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Background Abnormal karyotype is not infrequent in polycythemia vera (PV) and has been associated with shortened survival (BJH 2018;182:437). In the current study, we utilized a large institutional database, in order to describe the spectrum and prevalence of specific cytogenetic abnormalities in PV, during different stages of the disease, and examine their clinical, molecular, and prognostic correlates, in the context of previously established risk factors. Methods After institutional review board approval, we identified 669 cytogenetically-annotated patients with PV, evaluated at the Mayo Clinic between 1973 and 2023, and met the International Consensus Classification diagnostic criteria (Arber et al. Blood 2022;140;1200). Cytogenetic reporting was in accordance with the International System for Human Cytogenomic Nomenclature. Results i) Prevalence of cytogenetic abnormalities at diagnosis and at different stages of the disease 436 of 669 (65%) patients with PV underwent cytogenetic evaluation within one year of diagnosis, 116 (17%) within 1-10 years, 47 (7%) after 10 years, and 70 (10%) at time of fibrotic or leukemic transformation. In patients with cytogenetic information obtained within 1 year of diagnosis, karyotype was normal in 369 (85%), showed loss of Y chromosome (-Y) in 15 (3%), and other abnormalities in 52 (12%). Most common abnormalities included trisomy 9 (n=11; 3%), del(20q) (n=10; 2%), trisomy 8 (n=4; 1%), other sole abnormalities (n=16; 4%), and ≥ 2 abnormalities (n=11; 3%) including complex karyotype (n=4; 0.9%). Frequency of abnormal karyotype was higher with longer disease duration; 15% within a year of diagnosis, 28% in 1-10 years, 49% over 10 years after diagnosis, and 50% at time of fibrotic or leukemic transformation (p<0.01). Sole del (20q), single abnormalities excluding 20q-, +8, +9, or -Y, and ≥ 2 abnormalities including complex karyotype were progressively acquired; respective frequencies within a year of diagnosis, over 10 years after diagnosis, and at time of fibrotic or leukemic transformation were 2%/21%/10% (del (20q), 4%/17%/14% (single abnormality excluding 20q-, +8, +9, or -Y), 3%/11%/19% (≥ 2 abnormalities), 1%/ 2%,/10% (complex karyotype) (all p-values <0.01). In addition, sole +8 displayed highest frequency at fibrotic transformation compared to within a year of diagnosis (5% vs 0.9%; p=0.04). ii) Phenotypic and Molecular Correlates 436 patients with PV (51% males; median age; 64 years) who underwent cytogenetic testing within one year of diagnosis were examined for clinical and molecular correlations. Abnormal karyotype compared to normal karyotype, correlated with older age (median: 70 vs 63 years, p<0.01), male gender (64% vs 48%, p=0.02) and lower platelet count (370 vs 465 × 109/L, p<0.01). Sole del (20q) compared to normal karyotype was associated with older age (median: 76 vs 63 years, p<0.01); sole +8 exclusively occurred in patients ≥ 60 years (p=0.03); sole +9 was associated with venous thrombosis history (45% vs 17%, p=0.03). TET2 and SRSF2 mutations were found to cluster with normal karyotype (18% vs 4%; p=0.04 and 5% vs 0%; p=0.14 in normal vs abnormal karyotype, respectively). iii) Prognostic Impact of Cytogenetic Abnormalities At a median follow-up of 7.4 years (range; 0.04-42.4 years), a total of 163 (37%) patients have died, 50 (11%) and 14 (3%) underwent fibrotic and leukemic transformation, respectively. In univariate analysis for overall survival (OS), abnormal karyotype compared to normal karyotype, was associated with inferior survival (median OS; 10.5 vs 16.3 years, p<0.01). In multivariate analysis, the adverse impact of abnormal karyotype remained significant (p=0.02; HR 2.0, 95% CI 1.2-3.3), along with age ≥60 years, leukocyte count ≥15 × 109/L, and SRSF2 mutations. Abnormal karyotype was also an independent predictor of myelofibrosis-free survival (p<0.01; HR 3.7, 95% CI 1.8-7.8), along with SRSF2 mutations. Analysis of leukemia-free survival revealed abnormal karyotype to be a significant risk factor in univariate (p<0.01), but not multivariate analysis that included SRSF2 mutation (p>0.1). Conclusions The current study provides the distribution of cytogenetic abnormalities at different stages of PV and affirms the prognostic significance of abnormal karyotype at the time of diagnosis, highlighting the importance of incorporating cytogenetic studies in the diagnostic evaluation of PV.

