Mixed Phenotype Research Articles

Hematopoietic stem cell transplant for adults with mixed phenotypic acute leukemia.

6517 Background: Mixed phenotype acute leukemia (MPAL) is a rare acute leukemia subtype characterized by expression of lymphoid and myeloid lineage markers on blasts. Consequently, the role of hematopoietic stem cell transplant (HCT) consolidation is unclear. We conducted a retrospective study of MPAL patients treated at our institution integrating the impact of transplant status, chemotherapy, and initial response on survival. Methods: We evaluated patients with MPAL at Fred Hutchinson Cancer Center from 1/2005 – 9/2022, some of whom were previously reported (Huang et al, ASH 2023). Independent pathology review using 2022 WHO criteria verified MPAL diagnosis. Potentially transplant eligible patients were defined as those <75 years of age, diagnosed at least 6 months prior to data retrieval date, and alive 100 days after diagnosis with an evaluable induction response. Log-rank and Cox proportional hazard models were used to compare survival and adjust for baseline covariates. Survival was landmarked at 100 days. Results: Of 55 total patients, 42 were potentially HCT eligible and 30 received HCT (71.4%; 20/30 myeloablative). Patients who did not complete HCT were similar in age (median 46.2, range 18-70 vs median 50.3, range 24 – 74, p = ns) and had similar ECOG (ECOG 0-1 86.6% vs 75.0%, p = ns) at diagnosis. Nine of 12 non-transplanted patients had age-adjusted HCT-CI score ≤ 3. Patients receiving a HCT had better progression free survival (PFS) (p = 0.025) and were more likely to be alive at 48 months (61.6% vs. 32.4%, p = 0.011) with a median overall survival (mOS) of not reached (NR) (95% CI, 43 months – NR) compared to a mOS of 13 months (95% CI, 9 months – NR) for those not transplanted. Given that our prior work showed that the type of induction chemotherapy did not influence OS (p = 0.40) or remission status (p = 0.16) (Huang et al. ASH 2023), we investigated impact of remission status at the time of transplant. Differences in PFS and OS persisted when stratifying patients by induction response status with HCT status (PFS p = 0.0048; OS p < 0.0001). Patients who had a complete response (CR) followed by HCT had the longest mOS = NR (17 of 24 patients alive after 48 mo). Patients who had resistant disease (RD) and completed HCT had mOS = 43 months (95% CI, 43 months– NR). Patients who achieved CR but did not undergo HCT had mOS = 19 months (95% CI, 11 months – NR). Patients who had RD and did not undergo HCT had median OS of 6 months (95% CI, 6 – NR). The hazard ratio for death among patients who underwent HCT was 0.28 (95% CI 0.077 – 0.99, p = 0.049). Conclusions: HCT consolidation was significantly correlated with improved survival (p = 0.011). While potentially biased by selection and limited by small sample size, patients benefitted from undergoing a HCT regardless of response to prior therapy (p < 0.0001). This study suggests that patients with MPAL in CR after induction chemotherapy as well as those with less than complete responses should be considered for HCT consolidation.

Cytogenetic and molecular features of mixed-phenotype acute leukemia in US veterans and its potential impact on therapy.

e18535 Background: Mixed phenotype acute leukemia (MPAL) is a rare and heterogeneous subtype of acute leukemia. Per the 2022 WHO criteria, it is categorized based on its predominant immunophenotypic lineage (B vs. T) and the presence of a Philadelphia chromosome (Ph+) or a KMT2A rearrangement. While many other cytogenetic and molecular abnormalities have been described, due to the rarity of this disease, neither their incidence nor the role of targeted therapies has been well defined. We conducted a retrospective study of all veterans with MPAL diagnosed between 2000-2023 to examine the impact of different clinical, immunophenotypic, cytogenetic, molecular, and treatment strategies on overall survival (OS). Methods: Electronic medical record data from the Veterans Affairs Informatics and Computing Infrastructure database were used to identify 320 patients diagnosed with MPAL between 2000-2023 using the text utilization integration feature to query all notes. All patient charts were reviewed manually. Pathology reports and clinical notes were reviewed to collect data on diagnosis, treatment, and outcome. To date, 74 patients have been identified and included in this preliminary analysis. Cox regression analyses were used to compare OS between patients with different MPAL subtypes, cytogenetic, and molecular features as well as different treatment approaches. Multivariate analysis was performed with controls for age, race, and BMI. Results: In line with prior research, decreases in OS were seen among older patients > 65 yo compared to patients <30 yo and those who received an AML induction over ALL induction regimen with HR of 8.28 (p 0.04) and 2.02 (p 0.03) respectively. Patients who had a transplant had significantly improved OS with a HR of 0.19 (p <0.001). Of the 32 patients who had potentially targetable mutations, 14 received targeted maintenance therapy with either Ph+ or FLT3 inhibitors. Patients who got targeted maintenance therapy (including FLT3 inhibitors) had an OS of 55 months compared to 17 months for those who did not. 50% of those patients who did not get maintenance therapy went to transplant and 31% of patients who did get maintenance therapy went to transplant yet these patients still had improved OS. Conclusions: This preliminary data demonstrate that while MPAL is a complex and heterogeneous disease, a significant portion of patients have targetable mutations. Further studies evaluating targeted therapies such as FLT3, IDH, and menin inhibitors in MPAL patients may be warranted. [Table: see text]

