) and BCL2 inhibitors (venetoclax). METHODS: Multicenter retrospective study of 458 LPD patients treated with targeted therapies in real-life setting, in 18 Spanish institutions, from the time of their commercial availability to August 2020. RESULTS: Severe infections incidence was 23% during 17-month median follow-up; cumulative incidence was higher in the first 3-6 months of targeted drug treatment and then decreased. The most frequent etiology was bacterial (54%). Nine (6%) Invasive fungal infections (IFI) were observed, in its majority in chronic lymphocytic leukemia (CLL) patients treated predominantly with ibrutinib. Significant risk factors for severe infection were: severe lymphopenia (p = 0.009, OR 4.7, range 1.3-1.7), combined targeted treatment vs single agent treatment (p = 0.014 OR 2.2 range 1.1-4.2) and previous rituximab (p = 0.03 OR 1.8, range 1.05-3.3). Infection-related mortality was 6%. In 22% of patients with severe infections, definitive discontinuation of the targeted drug was observed. CONCLUSION: A high proportion of patients presented severe infections during follow-up, with non-negligible attributable mortality, but infection incidence is not superior to the one observed during the chemotherapy era. In selected cases with specific risk factors for infection, antimicrobial prophylaxis should be considered.

Risk Factors and Mortality of COVID-19 in Patients With Lymphoma: A Multicenter Study.

Patients with cancer are poorly represented in coronavirus disease 2019 (COVID-19) series, and heterogeneous series concerning hematology patients have been published. This study aimed to analyze the impact of COVID-19 in patients with lymphoma. We present a multicenter retrospective study from 19 centers in Madrid, Spain, evaluating risk factors for mortality in adult patients with COVID-19 and lymphoma. About 177 patients (55.9% male) were included with a median follow-up of 27 days and a median age of 70 years. At the time of COVID-19 diagnosis, 49.7% of patients were on active treatment. The overall mortality rate was 34.5%. Age >70 years, confusion, urea concentration, respiratory rate, blood pressure, and age >65 score ≥2, heart disease, and chronic kidney disease were associated with higher mortality risk (P < 0.05). Active disease significantly increased the risk of death (hazard ratio, 2.43; 95% confidence interval, 1.23-4.77; P = 0.01). However, active treatment did not modify mortality risk and no differences were found between the different therapeutic regimens. The persistence of severe acute respiratory syndrome coronavirus 2-positive polymerase chain reaction after week 6 was significantly associated with mortality (54.5% versus 1.4%; P < 0.001). We confirm an increased mortality compared with the general population. In view of our results, any interruption or delay in the start of treatment should be questioned given that active treatment has not been demonstrated to increase mortality risk and that achieving disease remission could lead to better outcomes.

Jumping Translocation in a Patient with Acute Leukemia and Fatal Evolution.

Jumping translocations are uncommon cytogenetic abnormalities in which a segment of a donor chromosome, often 1q, is transferred to two or more receptor chromosomes. We describe the case of a 64-year-old man with a history of acute myeloid leukemia associated with myelodysplastic syndrome, who presented with a relapse of the leukemia and, concomitantly, with the appearance of a jumping translocation involving chromosome 1q. The patient had a poor clinical course without the possibility of performing targeted treatment, and he died 5 months after relapse. Jumping translocations are a reflection of chromosomal instability, and they could be related to epigenetic alterations such as pericentromeric chromatin hypomethylation, telomere shortening, or pathogenic variants of the TP53 gene. The existing data suggests a poor clinical outcome, a high risk of disease progression, and an unfavorable prognosis. More molecular studies are required to gain an in-depth understanding of the genetic mechanism underlying these alterations and their clinical significance and to be able to apply an optimal treatment to patients.

Impact of hematologic malignancy and type of cancer therapy on COVID-19 severity and mortality: lessons from a large population-based registry study.

