Background Mild cognitive impairment (MCI) is a heterogeneous condition with variable progression to Alzheimer's disease (AD). Identifying MCI individuals at high risk for progression typically requires cerebrospinal fluid (CSF) biomarkers, magnetic resonance imaging (MRI), which are costly and invasive. Objective This study aimed to develop a cost-effective approach using routinely collected clinical data to identify a subgroup of MCI individuals at high risk for AD progression. Methods Analyses were conducted using the UK Biobank dataset, focusing on 1019 participants identified as having MCI, using the ICD-10 code F06.7 (mild neurocognitive disorder due to known physiological condition) in the absence of a dedicated diagnostic code for MCI. Participants (mean age = 71.7 years; 44% women) were characterized using routinely recorded demographic, comorbidity, and lifestyle data. A mixed-data clustering model was applied to classify individuals into subgroups. Clinical relevance of each cluster was evaluated using Kaplan-Meier survival analysis of MCI-to-AD progression over an average follow-up of 4.5 years. Results Three subtypes were identified with distinct progression risks: high-risk (HR), medium-risk (MR), and low-risk (LR). The HR subtype had significantly higher prevalence of hypertension (98%), cardiovascular disease (89%), diabetes (48%), and high cholesterol (67%) than MR and LR (p < 0.05). The HR group was younger on average but had greater comorbidity burden and higher likelihood of AD progression. Conclusions This study demonstrates the feasibility of using routinely collected data to identify high-risk MCI individuals. This approach offers a practical preliminary screening tool to prioritize individuals for targeted interventions and further specialized assessments.

BackgroundAlmost half of the dementia cases are preventable. Semaglutide treats several medical conditions that are risk factors for dementia.ObjectiveWe aim to investigate if semaglutide is associated with a decreased risk of dementia.MethodsWe conducted emulation target trials based on a nationwide population-based database of patient electronic health records (EHRs) in the US among 1,710,995 eligible patients with type 2 diabetes (T2D) comparing semaglutide with other antidiabetic medications. First-time diagnosis of Alzheimer's disease-related dementia (ADRD) including vascular dementia, frontotemporal dementia, Lewy body dementia and other dementias were examined using Cox proportional hazards and Kaplan-Meier survival analyses during a 3-year follow-up. Models were adjusted by propensity-score matching.ResultsWe show that semaglutide was associated with a significantly reduced risk of overall ADRD incidence with a hazard ratio ranging from 0.54 (0.49-0.59) compared with insulin, 0.67 (0.61-0.74) compared with metformin, to 0.80 (0.72-0.89) compared with older generation glucagon-like peptide-1 agonists (GLP-1RAs). The association varied for specific dementia types, with significantly reduced risk of vascular dementia and no evidence of associations with frontotemporal and Lewy body dementias.ConclusionsThese findings provide evidence supporting protective effects of semaglutide on dementias in patients with T2D. Future works are needed to establish the causal relationships through randomized clinical trials and to characterize the underlying mechanisms.

ABSTRACT:RAS pathway (RASp) mutations induce proliferative features, and promote transformation in chronic myelomonocytic leukemia (CMML). However, the unique clonal landscape and hierarchy of distinct RASp mutations remain unexplored. To characterize the landscape, architecture, and implications of unique RASp mutations in CMML, we evaluated a cohort of 814 patients with CMML. We identified 461 RASp mutations among 342 patients (42%). N/KRAS and CBL mutations were the most common, frequently involved the P-loop or RING domains, respectively, and frequently appeared as dominant events (63% and 65%, respectively). BRAF, NF1, and PTPN11 mutations spanned throughout the gene structure, and frequently appeared as subclonal events (75%, 64%, and 59%, respectively). CBL mutations frequently occurred in codominance with SRSF2 and multihit TET2, and were enriched for KIT mutations. PTPN11 mutations more frequently co-occurred with SETBP1 and DNMT3A mutations, and were infrequently codominant with TET2 or ASXL1. RASp mutations predicted for shorter overall survival (hazard ratio [HR], 1.55; 95% confidence interval [CI], 1.15-2.07; P = .0075) and leukemia-free survival (LFS; HR, 1.67; 95% CI, 1.26-2.20; P = .0011), influenced outcomes of myelodysplastic and TET2 mutant CMML, and cooperated with IDH2 and RUNX1 mutations to induce shorter LFS. These data set the bases for refined genomic classifications of CMML, and underscore the need to develop RAS-directed therapies for patients with CMML.

