• British Society for Haematology guidelines for the laboratory diagnosis of malaria

      Garg, Mamta (2022)
      The laboratory diagnosis of malaria depends on skilled examination of well-stained thick and thin blood films. Rapid diagnostic tests are a useful supplement and the use of nucleic acid-based testing in diagnostic laboratories should also be considered. These British Society for Haematology guidelines update the 2003 guidelines for malaria diagnosis. Training, quality control, incidental diagnosis, differential diagnosis and reference laboratory referral are considered.
    • A case series review of patients with Thrombocytopenia and Absent-Radii syndrome (TARS) and their management during pregnancy

      Halperin, Daniel; Myers, Bethan (2021)
      Bleeding diatheses due to platelet-related disorders can present challenges to treating clinicians, especially in the context of peri- and post-partum patients in the obstetric setting. Thrombocytopenia and Absent-Radii syndrome (TARS) is an inherited disorder characterized by reduced bone marrow platelet production, skeletal deformities affecting radii and other limbs; cardiac, renal, and other heterogeneous anomalies may occur. It is caused by the co-inheritance of a microdeletion and a nucleotide polymorphism in the RBM8A gene on chromosome 1. Bleeding phenotype is more severe than platelet numbers which might predict especially in infants but improves with age. There is minimal literature regarding the impact of pregnancy and puerperium. We describe the management of three pregnancies in the hematology-obstetrics clinic. As platelet counts normally decrease through pregnancy, close monitoring is required in TARS. No major bleeding was seen antenatally but two required platelet transfusions during labor. No other treatment definitely improves bleeding, although case reports of steroids claim variable success. Tranexamic acid may be helpful, and thrombopoietin agonists represent a potential future option.
    • Daratumumab plus bortezomib, melphalan, and prednisone versus bortezomib, melphalan, and prednisone in transplant-ineligible newly diagnosed multiple myeloma: frailty subgroup analysis of ALCYONE

      Garg, Mamta (2021)
      Background: In the phase 3 ALCYONE study, daratumumab plus bortezomib/melphalan/prednisone (D-VMP) versus bortezomib/melphalan/prednisone (VMP) significantly improved progression-free survival (PFS) and overall survival (OS) in transplant-ineligible, newly diagnosed multiple myeloma (NDMM) patients. We present a subgroup analysis of ALCYONE by patient frailty status. Patients and methods: Frailty assessment was performed retrospectively using age, Charlson comorbidity index, and baseline Eastern Cooperative Oncology Group performance status score. Patients were classified as fit (0), intermediate (1), or frail (≥2); a nonfrail category combined fit and intermediate patients. Results: Among randomized patients (D-VMP, n = 350; VMP, n = 356), 391 (55.4%) were nonfrail (D-VMP, 187 [53.4%]; VMP, 204 [57.3%]) and 315 (44.6%) were frail (163 [46.6%]; 152 [42.7%]). After 40.1-months median follow-up, nonfrail patients had longer PFS and OS than frail patients, but benefits of D-VMP versus VMP were maintained across subgroups: PFS nonfrail (median, 45.7 vs. 19.1 months; hazard ratio [HR], 0.36; P < .0001), frail (32.9 vs. 19.5 months; HR, 0.51; P < .0001); OS nonfrail (36-month rate, 83.6% vs. 74.5%), frail (71.4% vs. 59.0%). Improved greater than or equal to complete response and minimal residual disease (10-5)-negativity rates were observed for D-VMP versus VMP across subgroups. The 2 most common grade 3/4 treatment-emergent adverse events were neutropenia (nonfrail: 39.2% [D-VMP] and 42.4% [VMP]; frail: 41.3% and 34.4%) and thrombocytopenia (nonfrail: 32.8% and 36.9%; frail: 36.9% and 39.1%). Conclusion: Our findings support the clinical benefit of D-VMP in transplant-ineligible NDMM patients enrolled in ALCYONE, regardless of frailty status. Trial registration: ClinicalTrials.gov NCT02195479.
    • Guideline for the first-line management of Classical Hodgkin Lymphoma - A British Society for Haematology guideline