Introduction: Pembrolizumab Gemcitabine Vinorelbine and Liposomal Doxorubicin (P-GVD) has become a commonly employed regimen for the treatment of relapsed or refractory classical Hodgkin lymphoma (CHL), often preceding autologous stem cell transplant in patients with good response. The utilization of P-GVD is based on a 38-evaluable patient prospective trial (Moskowitz et al. JCO 2021). There remains minimal data regarding the efficacy of this regimen in a larger, real-world cohort. Methods:Patients with CHL who received P-GVD at 9 United States centers were retrospectively evaluated to estimate efficacy and safety, with a focus on the second line use of this regimen. Demographic and clinical data were extracted from medical records. Responses reflected Lugano criteria per local investigator assessment. Study objectives were overall response rates (ORR), complete response rates (CRR), progression free survival (PFS), overall survival (OS), and safety. PFS and OS were estimated by Kaplan-Meier. Results: Patients and Treatment: We identified 65 CHL patients who received P-GVD between July 2019 and June 2024, including 43 who were treated in the second line. Enrolled patients included a median age of 31 years (range 17-69 years), 36.9% (24) females, 75.4% (49) nodular sclerosing subtype, and 60% (39) stage III-IV at diagnosis. Patients included 64.6% (42) White and non-Hispanic, 15.4% (10) Hispanic or White and Hispanic, 10.8% (7) Black, 3.1% (2) Asian/Pacific Islander, 3.1% (2) other, and 3.1% (2) unknown. The median number of prior regimens was 1 (range 0-12); these included 3.1% (2) who had prior checkpoint inhibitor (CPI), 50.8% (33) who had prior brentuximab vedotin (BV), and 3.1% (2) who had prior transplant. Patients with primary refractory disease [no initial complete response (CR)] comprised 58.5% (38), and 35.4% (23) had relapse within 1 year of completing initial therapy. The median number of cycles received was 3 (range 1-5). Treatment in a community practice setting was noted for 21.5% (14) patients. Efficacy: At a median follow up of 17.7 months, ORR and CRR after P-GVD were 95% (62/65) and 74% (48/65), respectively. This increased to 95% ORR and 81% CRR when limited to 43 patients who were treated 2nd line. Responses by line included: 1st [n=1; 100% ORR and CRR]; 3rd [n=15; 100% ORR, 73.3% CRR], 4th [n=3; 100% ORR, 33.3% CRR], 5th [n=2; 100% ORR, 0% CRR], and 13th [n=1; SD]. CRR in BV- and CPI-exposed patients were 60.6% and 50%, respectively. Ninety one percent (39) of 2nd line treated patients proceeded to autologous transplant, as well as all 3 patients > age 60. Among 41 response-evaluable second-line patients, CR was achieved in 94% (30/32) who received 2 or 3 cycles of P-GVD, and 56% (5/9) who received 4 cycles of P-GVD. For the entire cohort at 12 and 24 months after starting P-GVD, PFS was 95% (95% CI: 89-100%), 91% (95% CI: 83-100%) and OS was 98% (95% CI: 94-100%), 94% (95% CI: 86-100%), respectively. Ninety five percent (41) of 2nd line transplanted patients remain alive and in remission at a median of 18.5 months (range 3.5-38.7 months). In the primary refractory subset, PFS and OS at 24 months were 91% and 97%, respectively. For the cohort with 2 or more prior lines of therapy, PFS and OS at 24 months were 91% and 94%, respectively. Safety: Immune-related adverse events (IRAEs) were reported in 20% (13) of patients, of which grade 3 IRAEs per NCCN criteria version 1.2024 comprised 38.5%; all others were grades 1-2. Non-IRAE grade 3-4 toxicities per CTCAE version 5.0 were reported in 22% (14) patients. Of 29 patients evaluable for treatment-related interruptions, 7% (2) had therapy discontinued, and 34% (10) had holds, dose reductions, and/or delays. No deaths were attributed to P-GVD treatment. Conclusions:This retrospective study confirms the utility of P-GVD in treating relapsed/refractory CHL in a real-world population. Though fewer CRs and higher rates of grade 3 adverse events were observed, the vast majority of patients were able to proceed to transplant, and outcomes to date are highly encouraging.

Background: Randomized studies in newly diagnosed (ND) AML patients with a FLT3 mutation show that adding 14 days of a FLT3 inhibitor to each intensive chemotherapy cycle improves overall survival. For patients unsuitable for intensive chemotherapy due to older age or comorbidities, the presence of a FLT3-ITD mutation is associated with high rates of early relapse and short remission duration. There is a limited survival benefit of venetoclax (ven) plus azacitidine (aza) compared to aza alone among patients with AML with a FLT3-ITD mutation. Accordingly, there is significant interest in adding FLT3-targeted therapy to lower intensity chemotherapy for patients with ND AML and FLT3 mutations, but to date no randomized data have demonstrated a survival benefit for this approach.Gilteritinib (gilt) is a highly potent and FLT3-selective oral tyrosine kinase inhibitor that is the standard therapy of relapsed/refractory FLT3-mutated AML, based upon superior survival in randomized studies. Gilt can also be combined safely with ven and/or aza, though myelosuppression remains a concern. Despite this, single institution studies of ven, aza, and gilt therapy of ND AML patients with FLT3-mutations show that this three-drug regimen is associated with near-universal complete remission rates (CR 90%, CRi 6%) and a majority of studied patients eliminate detectable FLT3-ITD (<1 x10-5) within four cycles by an ultrasensitive, amplicon-based NGS assay (Short, et al. JCO 2024). Rates of CR with full count recovery and survival appear superior to historical comparisons, but myelosuppression is potentially increased with the triplet regimen. These data demonstrate a need for multi-institutional comparative trials, not only to determine whether adding FLT3-inhibitors to ven and aza improve deep remission rates, but also to refine schedules of the triplet combination that improve tolerability for widespread use.Methods: This tier 1 MyeloMATCH substudy is an open-label, randomized phase 2 trial that will enroll ND AML patients older than age 60 or individuals judged by their treating physician to be best served by azanucleoside + ven-based therapy due to comorbidity. Subjects provide