Fragile X Syndrome (FXS) is a rare neurodevelopmental disorder caused by a trinucleotide repeat expansion on the 5’ untranslated region of the FMR1 gene. FXS is characterized by intellectual disability, anxiety, sensory hypersensitivity, and difficulties with executive function. A recent phase 2 placebo-controlled clinical trial assessing BPN14770, a first-in-class phosphodiesterase 4D allosteric inhibitor, in 30 adult males (age 18-41 years) with FXS demonstrated cognitive improvements on the NIH Toolbox Cognitive Battery in domains related to language and caregiver reports of improvement in both daily functioning and language. However, individual physiological measures from electroencephalography (EEG) demonstrated only marginal significance for trial efficacy. A secondary analysis of resting state EEG data collected as part of the phase 2 clinical trial evaluating BPN14770 was conducted using a machine learning classification algorithm to classify trial conditions (i.e., baseline, drug, placebo) via linear EEG variable combinations. The algorithm identified a composite of peak alpha frequencies (PAF) across multiple brain regions as a potential biomarker demonstrating BPN14770 efficacy. Increased PAF from baseline was associated with drug but not placebo. Given the relationship between PAF and cognitive function among typically developed adults and those with intellectual disability, as well as previously reported reductions in alpha frequency and power in FXS, PAF represents a potential physiological measure of BPN14770 efficacy.

Introduction: Somatic mutations in RAS pathway genes are present in ~30% of patients with chronic myelomonocytic leukemia (CMML) and are usually associated with myeloproliferative features. Whether mutations in distinct RAS pathway genes share similar genomic context and have similar implications remains unknown. Such an understanding might inform future studies evaluating genotype-specific therapies (RAS-codon specific and pan-RAS inhibitors) or cooperative leukemogenic mechanisms. This multicenter study examines the landscape of RAS pathway mutations (RASMT) in CMML. Methods: Newly diagnosed CMML patients evaluated at MD Anderson Cancer Center (n=449, 54%) and Mayo Clinic (n=383, n=46%) were included after IRB approval. Clonal relationships were tested using Pearson goodness-of-fit. Clones with the highest variant allele frequency (VAF), or with VAF close to 40% were defined as dominant, and those present at VAF <20% in the presence of another dominant clone were defined as subclonal. RASMT genes included NRAS, KRAS, BRAF, CBL, CBLC, PTPN11 and NF1. Standard statistical measures were applied. Results: Among 832 CMML patients (median age 71 years, 68% male) 387 (47%) had myeloproliferative (MP) and 445 (53%) had myelodysplastic (MD) CMML. A total of 350 (42%) patients had RASMT with higher prevalence among MP-CMML compared to MD-CMML (55% vs 31%, p<0.001). Among patients with RASMT, 262 (32%), 59 (7%), 20 (2%), 6 (<1%) and 3 (<1%) had 1, 2, 3, 4 or 5 distinct RASMT, respectively. Importantly, 17 patients harbored a RASmt as the only detectable somatic mutation, with KRASMT being the most frequent (n=10, 59%), including 3 (18%) patients with >1 RASMT. Among RASMT patients, a trend for higher hemoglobin (12.7 vs 10.8 g/dL, p=0.054) was observed in NF1MT CMML and lower WBC (14.4 vs 27.9 x109/L, p=0.067) values in CBLMT CMML. In total, 479 somatic mutations in RAS pathway genes were identified the most frequent involving NRAS (n=169, 35%) followed by CBL (n=120, 25%), KRAS (n=102, 21%), PTPN11 (n=38, 8%), NF1 (n=34, 7%), BRAF (n=11, 2%) and CBLC (n=5, 1%). Most frequently involved codons for each RAS pathway gene were as follows: G594 (n=3) for BRAF, R420 (n=22) and C404 (n=15) for CBL, G12 (n=41; G12D: 13, G12C: 3) and A146 (n=16) for KRAS, G12 (n=116; G12D: 74, G12C: 5) for NRAS and A72 (n=8) for PTPN11. CBLMT CMML had higher ASXL1 (58% vs 44%, p=0.011), BRAF (8% vs 4%, p=0.017), IDH2 (10% vs 4%, p=0.024), KIT (5% vs 1%, p=0.038), SRSF2 (54% vs 40%, p=0.004), TET2 (59% vs 47%, p=0.019) and ZRSR2 (10% vs 4%, p=0.014) co-mutation frequency. Of note, DNMT3A mutations were mutually exclusive of CBLMT CMML (0% vs 7%, p=0.007), and multihit TET2 was more common among TET2MT/CBLMT vs TET2MT/CBLWT CMML (82% vs 67%, p =0.039). CMML with NF1 mutations had higher BCORL1 (8% vs 0%, p=0.005), PTPN11 (15% vs 3%, p=0.014) and SETBP1 (23% vs 8%, p=0.022) mutational cooccurrences. NRASMT CMML was enriched for ASXL1 (53% vs 43%, p=0.042), ETNK1 (5% vs 1%, p=0.004), GATA2 (4% vs 1%, p=0.034) and STAG2 (4% vs 1 %, p=0.023) mutations but had lower JAK2 (1% vs 6%, p=0.036) and TP53 (1% vs 5%, p=0.022) mutational frequency. PTPN11 mutations were associated with higher NF1 (29% vs 7%, p=0.014), SETBP1 (27% vs 9%, p=0.01) and WT1 (6% vs <1%, p=0.014) mutational cooccurrences and lower SRSF2 (23% vs 42%, p=0.05) frequency. We evaluated RAS mutant clonal dominance in 253 (72%) RASMT patients. NRAS (63% vs 38%), KRAS (63% vs 38%) and CBL (65% vs 35%) mutations were more frequently observed as dominant clones while PTPN11 (44% vs 56%), BRAF (25% vs 75%) and NF1 (35% vs 65%) mutations were more frequently subclonal events, particularly of TET2, ASXL1 or SRSF2 mutant clones. Presence of a RAS pathway mutation was associated with shorter LFS (36 vs 53 months, HR 1.5, 95% CI 1.1-2.2, p=0.020), with presence of multiple RAS mutations shortening LFS further (53, 43, 27 months for 0, 1 and >1, p=0.017). PTPN11 mutations predicted for shorter OS (18 vs 39 months, HR 1.7, 95% CI 1.0-2.8, p=0.035) and LFS (14 vs 36 months, HR 1.9, 95% CI 1.2-2.9, p=0.008). Presence of >1 RAS pathway mutation was associated with shorter OS in ASXL1MT/TET2MT CMML compared to those with no RAS mutations (24 vs 53 months, HR 4.2, 95% CI 1.6-10.8, p=0.004). Conclusions: RAS mutations exhibit distinct genomic features and clonal hierarchy in CMML and cooperate with other somatic mutations to drive clinical outcomes. This study forms the foundation for assessment of novel RAS pathway inhibitors in CMML.