Determinant factors for first-line treatment choice and effectiveness in pediatric eosinophilic esophagitis: an analysis of the EUREOS EoE CONNECT registry

This study compared short-term effectiveness of proton pump inhibitors (PPI), swallowed topical corticosteroids (STC), and dietary therapies in reversing clinical and histological features in pediatric patients with eosinophilic esophagitits (EoE). Determinants for treatment choice and PPI therapy effectiveness were also assessed. A cross-sectional study analysis of patients under 18 years old recruited onto the multicenter EoE CONNECT registry was performed. Clinico-histological response was defined as symptomatic improvement plus a peak eosinophil count below 15 per high-power field after treatment. Effectiveness of first-line options used in monotherapy was compared. Overall, 393 patients (64% adolescents) receiving PPI, STC, or dietary monotherapy to induce EoE remission were identified. PPI was the preferred option (71.5%), despite STC providing the highest clinico-histological response rates (66%) compared to PPI (44%) and diet (42%). Logistic regression identified fibrotic features and recruitment at Italian sites independently associated to first-line STC treatment; age under 12 associated to dietary therapy over other options. Analysis of 262 patients in whom PPI effectiveness was evaluated after median (IQR) 96 (70–145) days showed that this effectiveness was significantly associated with management at pediatric facilities and use of high PPI doses. Among PPI responders, decrease in rings and structures in endoscopy from baseline was documented, with EREFS fibrotic subscore for rings also decreasing among responders (0.27 ± 0.63 vs. 0.05 ± 0.22, p < 0.001). Conclusion: Initial therapy choice for EoE depends on endoscopic phenotype, patient’s age, and patients’ origin. High PPI doses and treatment in pediatric facilities significantly determined effectiveness, and reversed fibrotic endoscopic features among responders.What is Known:• Proton pump inhibitors are widely used to induce and maintain remission in EoE in real practice, despite other first-line alternative therapies possibly providing higher effectiveness.What is New:• Proton pump inhibitors represent up to two-thirds of first-line monotherapies used to induce EoE remission in pediatric and adolescent patients with EoE. The choice of STC as first-line treatment for EoE was significantly associated with fibrotic features at baseline endoscopy and recruitment in Italian centers; age less than 12 years was associated with dietary therapy.• PPI effectiveness was found to be determined by use of high doses, attendance at pediatric facilities, presenting inflammatory instead of fibrotic or mixed phenotypes, and younger age. Among responders, PPI therapy reversed both inflammatory and fibrotic features of EoE after short-term treatment.

Phenotyping of chronic pain in breast cancer survivors: an original study using the cancer pain phenotyping (CANPPHE) Network multidisciplinary international guidelines

PurposeThe primary aim of this cross-sectional study is to examine the prevalence of pain phenotypes in breast cancer survivors (BCS). A secondary aim entails examining whether health related quality of life differs between the main pain phenotypes in BCS.MethodsBCS who experienced chronic pain were asked to complete the numeric pain rating scale for pain, Margolis pain diagram, and short form 36 (SF-36). Following administration of questionnaires and quantitative sensory examinations were applied. To determine the prevalence of the predominant type of pain, a recently proposed classification system by the Cancer Pain Phenotyping (CANPPHE) Network was used.ResultsOf the 86 female participants, 19 (22.09%) had dominant neuropathic pain, 18 (20.93%) had dominant nociceptive pain and 14 (16.28%) had dominant nociplastic pain. 35 participants (40.70%) were classified as having mixed pain. One-way ANOVA revealed a significant difference between the four pain groups for the SF-36 general health (F = 3.205, p = 0.027), social functioning (F = 4.093, p = 0.009), and pain (F = 3.603, p = 0.017) subscale scores.ConclusionThis study found that pain in BCS was mostly of mixed phenotype, followed by predominantly neuropathic and nociplastic pain. Furthermore, it was found that, compared to BCS with predominant neuropathic and nociceptive pain, BCS with predominant nociplastic pain have lower health related quality of life in the areas of bodily pain and social functioning.