      Bhuller, Kaljit (2022)
      This guideline was compiled according to the British Society for Haematology (BSH) process at BSH Guidelines Process 2016 (b-s-h.org.uk). The Grading of Recommendations Assessment, Development and Evaluation (GRADE) nomenclature was used to evaluate levels of evidence and to assess the strength of recommendations. The GRADE criteria can be found at http://www.gradeworkinggroup.org. Recommendations are based on a review of the literature using Medline, PubMed/Medline and Cochrane searches beginning from 2013 up to January 2021. The following search terms were used: [Hodgkin lymphoma OR Hodgkin disease] NOT non-Hodgkin; AND [chemotherapy OR radiotherapy]; AND [elderly]; AND [teenage OR adolescent OR young adult]; AND [pregnancy]. Filters were applied to include only publications written in English, studies carried out in humans, clinical conferences, congresses, clinical trials, clinical studies, meta-analyses, multicentre studies and randomised controlled trials. References pre-2013 were taken from the previous version of this guideline.1 Review of the manuscript was performed by the British Society for Haematology (BSH) Guidelines Committee Haematology Oncology Taskforce, the BSH Guidelines Committee and the Haematology Oncology sounding board of BSH.
    • Guidelines for the management of mature T- and natural killer-cell lymphomas (excluding cutaneous T-cell lymphoma): a British Society for Haematology Guideline

      Fox, Christopher; Ahearne, Matthew (2022)
      Methodology This guideline was developed according to the British Society for Haematology (BSH) process at http://www.b-s-h.org.uk/guidelines. The Grading of Recommendations Assessment, Development and Evaluation (GRADE) nomenclature was used to evaluate levels of evidence and to assess the strength of recommendations. The GRADE criteria are described at http://www.gradeworkinggroup.org. Literature review details Ovid MEDLINE and PubMed were searched for English language articles up to December 2020 using the keywords: peripheral T-cell lymphoma, T prolymphocytic leukaemia, large granular lymphocyte leukaemia, adult T-cell leukaemia lymphoma, anaplastic large cell lymphoma, peripheral T-cell lymphoma not otherwise specified, angioimmunoblastic T-cell lymphoma, extra-nodal natural killer (NK)/T-cell lymphoma, aggressive NK cell leukaemia, enteropathy associated T-cell lymphoma, monomorphic epitheliotropic intestinal T-cell lymphoma, hepatosplenic T-cell lymphoma. Review of the manuscript Review of the manuscript was performed by the BSH Guidelines Committee, Haematology Oncology Task Force and the members of Haematology Oncology sounding board of the BSH. It was also on the members section of the BSH website for comment.
    • Guidelines on the diagnosis and management of Waldenström macroglobulinaemia-A British Society for Haematology guideline

      Krishna, Rajesh (2022)
      Scope: The objective of this guideline is to provide healthcare professionals with clear guidance on the management of patients with Waldenström macroglobulinaemia. In individual patients, circumstances may dictate an alternative approach. Methodology: This guideline was compiled according to the British Society for Haematology (BSH) process at http://www.b-s-h.org.uk/guidelines/proposing-and-writing-a-new-bsh-guideline/. Recommendations are based on a review of the literature using Medline, Pubmed, Embase, Central, Web of Science searches from beginning of 2013 (since the publication of the previous guidelines) up to November 2021. The following search terms were used: Waldenström('s) macroglobulin(a)emia OR lymphoplasmacytic lymphoma, IgM(-related) neuropathy OR cold h(a)emagglutinin disease OR cold agglutinin disease OR cryoglobulin(a)emia AND (for group a only) cytogenetic OR molecular OR mutation OR MYD88 OR CXCR4, management OR treatment OR transfusion OR supportive care OR plasma exchange OR plasmapheresis OR chemotherapy OR bendamustine OR bortezomib OR ibrutinib OR fludarabine OR dexamethasone OR cyclophosphamide OR rituximab OR everolimus, bone marrow transplantation OR stem cell transplantation. The Grading of Recommendations Assessment, Development and Evaluation (GRADE) nomenclature was used to evaluate levels of evidence and to assess the strength of recommendations. The GRADE criteria can be found at http://www.gradeworkinggroup.org. Review of the manuscript was performed by the British Society for Haematology (BSH) Guidelines Committee Haemato-Oncology Task Force, the BSH Guidelines Committee and the Haemato-Oncology sounding board of BSH. It was also on the members section of the BSH website for comment. It has also been reviewed by UK Charity WMUK; these organisations do not necessarily approve or endorse the contents.
    • Hydroxyurea - a cost effective treatment in developing countries for Atypical Chronic Myeloid Leukemia (aCML): case report of two patients