This case-control study examines the incidence and risks of myeloid neoplasms in adults treated for B-cell lymphoproliferative disorders or multiple myeloma.

The current study was inspired by observations from exploratory analyses of an institutional cohort with chronic myelomonocytic leukemia (CMML; N = 398) that revealed no instances of blast transformation in the seven patients with plant homeodomain finger protein 6 (PHF6) mutation (PHF6MUT). A subsequent Mayo Clinic enterprise-wide database search identified 28 more cases with PHF6MUT. Compared with their wild-type PHF6 counterparts (PHF6WT; N = 391), PHF6MUT cases (N = 35) were more likely to co-express TET2 (89% vs. 45%; p < .01), RUNX1 (29% vs. 14%; p = .03), CBL (14% vs. 2%; p < .01), and U2AF1 (17% vs. 6%; p = .04) and less likely SRSF2 (23% vs. 45%; p < .01) mutation. They were also more likely to display loss of Y chromosome (LoY; 21% vs. 2%; p < .01) and platelets <100 × 109/L (83% vs. 51%; p < .01). Multivariable analysis identified PHF6MUT (HR 0.28, 95% CI 0.15-0.50) and DNMT3AMUT (HR 5.8, 95% CI 3.3-10.5) as the strongest molecular predictors of overall survival. The same was true for blast transformation-free survival with corresponding HR (95% CI) of 0.08 (0.01-0.6) and 9.5 (3.8-23.5). At median 20 months follow-up, blast transformation was documented in none of the 33 patients with PHF6MUT/DNMT3AWT but in 6 (32%) of 19 with DNMT3AMUT and 74 (20%) of 374 with PHF6WT/DNMT3AWT (p < .01). The specific molecular signatures sustained their significant predictive performance in the context of the CMML-specific molecular prognostic model (CPSS-mol). PHF6MUT identifies a unique subset of patients with CMML characterized by thrombocytopenia, higher prevalence of LoY, and superior prognosis.

Peritoneal tissue-resident macrophages have broad functions in the maintenance of homeostasis and are involved in pathologies within local and neighboring tissues. Their functions are dictated by microenvironmental cues; thus, it is essential to investigate their behavior in an in vivo physiological niche. Currently, specific peritoneal macrophage-targeting methodologies employ whole-mouse transgenic models. Here, a protocol for effective in vivo modulation of mRNA and small RNA species (e.g., microRNA) expression in peritoneal macrophages using lentivirus particles is described. Lentivirus preparations were made in HEK293T cells and purified on a single sucrose layer. In vivo validation of lentivirus effectivity following intraperitoneal injection revealed predominant infection of macrophages restricted to local tissue. Targeting of peritoneal macrophages was successful during homeostasis and thioglycolate-induced peritonitis. The limitations of the protocol, including low-level inflammation induced by intraperitoneal delivery of lentivirus and time restrictions for potential experiments, are discussed. Overall, this study presents a quick and accessible protocol for the rapid assessment of gene function in peritoneal macrophages in vivo.

