Ruff, Michael W.

Abstract PURPOSE Standard of care for craniopharyngiomas is surgery with or without radiotherapy (RT). Cohort A of Alliance A071601 evaluated the efficacy of BRAF/MEK inhibition with vemurafenib/cobimetinib in patients with previously untreated papillary craniopharyngiomas (PCP), which carry the BRAF V600E mutation. Cohort B is currently enrolling patients with recurrence after RT. In a correlative analysis, we examined changes in RT volumes after BRAF/MEK therapy in Cohort A. METHODS Previously unirradiated patients with BRAF-mutated PCP were treated with vemurafenib/cobimetinib. Sixteen patients had scans available before starting vemurafenib/cobimetinib (“pre-therapy”) and after completing therapy (“post-therapy”). Two patients went off study treatment after 8 and 9 days due to side-effects and were excluded for this analysis. Gross target volumes (GTV) were contoured on pre-therapy and post-therapy scans. On post-therapy scans, an additional target comprising gross disease and at-risk regions for microscopic residual disease (GTV-micro) was defined and considered the treatment volume. Clinical target volume (CTV) was a 5-mm uniform expansion on pre-therapy GTV and post-therapy GTV-micro. Volumes were independently reviewed by two radiation oncologists. Changes in volumes from pre- versus post-therapy were compared using the Wilcoxon signed rank test. RESULTS In 14 patients evaluated, 57% were female and median age at enrollment was 49.5 years (range 33-83). Median time on treatment was 8.9 months (range 4.0-18.0). Median GTV pre-therapy was 3.8 mL (range 0.2-23.4) versus 0.3 mL (range 0.0-3.2) post-therapy (p=0.0001) and 1.7 mL (range 0.1-8.0) post-therapy GTV-micro (p=0.0001). Median CTV pre-therapy was 13.7 mL (range 2.8-51.8) versus 9.1 mL (range 2.2-27.5) post-therapy (p=0.0001). All tumors abutted the optic chiasm pre-therapy, only 6 did post-therapy. CONCLUSIONS Vemurafenib/cobimetinib resulted in smaller RT volumes. BRAF/MEK inhibitors could reduce RT volumes and spare dose to surrounding normal structures. Enrollment to Cohort B of Alliance A071601 should be considered for patients with recurrent tumors after RT. SUPPORT https://acknowledgments.alliancefound.org

Abstract Medulloblastoma is a posterior fossa tumor rarely diagnosed in adults. Treatment includes craniospinal irradiation (CSI). Proton CSI is increasingly utilized to decrease radiation dose to normal tissues, despite the lack of randomized data demonstrating decreased toxicity compared to photon CSI. This single institution retrospective study of 39 adult medulloblastoma patients includes nineteen patients treated with photon CSI prior to 2015, and twenty treated with proton CSI thereafter. Median age was 28 years (range 18-66). Molecular subtype was most commonly sonic hedgehog (68%). The most common fractionation schedule was 36 Gy CSI in 20 fractions (85% of photon and 58% of proton patients) with a boost to 54-55.8 Gy (92%). Proton CSI delivered significantly lower mean doses to the cochleae (median 32 Gy vs. 44 Gy), lacrimal glands (8 vs. 36 Gy), lens (2 vs. 28 Gy), parotid glands (3 vs. 26 Gy), pharyngeal constrictors (6 vs. 15 Gy), esophagus (2 vs. 29 Gy), heart (0 vs. 14 Gy), lungs (1 vs. 7 Gy), liver (0.1 vs. 7 Gy), and skin (38 vs. 51 Gy) (all p< 0.001). Patients receiving proton CSI had significantly lower rates of acute dysphagia of any grade (5% vs. 35%, p= 0.044) and decreased median weight loss during radiation (+1.0 vs. -2.8 kg, p= 0.011). Acute hospitalization was associated with increased weight loss (p= 0.009). Median follow-up was 2.9 and 12.9 years for proton and photon patients, respectively, limiting late toxicity and outcome comparisons. At last follow-up five photon patients had died (two of progressive disease, three without recurrence ages 41-63) and 21% had experienced major cardiovascular events. At 10 years, 89% were alive and 82% were recurrence free. In conclusion, this study demonstrates dosimetric improvements with proton CSI, potentially leading to decreased acute toxicity including dysphagia and weight loss during treatment.