      Iqbal, Muhammad (2022)
      Introduction: Atypical chronic myeloid leukaemia (aCML) is a rare chronic myeloproliferative disorder with a poor prognosis. Case report: This case report presents two cases of male geriatric patients, both referred from primary care in rural areas and received at an urban clinic in a tertiary care hospital on separate instances. The first patient complained of low-grade fever (on/off), generalized body aches, rapid weight loss and shortness of breath for the last 2 months. The second patient arrived pale looking with symptoms of generalized body aches, dizziness and anorexia. Both patients were diagnosed to have aCML according to the World Health organization criteria. Management & outcome: Both the patients were from a low economic bracket and were treated with Hydroxyurea a relatively economic medicine successfully. The follow-up lasted for 12 months in both cases. No progression to acute myeloid leukaemia (AML) or relapse was observed. Discussion: This case report shows the promising results of Hydroxyurea in treating aCML and can be a cost effective alternate to other expensive treatments (allogeneic hematopoietic stem cell transplantation) and expensive medicines in lower and middle-income countries especially for resource-limited patients. These two cases show promising evidence for further studies to evaluate and conduct pharmaco-economic evaluations as well as clinical trials to compare hydroxyurea with other available alternative treatments for an affordable therapeutic option towards prevention of relapse and disease free survival after aCML.
    • Impact of major bleeding and thrombosis on 180-day survival in patients with severe COVID-19 supported with veno-venous extracorporeal membrane oxygenation in the United Kingdom: a multicentre observational study

      Isgro, Graziella; Yusuff, Hakeem (2022-02)
      Bleeding and thrombosis are major complications in patients supported with extracorporeal membrane oxygenation (ECMO). In this multicentre observational study of 152 consecutive patients (≥18 years) with severe COVID-19 supported by veno-venous (VV) ECMO in four UK commissioned centres during the first wave of the COVID-19 pandemic (1 March to 31 May 2020), we assessed the incidence of major bleeding and thrombosis and their association with 180-day mortality. Median age (range) was 47 years (23-65) and 75% were male. Overall, the 180-day survival was 70·4% (107/152). The rate of major bleeding was 30·9% (47/152), of which intracranial bleeding (ICH) was 34% (16/47). There were 96 thrombotic events (63·1%) consisting of venous 44·7% [68/152 of which 66·2% were pulmonary embolism (PE)], arterial 18·6% (13/152) and ECMO circuit thrombosis 9·9% (15/152). In multivariate analysis, only raised lactate dehydrogenase (LDH) at the initiation of VV ECMO was associated with an increased risk of thrombosis [hazard ratio (HR) 1·92, 95% CI 1·21-3·03]. Major bleeding and ICH were associated with 3·87-fold (95% CI 2·10-7·23) and 5·97-fold [95% confidence interval (CI) 2·36-15·04] increased risk of mortality and PE with a 2·00-fold (95% CI1·09-3·56) risk of mortality. This highlights the difficult balancing act often encountered when managing coagulopathy in COVID-19 patients supported with ECMO.
    • Implementation of Whole-Body MRI (MY-RADS) within the OPTIMUM/MUKnine multi-centre clinical trial for patients with myeloma

      Rennie, Winston (2022-07)
      Background: Whole-body (WB) MRI, which includes diffusion-weighted imaging (DWI) and T1-w Dixon, permits sensitive detection of marrow disease in addition to qualitative and quantitative measurements of disease and response to treatment of bone marrow. We report on the first study to embed standardised WB-MRI within a prospective, multi-centre myeloma clinical trial (IMAGIMM trial, sub-study of OPTIMUM/MUKnine) to explore the use of WB-MRI to detect minimal residual disease after treatment. Methods: The standardised MY-RADS WB-MRI protocol was set up on a local 1.5 T scanner. An imaging manual describing the MR protocol, quality assurance/control procedures and data transfer was produced and provided to sites. For non-identical scanners (different vendor or magnet strength), site visits from our physics team were organised to support protocol optimisation. The site qualification process included review of phantom and volunteer data acquired at each site and a teleconference to brief the multidisciplinary team. Image quality of initial patients at each site was assessed. Results: WB-MRI was successfully set up at 12 UK sites involving 3 vendor systems and two field strengths. Four main protocols (1.5 T Siemens, 3 T Siemens, 1.5 T Philips and 3 T GE scanners) were generated. Scanner limitations (hardware and software) and scanning time constraint required protocol modifications for 4 sites. Nevertheless, shared methodology and imaging protocols enabled other centres to obtain images suitable for qualitative and quantitative analysis. Conclusions: Standardised WB-MRI protocols can be implemented and supported in prospective multi-centre clinical trials. Trial registration NCT03188172 clinicaltrials.gov; registration date 15th June 2017 https://clinicaltrials.gov/ct2/show/study/NCT03188172.
    • Inguinal endometriosis: a systematic review