INTRODUCTION:Emerging preclinical evidence suggests that semaglutide, a glucagon-like peptide receptor agonist (GLP-1RA) for type 2 diabetes mellitus (T2DM) and obesity, protects against neurodegeneration and neuroinflammation. However, real-world evidence for its ability to protect against Alzheimer's disease (AD) is lacking. METHODS:We conducted emulation target trials based on a nationwide database of electronic health records (EHRs) of 116 million US patients. Seven target trials were emulated among 1,094,761 eligible patients with T2DM who had no prior AD diagnosis by comparing semaglutide with seven other antidiabetic medications. First-ever diagnosis of AD occurred within a 3-year follow-up period and was examined using Cox proportional hazards and Kaplan-Meier survival analyses. RESULTS:Semaglutide was associated with significantly reduced risk for first-time AD diagnosis, most strongly compared with insulin (hazard ratio [HR], 0.33 [95% CI: 0.21 to 0.51]) and most weakly compared with other GLP-1RAs (HR, 0.59 [95% CI: 0.37 to 0.95]). Similar results were seen across obesity status, gender, and age groups. DISCUSSION:These findings support further studies to assess semaglutide's potential in preventing AD. HIGHLIGHTS:Semaglutide was associated with 40% to 70% reduced risks of first-time AD diagnosis in T2DM patients compared to other antidiabetic medications, including other GLP-1RAs. Semaglutide was associated with significantly lower AD-related medication prescriptions. Similar reductions were seen across obesity status, gender, and age groups. Our findings provide real-world evidence supporting the potential clinical benefits of semaglutide in mitigating AD initiation and development in patients with T2DM. These findings support further clinical trials to assess semaglutide's potential in delaying or preventing AD.

Clonal cytopenia of undetermined significance (CCUS) is defined by a myeloid driver mutation in the context of otherwise unexplained cytopenia. CCUS has an inherent risk of progressing to myeloid neoplasm. However, it is unknown how exposure to previous cytotoxic therapy may impact the risk of progression and survival. We stratified patients with CCUS by prior exposure to DNA-damaging therapy. Of 151 patients, 46 (30%) had received cytotoxic therapy and were classified as therapy-related CCUS (t-CCUS), whereas 105 (70%) had de novo CCUS. A lower proportion of t-CCUS had hypercellular marrows (17.8% vs 44.8%, P = .002) but had higher median bone marrow blast percentages. After a median follow-up of 2.2 years, t-CCUS had significantly shorter progression-free survival (PFS, 1.8 vs 6.3 years; hazard ratio [HR], 2.1; P = .007) and median overall survival (OS; 3.6 years vs not reached; HR, 2.3; P = .007) compared with CCUS. Univariable and multivariable time-to-event analyses showed that exposure to cytotoxic therapy independently accounted for inferior PFS and OS. Despite the similarities in clinical presentation between CCUS and t-CCUS, we show that exposure to prior cytotoxic therapies was an independent risk factor for inferior outcomes. This suggests that t-CCUS represents a unique clinical entity that needs more stringent monitoring or earlier intervention strategies.

