Witzig, Thomas E.

Abstract Lymphoma is the sixth most commonly diagnosed cancer in the US. Genome-wide association studies have identified common variants associated with risk of specific lymphoma subtypes, but less is known about the contribution of rare inherited variants in the genetic architecture of lymphoma risk. CHEK2 is important to DNA repair and 2 small studies have found evidence of an association between CHEK2 variants and risk of lymphoma. Here, we investigated loss of function (LoF) variants in CHEK2 with risk of lymphoma (overall and subtypes). The study population included newly diagnosed lymphoma cases from Mayo who were enrolled in the Lymphoma SPORE Molecular Epidemiology Resource (MER). Controls were from the Mayo Clinic Biobank, from which we excluded a prior hematologic malignancy. Whole exome sequencing was performed by Regeneron on an Illumina NovaSeq panel (mean coverage of 48X). Variants were called using GATK v4 and variant annotation was performed using BioR. All LoF variants (nonsense, frameshift and consensus splice sites) in CHEK2 with a minor allele frequency < 0.5% were included in the analyses. Logistic regression was used to estimate odds ratio (OR) and 95% confidence intervals (CI) for the association between CHEK2mutation status and risk of lymphoma overall, and by lymphoma subtypes that had more than 5 mutation carriers, which included diffuse large B-cell (DLBCL), follicular (FL), and T-cell lymphoma (TCL). All analyses were adjusted for age (at diagnosis for cases and at enrollment for controls) and sex. A total of 4,852 lymphoma cases and 49,724 controls were included in this analysis. Median age for both cases and controls was 62 years (range 18-99 years). Males accounted for 57.5% (n=2,789) of the cases and 40.6% (n=20,186) of the controls. The DLBCL (23.6% of cases) and FL (23.3% of cases) subtypes were the most common; 7.6% of cases had TCL. A total of 407 (0.7%) individuals had a LoF variant. The frequency of LoF variants was 1.2% in lymphoma cases and 0.7% in controls. LoF variants were associated with increased risk of lymphoma overall (OR=1.77; 95%CI: 1.32-2.33), DLBCL (OR=2.07; 95%CI: 1.20-3.13), and TCL (OR=2.78; 95%CI: 1.178-5.50), but not FL (OR=1.31; 95%CI: 0.65-2.34). CHEK2 c.1100delC was the most frequently mutated variant in this population, accounting for 76% of all LoF variants. Individuals with a c.1100delC variant had an increased risk of lymphoma overall (OR=1.90; 95%CI: 1.37-2.58), DLBCL (OR=2.06; 95%CI: 1.09-3.54), and TCL (OR=3.17; 95%CI:1.24-6.58), but not FL (OR=1.05; 95%CI: 0.41-2.15). When restricted to cases, there was no significant difference in age at diagnosis (P=0.62) or sex (P=0.53) between LoF variant carriers and non-carriers. In this large lymphoma case-control study, CHEK2 LoF variants were associated with increased risk of lymphoma, demonstrating that rare inherited variants may play an important role in the etiology of lymphoma. Additional work is needed to investigate missense variants in CHEK2 and risk of lymphoma. Citation Format: Nicholas J. Boddicker, Raphael Mwangi, Dennis P. Robinson, Allison C. Rosenthal, Thomas M. Habermann, Andrew L. Feldman, Lisa M. Rimsza, Rebecca L. King, Melissa C. Larson, Brianna J. Gysbers, Stephen M. Ansell, Jithma P. Abeykoon, Grzegorz S. Nowakowski, Thomas E. Witzig, Anne J. Novak, Susan L. Slager, James R. Cerhan. Germline CHEK2 variants and risk of lymphoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5233.