BACKGROUND: Patients with cancer have been shown to have a higher risk of clinical severity and mortality compared to non-cancer patients with COVID-19. Patients with hematologic malignancies typically are known to have higher levels of immunosuppression and may develop more severe respiratory viral infections than patients with solid tumors. Data on COVID-19 in patients with hematologic malignancies are limited. Here we characterize disease severity and mortality and evaluate potential prognostic factors for mortality. METHODS: In this population-based registry study, we collected de-identified data on clinical characteristics, treatment and outcomes in adult patients with hematologic malignancies and confirmed severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection within the Madrid region of Spain. Our case series included all patients admitted to 22 regional health service hospitals and 5 private healthcare centers between February 28 and May 25, 2020. The primary study outcome was all-cause mortality. We assessed the association between mortality and potential prognostic factors using Cox regression analyses adjusted for age, sex, comorbidities, hematologic malignancy and recent active cancer therapy. RESULTS: Of 833 patients reported, 697 were included in the analyses. Median age was 72 years (IQR 60-79), 413 (60%) patients were male and 479 (69%) and 218 (31%) had lymphoid and myeloid malignancies, respectively. Clinical severity of COVID-19 was severe/critical in 429 (62%) patients. At data cutoff, 230 (33%) patients had died. Age ≥ 60 years (hazard ratios 3.17-10.1 vs < 50 years), > 2 comorbidities (1.41 vs ≤ 2), acute myeloid leukemia (2.22 vs non-Hodgkin lymphoma) and active antineoplastic treatment with monoclonal antibodies (2·02) were associated with increased mortality; conventional chemotherapy showed borderline significance (1.50 vs no active therapy). Conversely, Ph-negative myeloproliferative neoplasms (0.33) and active treatment with hypomethylating agents (0.47) were associated with lower mortality. Overall, 574 (82%) patients received antiviral therapy. Mortality with severe/critical COVID-19 was higher with no therapy vs any antiviral combination therapy (2.20). CONCLUSIONS: In this series of patients with hematologic malignancies and COVID-19, mortality was associated with higher age, more comorbidities, type of hematological malignancy and type of antineoplastic therapy. Further studies and long-term follow-up are required to validate these criteria for risk stratification.

Minimal residual disease evaluation by flow cytometry is a complementary tool to cytogenetics for treatment decisions in acute myeloid leukaemia.

The clinical utility of minimal residual disease (MRD) analysis in acute myeloid leukaemia (AML) is not yet defined. We analysed the prognostic impact of MRD level at complete remision after induction therapy using multiparameter flow cytometry in 306 non-APL AML patients. First, we validated the prognostic value of MRD-thresholds we have previously proposed (≥ 0.1%; ≥ 0.01-0.1%; and <0.01), with a 5-year RFS of 38%, 50% and 71%, respectively (p=0.002). Cytogenetics is the most relevant prognosis factor in AML, however intermediate risk cytogenetics represent a grey zone that require other biomarkers for risk stratification, and we show that MRD evaluation discriminate three prognostic subgroups (p=0.03). Also, MRD assessments yielded relevant information on favourable and adverse cytogenetics, since patients with favourable cytogenetics and high MRD levels have poor prognosis and patients with adverse cytogenetics but undetectable MRD overcomes the adverse prognosis. Interestingly, in patients with intermediate or high MRD levels, intensification with transplant improved the outcome as compared with chemotherapy, while the type of intensification therapy did not influenced the outcome of patients with low MRD levels. Multivariate analysis revealed age, MRD and cytogenetics as independent variables. Moreover, a scoring system, easy in clinical practice, was generated based on MRD level and cytogenetics.

Nonfatal pulmonary Trichoderma viride infection in an adult patient with acute myeloid leukemia: report of one case and review of the literature.