#760 Monogenic lupus with ANCA associated vasculitis-phenotype and ANCA positivity due to DNASE1L3 mutations: case report and review of the literature

Abstract Background and Aims Monogenic systemic lupus erythematosus (SLE) accounts for 7-10% of cases of early-onset SLE. DNASE1L3 is an extracellular nuclease that digests chromatin released from apoptotic cells and its deficiency, enhance autoantibody production and type I interferon responses and cause different autosomal recessive phenotypes including SLE and hypocomplementemic urticarial vasculitis syndrome (HUVS) [1]. In some cases patients can have a mixed SLE-AAV phenotype. Kidney involvement in patients with DNASE1L3 mutations is poorly characterised. Method We present the case report of a 7-years-old boy with monogenic SLE due to DNASE1L3 mutations who developed severe glomerulonephritis leading to kidney failure. Results The patient had a history of urticaria and arthralgia and presented with fever, haemolytic anaemia, lymphadenopathy, hepato-splenomegaly, erythematous- macular and necrotic skin lesions, and interstitial lung disease. He also had acute kidney injury (serum creatinine 5.4 mg/dL), nephrotic- range proteinuria (3.9 g/24 h) and microhaematuria. Immunological tests revealed low C3 and C4, positive anti-nuclear antibody (ANA) and myeloperoxidase-antineutrophil cytoplasmic antibodies (MPO-ANCA), with negativity of anti-extractable nuclear antigen antibodies (anti-ENA) and anti-double stranded DNA (anti-dsDNA). Kidney biopsy showed pauci-immune (mesangial C3+ and IgG+) necrotising crescentic glomerulonephritis. The patient proved refractory to standard immunosuppression with cyclophosphamide and steroids. Two years later, due to renal function decline, a new kidney biopsy was performed and showed diffuse necrotising glomerulonephritis with endo- and extra-capillary proliferation; intense mesangial and subendothelial full-house deposits were evident on immunofluorescence. The patient did not respond to other lines of immunosuppression (mycophenolate mofetil, tacrolimus, azathioprine, rituximab), including the JAK inhibitor ruxolitinib. Kidney failure progressed to end stage at the age of 15 years, when the patient began haemodialysis. The kidney presentation, immunological profile and the first kidney biopsy led to diagnosis of AAV. However, the diagnosis was questioned due to the refractory course, the systemic clinical presentation and the second kidney histology (consistent with lupus nephritis). High interferon signature was detected and in the suspicion of a monogenic lupus nephritis Whole Exome Sequencing was performed and revealed homozygous DNASE1L3 variants (c.290_291delCA p.T97Ifs*2); thus, a final diagnosis of DNASE1L3 monogenic SLE was made. DNASE1L3 serum levels were normal but the DNAse enzymatic activity was low. Conclusion Renal involvement is one of the most frequent manifestations of DNASE1L3-related lupus; in the literature 32 cases of DNASE1L3-associated lupus nephritis are described. ANCA are positive in about 55% of these cases and are associated with a mixed lupus-AAV phenotype. All cases are characterized by resistance to common immunosuppressors and poor renal prognosis [2]. DNASE1L3 regulates neutrophil extracellular trap (NET) clearance and when this function is reduced the permanence of NETs induces endothelial damage and autoantibody formation. This pathogenic pathway is shared both by SLE and by AAV. There is no association between specific mutations and antibody positivity or clinical manifestations.

Why am I grinding and clenching? Exploration of personality traits, coping strategies, oral parafunctional behaviors, and severe sleep bruxism in a polysomnographic study.

Causal relationships between psychopathological symptoms, personality traits, coping mechanisms, and sleep bruxism (SB) were studied in the past, giving inconsistent results mostly based on self-assessment evaluations. This polysomnography-based cross-sectional study aimed to explore the relationships between severe SB, personality traits (according to the Big Five model), and coping strategies with objective polysomnographic verification. The study included 66 participants divided into severe SB (SSB) (n=32) and no or mild SB (n=34) groups based on video-polysomnography performed in the sleep laboratory. Questionnaire assessment included the use of the Beck Depression Inventory, Beck Anxiety Inventory, Mini-COPE, International Personality Item Pool Big Five Markers 20-Item version, and Oral Behavior Checklist. Participants with SSB presented with fewer self-reported anxiety (p=0.008) and depressive (p=0.01) symptoms than the non- or mild-SB groups. The SSB group scored significantly higher in Big Five personal traits such as extraversion (p=0.007), emotional stability (p=0.013), and intellect (p=0.004), while regarding coping strategies, the SSB group was less likely to use negative strategies: self-distraction (p=0.036), denial (p=0.006), venting (p=0.03), behavioral disengagement (p=0.046), and self-blame (p=0.003), and turning to religion (p=0.041). The intensity of oral parafunctional behaviors was comparable in both groups (p=0.054). Emotional stability was a moderate protective factor (p=0.004), and the self-blame strategy was a strong risk factor (p<0.001) for increased oral parafunctional behavior intensity. Phasic activity negatively correlated with anxiety symptom severity (p=0.005), whereas tonic (p=0.122) and mixed (p=0.053) phenotypes did not. SB intensity was a protective factor against anxiety symptoms (p=0.016). In terms of psychopathology, severe sleep bruxers tend to present less severe anxiety and depressive symptoms, while some of their personality traits (extraversion, emotional stability, and intellect) were more strongly pronounced. SSB is possibly related to the lesser use of the "maladaptive" coping strategies and there were no specific coping strategies preferred by SSB participants, compared to the other group. These observations require further studies, as it should be determined whether SB (especially phasic activity) might be a form of a somatization/functional disorder. Further research should focus on the psychogenic background of oral parafunctional behaviors, which occur more often in less emotionally stable personalities and in people using self-blame coping strategies.