e14594 Background: Voyager-V1 (VV1) is an oncolytic vesicular stomatitis virus engineered to express interferon beta (IFNβ) to boost cellular antitumor immunity and sodium iodine symporter (NIS). A phase 1 trial investigating a single infusion of VV1 in patients with relapsed refractory multiple myeloma and T-cell lymphoma (TCL) showed promising and durable clinical activity, primarily in patients with TCL. VV1 infects tumor cells and macrophages in the lymph nodes and spleen, resulting in release of tumor antigens and IFNβ that activates and boost cellular antitumor immunity. We present the preliminary pharmacokinetic analysis in patients given one intravenous (IV) dose of VV1. Methods: Patients received 5e9 to 1.7e11 TCID50 (50% tissue culture infective dose) of VV1 given over 30 minutes. We also explored the impact of infusion duration (0.25, 0.5,1 and 3 hours) in 17 patients with solid tumors. The objectives were to determine the impact of viral dose on pharmacokinetics (PK) of VSV and IFNβ. For VSV and IFNβ PK, whole blood was collected in 30 patients on days 1, 2, 3, and 8. Dose levels were 1.7E10, 5E10, and 1.7E11 TCID50. The plasma concentration-time data were estimated with non-compartmental analysis using WinNonlin 8.3. Results: The half-life of VV1 is comparable (10-12h) between dose levels of 1.7e10, 5e10 and 1.7e11. Ideal infusion rate was found to be 30 minutes and provides the highest Cmax and AUC. Most peak concentrations for viremia and IFNβ were achieved between ‘end of infusion’ and 2.5 hours, 4.5 and 24 hours, respectively. Following infusion, a second concentration peak is found in over half the patients leading to highly variable terminal elimination half-life, specifically where the second peak is at 24 hours. There was no second peak for IFNβ levels. Peak plasma concentration and systemic exposure increased in proportion to increased dose, consistent with linear clearance. Conclusions: The 30 minute infusion rate was considered the best as its PK provides the highest Cmax and AUC with no significant CRS differences among other groups. IL-6 and IFNα markers at hour 4 indicate a likely CRS and hour 24 provides a possible prediction of the grade. Cmax and AUC were also highest at 1.7e11 TCID50. This is the recommended dose that will be used in the expansion arm in patients with TCL. Clinical trial information: NCT03017820 , NCT02923466 . [Table: see text]

7543 Background: Total metabolic tumor volume (TMTV) and maximum standardized uptake value (SUVmax) may improve upon standard 18F-fluorodeoxyglucose positron emission tomography/computed tomography (PET-CT)-based assessment of advanced DLBCL during treatment. We report the association of TMTV and SUVmax changes at interim PET-CT (PET2) on survival outcomes in patients with primary refractory DLBCL (prDLBCL) and early relapsing DLBCL (erDLBCL). Methods: Adult patients with prDLBCL (less than a complete response (CR) or progression by the end of treatment (EOT)) or erDLBCL (CR at EOT with relapse within 12 months) between 2005 and 2019 at Mayo Clinic Rochester were included. The % change (Δ) from baseline to PET2 (after 2 cycles of 1L therapy) of TMTV and SUVmax were measured. A PET segmentation threshold of 1.5 liver mean SUV + two standard deviations with a minimum volume constraint of 0.5 mL was utilized (MIM Software, Inc., Cleveland OH, USA) with manual input as needed. Functional splines analysis determined SUVmax and TMTV threshold for analysis. Results: 131 patients had complete PET-CT data (N=79 prDLBCL, N=52 erDLBCL). Baseline median TMTV and SUVmax were 767.4 cm3 (range 0.9-6840.4) and 20.5 (2.0-52.6), respectively. At a median follow up of 77.6 months, 81 patients had died. 2-year OS rate was 53% (95% CI: 42-67) for patients with a PET2 ΔSUVmax decline ≥65% compared to 15% (95% CI: 7-29) for patients with a PET2 ΔSUVmax decline <65%(p<0.001). PrDLBCL had a 2-year OS rate of 48% (95% CI: 31-74) for a ΔSUVmax ≥65% compared to 7% (95% CI: 2-21) for a ΔSUVmax <65%(p<0.001), which captured 58% of prDLBCL patients (N=46). Two-year OS rate was 43% (95% CI: 34-54) for patients with a ΔTMTV ≥75% compared to 5% (95% CI: 1-35) for patients with ΔTMTV <75% (p<0.001). All patients with a ΔTMTV <75% had prDLBCL. PrDLBCL had a 2-year OS rate of 28% (95% CI: 18-44) for a ΔTMTV ≥75%. The outcomes for ΔSUVmax <65% and ΔTMTV <75% at PET2 remained significant in a cox regression model adjusted for IPI and bulky disease (>10cm)(Table). Conclusions: A ΔTMTV decline <75% or a ΔSUVmax decline <65% at PET2 identified an ultra-high-risk subgroup of DLBCL with particularly poor outcomes, that captured a high percentage of prDLBCL patients, and was significant after adjusting for IPI and bulky baseline disease. These patients may benefit from clinical trials evaluating alternative PET-adapted treatment strategies, but further evaluation among all patients with DLBCL is needed. [Table: see text]

