Prenatal Management of Pregnancies at Risk of Fetal Neonatal Alloimmune Thrombocytopenia (FNAIT): Scientific Impact Paper No. 61.

WHAT IS IT?: Fetal neonatal alloimmune thrombocytopenia (FNAIT), also known as neonatal alloimmune thrombocytopenia (NAIT) or fetomaternal alloimmune thrombocytopenia (FMAIT), is a rare condition which affects a baby's platelets. This can put them at risk of problems with bleeding, particularly into the brain. One baby per week in the UK may be seriously affected and milder forms can affect one in every 1000 births. HOW IS IT CAUSED?: Platelets are blood cells that are very important in helping blood to clot. All platelets have natural proteins on their surface called human platelet antigens (HPAs). In babies, half of these antigens are inherited from the mother and half from the father. During pregnancy, some of the baby's platelets can cross into the mother's bloodstream. In most cases, this does not cause a problem. But in cases of FNAIT, the mother's immune system does not recognise the baby's HPAs that were inherited from the father and develops antibodies, which can cross the placenta and attack the baby's platelets. These antibodies are called anti-HPAs, and the commonest antibody implicated is anti-HPA-1a, but there are other rarer antibody types. If this happens, the baby's platelets may be destroyed causing their platelet count to fall dangerously low. If the platelet count is very low there is a risk to the baby of bleeding into their brain before they are born. This is very rare but if it happens it can have serious effects on the baby's health. HOW IS IT INHERITED?: A baby inherits half of their HPAs from its mother and half from its father. Consequently, a baby may have different HPAs from its mother. As the condition is very rare, and even if the baby is at risk of the condition we have no way of knowing how severely they will be affected, routine screening is not currently recommended. WHAT CAN BE DONE?: FNAIT is usually diagnosed if a previous baby has had a low platelet count. The parents are offered blood tests and the condition can be confirmed or ruled out. There are many other causes of low platelets in babies, which may also need to be tested for. As the condition is so rare, expertise is limited to specialist centres and normally a haematologist and fetal medicine doctor will perform and interpret the tests together. Fortunately, there is an effective treatment for the vast majority of cases called immunoglobulin, or IVIg. This 'blood product' is given intravenously through a drip every week to women at risk of the condition. It may be started from as early as 16 weeks in the next pregnancy, until birth, which would be offered at around 36-37 weeks. Less common treatments that may be considered depending on individual circumstances include steroid tablets or injections, or giving platelet transfusions to the baby. WHAT DOES THIS PAPER TELL YOU?: This paper considers the latest evidence in relation to treatment options in the management of pregnancies at risk of FNAIT. Specifically, we discuss the role of screening, when IVIg should be started, what dose should be used, and what evidence there is for maternal steroids. We also consider in very rare selected cases, the use of fetal blood sampling and giving platelet transfusions to the baby before birth. Finally, we consider the approaches to blood testing mothers to tell if babies are at risk, which is offered in some countries, and development of new treatments to reduce the risk of FNAIT.

Use of granulocyte transfusions among haematology units in England and North Wales.

OBJECTIVES: To establish the current use of granulocyte transfusions in haematology patients and explore interest in further research. BACKGROUND: Granulocytes may be used for the treatment of severe infection in neutropenic patients or for primary or secondary prophylaxis. Clinical utility of granulocyte transfusions is unclear, and recent studies have demonstrated equivocal outcomes. Pooled granulocytes are the main granulocyte product used in England and Wales, but there are no data on the patterns of use and little consensus on accepted indications. METHODS: A survey was distributed to UK hospitals delivering intensive chemotherapy. Clinical scenarios were posed, with further questions on clinician experience of using granulocytes, availability of the product, barriers to use and interest in further research. RESULTS: The response rate was 57%; 34·9% of all responses were from allogeneic stem cell transplant centres. Paediatric centres comprised 9·5% respondents, and 19% centres had access to apheresis granulocytes. Of respondents, 58·7% had used granulocytes in the last 3 years, 89·2% of whom used granulocytes to treat refractory infection. There was little consensus on use of granulocytes in the given clinical scenarios even when patients clearly met national guideline criteria. Paediatric centres were overall more likely to recommend granulocyte use. The most frequently identified barrier to use of granulocytes was lack of evidence of effect. Of the respondents, 75% indicated a willingness to participate in further research. CONCLUSION: There remains a lack of consistency about use of granulocytes, which is unsurprising given the lack of clinical data to support their efficacy. We did, however, demonstrate a willingness to participate in further research.