      Salta, Styliani (2022)
      Inguinal endometriosis is a very rare entity with uncertain pathophysiology, that poses several diagnostic and therapeutic challenges. This study aimed to summarize published literature on the diagnosis and treatment of this condition. Thus, a systematic literature search was conducted in PubMed/MEDLINE, Scopus and the Cochrane Library. An effort was made to numerically analyze all parameters included in case reports and retrospective analyses, as well. The typical and atypical features of this condition, investigations used, type of treatment and histopathology were recorded. More specifications about the surgical treatment, such as operations previously performed, type of surgery and treatment after surgery have been acknowledged. Other sites of endometriosis, the presence of pelvic endometriosis and the follow-up and recurrence have been also documented. Overall, the search yielded 61 eligible studies including 133 cases of inguinal endometriosis. The typical clinical presentation includes a unilateral inguinal mass, with or without catamenial pain. Transabdominal or transvaginal ultrasound was typically used as the first line method of diagnosis. Groin incision and exploratory surgery was the treatment indicated by the majority of the authors, while excision of part of the round ligament was reported in about half of the cases. Chemotherapy and radiotherapy were initiated in cases of coexisting endometriosis-related neoplasia. Inguinal recurrence or malignant transformation was rarely reported. The treatment of inguinal endometriosis is surgical and a long-term follow-up is needed. More research is needed on the effectiveness of suppressive hormonal therapy, recurrence rate and its relationship with endometriosis-associated malignancies.
    • Limitations of monitoring disease progression using circulating tumor DNA in lymphoma: an example from primary cutaneous DLBCL leg-type

      Walter, Harriet; Griffin, Yvette; Ahearne, Matthew; Saldanha, Gerald; Dyer, Martin (2022)
      No abstract available.
    • Polatuzumab vedotin plus obinutuzumab and lenalidomide in patients with relapsed or refractory follicular lymphoma: a cohort of a multicentre, single-arm, phase 1b/2 study