A hybrid protocol CLAG-M, a possible player for the first-line therapy of patients with mixed phenotype acute leukemia. A Polish Adult Leukemia Group experience.

Mixed-phenotype acute leukemia (MPAL) is a rare disease with poor prognosis. So far, no standard approach has been established as the "know-how" of MPAL is based only on retrospective analyses performed on small groups of patients. In this study, a retrospective analysis of the outcomes of adult MPAL patients included in the PALG registry between 2005 and 2024 who received the CLAG-M hybrid protocol as induction or salvage therapy was performed. Sixteen of 98 MPAL patients received CLAG-M: eight as first-line and eight as salvage therapy. In the first line, two patients achieved partial response (PR), and six achieved complete remission (CR), of whom four successfully underwent allogeneic hematopoietic stem cell transplantation (alloHSCT). Two patients who did not undergo alloHSCT promptly relapsed. Within the whole group, the overall response rate (ORR) was 75% (n = 12/16). With the median follow-up of 13 months, six out of eight patients remain in CR, however, two of them died due to acute graft versus host disease. Out of eight patients who received CLAG-M in the second line, four patients (50%) obtained CR. AlloHSCT was conducted in seven cases, six of which were in CR. Only two patients remained in CR at the time of the last follow-up. Tolerance to treatment was good. The median times for severe neutropenia and thrombocytopenia were 22 days (range, 16-24) and 17 days (range, 12-24), respectively. Overall, grade 3-4 infections were observed in 12 cases, and all infections presented successful outcomes. CLAG-M is an effective first-line salvage regimen for MPAL with an acceptable safety profile. Early achievement of CR with prompt alloHSCT allows for satisfactory disease control.

WHEN AND WHERE DOES A CARRIER OF HEREDITARY TRANSTHYRETIN AMYLOIDOSIS (ILE88LEU) SHOW THE FIRST SIGNS OF THE DISEASE? DIAGNOSTIC CHALLENGES OF A NOT ONLY CARDIOGENIC MUTATION

Abstract We present the case of a 58–year–old man with a familiar history (mother affected) positive for ATTR–v (Ile88Leu) amyloidosis. The mother initially had a pure cardiac phenotype and was firstly treated with tafamidis, but progression of the disease to the nervous system warranted a switch to patisiran. Our patient carried the Ile88Leu mutation and had a medical history notable for surgically treated left carpal tunnel syndrome (CTS) and untreated right CTS. Furthermore, he was affected by systemic arterial hypertension and was a habitual smoker. He has been followed at our Institution since 2018. His first echocardiogram showed normal left ventricular ejection fraction (LVEF 65%), mild hypertrophy (anterior septum: 12 mm), and borderline values of GLS (– 16%) without any specific pattern. Further exams were executed: Bone scintigraphy (BS): negative (Perugini score 0); Coronary artery ct scan: moderate stenosis of proximal LAD; Neurologic evaluation: no signs of peripheral neuropathy. The clinical picture remained unaltered in 2020 and 2021. However, during November 2022 the patient developed atrial fibrillation (AF) and presented to the ED for dyspnea and palpitations. Electrical cardioversion was performed with success but subsequent recurrences warranted AF ablation in April 2023. At that moment, the patient was re–evaluated. Basal ECG did not show any significant alteration (FIg.1)This time, echocardiogram showed mild worsening of hypertrophy (13 mm septum), GLS – 16% with apical sparing pattern, no other major variations (Fig.2). BS was repeated and revealed grade 1 Perugini score (Fig.3). Hence, a series of clinical question arose: Should we begin a specific treatment or is the disease in an excessively early stage that does not warrant treatment? Is AF linked to concomitant arterial hypertension or might it be an early marker of disease? We questioned the usefulness of endomyocardial biopsy in this specific setting. In this case, we repeated electromyography that showed mild axonal sensitive–motor polyneuropathy. Finally, we concluded for initial signs of systemic amyloidosis ATTR–v (Ile88Leu) type with mixed phenotype (neurological+cardiac). The neurological involvement allowed us to start a gene–silencer (Patisiran). In conclusion, a periodic neurological evaluation is necessary and has crucial therapeutic implications in all genotypes. Indeed, there is increasing evidence that Ile88Leu is a not only cardiogenic mutation.