A single unit transfusion policy reduces red cell transfusions in general medical in-patients.

BACKGROUND: The NICE guidelines for blood transfusion and the patient blood management recommendations state that a single unit of red cells should be the standard dose for patients with stable anaemia who are not bleeding. Studies have shown that changing clinical transfusion practice can be difficult and that many clinicians' order two units of blood as standard for patients needing a transfusion. AIM: A collaborative project between NHS Blood and Transplant and Kings College Hospital started in September 2014 to evaluate the impact of a single unit policy on blood usage. DESIGN METHODS: Training and education was undertaken for clinical staff on eight general medical wards and all staff working in the blood transfusion laboratory. We collected transfusion data for 12 months, (6 months before and after implementation). RESULTS: There was a decrease of 50% red cell unit usage between the two periods, equating to a unit cost saving of £28 670. The number of single unit transfusions, increased from 30 to 53% whilst the number of two units decreased from 65 to 43% (P < 0.001). DISCUSSION/CONCLUSION: This project has shown that transfusion practice can be changed and savings in blood usage can be achieved through the successful implementation of the single unit transfusions policy. Key to the implementation was engagement from key medical staff within the medical department in which the policy was implemented and support from the hospital transfusion team. Continued attention and training shall be needed to support these, and implement other, patient blood management recommendations.

Differentially expressed genes in adult familial myelodysplastic syndromes.

The precise genetic events leading to myelodysplastic syndromes (MDSs) and leukemic transformation remain poorly defined. Even less is known about adult familial MDS. We report an adult MDS family in whom enriched tissue-specific transcripts were derived by subtractive hybridization of cDNA from the mononuclear and CD34+ cells of affected and unaffected family members. These expression libraries were then hybridized to Genome Discovery arrays containing 18 404 genes and expressed sequence tags, and several clusters of differentially expressed genes were identified. A group of 21 genes was underexpressed (>5-fold) in affected vs unaffected family members, and among these were transcription factors and genes involved in myeloid differentiation, such as ZNF140 and myeloid nuclear differentiation antigen (MNDA). Another group of 36 genes was overexpressed (>5-fold), and these encoded proteins belonging to signaling pathways, such as Ras- and Fos-related genes. The top two genes downregulated in this MDS family, ZNF140 and MNDA, were similarly altered in another MDS family, and in some cases of sporadic MDS. Our data suggest that we have identified genes differentially expressed in adult familial MDS, and that alteration of some of these genes may also be important for the evolution of different stages or severity of sporadic MDS.

Frequent expression of HAGE in presentation chronic myeloid leukaemias.

Cancer testis (CT) antigens provide attractive targets for cancer-specific immunotherapy. Although CT genes are expressed in some normal tissues, such as the testis and in some cases placenta, these immunologically protected sites lack MHC I expression and as such, do not present 'self' antigens to T cells. To date, CT genes have been shown to be expressed in a range of solid tumours, but rarely in haematological malignancies. We have extended previous studies to investigate the expression of a comprehensive range of CT genes (MAGE-A1, -A3, -A6, -A12, BAGE, GAGE, HAGE,LAGE-1, NY-ESO-1 and RAGE) for their expression in a cohort of acute and chronic myeloid leukaemia patient samples. CT expression was not detected in 20 normal bone marrow or peripheral blood stem cell samples. In acute myeloid leukaemia (AML) nine of the 26 (35%) samples analysed expressed one or more of the CT genes with six of the samples (23%) expressing HAGE. In chronic myeloid leukaemia (CML) 24 of 42 (57%) presentation chronic myeloid leukaemia (CML) patient samples expressed one or more CT antigen with 23 expressing HAGE. We have shown that HAGE is frequently expressed in CML, and to a lesser extent in AML patient samples. This is the first demonstration of HAGE gene expression in myeloid leukaemia patients and the frequent expression of HAGE at disease presentation opens up the possibility of early immunotherapeutic treatments.