Trichoderma species have been recognized to be pathogenic in immunosuppressed hosts with increasing frequency. Trichoderma species are responsible for continuous ambulatory peritoneal dialysis associated peritonitis and infections in immunocompromised patients with a hematologic malignancy or solid organ transplantation. Trichoderma longibrachiatum is the most common species involved in these infections. We report the first case of nonfatal pulmonary infection caused by Trichoderma viride in leukemia patient. It had a successful answer to new antifungal agents as voriconazole and caspofungin. Trichoderma viride was isolated from pulmonary aspirate culture from a 54-year-old female who had received chemotherapy for acute myeloid leukemia. The minimal inhibitory concentrations for the organism were the following: amphotericin B (0.25 microg/mL) and voriconazole (2 microg/mL). Initially, she was treated unsuccessful with liposomal amphotericin B and voriconazole and caspofungin were added later. The patient is alive. We report one case along review of the literature.

Dose-adjusted EPOCH plus rituximab is an effective regimen in patients with poor-prognostic untreated diffuse large B-cell lymphoma: results from a prospective observational study.

This study was designed to assess the efficacy and safety of an infusional DA-EPOCH (dose-adjusted etoposide/vincristine/doxorubicin/bolus cyclophosphamide/prednisone) and rituximab (DA-EPOCH-R) regimen for patients with poor prognosis diffuse large B-cell lymphoma (DLBCL). Thirty-three patients, aged 21-76 years, with an age-adjusted International Prognostic Index (IPI) of 2 or 3, were enrolled, and 31/33 patients were evaluable for response. Consolidative radiation therapy was given to eight patients with bulky (> or =10 cm) disease at presentation. Overall, 26 patients (83.8%) achieved a complete remission (CR), four patients (12.9%) achieved a partial remission, and one patient (3.2%) died during induction. Two patients relapsed (7.6%) within 15 months. Grade 3-4 neutropenia developed in 52% of cycles and neutropenic fever in 14% of cycles (51% of patients). The estimates for event-free survival (EFS) and overall survival at 2 years were 68% and 75% respectively. The only factor related to poor EFS was the presence of three age-adjusted IPI-risk factors. We conclude that DA-EPOCH-R has clinically significant activity with a favourable toxicity profile for poor-prognostic DLBCL patients. The administration of DA-EPOCH-R as an outpatient regimen by using a single portable infusion pump may be a feasible alternative to improve the compliance and to reduce the total cost of this very effective regimen.

Changes in the natural history of progressive multifocal leukoencephalopathy in HIV-negative lymphoproliferative disorders: impact of novel therapies.

The aims of this study were to evaluate the clinical characteristics of HIV-negative patients affected by lymphoproliferative disorders (LPD) who developed progressive multifocal leukoencephalopathy (PML), to delineate the risk factors, and to analyze whether the new antineoplastic therapies are changing the natural history of this infectious disease. We retrospectively analyzed 46 cases with confirmed LPD-associated PML published from 1958 to 2004. Patients were stratified according to two different time periods: group A included patients diagnosed before 1989, and group B included patients diagnosed since 1990, after introduction of purine analogues. Group A patients (n = 22) had received alkylating agents and/or radiotherapy, and the majority (63.6%) had advanced Hodgkin disease. At univariate analysis, uncontrolled Hodgkin disease was the only risk factor for PML. In group B patients (n = 24), the most frequent treatments received were purine analogues (58.3%) and high-dose therapy with hematopoietic stem cell transplantation (33.3%; HDT/HSCT). B-cell chronic lymphocytic leukemia (45.8%) and aggressive non-Hodgkin lymphoma (24.9%) were the most frequent underlying LPDs. Patients treated with purine analogues were more likely to have active LPD, lower CD4 cell counts, and to be older and male than were HSCT recipients. The median interval from purine analogues or HDT/HSCT to PML was 11 months. In HDT/HSCT recipients, this interval was delayed for 10 months when peri-transplantation rituximab was used. Univariate analysis identified age >55 years, male sex, and CD4 cell counts

Iatrogenesis or bad luck? relapse of an LMP1-positive follicular lymphoma after immunosuppression for hepatitis-associated aplastic anaemia.

