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Introduction: Despite the improved clinical outcomes with novel therapies in patients with chronic lymphocytic leukemia (CLL), Richter transformation (RT) of CLL remains a major clinical challenge. Our group showed a selective response (~40%) with the PD-1 blocking antibody pembrolizumab in patients with RT, particularly after prior exposure to ibrutinib (Ding et al, Blood, 2017). It has been established that PD-1 inhibits cytotoxic T-cell functions via the interaction with its ligands, PD-L1 and PD-L2. However, the role of PD-1 in tumor B-cells is understudied. Here we show the expression and functional implications of the PD-1 signaling axis in B-cell pathobiology. Methods: CLL-involved lymph node (LN) (n=26) and RT (with diffuse large B-cell lymphoma [DLBCL] histology) -involved LN (n=20) samples were tested for PD-1 expression by immunohistochemistry (IHC) (mouse clone NAT105, Abcam). Then, PD-1 expression was examined in 10 lymphoma cell lines and 1 CLL-like cell line (MEC-1) by both flow cytometry (FC) and Western blot (WB) analysis. PD-1 overexpression using pLEX-lentiviral packaging system (Thermo Scientific) was evaluated for its impact on in vitro cell cycle regulation and in vivo tumor growth. CRISPR/CAS9 mediated PD-1 knockdown evaluated on cell growth in MEC-1 and Mino cells. Role of PD-1 was further examined on the regulation of cell signaling pathways using human phospho kinase array kit (R&D) and WB analysis. Additionally, gene expression signatures were evaluated by RNA-seq using pLEX-lentiviral-PD-1 transfected DLBCL cell lines OCI-LY7 and OCI-LY19. Results: PD-1 expression was significantly increased in RT-LN compared to CLL-LN (mean ± SEM in RT vs. CLL, 30.6% ± 4.7% vs. 11.5% ± 2.8%, p<0.001) as determined by IHC. PD-1 expression was highest in patients with RT where the last prior CLL therapy was ibrutinib. To test the role of PD-1 in tumor B-cells, its expression was assessed in lymphoma and CLL cell lines. The expression of PD-1 was found to be variable by both FC and WB analysis, but at very low-levels in DLBCL cell lines OCI-LY7 and OCI-LY19. Constitutive lentiviral (pLEX-PD-1)-mediated overexpression of PD-1 in OCI-LY7 and OCI-LY19 cells led to increased cell growth (1.4 and 1.9 fold, respectively, compared to original lines after 4 days of culturing of equal number of cells), which was further confirmed by cell cycle analysis which showed an increase in S phase by 20.4% (p<0.05) and 24.5% (p<0.05) in OCI-LY7 and OCI-LY19 cells, respectively. When the luciferase+ PD-1 overexpressing OCI-LY7 cells (1X10 6) were injected intravenously into NSG mice, increased tumor growth (p<0.01) was observed by bioluminescence imaging after 22 days of follow-up. PD-1 knockdown in MEC-1 and Mino cells (high basal PD-1 expression) led to significant inhibition of cell growth (85% and 79.5%, respectively, p<0.05). Using phospho-kinase array, an overall decrease of phosphorylation was detected on multiple sites of p53 (S392, S15, S46) and Chk-2 (T68) in PD-1 overexpressing OCI-LY7 and OCI-LY19 cell lines. This finding was further confirmed by WB analyses. In addition, decreased total p53 and increased phosphorylation on both SHP-1 and SHP-2 were found in PD-1 overexpressing OCI-LY7 and OCI-LY19 cell lines. SHP-1 (TPI-1) and SHP-1+SHP-2 (TPI-1+TNO155) inhibitor treatment promoted OCI-LY19 cell death and led to recovery of phosphorylation on ATM, Chk-2 and p53 in these cells. RNA sequencing data in PD-1 overexpressing OCI-LY7 and OCI-LY19 cells revealed increased expression of genes that promote cell cycle progression and DNA replication. Conclusion: Our data showed robust PD-1 expression in patients with Richter transformation from CLL to DLBCL. PD-1 overexpression in DLBCL lymphoma cell lines enhanced cell proliferation in vitro and in vivo. Further investigation identified that PD-1 modified the phosphorylation/function of SHP phosphatases and thereby regulated p53 pathways ( Figure 1). These findings indicate that RT with DLBCL cells have the ability to “hijack” the PD-1 pathway which results in downregulation of p53 mediated DNA repair. This novel finding provides for new strategies to further evaluate the interaction of PD-1 signals with DNA repair pathways and suggest potential novel targets for RT.