SCARY ECG IN TAKOTSUBO CARDIOMYOPATHY

Abstract Takotsubo cardiomyopathy (TTC) is characterized by a transient ventricular wall motion abnormality and shares common features with acute coronary syndrome (ACS) with a wide spectrum of triggers, such as both emotional or physical, activating common toxic catecholamines–mediated pathways. TTC might be associated with severe clinical complications. We report a case of a 74–year female patient with circulatory shock with mixed phenotype, distributive due to sepsis and cardiogenic due to TTC. The patient presented to the emergency department with severe hypotension, pathological qSOFA score and clinical and laboratory findings consistent with severe sepsis. ECG showed diffuse lambda ST–segment elevation, also called “shark fin”(fig.1). According to hemodynamically unstable scenario and ECG pattern, emergency coronary angiography was performed, showing absence of coronary artery disease(fig. 2). Echocardiography revealed severe LV systolic disfunction (EF 20–25%) with apical ballooning and severe systolic anterior motion (SAM) of the anterior mitral leaflet eliciting dynamic LVOTO with peak gradient greater than 90 mmHg; fig. 3) and severe MR. Patient was treated with cautious fluid administration, hemotrasfusion and empiric large spectrum antibiotic therapy. According to TTC working diagnosis, even if septic state was thought to contribute to cardiogenic shock, norepinephrine was not administered due to concerns for LVOTO and SAM. After overcoming of dramatic first phase and after hemodynamic steadiness was achieved, beta blockade and ACEi were progressively administered. Seriate transthoracic echocardiogram revealed progressive improvement of LV systolic function, regression of apical ballooning, SAM and LVOTO. ECG monitoring showed regression of ischemic ST elevation. ST elevation has been described in about 56% of TTC cases, however with amplitude tipically lower than in STEMI. Lambda–wave ST elevation has been rarely reported. Vasospasm, local intermittent microvascular constriction, catecholamine–induced metabolic toxicity, transmural systolic wall tension and its the mechano–electric feedback can explain scaring ECG–pattern. Despite its rare prevalence in TTC “shark fin” pattern appears to relate to adverse outcome and it should be used as an early marker of potential further clinical deterioration, suggesting more intensive monitoring to avoid delays in possible therapy escalation.

Treatment characteristics of patients with hereditary transthyretin amyloidosis: a cohort study

BackgroundThere are novel medications approved for the treatment of hereditary transthyretin amyloidosis (ATTRv), classified as transthyretin (TTR) stabilizers or gene silencers. While many patients may be on both classes of medications, there is no data available on the safety and efficacy of combination therapy.ObjectivesTo describe ATTRv patient and TTR-targeted therapy characteristics in a US cohort, and compare outcomes with combination therapy versus monotherapy.MethodsWe performed a retrospective cohort study with electronic health record data of patients with ATTRv seen at a single institution between January 2018 and December 2022. We collected data on symptomatology, gene mutation, disease severity, ATTRv treatment, hospitalizations, and mortality.ResultsOne hundred sixty-two patients with ATTRv were identified. The average age at diagnosis was 65 years. 86 patients (53%) had the V122I variant. 119 patients were symptomatic, of whom 103 were started on ATTRv-specific treatment. 41 patients (40%) had cardiomyopathy only, and 53 (51%) had a mixed phenotype of cardiomyopathy and neuropathy. 38 patients (37%) received therapy with both a gene silencer and protein stabilizer. 9 patients (15%) in the monotherapy group had two or more cardiac hospitalizations after starting treatment, compared to 3 patients (9%) on combination therapy (p=0.26). The adjusted hazard ratio of all-cause mortality for the patients on combination therapy compared to monotherapy was 0.37 (0.08-1.8, p=0.21).ConclusionsWhile the efficacy is unproven, over one-third of patients with ATTRv are on both a stabilizer and a silencer. There were no safety issues for combination therapy. There was a trend towards improved hospitalizations and survival in patients in the combination group but this was not statistically significant. Larger studies with longer follow-up are necessary to determine benefit of combination therapy.

Reproducible preclinical models of androgen receptor driven human prostate cancer bone metastasis.