Abstract While dose escalation of radiotherapy (DERT) has failed to improve overall survival (OS) or progression-free survival (PFS) for glioblastoma in previous studies, a recent phase II clinical trial utilizing 18F-DOPA-PET-directed DERT demonstrated improved PFS in MGMT-unmethylated patients and OS in MGMT-methylated patients compared to historical controls. This planned secondary analysis sought to determine 1) how 18F-DOPA-PET changes RT volumes beyond standard MRI-planning, 2) which patients benefit most and least from this protocol, 3) which are mostly likely to experience clinically significant radionecrosis after DERT, and 4) patterns of failure after DERT. For 69 evaluable patients, median MRI-defined, PET-defined, and combined low-dose gross tumor volumes (GTV51) were 54 cc (range 9-248), 23 cc (0.4-179), and 62 cc (10-260), respectively. Median MRI-defined, PET-defined, and combined high-dose GTVs (GTV76) were 32 cc (range 4-136), 6 cc (0.1-138), and 34 cc (4-162), respectively. 18F-DOPA-PET resulted in a median volumetric expansion of 13% (0-243%) and 5% (0-217%) from MRI-defined low-dose and high-dose GTV’s, respectively. Central failures ( >95% of recurrence tumor volume) occurred within the 76 Gy, 60 Gy, and 51 Gy isodose lines in 32 (46%), 60 (87%), and 64 (93%) patients, respectively. Recursive partitioning analysis stratified patients by OS and PFS. Patients with 18F-DOPA-PET-defined GTV76 > 7.8cc, MRI-defined GTV76 > 42.7cc, and MGMT promotor-unmethylated tumors had the shortest OS, while patients with smaller PET and MRI-defined tumors had the longest OS (median 10.4 vs. 64.6 months, p< 0.001). Similarly, PFS was worst in patients with 18F-DOPA-PET-defined GTV76 > 2.17 cc who had biopsy only (median PFS 3.2 months, p< 0.001). Patients with 18F-DOPA-PET-defined GTV51 > 50 cc had the highest risk of grade 3+ radionecrosis (p< 0.001). In conclusion, larger 18F-DOPA-PET and MRI-defined tumor volumes were associated with worse outcomes, and 18F-DOPA-PET-directed DERT appears to reduce risk of central recurrence in high-dose volumes.

Abstract Although chimeric antigen receptor T (CART)–cell therapy has been successful in treating certain hematologic malignancies, wider adoption of CART-cell therapy is limited because of minimal activity in solid tumors and development of life-threatening toxicities, including cytokine release syndrome (CRS). There is a lack of a robust, clinically relevant imaging platform to monitor in vivo expansion and trafficking to tumor sites. To address this, we utilized the sodium iodide symporter (NIS) as a platform to image and track CART cells. We engineered CD19-directed and B-cell maturation antigen (BCMA)–directed CART cells to express NIS (NIS+CART19 and NIS+BCMA-CART, respectively) and tested the sensitivity of 18F-TFB-PET to detect trafficking and expansion in systemic and localized tumor models and in a CART-cell toxicity model. NIS+CART19 and NIS+BCMA-CART cells were generated through dual transduction with two vectors and demonstrated exclusive 125I uptake in vitro. 18F-TFB-PET detected NIS+CART cells in vivo to a sensitivity level of 40,000 cells. 18F-TFB-PET confirmed NIS+BCMA-CART-cell trafficking to the tumor sites in localized and systemic tumor models. In a xenograft model for CART-cell toxicity, 18F-TFB-PET revealed significant systemic uptake, correlating with CART-cell in vivo expansion, cytokine production, and development of CRS-associated clinical symptoms. NIS provides a sensitive, clinically applicable platform for CART-cell imaging with PET scan. 18F-TFB-PET detected CART-cell trafficking to tumor sites and in vivo expansion, correlating with the development of clinical and laboratory markers of CRS. These studies demonstrate a noninvasive, clinically relevant method to assess CART-cell functions in vivo.