Introduction Somatic mutations in TET2 are present in ~60% of chronic myelomonocytic leukemia (CMML) patients and are associated with improved median overall (mOS) and acute leukemia free survival (mLFS). Multiple TET2 mutations are detected in ~50% of such cases and are thought to confer additional benefit. Whether these benefits apply to the molecularly distinct myelodysplastic (MD) and myeloproliferative (MP) CMML subtypes is unknown. This study assesses the impact of the number and type of TET2 mutations across the MD and MP-CMML subtypes. Methods CMML patients seen at Mayo Clinic (n = 400, 47%) and MD Anderson Cancer Center (n = 449, 53%) were included with IRB approval. Analyses considered the clinical and molecular data at diagnosis or first referral. Truncating (frameshift or nonsense) mutations in TET2 were further annotated by location in either the proximal genome targeting domain (aa 1 - 1128) or distal catalytic domain (aa 1129 - 1936) as described (Coltro, 2020). Categorical variables were compared by Pearson Chi squared and continuous by Mann-Whitney U or Kruskal-Wallis tests. Univariate and multivariate analyses utilized Cox proportional hazards models with continuous variables binarized by receiver operator curves. Kaplan-Meier analyses were censored at hematopoietic cell transplantation (HCT). P < 0.05 was considered significant. Calculations used BlueSky (v10.3.1) or Prism (v10.2.3). Results In a cohort of 849 CMML patients (median age 71 years, 32% female, 90% white), 454 (54%) were classified as MD-CMML and 394 (46%) as MP-CMML. Cumulatively, 664 somatic TET2 mutations were identified across the MD (n = 415, 63%) and MP-CMML (n = 249, 38%) subgroups, with ≥1 TET2 mutation in 57% vs 44% (p < 0.001) and ≥2 TET2mutations in 33% vs 20% (p = 0.001) , respectively. The median (range) number of TET2 mutations in MD vs MP-CMML was 1 (0 - 4) vs 0 (0 - 3; p < 0.001). Truncating frameshift or nonsense mutations comprised 86% vs 78% of mutations in each group with hypomorphic missense mutations comprising 13% vs 20% (p = 0.03). Proximal (48% vs 43%) and distal (38% vs 35%) truncations were evenly distributed between MD and MP-CMML (p = 0.91). In both the MD and MP subgroups as well as the overall cohort (p < 0.01 each), increasing number of TET2 mutations (0 vs 1 vs ≥2) was associated with higher median hemoglobin (10.3 vs 11.4 vs 12.0 g/dL), fewer mean peripheral blasts (1.3% vs 0.5% vs 0.3%), and a higher proportion of normal karyotypes (60% vs 76% vs 79%). Likewise, co-mutations were increasingly frequent in SRSF2 (36% vs 45% vs 53%) and less frequent in IDH2 (9% vs 1% vs 1%) and SETBP1 (14% vs 4% vs 0%) throughout the cohort. TET2 mutations were mutually exclusive with ETV6, GATA2, IDH1, and WT1 mutations. Uniquely, MD-CMML cases with TET2 mutations (0 vs 1 vs ≥2) were enriched for co-mutations in ZRSR2 (5% vs 13% vs 9%) and had fewer co-mutations in both ASXL1 (47% vs 34% vs 28%) and U2AF1 (11% vs 6% vs 3%). In MP-CMML, co-mutations in CBL (11% vs 20% vs 27%) were increased while co-mutations in BCOR (6% vs 1% vs 0%) were decreased. Accordingly, the favorable ASXL1WT/TET2MT genotype was more common in MD (39% vs 22%), while the adverse ASXL1MT/TET2WT genotype was more common in MP-CMML (20% vs 33%; p < 0.001). In multivariate analyses, the presence of ≥2 TET2 mutations was favorably associated with OS and LFS in both MD and MP-CMML (hazard ratios 0.53 - 0.67, p < 0.04) independent of several clinical and genetic features including ASXL1 mutations and abnormal karyotype. Beyond the number of TET2 mutations, however, neither the mutation class (truncating vs hypomorphic) nor location (proximal vs distal) further affected OS or LFS. Likewise, although the median variant allele fraction (VAF) was lower in MD vs MP-CMML (42% vs 46%, p < 0.001), VAF did not impact survival. Accordingly, the mOS for patients with 0, 1, or ≥2 TET2 mutations was 29, 58, and 75 months in MD-CMML and 19, 26, and 39 months in MP-CMML, respectively (p < 0.001 each). The mLFS was similarly improved to 26, 52, and 67 months in MD-CMML (p < 0.001) and 16, 22, and 31 months in MP-CMML (p = 0.003). CONCLUSIONS This multicenter study demonstrates that the favorable impact of multiple TET2 mutations in CMML extends to both the MD and MP subtypes. In MD-CMML, multiple TET2 mutations segregate against adverse ASXL1 mutations while, in MP-CMML, they appear to offset the impact of adverse RAS pathway mutations. Thus, multiple TET2 mutations may identify a lower-risk subgroup of MP-CMML patients.

Fragile X syndrome (FXS) is a rare neurodevelopmental disorder caused by a CGG repeat expansion ≥ 200 repeats in 5' untranslated region of the FMR1 gene, leading to intellectual disability and cognitive difficulties, including in the domain of communication. A recent phase 2a clinical trial testing BPN14770, a phosphodiesterase 4D inhibitor, showed improved cognition in 30 adult males with FXS on drug relative to placebo. The initial study found significant improvements in clinical measures assessing cognition, language, and daily functioning in addition to marginal improvements in electroencephalography (EEG) results for the amplitude of the N1 event-related potential (ERP) component. EEG results suggest BPN14770 improved neural hyperexcitability in FXS. The current study investigated the relationship between BPN14770 pharmacokinetics (PK) and the amplitude of the N1 ERP component from the initial data. Consistent with the original group-level finding in period 1 of the study, participants who received BPN14770 in the period 1 showed a significant correlation between N1 amplitude and serum concentration of BPN14770. These findings strengthen the validity of the original result, indicating that BPN14770 improves cognitive performance by modulating neural hyperexcitability. This study represents the first report of significant correlation between a reliably abnormal EEG marker and serum concentration of a novel pharmaceutical in FXS.