Preclinical models recapitulating the metastatic phenotypes are essential for developing the next-generation therapies for metastatic prostate cancer (mPC). We aimed to establish a cohort of clinically relevant mPC models, particularly androgen receptor positive (AR+) bone metastasis models, from LuCaP patient-derived xenografts (PDX) that reflect the heterogeneity and complexity of mPC. PDX tumors were dissociated into single cells, modified to express luciferase, and were inoculated into NSG mice via intracardiac injection. The progression of metastases was monitored by bioluminescent imaging. Histological phenotypes of metastases were characterized by immunohistochemistry and immunofluorescence staining. Castration responses were further investigated in two AR-positive models. Our PDX-derived metastasis (PDM) model collection comprises three AR+ adenocarcinomas (ARPC) and one AR- neuroendocrine carcinoma (NEPC). All ARPC models developed bone metastases with either an osteoblastic, osteolytic, or mixed phenotype, while the NEPC model mainly developed brain metastasis. Different mechanisms of castration resistance were observed in two AR+ PDM models with distinct genotypes, such as combined loss of TP53 and RB1 in one model and expression of AR splice variant 7 (AR-V7) expression in another model. Intriguingly, the castration-resistant tumors displayed inter- and intra-tumor as well as organ-specific heterogeneity in lineage specification. Genetically diverse PDM models provide a clinically relevant system for biomarker identification and personalized medicine in metastatic castration-resistant prostate cancer.

Clonotypic VDJ Rearrangements in Mixed Phenotype Acute Leukemia can be Successfully Utilized to Track Minimal Residual Disease.

Multiplex polymerase chain reaction (PCR) and next-generation sequencing (NGS) can both be used to identify a neoplastic clonotype by targeting CDR3 and assessing rearrangements in IgH, IgK, IgL, TCR-β, and TCR-gamma loci. The clonotypic sequence can be robustly used to track minimal residual disease (MRD). The ability to track MRD by NGS in mixed phenotype acute leukemia (MPAL) is unknown and warrants investigation. Institutional Review Board (IRB) approval was obtained. Central Moffitt Cancer Center (MCC) database was searched to locate any patients with MPAL from over 600,000 entries. Patient charts were manually curated to identify those with clonoSEQ data, and clinical data was procured from the electronic medical record (EMR). Twenty-nine patients with MPAL were identified. Only 2 patients with clonoSEQ testing were found. Both demonstrated a B/myeloid phenotype, and both were bilineal. NGS (clonoSEQ) identified 4 dominant (IGH) (patient A; 8/2019) and 2 dominant sequences (patient B; 10/2019), respectively. In both patients, clonoSEQ testing successfully tracked minimal residual disease and mirrored clinical disease burden. This report is the first to confirm the utility of NGS-based MRD tracking in patients with MPAL and shows increased sensitivity of NGS over MRD flow cytometry.

Intellectual Developmental Disorder with Speech Delay and Axonal Peripheral Neuropathy (IDDSAPN): A Mixed Phenotype.

Intellectual Developmental Disorder with Speech Delay and Axonal Peripheral Neuropathy (IDDSAPN): A Mixed Phenotype.

A survey of severe asthma in Canada: results from the CASCADE practice reflective program.

Since the last guidance was published by the Canadian Thoracic Society, there have been several advances in the clinical management of severe asthma. To gain a better understanding of the current standards of care and treatment patterns of patients, the CASCADE practice reflective program was established to conduct a real-world analysis of severe asthma management among specialists in Canada with a goal of identifying areas of opportunity to enhance patient management and outcomes. The CASCADE program was a two-part practice reflective and assessment program delivered through an on-line portal for selected specialists (Respirologists and Allergists) in Canada. The program consisted of a one-time overview survey of physician practice to establish overall practice parameters, followed by a review of at least 5 severe asthma patients to establish the current landscape of severe asthma management. The program collected practice overview surveys from 78 specialists (52 Respirologists, 24 Allergists, and 2 General practice physicians with an interest in respiratory disease) in 8 provinces. Practices included a variety of types in both large metropolitan centres and smaller regional settings. There were 503 patients reviewed and included in the program. Most (65%) patients were currently using a biologic treatment, 30% were biologic naive, and 5% had used a biologic treatment in the past. Most patients (53%) were reported to have mixed allergic and eosinophilic phenotypes, despite a perception that allergic, eosinophilic and mixed phenotypes were evenly balanced in the physician practice. Overall, patients currently treated with biologic agents had parameters suggesting higher control and were more satisfied with treatment. However, there was less than optimal treatment satisfaction for more than half of all patients, particularly for those patients not treated with a biologic agent. Phenotyping is hampered by poor availability for several assessments, and the full range of treatments are not currently fully utilized, partly due to physician familiarity with the agents and partly due to prescribing restrictions. Even when treated with biologic agents, patient satisfaction can still be improved.

Clinicopathological and Immunophenotypic Characteristics of Series of ZNF384 Re-arranged Acute Leukemias.