e19556 Background: CD19 chimeric antigen receptor T cell therapy possesses unique side effects including cytokine release syndrome (CRS) and immune effector cell associated neurotoxicity syndrome (ICANS). Age is a major risk factor for ICANS. However, whether ICANS in older patients is different compared to younger patients is unknown. Herein, we report clinical course, outcomes and risk factors for ICANS in older patients with large B cell lymphoma (LBCL) treated with axicabtagene ciloleucel (axi-cel). Methods: We comprehensively reviewed detailed clinical courses of ICANS in 78 adult patients with LBCL treated with axi-cel between June 2016 and October 2020. Incidence, manifestation, risk factors, treatment, and outcomes of ICANS were compared between patients age ≥60 (n=32) and <60 (n=46) years old. Results: Baseline characteristics were comparable between older and younger patients except higher proportion of high international prognostic index and underlying cerebral microvascular disease in older patients. ICANS was observed in 16 patients in the older and 24 patients in the younger age group, with a 30-day incidence of 52% and 50%, respectively. Median time to CRS and ICANS were similar between 2 age groups. The most common initial neurological findings included aphasia, dysgraphia and encephalopathy in both age groups. Table summarizes the characteristics, clinical course and interventions of ICANS in older and younger patients. In Cox regression model, the presence of CRS was the only factor associated with ICANS in both age groups. Age, history of central nervous system involvement and cerebral microvascular disease were not associated with ICANS. Importantly, all patients had complete resolution of ICANS. No elderly patients in our cohort experienced seizure as a manifestation of ICANS. Conclusions: In our study, older age was not a risk factor for ICANS. CRS was the only factor associated with ICANS in both younger and older patients. Incidence, clinical course and neurological outcomes of ICANS in older patients treated with axi-cel were comparable to younger patients. [Table: see text]

2000 Background: Craniopharyngiomas, a rare brain tumor along the pituitary-hypothalamic axis, can cause significant clinical sequelae. Surgery and radiation, the only effective treatments, can cause significant morbidity. Genetic analysis of craniopharyngiomas revealed that 95% of papillary craniopharyngiomas (PCP) have BRAF V600E mutations (Brastianos et al. Nature Genetics 2014). We evaluated the efficacy of BRAF/MEK inhibition in patients (pts) with previously untreated PCP. Methods: Eligible pts without prior radiation whose PCP screened positively for BRAF mutations were treated with oral vemurafenib/cobimetinib in 28-day cycles. The primary endpoint of response rate (RR) based on centrally determined volumetric data was evaluated in 16 pts, where a partial response was defined as >20% decrease in volume. This single arm, Simon two-stage phase 2 trial had 89% power to detect a true RR of at least 30% (vs. the null RR 5%; alpha=0.04). In this design, 3 or more observed volumetric responses in 16 evaluable pts would be considered promising activity. Results: In the 16 pts evaluated, 56% were female, and the median age was 49.5 years. Median follow-up was 22 months (95% CI: 16-26.5) and median number of treatment cycles was 8. Three patients progressed after therapy was discontinued and none have died. Based on volumetric response criteria, 14 of 15 pts with volumetric data available for central review had response to therapy (93%; 95% CI: 68% to 99.8%). Of 16 patients evaluable based on local review, 15 had response to therapy (93.75%; 95% CI: 70% to 99.8%). The median tumor reduction was -83% (range: -52% to -99%). The one nonresponder received 2 days of treatment before coming off therapy due to toxicity. Median progression-free survival was not reached. Grade 3 toxicities at least possibly related to treatment occurred in 12 pts (rash in 6 pts). Grade 4 toxicities were observed in two pts: hyperglycemia (n=1) and increased CPK (n=1). Three pts discontinued treatment for adverse events. Conclusions: Vemurafenib/cobimetinib resulted in an objective response in all pts who received 1 or more cycles of therapy. Our study indicates that BRAF/MEK inhibitors could be a powerful tool in the treatment of previously untreated PCP and warrants further evaluation in larger studies. A second arm of this study is enrolling pts with progressive PCP after prior radiotherapy. Support: U10CA180821, U10CA180882; U24CA196171, U10CA180868 (NRG); Genentech; https://acknowledgments.alliancefound.org. Clinical trial information: NCT03224767.