The "G-CSF test": the response to a single dose of granulocyte colony-stimulating factor predicts mobilization of hemopoietic progenitors in patients with hematologic malignancies.

A significant proportion of patients with hematologic malignancies fail to mobilize sufficient hemopoietic progenitor cells (HPC), thereby restricting wider application of autologous transplantation. It would be of considerable use to develop a test that could be used prospectively to assess an individual patient's capacity to mobilize HPC. Twenty-two patients with lymphoma, myeloma, and chronic lymphocytic leukemia were given a single dose of 12 microg/kg SC of granulocyte colony-stimulating factor (G-CSF). Blood colony-forming unit granulocyte-macrophage (CFU-GM) and CD34+ cells were scored prior to the test dose, and 72, 96, and 120 hours later. The patients were then mobilized with a standard cyclophosphamide and G-CSF regimen and had blood stem cells harvested. Patients were categorized as good, poor, or intermediate mobilizers on the basis of the CFU-GM/CD34+ cell harvest content and the number of aphereses required to reach established threshold counts. The outcome of cyclophosphamide/G-CSF mobilization was correlated with the response to the test dose of G-CSF. Good mobilizers had significantly higher peak CFU-GM values and CFU-GM increment in response to the test dose of G-CSF compared to intermediate and poor mobilizers. A peak CFU-GM count of > or = 250/mL identified the good mobilizers; conversely, all poor mobilizers had a peak CFU-GM count of <102/mL. An increment in CD34+ cells counts of > or = 2.5/microL was only observed in good mobilizers. The "G-CSF" test is a reliable test that can be used successfully for the assessment of mobilizable HPC in patients with hematologic malignancies. It can also be used to stratify patients enrolled in trials of mobilizing agents.

Single-donor granulocyte transfusions for improving the outcome of high-risk pediatric patients with known bacterial and fungal infections undergoing stem cell transplantation: a 10-year single-center experience.

Bacterial and fungal infections remain a significant cause of transplant-related mortality following allogeneic stem cell transplantation (SCT). Granulocyte transfusions (GTs) may reduce the neutropenic period after SCT and prevent further progression of the existing infection (that is, therapeutic GT) in addition to standard antibacterial and antifungal therapy. A retrospective analysis was performed on 28 consecutive pediatric SCT recipients who received at least one dose of GT between March 2003 and November 2013 at a single institution. All donors were conditioned with G-CSF+dexamethasone with harvest performed 12-18 h later. Indications for SCT were acute leukemia in 46% (13/28) and severe aplastic anemia in 21% (6/28). The main indications for GT were invasive fungal disease in 50%, bacterial infection in 21% and co-morbidities with predicted reduced tolerance to sepsis in 18% (5/28). The median number of GT was 6 (range 1-14) with a median dose of 3.56 × 10(10) granulocytes infused. The median increment in ANC was 1.06 × 10(9)/L and correlated with the granulocyte dose infused. Adverse reactions observed were mild and infrequent. Sixty-four percent of patients (18/28) are alive with only 2 of the 10 deaths being related to progression of infection. In addition there was a low overall incidence of grade 3-4 acute mucositis and a very low incidence (7%) of acute GvHD grade 3-4. Single-donor GTs afford protection to children undergoing SCT at additional risk of infection and may reduce the overall incidence of severe GvHD.

The myelodysplastic syndromes: towards a functional classification.