Rearrangement involving ZNF384 gene (ZNF384-r) is recently being described in acute leukemias. We present the clinic-pathological and immunophenotypic findings in a series of five cases of acute leukemia with ZNF384-r reported in our Institute between September 2020 to September 2023. Notably, while TCF3::ZNF384 fusion was the most frequently encountered abnormality, the fusion partner was not identified in two patients with ZNF384-r BCP-ALL. Immunophenotypically, patients presenting as B-cell precursor acute lymphoblastic leukemia (BCP-ALL) had a distinct profile characterized by weak or absent CD10 expression and the presence of myeloid markers such as CD13/CD33. Our findings underscore the importance of recognizing the distinct immunophenotypic features of ZNF384-r leukemias, particularly in cases presenting with atypical BCP-ALL or B/Myeloid mixed phenotype acute leukemia phenotypes. Moreover, these findings highlight the necessity for tailored diagnostic algorithms in clinical laboratories to facilitate the timely and accurate diagnosis of this clinically relevant leukemia subtype.

Genomic and Pathologic Profiling of Very Well-Differentiated Gastric Adenocarcinoma of Intestinal Type: A Study With Emphasis on Diffuse-Type Transformation.

Very well-differentiated adenocarcinoma of intestinal type is a distinct subtype of gastric cancer characterized by anastomosing glands with a hand-in-hand pattern and low-grade cytologic atypia resembling intestinal metaplasia. This is a slow-growing neoplasm with an indolent clinical course; however, a subset demonstrates transformation into adenocarcinoma with higher-grade histology, typically diffuse-type carcinoma, and behaves aggressively. This study aimed to better characterize the genomic and pathologic features, with a focus on factors associated with diffuse-type transformation. A total of 58 cases with (n=31) and without (n=27) diffuse-type transformation were analyzed for molecular and pathologic features. First, comprehensive deep DNA sequencing was conducted in 18 cases (discovery cohort), followed by a digital droplet polymerase chain reaction of hot spot RHOA mutations in 40 cases (validation cohort). In total, RHOA mutations were the most common alteration (34%), followed by loss of ARID1A (12%), p53 alterations (10%), and CLDN18 :: ARHGAP26/6 fusions (3.4%). FGFR2 amplification was identified in an advanced case with a p53 alteration. Altered p53 expression was recognized only in higher-grade components and was significantly associated with advanced disease ( P =0.0015) and diffuse-type transformation ( P =0.026). A mixed mucin phenotype was also strongly correlated with advanced disease ( P <0.001) and diffuse-type transformation ( P <0.001). Decreased E-cadherin expression was frequently observed (74%) in poorly cohesive components. This study demonstrated that a subset of RHOA -mutant diffuse-type gastric cancers develops through the transformation of very well-differentiated adenocarcinoma of intestinal type. Our observations suggest a mixed mucin phenotype as a risk factor and alterations in p53 and E-cadherin as drivers of diffuse-type transformation.

Allogeneic hematopoietic cell transplantation for acute myeloid leukemia with BCR::ABL1 fusion.

BCR::ABL1 fusion is found in<1% of de novo acute myeloid leukemia (AML) cases and confers a poor prognosis. This Japanese nationwide survey analyzed patients with AML (n=22) and mixed phenotype acute leukemia (MPAL) (n=10) with t(9;22) or BCR::ABL1 who underwent allogeneic hematopoietic cell transplantation (allo-HCT) between 2002 and 2018. The 3-year overall survival (OS) rates were 81.3% and 56.0%, respectively (p=0.15), and leukemia-free survival (LFS) rates were 76.2% and 42.0%, respectively (p=0.10) in patients with AML and MPAL. The relapse rates were 9.5% and 14.0% (p=0.93), and the non-relapse mortality (NRM) rates were 14.3% and 44.0%, respectively (p=0.10) in patients with AML and MPAL. One in 17 patients with AML, with pre-transplant tyrosine kinase inhibitors (TKI), and three in five patients with AML, without pre-transplant TKI, did not achieve complete remission (CR) before allo-HCT (p=0.024). Among the 20 patients with known disease status after allo-HCT, 95.0% were in hematological or molecular CR. None of the four patients who received post-transplant TKI for prophylaxis or measurable residual disease relapse experienced hematological relapse. In conclusion, our results suggest that pre-transplant TKI could improve disease status before allo-HCT. Moreover, allo-HCT resulted in high OS, high LFS, low relapse, and low NRM rates in patients with AML with BCR::ABL1.