7530 Background: Oncolytic virotherapy is a novel immunomodulatory therapeutic approach currently under study for relapsed refractory (R/R) hematologic malignancies. The engineered Vesicular Stomatitis Virus VSV-IFNβ-NIS used in this study encodes interferon beta (IFNβ) and sodium iodine symporter (NIS). Virally-encoded IFNβ serves as an index of viral proliferation and enhances host anti-tumor immunity. Herein, we report the updated results of the phase 1 clinical trial NCT03017820 of a one-time administration of systemic VSV-IFNβ-NIS among patients (pts) with R/R multiple myeloma (MM), T cell Lymphoma (TCL) and Acute myeloid Leukemia (AML) with particular focus on the TCL cohort treated at the highest dose level (DL4) without Ruxolitinib. Methods: VSV-IFNβ-NIS was administered as a single IV dose following a classical 3+3 phase I trial, starting at 5e9 TCID50 DL1 through 1.7e11 TCID50 DL4. A dose expansion cohort of 8 additional R/R TCL pts was added at DL4 in view of an initial signal of activity. The primary objective was to determine the maximum tolerated dose of VSV-IFNβ-NIS as a single agent. Secondary objectives were determination of safety profile and preliminary efficacy of VSV-IFNβ-NIS. Adverse events (AEs) are reported based on CTCAE V4; cytokine release syndrome (CRS) grading was based on Lee (Blood 2014) criteria. Results: To date, a total of 43 pts received VSV-IFNβ-NIS: MM (N=21), TCL (N=21) and AML (N=1). In the TCL cohort: at DL4 Group A (no Ruxolitinib), 12 pts were registered, but only 11 pts were treated (mycosis fungoides (MF) N=3; ALK- ALCL N=1; PTCL-NOS N=2; AITL N=4; Nodal PTCL, TFH N=2; ages 33-79). These pts received a median of 3 lines of therapy (range: 1-5). No dose limiting toxicities were observed. Grades 3 and higher AEs were hematologic with transient lymphopenia (63.6%), neutropenia (36.4%) being the most common. The non-hematologic AEs observed included unique toxicities such as CRS grades 1-2 (81.8%); the transient transaminitis grade 1-2 (36.4%) and grade 3 (27.3%) along with thrombocytopenia grade 1-2 (63.6%) and grade 3 (18.2%) were a reflection IFNβ toxicity. All the AEs observed were short lived. Responses were seen in pts with PTCL. At DL4, in pts with systemic PTCL (N=9), there were 5 responses including 3 complete responses (CR) with two long-lasting at 24 months, an ongoing 15 month CR, and a 5 month CR, and 2 partial responses (PR) lasting 5 months in one pts and 6 months in the other. Conclusions: These updated results of single agent VSV-IFNβ-NIS which include pts with PTCL treated at the highest dose level and a dose expansion cohort demonstrates not only the safety of VSV-IFNβ-NIS but also a strong efficacy signal in pts with PTCL with impressive durable responses. Two expansion cohorts at DL4 have been added (20 additional PTCL, 10 B-cell lymphoma pts) to obtain additional data on tolerability and efficacy and biomarkers of response in these pts. Clinical trial information: NCT03017820 .

Immune checkpoint inhibitors (ICIs) and brentuximab vedotin (BV) are novel agents for classic Hodgkin lymphoma, including relapse after autologous stem cell transplant (ASCT). However, their impact on survival post-ASCT relapse, in comparison with conventional therapy, is less known due to the lack of randomized controlled trials. Clinical characteristics and outcomes of 115 patients with relapse (or progression) after ASCT are studied. After a median follow-up of 8.59 years from post-ASCT relapse, the median progression-free survival (PFS) and overall survival (OS) were 0.91 and 5.07 years, respectively. Median lines of therapy after post-ASCT relapse was 2 (range, 1–12). The median PFS was not reached (NR) versus 1.11 versus 0.50 versus 0.85 versus 0.78 years (P = 0.006) and OS was NR versus 7.60 versus 3.08 versus 3.51 versus 3.17 years (P = 0.28) in patients first treated with ICIs versus BV versus investigational agents versus chemotherapy versus radiation therapy (RT). First-line treatment with novel agents (ie, ICIs and BV) was associated with superior outcomes compared with investigational agents and chemotherapy/RT with a median PFS of 1.65 versus 0.50 versus 0.79 years (P = 0.003) and a median OS of 7.60 versus 3.08 versus 3.32 years (P = 0.08). Regardless of lines of therapy, the treatment with ICIs had the most favorable outcome with a median PFS and OS of 3.98 and NR years, respectively. Allogeneic stem cell transplant (allo-SCT) was done in 23 patients (20%), and the median post-allo-SCT PFS and OS were 1.31 and 2.35 years, respectively. In conclusion, survival following post-ASCT relapse improves significantly when patients receive novel agents.