INTRODUCTION Chronic lymphocytic leukemia (CLL) and SLL represent a single entity in current classifications per the WHO-HAEM5 and ICC. Both are characterized by malignant B cells with an identical immunophenotype, but they are distinguished by the predominant site of disease burden, such as peripheral blood for CLL and nodal compartments for SLL. Most clinical trials and retrospective studies involving CLL/SLL do not separate out SLL patients. In this study, we report reasons for treatment initiation and outcomes in a cohort of SLL patients seen at our institution. METHODS Using the Mayo Clinic CLL Database, we retrospectively identified patients with SLL who received first line therapy between 12/1995 and 2/2023. Patients were classified as having SLL if they met at least one of the following criteria for diagnosis: 1) presence of lymphadenopathy or splenomegaly (either by exam or scans) and an absolute clonal B cell count of < 5 x 109/L; or 2) presence of lymphadenopathy or splenomegaly (either by exam or scans) and an absolute lymphocyte count of < 11 x 109/L (if absolute clonal B cell count was not available). Although the presence of anemia (hemoglobin < 11 g/dL) and thrombocytopenia (platelets < 100 x 109/L) are considered as exclusion criteria for a diagnosis of SLL, patients were included if these cytopenia were due to autoimmune hemolytic anemia (AIHA) or immune thrombocytopenia (ITP) with < 50% bone marrow involvement by SLL (bone marrow biopsy confirmed). Patients treated for SLL with extranodal involvement were also included. Indications for SLL treatment were abstracted by chart review and included one of the following: 1) symptomatic or progressive lymphadenopathy and/or splenomegaly, 2) autoimmune cytopenias, 3) extranodal/direct organ infiltration; and 4) other (e.g., cryoglobulinemia, paraneoplastic, non-infiltrative renal disease such as minimal change disease). Overall survival (OS) was defined as the time from treatment start until date of death or last known to be alive. Treatment-free survival (TFS) was defined as the time from treatment start until the earliest of date of next treatment, or death. Kaplan-Meier method was used to display OS and TFS. Cox proportional hazards regression models were used to estimate associations of factors with time-to-event outcomes. RESULTS We identified 182 patients with SLL who received first line treatment. The median age at treatment start was 66 years, and 132 (73%) were male. Unmutated IGHV was identified in 60/82 (73%) patients and peripheral blood fluorescent in situ hybridization (FISH) assessed pre-treatment start showed del13q in 10%, trisomy 12 in 43%, del11q in 7%, del17p in 8%, and no abnormalities in 33%. TP53 disruption (del17p and/or TP53 mutation) was present in 7/89 (8%) patients prior to first therapy. The median follow-up after diagnosis was 6.9 years. The indications for treatment were grouped as follows: symptomatic or progressive lymphadenopathy/splenomegaly (n=130), extranodal/direct organ infiltration (n=30), AIHA/ITP (n=10), and other (n=12). For the entire cohort, the median TFS was 2.9 years and the median OS was 12.3 years. The median TFS by the treatment indication groups were 3.3, 5.1, 1.6, and 5.3 years, respectively (Figure 1). The median OS by treatment indication groups were 11.1, 10.5, 8.8, and 19.9 years, respectively. Among the 160 patients with symptomatic or progressive lymphadenopathy/splenomegaly or extranodal disease, the treatments administered include: mAb alone (n=35), chemotherapy alone (n=33), chemoimmunotherapy (n=57), targeted agent (n=33; Bruton tyrosine kinase inhibitor [BTKi] based n=28; venetoclax-based n=3, BTKi + venetoclax n=2), and other (n=2). The median TFS by treatment groups were 2.4, 2.0, 3.8, and 6.0 years (Figure 2), respectively (excluding the small “other” subgroup of 2 patients). The median OS for these groups were 9.5, 9.7, 15.0, and not evaluable, respectively. Only older age at treatment start predicted for shorter TFS (HR 1.02; 95%CI 1.002-1.038; p=0.03) and shorter OS (HR 1.05; 95%CI 1.03-1.07; p<0.01). CONCLUSIONS In this study of patients with SLL who received therapy, the most frequent indications for treatment included symptomatic lymphadenopathy and/or splenomegaly, followed by extranodal disease. Trisomy 12 was the most frequently detected abnormality on peripheral blood FISH testing. Treatment with novel agents achieved durable TFS and OS.