A 55-year-old man suffered a cutaneous relapse of an LMP1-positive follicular lymphoma after treatment with antithymocyte globulin and cyclosporine A (CSA) for a hepatitis-associated aplastic anaemia (AA). Rituximab was not effective, so CSA was tapered off. Lymphoma masses did not regress but AA relapsed. A second remission of both lymphoma and AA was achieved with high-dose cyclophosphamide, but the patient died of a bilateral pneumonia. The relationships between immunosuppression, viral reactivation and tumour growth are discussed. The use of rituximab and lamivudine in immunodepressed patients is also commented.

Spontaneous gas gangrene in malignant lymphoma: an underreported complication?

We report a case of spontaneous gas gangrene (SGG), the most rapidly progressive form of clostridial infection, in a patient with non-Hodgkin's lymphoma (NHL). We review the literature and examine the association between these two entities. A 43-year-old man with NHL developed fatal C. perfringens-associated SGG and massive hemolysis during induction chemotherapy. Although patients with NHL usually have several risk factors of SGG, such as bowel involvement or neutropenia, only two cases have been described previously in detail. Common features of all reports are a delayed diagnosis and a fatal outcome. Awareness of this condition should result in prompt antibiotic therapy at the onset of typical presenting symptoms in any lymphoma patient, especially if risk factors are present.

Elderly haematological patients with chemotherapy-induced febrile neutropenia have similar rates of infection and outcome to younger adults: a prospective study of risk-adapted therapy.

We prospectively evaluated 131 consecutive episodes of fever and chemotherapy-induced neutropenia in 85 adults with haematological malignancies to determine whether older patients (aged < 60 years) have different causes of fever and outcome than younger adults (aged < 60 years). Patients were stratified into high-risk and low-risk groups according to previously published criteria. High-risk patients received ceftazidime plus amikacin and low-risk patients received ceftazidime alone. All patients were hospitalized until fever and neutropenia resolved. Ninety one high-risk episodes were documented: 56 occurring in older patients (mean age 69 years) and 35 in younger adults (mean age 45 years). Non-Hodkgin's lymphoma and acute myeloid leukaemia were the most frequent underlying neoplasias in both age groups. Intensity of chemotherapy was similar in both age groups. Mean neutrophil count at entry, median duration of neutropenia, rate of documented infection, incidence of bacteraemia, response to therapy, overall mortality and infectious mortality were similar in the two high-risk age subgroups. The elderly subgroup had a trend to have more Gram-negative infections and the younger patients more Gram-positive infections. In addition, 40 low-risk episodes were registered: 29 in elderly patients (mean age 68 years) and 11 in younger patients (mean age 44 years). Elderly low-risk patients had more concurrent diseases that younger ones (P = 0.124). Mean neutrophil count at entry, median duration of severe neutropenia and rate of response were similar in the two age subgroups. All low-risk patients survived. In conclusion, elderly haematological cancer patients with febrile neutropenia show similar rates of infection and outcome to younger ones.

Apoptotic rate: a new indicator for the quantification of the incidence of apoptosis in cell cultures.

BACKGROUND: Late apoptotic cells divide into apoptotic bodies and are missed by current detection methods. This results in an artificially low apoptotic index (AI). METHODS: This study proposes a flow cytometry-based ratiometric method that uses an internal reference standard of microbeads combined with fluorescein-annexin V binding and 7-aminoactinomycin D to enumerate viable, necrotic, and early and late apoptotic cells within specific subsets of a heterogeneous culture. RESULTS: In the absence of cell growth, the number of apoptotic cells that undergo fragmentation into apoptotic bodies in culture can also be determined accurately by this method. This information can then be used to obtain the apoptotic rate (AR), a new indicator of apoptosis that calculates the proportion of cells that have undergone apoptosis with respect to the total number of seeded cells. The main limitation of the method is that the AR is only suitable for the study of apoptosis in noncycling cells. CONCLUSIONS: This study reveals the superiority of the proposed method over the widely used Nicoletti method and current annexin-V binding methods. The AI did not reflect the true incidence of lymphocyte apoptosis, neither in response to lectins or phorbol esters, nor to serum deprivation. AR was more sensitive than AI, detecting apoptosis at lower concentrations of cell death inducers in all the subsets studied.