samples to the central MyeloMATCH Master Screening and Reassessment Protocol (MSRP, NCT05564390 ) and are assigned to the MM1OA-EA02 substudy (NCT06317649) based upon the presence of a FLT3-ITD or FLT3-TKD (D835) mutation by NGS. Subjects are randomized to one of three arms (regimens 1, 2, or 3), stratified by age (<70 years vs. older) and FLT3-ITD burden (ITD VAF>33% vs. lower VAF or D835). All study arms contain aza and ven administered in 28-day cycles with two study arms also including gilt. Regimen 1 is ven and aza administered as per the pivotal VIALE-A study. Regimen 2 adds gilt 80 mg once daily on days 1-28 and regimen 3 adds gilt on days 8-21 only. A marrow biopsy is performed during the first cycle (day 14-21, depending on study arm) and all chemotherapy is held until count recovery if a morphologic leukemia-free state or aplasia is seen. Patients receive up to two induction cycles to achieve remission and then receive ongoing consolidation cycles of therapy with a similar pattern of gilt administration up to 2 years. To minimize risk of cumulative myelosuppression, during consolidation azacitidine administration is reduced to days 1-5 on regimens 2 and 3 and ven limited to days 1-7 on regimen 2 and days 1-14 on regimen 3. For all arms, treatment-related adverse events prompt further reduction in ven, aza, or gilt dose or duration. Response will be measured after cycles 2 and 4, including centrally-performed MRD by flow cytometry. The primary endpoint is the rate of MRD(-) CR (<1 x 10-3) after up to 4 cycles. Key secondary endpoints include safety data, response rate, EFS, and OS. The study is powered to detect a 25% absolute improvement in MRD(-) CR rate in either triplet arm from a target accrual of 147 patients. A safety run-in for regimens 2 and 3 to confirm tolerability and feasibility will be performed after 20 subjects enroll to these arms; an interim futility analysis will be performed once response assessment of 50% of all planned accrual occurs. Study was activated on June 6, 2024 and is expected to continue until December 2025.

Background: Chromosomal rearrangements involving the KMT2A gene and mutations involving the nucleophosmin (NPM1) gene have been associated with response to menin inhibitors in multiple studies of acute myeloid leukemia (AML) patients characterized by these genetic alterations. Both alterations are more common in younger AML patients and those with NPM1 mutations historically perform better. However, the prevalence and outcomes of older (≥60 years) AML patients with these alterations who are treated with current therapies is less well understood. We present results from the Beat AML Master Trial (BAMT) that analyze the outcomes of this large, but less common subset of patients in an older AML population. Methods: Patients were consented and enrolled on this precision medicine-based Beat AML Master Trial (NCT03013998) from November 2016 to June 2024. Patients were included in this analysis if they harbor either the classic NPM1 mutation or a KMT2A rearrangement (over 10 different gene partners). Additionally, 2 patients with a NUP98 rearrangement were included with the KMT2A cohort as they are known to have similar biology in terms of menin dependence. These patients were treated with the following therapies: a sub-study specific targeted therapy for the dominant clone, venetoclax + hypomethylating agent (Ven/HMA), intensive chemotherapy (IC), alternative non-intensive therapy (NIT), or no therapy. Clinical/genetic characteristics and median overall survival (OS) were analyzed in each group separately. Overall survival (OS) was calculated from master consent and estimated using the Kaplan-Meier method. Results: Of 1096 AML patients enrolled on the BAMT, 246 patients had either an NPM1 (N=207, 18.9%) or KMT2A (N=39, 3.6%) genetic alteration. The baseline characteristics for the NPM1 cohort were median age 72 years (range: 60-92), 50% female, and ECOG performance status: 0-16%, 1-58%, 2-25% and 3-1%. Additional disease characteristics include treatment related AML at 11% and complex karyotype (≥3 abnormalities) at 3%. Median OS for the entire NPM1 cohort was 18.4 months (95% CI: 14.2-24.7). Patients were treated on a sub-study (35%), Ven/HMA (19%), IC (26%), NIT (8%) or no treatment (9%). Between treatment groups, median OS was 21 months (95% CI: 12.8-31.7) for those treated on a sub-study, 22 months (95% CI: 12.6-not reached (NR)) for Ven/HMA, 41.6 months (95% CI: 18.2-NR) for IC, and 6.3 months (95% CI: 1.3-23) for NIT, which was significantly different (p<0.0001). The baseline characteristics for the KMT2A cohort were median age 69 years (range: 60-88), 44% female, and ECOG performance status: 0-19%, 1-45%, 2-35% and 3-0%. Treatment related AML occurred in 23% of patients and complex karyotype in 31%. Median OS for the KMT2A cohort was 6.5 months (95% CI: 3.6-31.6). Patients were treated on a sub-study (38%), Ven/HMA (15%), IC (15%), NIT (15%), or no treatment (5%). Between treatment groups, median OS was 31.6 months (95% CI: 3.2-NR) for sub-study, 5.0 months (95% CI: 3.0-NR) for Ven/HMA, 12.7 months (95% CI: 4.9-NR) for IC, and 2.9 months (95% CI: 1.5-NR) for NIT, which was also significantly different (p<0.0001). Conclusions: In this cohort of older AML patients who possess NPM1 or KMT2A mutations, initial demographics and clinical characteristics upon presentation are as expected. Within each cohort and depending on the type of therapy received, there were interesting differences in survival. Median OS was the highest for the NPM1 cohort that received IC while the KMT2A cohort had the highest median OS when enrolled on a KMT2A targeted sub-study; a finding that supports precision-based medicine for this high-risk KMT2A group characterized by poor outcomes. As menin inhibitor combinations with induction chemotherapy or Ven/HMA move to the frontline setting, we anticipate seeing significant improvement in outcomes for AML patients in both of these genomic subsets. Further multivariable analysis will be performed to validate treatment effect on OS. Our analyses for these two cohorts also provide valuable outcome benchmarks for the design of future precision oncology studies that seek to improve survival in these subsets of older AML patients.