      McMillan, Andrew; Miall, Fiona (2021)
      Background: Obinutuzumab with polatuzumab vedotin or lenalidomide showed tolerability and activity in phase 1b/2 trials that recruited patients with relapsed or refractory follicular lymphoma. We aimed to examine whether the novel polatuzumab vedotin-obinutuzumab-lenalidomide (Pola-G-Len) combination might enhance antitumour response in patients with relapsed or refractory follicular lymphoma. Methods: This multicentre, single-arm phase 1b/2 study tested Pola-G-Len in patients with relapsed or refractory follicular lymphoma, and polatuzumab vedotin in combination with rituximab and lenalidomide in patients with relapsed or refractory diffuse large B-cell lymphoma. Here we report the results from the cohort of patients with follicular lymphoma. The trial was done in 18 cancer centres across three countries (Spain, UK, and USA). Patients (≥18 years old) with CD20-positive relapsed or refractory follicular lymphoma (excluding grade 3b) and Eastern Cooperative Oncology Group performance status of 2 or less who had previously received anti-CD20-containing chemotherapy were eligible for inclusion. During the dose-escalation phase, patients received six 28-day cycles of induction treatment with intravenous obinutuzumab 1000 mg (all cohorts), and intravenous polatuzumab vedotin and oral lenalidomide (Celgene, Summit, NJ, USA) in the following doses: 1·4 mg/kg polatuzumab vedotin and 10 mg lenalidomide (cohort 1); 1·8 mg/kg polatuzumab vedotin and 10 mg lenalidomide (cohort 2); 1·4 mg/kg polatuzumab vedotin and 15 mg lenalidomide (cohort 3); 1·8 mg/kg polatuzumab vedotin and 15 mg lenalidomide (cohort 4); 1·4 mg/kg polatuzumab vedotin and 20 mg lenalidomide (cohort 5); and 1·8 mg/kg polatuzumab vedotin and 20 mg lenalidomide (cohort 6). Polatuzumab vedotin was administered on day 1, lenalidomide on days 1-21, and obinutuzumab on days 1, 8, and 15 of cycle one and day 1 of cycles two to six of each 28-day cycle. During phase 2 (dose expansion phase), patients received six cycles of induction with Pola-G-Len at the recommended phase 2 dose established during dose-escalation. Patients who had a response or stable disease at the end of induction were eligible to enter the maintenance phase, in which they received obinutuzumab for 24 months at 1000 mg on day 1 of every other 28-day cycle for a total of 12 doses, and lenalidomide for 12 months at 10 mg on days 1-21 of each 28-day cycle for a maximum of 12 cycles. The primary activity endpoint was complete response at the end of induction. Adverse events were monitored throughout the study. The primary safety objective was to determine the maximum tolerated dose of Pola-G-Len. Analyses were in the safety population, which included all patients that received at least one dose of any of the component drugs (ie, all patients who entered the induction phases in both the escalation and expansion phases), and activity-evaluable population, which included all patients who received at least one dose of any of the component drugs at the recommended phase 2 dose (ie, all patients who received the recommended phase 2 dose in the dose escalation investigation and all patients who entered induction in the dose expansion investigation). This ongoing trial is registered at ClinicalTrials.gov, NCT02600897. Findings: Between March 24, 2016, and August 23, 2018, 56 patients (33 [59%] men and 23 [41%] women; 49 [88%] non-Hispanic or Latino) were enrolled. Two of four patients in cohort 2 reported dose-limiting toxicity events during dose escalation (one patient had grade 4 amylase and lipase elevation and one patient had grade 4 neutropenia and grade 3 thrombocytopenia), and there were no dose-limiting toxicities observed in cohorts 3 or 5; therefore, the recommended phase 2 dose for the dose-expansion was 1·4 mg/kg polatuzumab vedotin plus 20 mg lenalidomide. 46 (82%) patients were included in the activity-evaluable population. After a median follow up of 26·7 months (IQR 22·2-31·3) the objective response rate was 76% (90% CI 64-86) and complete response rate was 63% (90 CI 50-75). After a median follow-up of 27·0 months (IQR 18·7-34·0), the most common grade 3-4 adverse events were neutropenia (31 [55%] of 56 patients) and thrombocytopenia (14 [25%] patients). 61 serious adverse events were reported in 35 (63%) patients; the most common of which were febrile neutropenia (five [9%] patients; a sixth patient had febrile neutropenia, but this was not considered serious by the investigator), pneumonia (four [7%] patients), and pyrexia (four [7%] patients). One fatal adverse event (grade 5 septic shock) occurred in a patient who had discontinued study treatment due to disease progression and had initiated a new anti-lymphoma tyrosine kinase inhibitor treatment. This was not considered related to study treatment by the investigator. Interpretation: Pola-G-Len showed high complete response rates, although it did not reached the prespecified threshold for activity, in patients who were heavily pretreated with refractory follicular lymphoma. Our findings compare favourably with available therapies and support future investigation of Pola-G-Len in a larger patient population. Funding: Genentech/F Hoffmann-La Roche.
    • Predictors of adverse outcome in the first and second waves of the COVID-19 pandemic: results from a UK centre