The BCL6-corepressor (BCOR) is a tumor-suppressor gene located on the short arm of chromosome X. Data are limited regarding factors predicting survival in BCOR-mutated (mBCOR) acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). We evaluated 138 patients with mBCOR myeloid disorders, of which 36 (26.1%) had AML and 63 (45.6%) had MDS. Sixty-six (47.8%) patients had a normal karyotype while 18 (13%) patients had complex karyotype. BCOR-mutated MDS/AML were highly associated with RUNX1 and U2AF1 co-mutations. In contrast, TP53 mutation was infrequently seen with mBCOR MDS. Patients with an isolated BCOR mutation had similar survival compared to those with high-risk co-mutations by European LeukemiaNet (ELN) 2022 criteria (median OS 1.16 vs. 1.27 years, P=0.46). Complex karyotype adversely impacted survival among mBCOR AML/MDS (HR 4.12, P<0.001), while allogeneic stem cell transplant (alloSCT) improved survival (HR 0.38, P=0.04). However, RUNX1 co-mutation was associated with an increased risk of post-alloSCT relapse (HR 88.0, P=0.02), whereas melphalan-based conditioning was associated with a decreased relapse risk (HR 0.02, P=0.01). We conclude that mBCOR is a high-risk feature across MDS/AML, and that alloSCT improves survival in this population.

Recent interest in the biology and function of peritoneal tissue resident macrophages (pMΦ) has led to a better understanding of their cellular origin, programming and renewal. The programming of pMΦ is dependent on microenvironmental cues and tissue specific transcription factors, including GATA6. However, the contribution of microRNAs remains poorly defined. We conducted a detailed analysis of the impact of GATA6-deficiency on microRNA expression in mouse pMΦ. Our data suggest that for many of the pMΦ, microRNA composition may be established during tissue specialization, and that the effect of GATA6 knockout is largely unable to be rescued in the adult by exogenous GATA6. The data are consistent with GATA6 modulating the expression pattern of specific microRNAs, directly or indirectly, and including miR-146a, -223, and -203 established by the lineage-determining transcription factor PU.1, to achieve a differentiated pMΦ phenotype. Lastly, we showed a significant dysregulation of miR-708 in pMΦ in the absence of GATA6 during homeostasis and in response to LPS/IFN-γ stimulation. Overexpression of miR-708 in mouse pMΦ in vivo altered 167 mRNA species demonstrating functional downregulation of predicted targets, including cell immune responses and cell cycle regulation. In conclusion, we demonstrate dependence of the microRNA transcriptome on tissue-specific programming of tissue macrophages as exemplified by the role of GATA6 in pMΦ specialization.

7038 Background: While chimeric antigen receptor T-cell (CAR-T) therapy has transformed treatment for hematological malignancies, severe (≥ grade 3,Gr 3, CTCAE) cytopenias post CAR-T and therapy related myeloid neoplasms (t-MN) are particularly difficult to manage. Survival after diagnosis of t-MN is dismal. Management of ≥Gr 3 cytopenias that do not recover by month 3 after CAR-T require frequent monitoring and transfusion support. Growth factor and thrombopoietin mimetics have variable success, with stem cell boost emerging as having more consistent success. Evaluation for CAR-T eligibility, prior to leukapheresis, is the critical timepoint where factors identified at this time could inform risks and benefits of CAR-T versus alternative treatment and for consideration of stem cell collection in patients who may benefit from stem cell boost to treat severe cytopenia post CAR-T. Methods: We conducted a retrospective analysis comparing clinical data of patients (pts) with lymphoma (NHL) and multiple myeloma (MM) treated with FDA-approved and investigational CAR-T between 01/2016 – 06/2022 at Mayo Clinic. Prolonged cytopenias were defined as hemoglobin (Hg)<8 g/dL, absolute neutrophil count (ANC)<0.5 x 109/L, and/or platelet count (PLT)<50 x 109/L at 3 months after CAR-T. Pts with progressive disease were excluded from cytopenia analysis. Logistic regression to identify factors correlated with cytopenia and t-MN. Results: Among the 186 pts who received CAR-T, 42 (22%) patients developed severe cytopenia (26/102, 25%, in NHL; 16/84, 19% in MM); 15/186 (8%) had more than 1 concurrent cytopenia. Univariate analysis identified that baseline ferritin and CAR-HEMATOTOX score correlated with cytopenia. Multivariate analysis showed that advanced age (≥ 65 years) (HR 2.69, 95%CI 1.17 – 6.40, P = 0.02) and thrombocytopenia (HR 4.01, 95%CI 1.77 – 9.34, P = 0.001) were associated with severe cytopenia. Twenty (10.7%) patients [8 (40%) males] developed t-MN at a median of 9.5 months (IQR 4.8 – 19.3 months) after CAR-T (14/102, 13% in NHL; 6/84, 7% in MM). Univariate analysis identified ferritin and CAR-HEMATOTOX score correlated with t-NM. Multivariate logistic regression showed advanced age (HR 5.03; 95%CI 1.59-18.7, P= 0.009), hemoglobin ≤ 10 g/dl (HR 3.63, 95%CI 1.04-13.30, P = 0.04), and thrombocytopenia (HR 4.06, 95%CI 1.32-14.2, P 0.02) are significantly associated with development of post-CART t-MN. Conclusions: Advanced age, anemia and thrombocytopenia at the time of evaluation for CAR-T eligibility, prior to leukapheresis, are associated with the development of post-treatment severe cytopenia and t-MN. Larger, multi-center studies are needed to validate these findings.