Background: Erdheim-Chester disease (ECD) manifests as multi-organ infiltration of clonal histiocytes with mutations primarily in the mitogen-activated protein kinase (MAPK) pathway. The revised classification of histiocytosis (2016) categorizes ECD with typical bilateral, symmetric osteosclerosis of the lower extremity long bones as classical (C) and as non-classical (NC) if otherwise. The clinical and biological significance of this classification is unclear. This study compared the clinical and genotypic findings of the 2 subtypes within a large cohort of ECD patients. Methods: We included patients seen at our institution since 1990 meeting the following criteria: A) had an ECD diagnosis based on the revised criteria by Haroche J, et al. (Blood 2020;135:1311-1318); B) completed whole-body imaging studies including long bones of the legs. Classification as C-ECD required symmetric osteosclerosis of the distal femur and/or proximal tibia/fibula. Results: A total of 145 patients were enrolled, comprising a male-to-female ratio of 2:1 (66.2% male). C-ECD was diagnosed in 77.2% of cases, and NC-ECD in 22.8%. The median age at diagnosis was similar with 58.5 years (range, 18-82) in C-ECD and 55 years (range, 20-81) in NC-ECD, (p=0.65). The most frequently affected systems included skeletal (81.4%), renal (60.0%), vascular (45.5%), cardiac (40.0%), central nervous (38.6%), sinus (37.2%), pulmonary (36.6%), and adrenal (32.4%). The numbers of organs involved were significantly higher in C-ECD compared to NC-ECD (median 5 vs 3, p<0.01). Higher rates of skeletal (100% vs. 18.2%, p<0.01), adrenal gland (37.5% vs. 15.2%, p=0.02), and sinus (44.6% vs. 12.1%, p<0.01) involvement were observed in C-ECD. Arginine vasopressin deficiency (AVP-D) was exclusively observed in C-ECD (20.7% vs. 0%, p<0.01). NC-ECD had a higher rate of cardiac involvement (69.7% vs. 31.3%, p<0.01). No significant differences were found in other systems. The overall median time between onset of the first symptoms/signs to diagnosis was 15 months with no significant difference between C-ECD and NC-ECD (17.5 vs. 12 months, p=0.18). The median time to diagnosis of ECD in patients with AVP-D was 38.5 months, compared to 12 months without AVP-D (p=0.09). Genetic testing was conducted in 86.2% of patients and B-Raf proto-oncogene (BRAF) alterations were identified in 59.2% of the tested cases. BRAFV600E was more prevalent in C-ECD (61.3% vs. 21.9%, p<0.01), whereas non-V600 mutations were more commonly found in NC-ECD (12.5% vs. 1.1%, p<0.01). There was no significant difference in the frequency of BRAF fusions. NC-ECD patients exhibited a higher rate of MAP2K1 mutations (25.0% vs. 5.4%, p<0.01). Other frequently mutated genes included KRAS (4.0%), NRAS (2.4%), and CSF1R (2.4%), although no significant difference was observed in their frequency between C-ECD and NC-ECD. Testicular involvement appeared to be more common in patients with MAP2K1 mutations (23.1% vs. 8%, p=0.08). Two patients with pathogenic CSF1R mutations both had central nervous system (CNS) involvement. Among C-ECD patients, BRAFV600E conferred a higher rate of CNS involvement (50.9% vs. 25%, p=0.01). No significant difference was observed in NC-ECD patients (71.4% vs 36%, p=0.09). Among NC-ECD cases, MAP2K1 mutations conferred a reduced risk of cardiac involvement (37.5% vs. 79.2%, p=0.03) and no such difference was observed in patients with C-ECD (40% vs 34.1%, p=0.79). The median numbers of organs involved in patients with BRAF, MAP2K1, and KRAS/NRAS mutations were 5, 3, and 3, respectively, with no significant difference. Conclusions: C-ECD affects more organs and more frequently involves bones, adrenal glands, and sinuses. AVP-D was exclusively observed in C-ECD. Conversely, NC-ECD showed a higher prevalence of cardiac involvement. BRAFV600E was more commonly identified in C-ECD, whereas non-V600 mutations and MAP2K1 mutations were more prevalent in NC-ECD. Among C-ECD patients, BRAFV600E was correlated with a greater likelihood of CNS involvement. Conversely, NC-ECD patients with MAP2K1 mutations had a reduced risk of cardiac involvement. This study highlights the distinct clinical and molecular features of C-ECD and NC-ECD, underscoring the importance of subclassifying ECD into classical and non-classical forms. Further studies are needed to explore differences in the clinical outcomes between the 2 forms.