The application of targeted RNA sequencing for the analysis of fusion genes, gene mutations, IKZF1 intragenic deletion, and CRLF2 overexpression in acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) is characterized by highly genetic heterogeneity, owing to recurrent fusion genes, gene mutations, intragenic deletion, and gene overexpression, which poses significant challenges in clinical detection. RNA sequencing (RNA-seq) is a powerful tool for detecting multiple genetic abnormalities, especially cryptic gene rearrangements, in a single test. Sixty samples (B-ALL, n = 49; T-ALL, n = 9; mixed phenotype acute leukemia (MPAL), n = 2) and 20 controls were analyzed by targeted RNA-seq panel of 507 genes developed by our lab. Of these, 16 patients were simultaneously analyzed for gene mutations at the DNA level using a next-generation sequencing panel of 51 genes. Fusion genes, CRLF2 expression, and IKZF1 intragenic deletion were also detected by reverse transcription-polymerase chain reaction (RT-PCR). Karyotype analysis was performed using the R-banding and G-banding technique on bone marrow cells after 24 hours of culture. Partial fusion genes were analyzed using fluorescence in situ hybridization (FISH). Compared with the results of Karyotype analysis, FISH, and RT-PCR, the detection rate of fusion genes by targeted RNA-seq increased from 48.3% to 58.3%, and six unexpected fusion genes were discovered, along with one rare isoform of IKZF1 intragenic deletion (IK10). The DNA sequencing analysis of 16 ALL patients revealed that 96.2% (25/26) of gene mutations identified at the DNA level were also detectable at the RNA level, except for one mutation with a low variant allele fraction. The detection of CRLF2 overexpression exhibited complete concordance between RT-PCR and RNA-seq. The utilization of RNA-seq enables the identification of clinically significant genetic abnormalities that may go undetected through conventional detection methods. Its robust analytical performance might bring great application value for clinical diagnosis, prognosis, and therapy in ALL.

13C Metabolic Flux Analysis in Chondrocytes Reveals a Novel Switch in Metabolic Phenotype.

Chondrocytes are typically known for their anaerobic metabolism both in vivo and under culture conditions in vitro. However, chondrocytes have been shown to display greater biosynthetic activity when subjected to conditions that elicit aerobic metabolism. We have previously shown that tissue formation by chondrocytes can be upregulated by controlling nutrient availability and that this response arises from changes in glucose metabolism. The aim of the present study was to further characterize these changes through 13C-metabolic flux analysis (13C-MFA), as well as to determine the most optimal response. Primary bovine chondrocytes were grown in scaffold-free high-density tissue culture. [U-13C] glucose labeling experiments were combined with a tissue-specific metabolic network model to carry out 13C-MFA under varying levels of nutrient availability. 13C-MFA results demonstrated that when subjected to increasing nutrient availability, chondrocytes switch from a predominately anaerobic to a mixed aerobic-anaerobic phenotype. This metabolic switch was attributed to the saturation of the lactate fermentation pathway and metabolite overflow toward the tricarboxylic acid cycle. This effect appears to be similar to, but the inverse of, the Crabtree effect ("inverse Crabtree effect"). The relationships between metabolic flux and nutrient availability were then utilized to identify culture conditions that promote enhanced tissue formation. This novel metabolic effect presents a simple but effective approach for enhancing the biosynthetic response of chondrocytes-a key requirement to develop functional engineered cartilaginous tissue for joint resurfacing.

Real-World Effectiveness of High-Dose Tafamidis on Neurologic Disease Progression in Mixed-Phenotype Variant Transthyretin Amyloid Cardiomyopathy.

Transthyretin amyloidosis (ATTR) is a progressive, heterogeneous rare disease manifesting as ATTR polyneuropathy (ATTR-PN), ATTR cardiomyopathy (ATTR-CM), or a mixed phenotype. Tafamidis meglumine (20mg po qd) is approved in some markets to delay neurologic progression in ATTR-PN, while high-dose tafamidis (80/61mg po qd) is approved worldwide to reduce cardiovascular mortality and cardiovascular-related hospitalization in ATTR-CM. The objective of this study was to assess the real-world benefit of high-dose tafamidis for delaying neurologic progression in patients with mixed-phenotype variant ATTR-CM (ATTRv-CM). This exploratory, retrospective, observational cohort study evaluated anonymized electronic medical records and included adult patients with mixed-phenotype ATTRv-CM treated with high-dose tafamidis for at least 6months. Neurologic assessments included the Medical Research Council (MRC) Scale for Muscle Strength, Neuropathy Impairment Score (NIS) muscle weakness subscale, and Polyneuropathy Disability (PND) instrument. Modified body mass index (mBMI) was also assessed. Patients (N = 10) started tafamidis treatment an average of 3.8months after diagnosis, with an average treatment duration of 20.8months. Seven of 10 patients demonstrated normal muscle strength on the MRC scale throughout the study, and 9 of 10 patients had no decline in muscle strength during the post-treatment period. The NIS muscle weakness subscale score was ≤ 60 for all patients in the study at all time points, suggesting normal function to mild impairment. Six of 10 patients had no change in walking capacity as measured by the PND instrument at pre- and post-assessments, while one-third of patients had a decrease in PND stage (signaling improvement) from pre- to post-assessment. mBMI remained relatively stable throughout the study. This is the first real-world study to demonstrate the potential value of high-dose tafamidis for delaying neurologic disease progression in patients with mixed-phenotype ATTRv-CM. The findings underscore the importance of multidisciplinary assessment for patients with ATTR amyloidosis. ClinicalTrials.gov: NCT05139680.