The French-American-British classification of myelodysplastic syndromes (MDS) has contributed greatly to better communication and conduct of clinical trials. However, the advent of novel cytogenetic, immunological and molecular techniques in recent years warrant some alterations to this purely morphological classification. This review aims at highlighting the advances which reflect more closely the unique biological and clinical features of various subtypes of MDS. We propose a comprehensive classification of MDS, to include the newly defined categories, as well as those not included in previous classifications, such as the therapy-induced and hereditary MDS. We hope that this classification will help in focusing attention on the biological features of MDS, the understanding of which will be crucial to combat this disease.

Acute erythremic myelosis (DiGuglielmo) following atypical aplastic anemia.

A patient presented with acute erythromyelosis (DiGuglielmo) which was developed after 3 yr of aplastic anemia. Aplastic anemia differed from the classical form, since erythroid cells and megakaryocytes were relatively preserved in the bone marrow. Treatment with androgens induced the increase of hematocrit and reticulocyte as well as general improvement. The sudden appearance of hemorrhagic syndrome due to thrombocytopenia was associated with aggravation of anemia and granulocytopenia. In the bone marrow, giant multinuclear proerythroblasts with bizarre nuclear morphology and PAS positivity with coarse granules was found. Serum erythropoietin (Ep) level was high. Bone marrow cells culture in vitro revealed two types of erythroid colonies: typical and giant multinuclear cells, both benzidine-positive. The number of colonies was irrespective to the Ep dose. "Autonomous" Ep independent growth of these colonies was also demonstrated. The number of colonies was more than 3 times higher per number of cells seeded when compared to normals, which indicated malignant proliferation and Ep independent growth. Treatment with 6-mercaptopurine and transfusions was without effect and the patient died after 15 days with signs of cerebral bleeding.

Idiopathic hyperammonaemia syndrome following allogeneic peripheral blood progenitor cell transplantation (allo-PBPCT).

An idiopathic hyperammonaemia syndrome has been reported in, and following chemotherapy for, various haematological malignancies as well as following bone marrow transplantation. It should be considered in the differential diagnosis of any neurological deterioration, and we describe a further case associated with an allogeneic peripheral blood progenitor cell transplant (allo-PBPCT).

Blast counts in blood progenitor cell (BPC) correlate with CD34+ cells and CFU-GM and are useful predictor of haemopoietic recovery after autologous BPC transplantation collections.

Assessment of the quality of blood progenitor cell (BPC) collections is based mainly on CD34+ cell enumeration by flow cytometry, or scoring of granulocyte-macrophage colony-forming cells (CFU-GM). A minimum cell dose for haemopoietic recovery can be defined by both assays; however, the CFU-GM assay can not be used for 'real-time' decisions, whereas CD34+ cell scoring requires facilities and expertise which are not universally available. We have investigated the possibility of using morphologically defined blast cells within BPC harvests as a surrogate marker of harvest haemopoietic stem/progenitor cell content, as well as their correlation with CD34+ cells and CFU-GM within the harvests. We have found that blast counts correlate strongly with both CD34+ cell counts and CFU-GM within BPC harvests, as well as with time to granulocyte and platelet recovery after autologous BPC transplantation (ABPCT). Furthermore, we have defined a threshold value of 1.3 x 10(6)/kg blasts, above which there is a high probability of rapid haemopoietic recovery after ABPCT. We conclude that blast count is a simple, rapid and reliable method of assessing BPC harvest quality.

Myelosuppressive chemotherapy to mobilize normal stem cells in chronic myeloid leukemia.