Background: Clonal cytopenia of undetermined significance (CCUS), a precursor of myeloid neoplasms (MN), is characterized by one or more persistent cytopenias, and acquired mutations in myeloid malignancy-associated genes. ZRSR2, located at Xp22.2, is one of the MN-associated genes and a member of the spliceosome family of genes. Aim: Describe the clinical characteristics, pattern of progression, and genetics of ZRSR2 mutation (ZRSR2m)-harboring CCUS patients. Methods: Next-generation sequencing (NGS) was performed for 9320 patients between 2016-2023 at Mayo Clinic. After obtaining IRB approval, clinical characteristics and results of bone marrow morphology, cytogenetics, and molecular studies were abstracted. Overall Survival (OS) was calculated using Kaplan-Meier, from the date of NGS until death or last follow-up. The ZRSR2m MN control cohort included 125 patients with myelodysplastic syndrome (MDS), myeloproliferative neoplasm (MPN), and MDS/MPN overlap. BlueSky Statistics V10.3.1 was used for statistical analysis. Results: ZRSR2m was detected in 39 CCUS patients, all of whom were males. Concurrent hematological neoplasms were identified in 8 patients (21%). Five (13%) had plasma cell proliferative disorders including 3 multiple myeloma, 1 MGUS, and 1 AL amyloidosis. There were also 2 patients with chronic lymphocytic leukemia and 1 with VEXAS syndrome. Prior chemotherapy, with or without radiotherapy, was given to 4 patients (10.3%), whereas 13 patients (33%) had received previous immunotherapy. Abnormal cytogenetics were detected in 11 patients (29%), with the most common abnormality being +8 (n=5, 13%). In 23 patients (59%), an absolute monocyte count of ≥0.5 x 10^9/L was documented which could be classified as clonal cytopenia with monocytosis of unknown significance (CCMUS). ZRSR2 median gender-corrected VAF among CCUS patients was 32% (range, 1-45). Isolated ZRSR2m was found in 6 patients (15%), and 1 (3%) had multiple ZRSR2m. The median number of co-mutations was 1 (range, 0-3). Sequential NGS, available in 13 patients, showed that 1 patient developed 3 ZRSR2 mutations and 2 ZRSR2 VUS. TET2 was the most common co-mutation (n=25, 64%). Additionally, TET2 was an isolated co-mutation in 18 patients (46%). ASXL1 co-mutation was the second most common (n=8, 20.5%), while other less common co-mutations were IDH1, IDH2, SRSF2, SF3B1, TP53, GATA2, and KDM6A. Pre-ZF1 was the most commonly mutated region in ZRSR2 mutations (n=23, 59%). With a median follow-up of 32 months, 9 (23%) of the ZRSR2m CCUS patients progressed to MNs, none progressed to AML. Five (55.6%) progressed to MDS, 4 were subclassified as low blast (LB) and 1 as increased blast-1 (IB1). Four (44.4%) progressed to MDS/MPN overlap syndrome, with 3 subclassified as chronic myelomonocytic leukemia (CMML). None of the patients with isolated ZRSR2m progressed to MDS or CMML. None of the patients who received prior immunotherapy progressed to MDS. Thirty-six (92.3%) patients had high-risk CCUS (Malcovati et al, Blood 2017) while 3 had intermediate risk with no low-risk patients. None of the intermediate-risk patients progressed to MDS, while 5 patients of the high-risk (13.9%) progressed to MDS (p=1). Twelve (31%) patients died, and the median overall survival (mOS) was 56 months. Higher hemoglobin levels improved OS (HR=0.59, p=0.02), while higher adjusted ZRSR2 VAF worsened OS (HR=1.05, p=0.017). Patients with TET2 as isolated co-mutations showed better mOS than the rest of the cohort (not reached vs 41 months, p= 0.03). ZRSR2m CCUS patients showed a trend suggestive of better mOS than ZRSR2m with MNs including AML (56 vs 30 months, p=0.14), and with excluding AML (56 vs 39 months, p=0.24). On multivariate analysis, both Hgb (HR=0.61, p=0.03) and isolated TET2 co-mutation (HR=0.19, p=0.04) retained statistical significance. mOS was not affected by prior history of treatment, number of co-mutations, isolated ZRSR2, ASXL1 co-mutation, or MDS progression. Conclusion: ZRSR2m CCUS cohort was exclusively males. There was a significant number of patients with concurrent hematological malignancy or a history of prior chemo- and/or immuno-therapy (36%). Most patients had high-risk CCUS but showed a low rate of progression to MNs, especially in those with isolated ZRSR2. Additionally, patients with isolated TET2 co-mutation had better survival, indicating that TET2 confers a favorable prognosis among ZRSR2m CCUS patients.