      Martin, Christopher; Pan, Daniel; Hills, George; Modha, Deborah; Patel, Prashanth; Jenkins, David; Barton, Linda; Jones, William; Brunskill, Nigel; Haldar, Pranab; et al. (2022)
      Background/aims: Data concerning differences in demographics/disease severity between the first and second waves of COVID-19 are limited. We aimed to examine prognosis in patients presenting to hospital with COVID-19 amongst different ethnic groups between the first and second waves in the UK. Methods: In this retrospective cohort study, we included 1763 patients presenting to a regional hospital centre in Leicester (UK) and compared those in the first (n = 956) and second (n = 807) waves. Admission National Early Warning Scores, mechanical ventilation and mortality rate were lower in the second wave compared with the first. Results: Thirty-day mortality risk in second wave patients was approximately half that of first wave patients [adjusted hazard ratio (aHR) 0.55, 95% confidence interval (CI) 0.40-0.75]. In the second wave, Black patients were at higher risk of 30-day mortality than White patients (4.73, 1.56-14.3). Conclusion: We found that disporportionately higher risks of death in patients from ethnic minority groups were not equivalent across consecutive waves of the pandemic. This suggests that risk factors for death in those from ethnic minority groups are malleable and potentially reversible. Our findings need urgent investigation in larger studies.
    • Timing of high dose methotrexate CNS prophylaxis in DLBCL: a multicenter international analysis of 1,384 patients

      Martinez-Calle, Nicolás; Ahearne, Matthew; Miall, Fiona; Fox, Christopher (2022)
      Prophylactic high-dose methotrexate (HD-MTX) is often used for diffuse large B-cell lymphoma (DLBCL) patients at high risk of central nervous system (CNS) relapse, despite limited evidence demonstrating efficacy or the optimal delivery method. We conducted a retrospective, international analysis of 1,384 patients receiving HD-MTX CNS prophylaxis either intercalated (i-HD-MTX) (n=749) or at the end (n=635) of R-CHOP/R-CHOP-like therapy (EOT). There were 78 CNS relapses (3-year rate 5.7%), with no difference between i-HD-MTX and EOT; 5.7% vs 5.8%, p=0.98, 3-year difference: 0.04% (-2.0% to 3.1%). Conclusions were unchanged on adjusting for baseline prognostic factors or on 6-month landmark analysis (n=1,253). In patients with high CNS international prognostic index (n=600), 3-year CNS relapse rate was 9.1% with no difference between i-HD-MTX and EOT. On multivariable analysis, increasing age and renal/adrenal involvement were the only independent risk factors for CNS relapse. Concurrent intrathecal prophylaxis was not associated with reduction in CNS relapse. R-CHOP delays of ≥7 days were significantly increased with i-HD-MTX versus EOT, with 308/1573 (19.6%) i-HD-MTX treatments resulting in delay to subsequent R-CHOP (median 8 days). Increased risk of delay occurred in older patients when delivery was later than day 10 in the R-CHOP cycle. In summary, we found no evidence that EOT delivery increases CNS relapse risk versus i-HD-MTX. Findings in high-risk subgroups were unchanged. Rates of CNS relapse in this HD-MTX-treated cohort were similar to comparable cohorts receiving infrequent CNS prophylaxis. If HD-MTX is still considered for certain high-risk patients, delivery could be deferred until R-CHOP completion.
    • Unexplained peripheral blood eosinophilia with gastrointestinal symptoms

      Wardlaw, Andrew; Myers, Bethan; Rathbone, Barrie; Siddiqui, Salman; Wurm, Peter (2021)
      No abstract available.
    • Von Willebrand factor assays in patients with acquired immune thrombotic thrombocytopenia purpura treated with caplacizumab

      Hopkins, Barbara; Mensah, Patrick (2022)
      Acquired immune thrombotic thrombocytopenic purpura (iTTP) is a rare disease with a poor prognosis if undiagnosed. It is caused by autoantibody production to the von Willebrand factor (VWF) cleaving protease, A disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13). Caplacizumab, an immunoglobulin directed to the platelet glycoprotein Ibα receptor of VWF, has been reported to induce quicker resolution of iTTP compared to placebo. The laboratory measurement of VWF activity was significantly reduced in clinical trials of caplacizumab. Several VWF assays are available in the UK and this study investigated whether differences in VWF parameters were present in 11 patients diagnosed with iTTP and treated with daily caplacizumab. Chromogenic factor VIII activity, VWF antigen, collagen binding activity, VWF multimers and six VWF activity assays were measured prior to caplacizumab therapy and on several occasions during treatment. VWF antigen and collagen binding activity levels were normal or borderline normal in all patients. Ultra-large molecular weight multimers were present in all patients following treatment. VWF activity assays were normal or reduced during treatment, but this was reagent and patient dependant. In the unusual scenario of a caplacizumab-treated patient requiring measurement of VWF activity, it is important that laboratories understand how their local reagents perform as results cannot be predicted.