Background: Previously published risk variables for chronic myelomonocytic leukemia (CMML) included age, gender, CMML subtype, red blood cell (RBC) transfusion status, hemoglobin, leukocyte, monocyte, lymphocyte and platelet counts, immature myeloid cells, peripheral blood (PB) or bone marrow blast percentage, and mutations including ASXL1, RUNX1, NRAS, and SETBP1. Recent studies have reported on additional prognostically favorable (TET2) or unfavorable (DNMT3A, ASXL1, NRAS, SETBP1) mutations. In the current study, we took advantage of a clinically well-characterized and molecularly annotated cohort of CMML patients, in order to revisit the prognostic interaction between the aforementioned risk factors, with emphasis on mutations. Methods: The current study was conducted under an institutional review board approved protocol that allowed retrospective collection and analysis of data from Mayo Clinic patient records. Mutations were screened by multi-gene next-generation sequencing (NGS). Diagnostic criteria were according to the International Consensus Classification (Blood 2022; 140:1200). Conventional statistical methods were employed (JMP Pro 17.0.0 software SAS Institute, Cary, NC, USA). Survival analysis was censored for allogeneic stem cell transplantation (ASCT). Receiver Operating Characteristic (ROC) plots and Akaike Information Criterion (AIC) were used to compare predictive performance of the new model (MMMv2) vs. the Mayo Molecular Model (MMM) vs. the CMML-specific prognostic scoring system (CPSS). Results: The study cohort included 416 patients with CMML that were fully annotated for mutations. Cytogenetic information was available in 404 patients. At a median follow-up of 18 months, 302 (72%) deaths, 72 (18%) leukemic transformations, and 44 (11%) ASCTs were documented. Age-adjusted multivariable analysis (MVA) of mutations identified the following as risk factors: DNMT3A (HR 5.5; p<0.01), PHF6 (HR 0.3; p<0.01), TET2 (HR 0.6; p<0.01), PTPN11 (HR 2.5; p=0.01), NRAS (HR 1.6; p<0.01), ASXL1 (HR 1.3; p=0.02), and SETBP1 (HR 1.8; p=0.01). A separate MVA of clinical and cytogenetic variables identified RBC transfusion need, leukocyte count ≥13 x 109/L, peripheral blood blast ≥1%, abnormal karyotype, older age, and male sex to be independently associated with inferior survival (p<0.04 in all instances). All-inclusive MVA identified the following as independent risk factors for survival (p <0.01): DNMT3A mutation (HR 3.7; 95% CI 2.0-6.7), absence of PHF6 mutation (HR 2.8; 1.3-5.8), RBC transfusion need (HR 2.3; 1.7-3.1), leukocyte count ≥13 x 109/L (HR 2.0; 1.5-2.6), abnormal karyotype (HR 1.6; 1.2-2.2), and absence of TET2 mutation (HR 1.4,1.1-1.8). An HR based prognostic model was subsequently constructed by assigning 3 points to DNMT3A mutation, 2 points each for RBC transfusion need, leukocyte count ≥13 x 109/L, and absence of PHF6 mutation and one point each for abnormal karyotype and absence of TET2 mutation, resulting in a 4-tiered survival risk model (MMMv2): low risk (0-2 points; N=103) with median survival of 74 months; intermediate-1 (3 points; N=65) with median 37 months; intermediate-2 (4-5 points; N=126) with median 19 months; and high (6 points; N=88) with median 12 months (p<0.01). Independent genetic risk factors associated with inferior leukemia-free survival included DNMT3A (HR 5.5), PHF6 (HR 0.2), abnormal karyotype (HR 2.0), PTPN11 (HR 5.2), ASXL1 (HR 2.0), and BCOR (HR 8.7). Leukocyte count ≥13 x 109/L (p=0.01) but not RBC transfusion need (p=0.2) was additionally significant. However, presence of immature myeloid cells (p<0.01) and circulating blasts ≥1% (p<0.01) were prognostically more relevant than leukocyte count ≥13 x 109/L and independently predicted leukemic transformation, along with DNMT3A, PTPN11 and ASXL1 mutations. Predictive performance was superior for MMMv2 (AUC 87, AIC 137) vs. MMM (AUC 74, AIC 194) vs. CPSS (AUC 76, AIC 177). Conclusion: The revised Mayo Molecular Model (MMMv2) provides a contemporary risk model for CMML that recognizes DNMT3A as the most unfavorable and PHF6 as the most favorable mutation and confirms the mutation-independent prognostic contribution from red cell transfusion need and sustained leukocytosis (≥13 x 109/L). Mutations were also important in predicting leukemic transformation, notably DNMT3A, PTPN11 and ASXL1.