Background: Multiple Myeloma (MM) is predominantly a disease of older adults. However, older patients vary widely in physiological status and treatment tolerance. The simplified frailty index (Facon 2020) incorporates age, ECOG performance status, and comorbidities (Charleston Comorbidity Index [CCI]) to classify patients as frail or non-frail. While frailty is associated with increased toxicity with conventional systemic therapies, its impact on outcomes of older patients treated with bispecific antibodies (BsAbs) has not been established. Thus, we conducted a retrospective study to evaluate the efficacy and toxicity of BsAbs in older patients based on frailty. Methods: This is a single institution study. We included patients with MM ≥65 years who initiated treatment with a BsAb at Mayo Clinic in Rochester, MN from August, 2018 to January, 2024. Patients who received a BsAb on trial or as standard of care were included. For patients who received 2 different BsAbs at different timepoints, we included the first treatment only. We grouped patients into frail and non-frail based on the simplified frailty index (score ≥2 = frail). We evaluated the overall response (ORR) and ≥very good partial response (VGPR) rates, progression-free (PFS) and overall (OS) survival, early mortality (within 90 days) including early non-relapse mortality (NRM), and toxicities including cytokine release syndrome (CRS), immune effector cell associated neurotoxicity syndrome (ICANS) and healthcare utilization. P values were insignificant (>0.05) unless otherwise stated. PFS and OS were calculated from the day of BsAb start. Results: A total of 99 patients age 65-89 years were included. The products used were teclistamab (N=43), talquetamab (N=2), and investigational agents (N=54); 67% were ≥70 and 29% were ≥75 years; 39% were female, 95% were non-Hispanic White and 4% were Black/African American. Seventy patients (71%) were considered frail and 29 (29%) were non-frail. At the time of BsAb start, a higher proportion of frail patients had platelets <75 x 10^9/L (30% vs 8%, P=0.04). Otherwise, there was no significant difference between frail and non-frail patients in age (median: 72 vs 70), hemoglobin <8 g/dL (19% vs 12%), absolute neutrophil count <1 x 10^9/L (6% vs 8%), presence of high-risk FISH (70% vs 60%), ISS stage 3 (32% vs 41%), high LDH (58% vs 61%), presence of extramedullary disease (39% vs 25%), serum creatinine ≥2 mg/dL (15% vs 8%), bone marrow plasma cell% (20% in both), triple- (83% vs 90%) and penta-class refractoriness (63% vs 48%), number of prior lines of therapy (median: 6 in both), prior exposure to BCMA-directed therapy (24% vs 21%, P=0.80) and prior autologous stem cell transplant (73% vs 86%). Step-up dosing was used in 81% vs 48% of patients (P=0.001), and treatment was initiated in the outpatient setting in 46% vs 28% (P=0.12) in frail and non-frail patients, respectively. The median follow-up was 1.9 (95%CI: 1.0-2.3) years. There was no significant difference between frail and non-frail patients in ORR (52% vs 62%), ≥VGPR (46% vs 48%), 1-year PFS (36% vs 48%, P=0.26), or 1-year OS (57% vs 61%, P=0.30), respectively. Age and CCI alone were not associated with PFS or OS, but ECOG >1 was associated with decreased PFS (hazard ratio: 3.1, P=0.001) and OS (hazard ratio: 3.1, P<0.001). There was no significant difference between frail vs non-frail patients in grade 3/4 thrombocytopenia (39% vs 17%, P=0.06), grade 3/4 neutropenia (27% vs 35%), total days of hospitalization (median: 8 vs 5, P=0.35), number of unplanned admissions in 6 months (median: 1 in both), or total hospital days for unplanned admissions (1 vs 2 days). There was no significant difference between frail and non-frail patients in early mortality (23% vs 10%, P=0.17), including early NRM (4% in both), CRS (59% vs 66%), grade 3-4 CRS (3% vs 7%), or CRS recurrence (21% vs 10%). ICANS occurred in 5 (9%) frail patients and 1 non-frail patient (grade 2); among frail patients, grades were: 1 in 3 patients, 3 in 1 patient, and 4 in 1 patient. Similarly, there was no significant difference in CRS/ICANS between the 2 groups when patients with and without step-up dosing were analyzed separately. Conclusion: Frailty, defined using the simplified frailty index, was not significantly associated with inferior efficacy or increased toxicity with BsAb therapy among older patients with MM. Poor performance status predicted inferior PFS and OS, while age and comorbidity scores did not.