Ten patients with Ph+ chronic myeloid leukemia (CML) were treated with idarubicin, cytarabine and etoposide followed by G-CSF to harvest Ph-negative progenitor cells. Six were in first chronic phase (CP1), and four beyond CP1. Between two and six aphereses (median 3, total 36) were performed starting 9-26 days (median 14.5) after chemotherapy when the leukocyte count was 0.6-4.7 x 10(9)/l (median 1.2). 1.3-3.6 x 10(8) mononuclear cells/kg (median 2.8), 0-128.4 x 10(4) CFU-GM/kg (median 1.2; seven patients) and 0.3-25.1 x 10(6) CD34+ cells/kg (median 9.8; seven patients) were collected. Seven of 27 harvests showing metaphases were 100% Ph-negative, 11 partially Ph-negative, and nine were 100% Ph+. All three patients with 100% Ph-negative collections were in CP1 and within 4-26 months of diagnosis. Four of six CP1 patients showed significant cytogenetic response compared with none of four beyond CP1 (P = 0.036). The absolute neutrophil count remained < 0.5 x 10(9)/l for 9-44 days (median 15.5) following chemotherapy. Four patients (three Ph-negative) were autografted after 16 mg/kg busulfan (n = 2) or 200 mg/m2 melphalan (n = 2). One of the three patients receiving Ph-negative cells died of graft failure, and two are alive with 15% and 50% Ph-negative cells at 15 and 11 months on interferon-alpha. We conclude that it is possible to harvest Ph-negative cells after myelosuppressive chemotherapy in some CML patients treated early in the course of CP1. However, in view of lack of consistent response, investigation of alternative approaches is necessary.

A case of chronic myeloid leukemia without Ph1 translocation.

A case of chronic myeloid leukemia (CML) with a short survival (11 months) is described. Cytogenetic peculiarities of the bone marrow cells analyzed by G-banding consisted of a Ph1 chromosome with no translocation and a translocation t(5;13). Hematological characteristics were marked leukocytosis and massive splenomegaly. After treatment with busulfan complete hematological remission was achieved, followed by the appearance of a normal clone. However, 6 months later the patient entered the blastic crisis and a hyperdiploid clone appeared. The usual chemotherapy was given, but the patient responded only partially and died with a prevalence of pathologic myeloblasts in the bone marrow, corresponding to progression of the hyperdiploid clone.

Autologous blood stem cell transplantation in hematological malignancies.

Autologous blood stem cell transplants (ABSCT) are increasingly used for the treatment of haematological malignancies. The use of hemopoietic growth factors, in conjunction with stem cell mobilization by chemotherapeutic agents, has permitted successful harvests requiring only a few leukaphereses; cells mobilized in this manner contain a relatively large number of committed precursors of all lineages, as well as early progenitor cells capable of maintaining long-term haemopoiesis. Haematological recovery after ABSCT is rapid, thereby significantly shortening the period of post-chemotherapy neutropenia and thrombocytopenia. Furthermore, blood-derived grafts may contain fewer malignant cells than the bone marrow cells. The preliminary results have been so encouraging that it is envisaged that in myeloma, Hodgkin's disease and non-Hodgkin lymphomas, ABSCT may eventually replace autologous marrow transplantation.

Drug-induced agranulocytosis: bone marrow granulocytic progenitor cells.

Bone marrow CFU-GM and cluster forming cells were studied in ten patients in different stages of drug-induced agranulocytosis using a methylcellulose cell culture technique in vitro. In the aplastic phase of the disease (A), the number of both CFU-GM and cluster forming cells was decreased in comparison to normal values. In the regenerative phase of the disease (R), the number of both granulocytic progenitors increased but did not reach normal values. In patients considered to be recovered from acute agranulocytosis (Rec), a decreased number of progenitors persisted indicating residual damage at this granulocytic cell level. It is suggested that agranulocytosis is due to isolated damage of granulocytic cells and predictable cascade of events within different cell compartments could be used as an in vivo model for investigation of the regulation of granulopoiesis.

VAPEC-B chemotherapy in the treatment of aggressive non-Hodgkin's lymphoma: a retrospective analysis of 45 patients.