Abstract Background The BioFire® FilmArray® Gastrointestinal Pathogen Panel (GIPP) tests for 22 potential pathogens in stool, including Clostridioides difficile. C. difficile nucleic acid is detected via nested polymerase chain reaction (PCR) targeting toxin A (TcdA) and/or toxin B (TcdB) genes. While PCR-based diagnostic methods are highly sensitive, they lack specificity to differentiate colonization from C. difficile infection (CDI). Mayo Clinic Arizona (MCA) uses enzyme immunoassay (EIA) testing for toxin A/B and glutamate dehydrogenase antigens with reflex to PCR for discordance as the primary algorithm for diagnosing CDI; however, GIPP PCR testing circumvents this algorithm. When used in patients with low pretest probability for CDI, clinical false positives can lead to inappropriate diagnosis and treatment of colonization. GIPA Results in the Electronic Medical Record Pre-Intervention MCA Standalone C. difficile Testing Algorithm Abbreviations – EIA, enzyme immunoassay; GDH, glutamate dehydrogenase; NAAT, nucleic acid amplification test Methods We convened a multidisciplinary quality improvement team to reduce the detection and treatment of C. difficile colonization secondary to GIPP test results. Surveys were sent to providers and nurses as part of a formal gap analysis and included questions around ordering patterns, knowledge of appropriate tests, and associated costs. Baseline data was compiled from GIPP orders in MCA from 1/1/2022-9/30/2022. Results There were 1978 GIPP orders for 1694 patients during the study period (∼7 GIPPs/day). Most orders (65%) originated from ambulatory settings. The most common positive target was C. difficile (355/723, 49%), which resulted in treatment for suspected CDI in 94.9% (337/355) of cases. Of the 49 providers who responded to the survey, 86% ordered the GIPP at least monthly. Testing for C. difficile was the 2nd most common reason for ordering the GIPP. Most providers underestimated the cost of the panel and were unfamiliar with the differences between the GIPP and EIA testing algorithm. Nurses sought clarification on stool samples suitable for GIPP testing. Following education and modifications to ordering in the electronic health record, the decision was made to remove the C. difficile target from the GIPP. Provider Understanding of C. difficile Testing Methods GIPA Results in the Electronic Medical Record Post-Intervention Conclusion Multiplex PCR panels are comprehensive but require careful implementation to facilitate diagnostic stewardship. Facilities with robust standalone C. difficile testing algorithms may consider removing the C. difficile target from the GIPP. Disclosures All Authors: No reported disclosures

Background Recruitment for clinical studies is challenging. To overcome barriers, investigators have previously established call-to-entry rates to assist in planning. However, rates specific to low-income minority populations are needed to account for additional barriers to enrolment these individuals face. Objective To obtain a call-to-entry rate in a low-income uninsured Hispanic population with chronic disease. Methods We used data from four of our randomised clinical studies to determine the call-to-entry rate for individuals (n=1075) with or at risk for type 2 diabetes: participants needed/potential participants contacted=recruitment rate (yield). Research staff contacted potential participants to enrol in a study that evaluated 6 month diabetes programmes at community clinics from 2015 to 2020. We recorded call-to-entry rates, reasons for declining the study, show rates, and attrition. Results The call-to-entry rate was 14.5%. Forty per cent of potential participants could not be contacted, and 30.6%, 19.1%, and 5.4% responded yes , no , and maybe , respectively. No show percentages were 54% for yes and 91.4% for maybe responders. The majority (61.6%) declined due to inability to attend; reasons to decline included work (43%), eligibility (18%), transportation (10%), out of town (9%), did not think they needed the programme (7%) and other/unknown (14%). Being a physician predicted inability to reach participants (adjusted OR 2.91, 95% CI 1.73 to 4.90). Attrition was 6.8%. Conclusions We described a call-to-entry rate and detailed recruitment data, including reasons to decline the study. This valuable information can assist investigators in study planning and overcoming enrolment barriers in low-income populations. Telehealth-based or strategies that limit transportation needs may increase participant involvement. Trial registration number NCT03394456 .