Background: Immunosuppressive therapy (IST) is considered the standard front-line treatment for acquired severe aplastic anemia (SAA) for pts >40 years. For pts aged <40 years who have a suitable human leukocyte antigen (HLA)-matched sibling donor (MSD), hematopoietic stem cell transplantation (HSCT) is recommended. This recommendation is based on the toxicity associated with alternative donors; haploidentical (HAPLO) and unrelated donor (URD) HSCT for patients who lack a MSD. However, the hematopoietic response after IST is ~ 70-80%, even after augmentation with eltrombopag in the front-line setting, and failure-free survival (FFS, defined as patients alive and in remission without clonal disease more than 10 years after IST) remains < 40%. HSCT is often employed as a second line salvage therapy for those who do not respond to IST or have disease recurrence. However, outcomes after HSCT are often compromised because of IST-related complications such as clonal evolution, development of donor specific antibodies, infectious adverse events and organ dysfunction. In contrast, a successful allogeneic HSCT leads to relatively prompt hematopoietic and immune reconstitution and mitigates the risk of secondary clonal disease. Innovations in recent decades, such as improved graft versus host disease (GVHD) prophylaxis and supportive care, have steadily reduced the morbidity and mortality of adults undergoing HSCT from unrelated or mismatched donors and expanded the donor pool such that almost all pts with SAA have a potential donor available. The use of post-transplant cyclophosphamide (PTCy) is now a well-established approach to facilitate engraftment and mitigate the risk of GVHD. It was successfully used in relapsed/refractory SAA pts using HAPLO donors in the multi-center BMT CTN #1502 trial (NCT02918292). Similar PTCy based approaches have been effective in the upfront setting with HAPLO and URD in small single center studies. Opened in July 2024, the CureAA trial (NCT06517641), seeks to increase donor options for SAA pts who have not yet received IST by using PTCy based GVHD prophylaxis for HSCT using HAPLO or URD. Study Design and Methods: This BMT CTN study (#2207) is a prospective multi-center phase II trial to assess efficacy of HSCT using HAPLO or URD bone marrow with a reduced intensity, thymoglobulin (ATG)-containing preparative regimen and PTCy GVHD prophylaxis in previously untreated SAA pts. This trial has two parallel cohorts for each donor type, HAPLO and URD, with an accrual goal of 30 participants in each cohort. All will receive ATG (0.5mg/kg on day -9 and 2mg/kg on days -8 and -7), cyclophosphamide (14.5mg/kg on days -6 and -5), fludarabine (30mg/m2 on days -6 to -2), 400 cGy total body irradiation (day -1 in a single fraction), a marrow graft (goal >2.5 x 108 nucleated cells per kg of recipient) on day 0 and PTCy (50mg/kg on days +3 and +4). Additionally, participants receive tacrolimus (until day +180) and mycophenolate mofetil days +5-35. The primary objective of the trial is to estimate the GVHD-free failure-free survival (GFFS) rate at 1 year after initiation of conditioning. GFFS is a composite endpoint of survival without grade III-IV acute GVHD, chronic GVHD requiring immunosuppression, or primary or secondary graft failure requiring second definitive therapy within 1 year after initiation of conditioning. Secondary objectives will evaluate patient survival, engraftment, transfusion dependence, cumulative incidence of GVHD and hematologic response. Health related quality of life, healthcare utilization and financial toxicity data will also be collected. These endpoints will allow clinical comparison with current results achieved with upfront IST in other studies. We hypothesize that this HSCT approach will improve survival without significant transplant-associated morbidity by using a low toxicity regimen that reduces rates of GVHD and provides rapid hematologic recovery without graft failure, thereby making HAPLO and URD HSCT a viable option for initial upfront treatment of SAA. The ability to use mismatched donors will also ensure that a HSCT option is more widely available to all racial/ethnic groups. This collaborative study was funded by grants U10HL069294, U24HL138660 to the BMT CTN from the NHLBI/ NCI, along with funding from Sanofi. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.

Background: A deeper understanding of the cytogenetic features has enabled robust risk stratification for patients with newly diagnosed multiple myeloma (NDMM). However, heterogeneity persists in the currently available prognostic tools and there is further room for personalization. We assess the utility of PC-PRO, a test to measure the proportion of monotypic plasma cells in the synthetic phase (S-phase) of cell cycle, as a prognostic marker for NDMM. Methods: Patients with active MM diagnosed between January 2013 and January 2023 with a flow cytometry-based S-phase assessment at diagnosis were included. Flow cytometric immunophenotyping was performed using the following antibodies: CD19, CD38, CD45, CD138, cytoplasmic kappa and lambda immunoglobulin, and DAPI (4',6-diamidino-2-phenylindole). The percentage of plasma cells in S-phase was determined by measuring the proportion of cells with DNA content of the monotypic plasma cells between the G0/G1 and G2/M peaks. Plasma cell clonality was detected through demonstrating CD38 and CD138 positivity along with immunoglobulin light chain restriction, abnormality of CD19/ CD45 expression and/or ploidy difference by DAPI staining. High-risk cytogenetic abnormalities (HRCA) by FISH were defined as follows: presence of deletion 17p (del17p), t(4;14), t(14;16), t(14;20), 1q gain/amplification, 1p deletion (when available). Additionally, the second iteration of the revised ISS (R2-ISS), incorporating both laboratory and cytogenetic features as well an adaptation of the recently proposed International Myeloma Working Group (IMWG) Classification for high-risk disease, was utilized for risk stratification. All time-to-event analyses were performed using the Kaplan Meier method and compared using the log-rank test. Univariable and multivariable Cox proportional hazards analyses were performed to assess the independent prognostic impact of S-phase. Results: A total of 823 patients were included in the study. The median follow-up was 6 years [95% Confidence interval (CI): 5.5-6.3 years] and estimated median overall survival (OS) was 8.3 years. The median S-phase value for the cohort was 0.8 % (interquartile range: 0.5-1.4%), with 135 (16%) patients harboring an S-phase of ≥2% at diagnosis. Applying the R2-ISS prognostic model to our cohort, 11% (79/706) patients were stratified to the R2-ISS high-risk cohort; 93 patients (out of 788, 12%) had 2 or more HRCA at diagnosis. Patients with a S-phase ≥2% at diagnosis had a median OS of 3.9 years (95% CI: 2.9-5.7) vs. 9.2 years [95% CI: 7.9-not reached (NR)] for patients with a S-phase <2% [Hazard ratio (HR): 2.2 (95% CI: 1.7-2.8), p<0.0001]. The median OS was inferior for cohorts with an increasing S-phase value: S-phase of 0.1-1%: HR: 1 (standard), S-phase 1.1-3%: HR 1.7 (95% CI: 1.3-2.1), S-phase >3%: HR 3.0 (95% CI: 2.2-4.1)]. On a multivariable analysis including S-phase ≥2%, age at diagnosis of MM (>65 years) and R2-ISS high-risk status, S-phase ≥2% was an independent predictor of inferior OS [hazard ratio (HR) 1.9 (95% CI: 1.4-2.6), p<0.0001]. Age >65 years [HR 1.9 (95% CI: 1.5-2.5), p<0.0001] and R2-ISS high-risk status [HR 1.8 (1.3-2.6), p=0.001] were also independent predictors of OS. Within the sub cohort of patients with 2 or more of the listed high-risk cytogenetic features (n=93, 12%), a S-phase ≥2% (n=24) was associated with a worse median OS of 1.6 years (95% CI: 0.7-NR) vs. 6.2 years (95 % CI:3.3-NR) for S-phase <2% (p=0.002). Conclusion: Flow cytometry-based determination of the proportion of monotypic plasma cells in S-phase is an easy-to-study and powerful prognostic tool for NDMM. A S-phase of ≥2% at diagnosis is a predictor of OS independent of age and R2-ISS high-risk stage. An S-phase of ≥2% was able to identify a distinct group with markedly inferior outcomes even within the sub cohort of patients with 2 or more high-risk cytogenetic features.