Abstract Introduction: Rapid cell proliferation requires intact and faithful DNA damage repair mechanisms. DNA polymerase theta (POLQ) plays a key role in repairing DNA double-strand breaks through the microhomology-mediated end-joining (MMEJ), which is one of the three main pathways involved in repairing replication-induced double-strand breaks. Limited data have suggested that concurrent depletion of POLQ and ataxia-telangiectasia mutated (ATM) could be embryonic lethal. Hence, this phenomenon has the potential to be exploited for therapeutic benefit in cancers where ATM mutations are commonly seen. Mantle cell lymphoma (MCL) is a non-Hodgkin lymphoma marked by (11;14) translocation with ATM alterations seen in 40-50% of patients. There is an unmet need to find novel therapeutic strategies, especially in relapsed and/or refractory (R/R) MCL. Here, we investigated whether targeting POLQ and ATM could be a potential therapeutic strategy in MCL. Methods: In vitro studies were conducted by using MCL cell lines. CRISPR-Cas9 system was used to genetically deplete POLQ and ATM genes and sgRNAs co-expressing fluorescence markers were used to track the cell population with respective genotypes over time. Cell viability was assessed by CellTiter-Glo assay and flow cytometry. All cell lines were profiled for ATM expression and activity. A p-value of &lt; 0.05 was considered statistically significant. The combination index of &lt;1 was defined as synergistic. Results: CRISPR-Cas9-mediated depletion of POLQ significantly decreased cell proliferation in multiple MCL cell lines. In particular, Granta-519, which possesses a single copy of kinase-dead ATM that is reduced in expression, was most sensitive to POLQ depletion. Concurrent genetic depletion of ATM and POLQ resulted in a synergistic antiproliferative effect in ATM-proficient MCL cell lines. Subsequently, this cellular phenotype caused by the genetic intervention was recapitulated by using two POLQ inhibitors (novobiocin and ART558) and an ATM inhibitor (AZD0156). In vitro, single-agent treatment with novobiocin or ART558 caused a significant cytotoxic effect at physiologically relevant concentrations in ATM-deficient cells and co-treatment of novobiocin or ART558 with AZD0156 was synergistic in killing ATM-proficient MCL cells. Importantly, POLQ inhibitors significantly decreased the cell viability of MCIR1, which is an ibrutinib-resistant MCL cell line. Mechanistically, novobiocin or ART558 treatment induced gH2AX and cleaved PARP upregulation, which was further enhanced by ATM depletion, suggesting that co-inhibition of POLQ and ATM caused apoptosis due to the accumulation of unrepaired DNA damage. Conclusion: POLQ is a promising target in MCL, especially in ATM-deficient setting. In ATM-proficient MCL, targeting ATM and POLQ is synergistic. Our data has the potential to uncover novel biomarker-driven drug therapy of POLQ inhibitors in R/R MCL. Citation Format: Jithma Prasad Abeykoon, Shuhei Asada, Kalindi Parmar, Xiaosheng Wu, Thomas Witzig, Geoffrey Shapiro, Alan D. D'Andrea. Targeting DNA polymerase theta and ATM leads to synergistic killing of mantle cell lymphoma cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6208.

Key Points Urinary TIMP2×IGFBP7 (uTIMP2×IGFBP7) concentrations experienced a rapid and sustained decline after high-dose methotrexate (MTX) exposure. uTIMP2×IGFBP7 kinetics and concentrations after high-dose MTX exposure demonstrated no utility in predicting AKI.

Abstract Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can result in a cytokine storm that is associated with poor outcomes in patients. The activation of a NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome following SARS-CoV-2 infection has been shown to play a major role in inflammatory immune responses. In this study, we show that open reading frame 8 (ORF8) protein is abundantly secreted as a glycoprotein in vitro. The glycosylated ORF8 stimulates human CD14+/CD16+ monocytes to cause upregulation of proinflammatory cytokine production and cell surface marker expression within 24 hours. The data suggests that stimulation of human monocytes by ORF8 is not receptor mediated. Rather, ORF8 is likely phagocytosed by human monocytes. ORF8 then stimulates the monocytes by independently binding to the NACHT and LRR domains of the NLRP3 protein. Pharmacologic inhibition of NLRP3 significantly diminished the production of pro-inflammatory cytokines from ORF8-stimulated human monocytes. Finally, this study shows that the ORF8 protein is also secreted in patients with newly diagnosed coronavirus disease 2019 (COVID-19). Levels of ORF8 in the blood of patients diagnosed with COVID-19 correlated with disease mortality and trajectory of disease. ORF8 stimulation of monocytes causes pro-inflammatory cytokine production that leads to the development of severe COVID-19.