Background: Chimeric antigen receptor T-cell (CAR-T) therapy has shown substantial promise in treatment of hematological malignancies, including non-Hodgkin Lymphoma (NHL). A subset of patients develop prolonged cytopenia, comprising quality of life, increasing healthcare utilization, and imparting considerable morbidity and mortality. Therapy-related myeloid neoplasms (t-MN), typically seen in patients who receive DNA-damaging agents such as radiotherapy and alkylators, have also been observed in CAR-T recipients. CAR-Hematotox score predicts the development of hematotoxicity in CAR-T recipients using laboratory parameters obtained at lymphodepleting (LD) chemotherapy, a timepoint when the patient has already gone through leukapheresis thus committed to the CAR-T process. Therefore, it is critical to identify patients at the highest risk of prolonged cytopenia and/or t-MN prior to apheresis, making it possible for a better triage of those who will proceed with CAR-T and those who might benefit from other strategies. Methods: A retrospective analysis was conducted in NHL patients who received commercial CAR-T products between January 2018 and March 2024 at Mayo Clinic Rochester, Arizona and Florida. Clinical and laboratory data including age, hemoglobin (Hb), absolute neutrophil count (ANC), platelet (PLT), ferritin, and LDH, obtained at the initial CAR-T evaluation, typically within 1 week of leukapheresis, were used for analysis. Prior lines of therapies including autologous stem cell transplant (ASCT) were collected. Prolonged cytopenias were defined as Hb < 8 g/dL, ANC < 0.5 x 109/L, and/or PLT < 50 x 109/L at 3 months after CAR-T. t-MN was diagnosed per WHO 2016. For prolonged cytopenia, the optimal cutoff points for continuous variables were determined by maximizing the sum of sensitivity and specificity. Logistic regression was then used to determine the factors associated with cytopenia at 3 months post-CAR-T. For t-MN, maximally selected log-rank statistics was used to determine the optimal cutoffs. We used competing risk analysis to determine the factors associated with development of t-MN. Death without t-MN was considered a competing risk. For both analyses, the factors with P < 0.1 in univariate analysis (UVA) were included in multivariate analysis (MVA). Results: Among the 387 NHL patients, with a median follow-up of 11 months (95% CI 8.7 - 12.2 months), 56 (14.4%) developed (41, 10.5% with 1 cytopenia; 14, 3.6% with 2 and 1, 0.2% with cytopenia in all 3 lineages), 20 (5.2%) developed and 315 (81.2%) did not have either. Factors associated with prolonged cytopenia: were age >, PLT ≤ 150, ferritin ≥ 600, and LDH ≥ 250. Univariate analysis identified baseline Hb, ANC, PLT and ferritin to be significantly associated with prolonged cytopenia. MVA identified Hb ≤ 11 (HR = 1.98; 95% CI 1.03-3.85, P = 0.042) and PLT ≤ (HR = 2.56; 95% CI 1.39-4.78; P = 0.003) to be associated with the development of prolonged cytopenia. Factors associated with t-MN:were age > 65 years, Hb ≤ 9 g/dl, ANC < 3.8, PLT ≤ 80, Univariate analysis identified age, Hb, ANC, PLT, ferritin, and ≥ 3 prior lines of treatment to be significantly associated with t-MN. On MVA, age ≥ 65 (HR = 3.91; 95% CI 1.52-10.03; P = 0.004), Hb ≤ 9 (HR = 3.49; 95% CI 1.38-8.85; P = 0.008), ferritin ≥ 300 (HR = 3.06 ; 95% CI 1.08-8.67; P = 0.035) and ≥ 3 prior lines of therapy (HR = 3.25; 95% CI 1.33-7.94 ; P = 0.010) were associated with t-MN development. Association of prolonged cytopenia with t-MN development: Prolonged cytopenia was noted in 45% of patients who developed t-MN and 15% of those who did not (P = 0.002). Cumulative incidence of t-MN at 1 year was 3.3% and the presence of prolonged cytopenia was associated with a higher 1of t-MN (HR 8.93, 95% CI 2.15 - 37.1, P = 0.003). Conclusions: Readily accessible clinical and lab data such as blood count and inflammatory markers at the time of CAR-T evaluation were associated with the development of prolonged cytopenias. Similarly, advanced age, anemia, high ferritin levels and history of ≥3 lines of therapy were associated with the development of t-MN. These findings, when validated in larger dataset, can be used for patient counseling and therapy selection by identifying patients at high-risk of developing hematotoxicity and/or t-MN development.