518 Background: Oncology has entered the digital era, accompanied by changes in patient interactions and expectations involving telemedicine. While much emphasis is appropriately placed on digital interactions with patients, little is known about modern non-digital interactions with patients in oncology practices. Methods: We analyzed Medical Oncology Pager (MOP) data from 7/1/18 – 4/14/23, during which patients had access to the electronic health record and messaging capabilities. The MOP for Mayo Clinic (Rochester) is covered by the medical oncology fellows, including after hours (6PM – 7AM). It covers consults and direct patient calls after hours and during weekends, in addition to critical results back up and inpatient oncology matters at all times. We also analyzed Medical Oncology Call Center (MOCC) data since 6/1/22, which takes inbound patient calls during working hours (7AM – 6PM) and is staffed by appointment office employees. Results: The MOP received a total of 17,679 pages during the analyzed period for a mean of 10.1 pages per day. Of these, 6,319 pages occurred after hours. The most after-hours pages were received on Fridays (n=1,091) and the least on Sundays (n=701); of those years with fully available data, 2022 had the highest volume (n=1,405) (Table 1). When comparing the MOP activity pre-pandemic (7/1/18 – 3/10/20) and intra-pandemic (3/11/20 – 4/14/23), the mean number of after-hours pages per day was relatively stable with 3.47 pre-pandemic and 3.69 intra-pandemic. Since 6/1/22, the MOCC has averaged 6,464 inbound calls per month during working hours, each of which had an average talk time of approximately five minutes. The majority of inbound calls occur on Monday mornings between 9 – 11AM, with over 6,500 recorded throughout the previous year. Conclusions: Telemedicine is multi-faceted, comprised of both digital and non-digital patient interactions. We show that many oncology patients utilize telephone communication with the healthcare system despite increased availability of digital capabilities. These data have implications for addressing the digital divide for those with limited digital access, serving older patient populations unfamiliar with digital technologies, healthcare navigation, trainee responsibilities, and resource allocation. Further work and standardization in non-digital telemedicine is necessary as it remains a pillar of modern healthcare delivery and oncology practices.[Table: see text]

Aim. To evaluate retinal vascular perfusion and density by optical coherence tomography angiography (OCTA) before, during, and after hypoglycemia in individuals with diabetes mellitus with or without diabetic retinopathy (DR). Methods. A focused clinical history was performed, followed by an ophthalmological examination to document retinopathy status. OCTA was performed at baseline, at hypoglycemia, and at glucose normalization. Eye tracking and eye alignment devices on the platform were used to obtain a macular thickness cube (512 × 128) and vascular perfusion and density protocols of 3 × 3 mm. Retinal vascular reactivity was analyzed with superficial plexus vascular perfusion and density protocols on OCTA. Results. Fifty-two participants encompassing 97 eyes fulfilled the eligibility criteria. Their mean age was 42.9 ± 15.1 years (range, 22 to 65), and 20 (38.2%) were men. We found a statistically significant difference in vascular perfusion and density when comparing all groups at baseline. The controls had higher vascular perfusion and density values than the cases. Vascular perfusion and density were significantly reduced in all groups during the hypoglycemia episode, except for vascular density in DR cases. Conclusion. Acute hypoglycemia significantly alters the retinal vascularity in DM patients with and without DR, suggesting that repeated episodes of acute hypoglycemia could exacerbate retinopathy in the long term.

#LGBTQ+ people with cancer are invisibilized: A call for #intersectional cancer research (link here) #healthequity

Total knee arthroplasty (TKA) is a treatment option for individuals with symptomatic osteoarthritis who have failed conservative therapy. In this manuscript the authors describe the pathophysiology of post-operative edema and explore the patient dependent factors potentially contributing to lymphatic dysfunction and thus directly influencing the TKA postoperative course. A proposed multimodal perioperative protocol is presented that focuses on identifying limb edema/lymphedema preoperatively, intraoperative technique changes that may decrease swelling post-TKA.