Chronic lymphocytic leukemia (CLL), one of the most common types of leukemia among adults in the US, is characterized by the accumulation of CD5+CD19+ monoclonal B cells. Current clinical treatment approaches for progressive CLL patients focus on novel agents targeting Bruton Tyrosine Kinase (BTK) and Bcl-2. Although clinical trials combining novel agent have shown significant efficacy with high overall response and, in some cases, achievement of high rates of undetectable measurable residual disease (MRD), CLL remains incurable with a majority of patients eventually experiencing relapse of their disease. Therefore, it remains of high interest to expand therapeutic options. Transcriptional chemical inducers of proximity (TCIPs) are small molecules designed and developed by Dr. Gerald R. Crabtree at Stanford University. TCIPs are a new class of anticancer drugs that recruit endogenous cancer drivers or a downstream transcription factor to the promoters of cell death genes and activate their expression, resulting in apoptosis of cancer cells. Current TCIPs are produced by covalently linking small molecules that bind B-cell lymphoma 6 (BCL6) to those that bind to transcriptional activators, such as Bromodomain-containing protein 4 (BRD4) or cyclin-dependent kinases 9 (CDK9). The Stanford team has demonstrated TCIPs generate remarkable killing in diffuse large B cell lymphoma (DLBCL) cell lines, including chemotherapy-resistant, TP53-mutant lines, with a half maximal effective concentration (EC50) of 1-10nM at 72hours (h) and exhibited cell-specific and tissue-specific effects (Gourisankar et al, 2023). We have initiated TCIPs cytotoxicity assessment in primary leukemic cells from untreated CLL patients. Testing two types of TCIPs with a flow cytometry-based apoptosis assay, we found time and dose-dependent drug-associated killing in primary CLL B cells collected from untreated CLL patients over a 24-72h interval. The IC50 for BRD4-BCL6 TCIP JWZ-7-133 at 72h was 560nM while CDK9-BCL6 TCIP BAK-04-232 at 72h was 10nM. After 24, 48, and 72h exposure of CLL B cells to TCIP JWZ-7-133, the IC50 was 1050, 820 and 560nM, respectively. We also found that TCIP JWZ-7-133 exhibited a similar ability to inhibit leukemic cell viability with an IC50 of 1070nM using a cell viability assay, compared to a flow cytometry-based cell death assay (24h IC50 1050nM). CDK9-BCL6 TCIPs are designed and produced by covalently linking BI-3812 (binding to BCL6) and SNS-032 (binding to CDK9) with a rigid linker. BI-3812 is a potent and efficacious BCL6 inhibitor, inhibiting the BTB domain of BCL6, and SNS-032 is a potent and selective inhibitor of CDK9. When CLL B cells were treated with only BI-3812 or SNS-032 or TCIP BAK-04-232 as single agents for 72h, we found that TCIP BAK-04-232 had superior and significant killing effects compared to the single agents, with an IC50 of 10nM. Specifically this killing activity was more than 10-fold compared to SNS-032 alone (IC50 over 100nM), and more than 1,000-fold compared to BI-3812 alone (IC50 over 10,000nM). There remains an urgent to identify new categories of potent drug, inhibitor, or small molecules to expand potential clinical treatment options for patients with CLL. The results presented here indicate that TCIPs targeting BCL6 and BRD4/CDK9 have a strong capacity to induce high levels of cell death in CLL B cells at nanomolar concentrations, and this remarkable killing effect may be due to its unique gain-of-function drug mechanism. TCIPs provide a completely new drug direction and strategy for the design of anticancer drugs: to rewire cancer drivers to activate apoptosis in cancer cells. This study is the first of a kind demonstration of TCIPs in robustly targeting CLL B cells. Further studies to fully validate the preclinical activity of TCIPs and investigate the molecular mechanism(s) for TCIP mediated killing of leukemic B cells will help to define the role of TCIPs as a novel therapeutic approach in CLL.