We performed a retrospective analysis on 45 patients who, between January 1989 and October 1993, received VAPEC-B chemotherapy for high and intermediate grade non-Hodgkin's lymphoma. The aim was to assess response and tolerance to treatment. The weekly regimen consisted of: doxorubicin 35 mg/m2 i.v. weeks 1,3,5,7,9,11; cyclophosphamide 350 mg/m2 i.v. weeks 1, 5, 9; etoposide 100 mg/m2 p.o. daily for 5 days, weeks 3, 7, 11; vincristine 1.4 mg/m2 i.v. (2 mg max.) weeks 2, 4, 6, 8, 10; bleomycin 10 mg/m2 i.v. weeks 2, 6, 10; methotrexate 12.5 mg i.t. weeks 1, 5, 9; prednisolone 50 mg p.o. daily for 6 weeks, reduced to 25 mg daily for 6 weeks. The patients treated were aged 22-71 years, 34 (75%) had high grade (Working Formulation) non-Hodgkin's lymphoma (NHL); 11 (24%) had intermediate grade NHL; 25 had Stage III/IV disease; and 14 (31%) had marrow involvement. The majority of patients (76%) received VAPEC-B as first line chemotherapy; the remainder received it for relapsing disease. Follow-up time from completion of VAPEC-B chemotherapy ranged from 6 months to 50 months (median 25). VAPEC-B, as first line therapy, induced a complete response (CR) and partial response (PR) in 79% and 18% respectively, whilst 3% had no response to treatment. VAPEC-B used for relapsing disease produced CR and PR in 64% and 27% respectively, whilst 9% failed to respond. Six patients in PR and five patients in CR have subsequently undergone an autologous bone marrow transplant or a peripheral blood stem cell transplant. In the group who received VAPEC-B first line but did not proceed to transplant (27 patients), five relapsed (three with CNS disease who had not had CNS prophylaxis). Tolerance to treatment was measured by WHO toxicity scores. The haemoglobin (Hb) toxicity median score for all patients was grade 1 (Hb 9.5-10.9 g/dl), and the white cell count (WCC), toxicity score was grade 2 (WCC 2.0-2.9 x 10(9)/l). No platelet toxicity was observed. Ten per cent of patients suffered grade 3 severity infections requiring antibiotics and there was one treatment related death. The majority of patients received VAPEC-B on time, however, 24% patients had a 2-week delay. VAPEC-B chemotherapy is an effective regimen for malignant lymphoma, either as a first line or as a salvage treatment. Although chemotherapy was given weekly, the tolerance to treatment was acceptable, thus making this short regimen a good alternative to CHOP chemotherapy.

Allogeneic haematopoietic SCT for chronic myelomonocytic leukaemia: a single-centre experience.

Haematopoietic SCT (HSCT) offers the only potentially curative option in chronic myelomonocytic leukaemia (CMML). In this study, we report on single-centre results of 18 patients with CMML who have undergone allogeneic HSCT. The median age of patients was 54 years. Seven patients had AML, which had transformed from CMML. Overall, 11 patients received stem cells from an unrelated donor. A total of 15 patients received a T-cell-depleted fludarabine/BU-based reduced-intensity conditioning HSCT. The actuarial 3-year OS, non-relapse mortality (NRM) and relapse incidence for the cohort was 31±11%, 31±14% and 47±13%, respectively. Patients with favourable cytogenetics had a 3-year disease-free survival of 65±17%, whereas none of the seven patients with intermediate or poor risk cytogenetics survived beyond 2 years (P<0.01). No patients with favourable risk cytogenetics died from NRM causes, while the 2-year NRM for the intermediate/poor risk cytogenetics subgroup was 71±22% (P<0.02). In terms of disease status at transplantation, patients who had <5% BM blasts had a 3-year disease-free survival of 46.9±19% compared with those with >5% blasts at the time of transplantation (that is, 20.0±13%). Recipient age, type of conditioning regimen or stem cell dose did not have a significant impact on overall outcomes. Our data support existing evidence that allogeneic HSCT is a feasible therapeutic option for CMML, with the ability to attain long-term remission among patient subgroups.