Abstract Study Objective To determine whether there is a drug‐drug interaction precluding the concomitant use of levetiracetam and high‐dose methotrexate (HDMTX). Design Retrospective analysis. Setting Large academic tertiary care medical center. Patients Adult lymphoma patients who received HDMTX as a 4‐h infusion with or without concomitant levetiracetam. Measurements and Main Results Generalized estimating equations clustered on patient were used to assess each outcome. The primary outcome was the incidence of delayed MTX elimination (MTX level >1 µmol/L at 48 h). Secondary outcomes included incidence of acute kidney injury (AKI) and hospital length of stay (LOS). The 430 included patients receiving 1993 doses of HDMTX had a median (IQR) age of 66 (57.5, 72.6) years, 88 (20.5%) received concomitant levetiracetam with at least one dose of MTX, 267 (62.1%) were male, and 397 (92.3%) were Caucasian. HDMTX doses ranged from 1 to 8 g/m 2 . The most common lymphoma diagnoses were systemic diffuse large B‐cell lymphoma (DLBCL; 58.5%) and systemic DLBCL with central nervous system (CNS) involvement (32.8%). Rates of delayed elimination with and without levetiracetam were 13.4% and 16.3%, respectively (OR = 0.80, 95% CI 0.47–1.34, p = 0.39). AKI occurred in 15.6% and 17.0% of patients with and without concomitant levetiracetam, respectively (OR = 0.83, 95% CI 0.52–1.33, p = 0.28). The median LOS with and without levetiracetam was 4.2 and 4.1 days, respectively ( p = 0.039). On multivariable analyses, only age, body surface area, diagnosis of systemic DLBCL with CNS involvement, serum creatinine, hemoglobin, total bilirubin, and dose of HDMTX were associated with delayed elimination. Conclusions High‐dose methotrexate administered with concomitant levetiracetam was not associated with increased risk for delayed MTX elimination or AKI. These results support that levetiracetam and HDMTX are safe for coadministration.

Posterior fossa (PF) diffuse gliomas in pediatric patients frequently harbor the H3 K27M mutation. Among adults, PF diffuse gliomas are rare, with limited data regarding molecular features and clinical outcomes. We identified 28 adult PF diffuse glioma patients (17 males; median: 50 y, range: 19 to 78 y), with surgery performed at our institution (13 brainstem; 15 cerebellum). Histologic subtypes included anaplastic astrocytoma (n=21), glioblastoma (n=6), and diffuse astrocytoma (n=1). Immunohistochemistry was performed for H3 K27M (n=26), IDH1-R132H (n=28), and ATRX (n=28). A 150-gene neuro-oncology-targeted next-generation sequencing panel was attempted in 24/28, with sufficient informative material in 15 (51.7%). Tumors comprised 4 distinct groups: driver mutations in H3F3A (brainstem=4; cerebellum=2), IDH1 (brainstem=4; cerebellum=4), TERT promotor mutation (brainstem=0; cerebellum=3), and none of these (n=5), with the latter harboring mutations of TP53 , PDGFRA , ATRX , NF1 , and RB1 . All TERT promoter–mutant cases were IDH-wild-type and arose within the cerebellum. To date, 20 patients have died of disease, with a median survival of 16.3 months, 1-year survival of 67.5%. Median survival within the subgroups included: H3F3A =16.4 months, IDH mutant=113.4 months, and TERT promoter mutant=12.9 months. These findings suggest that PF diffuse gliomas affecting adults show molecular heterogeneity, which may be associated with patient outcomes and possible response to therapy, and supports the utility of molecular testing in these tumors.

Chimeric antigen receptor T (CART) cells are engineered with an artificial receptor which redirects T cells to recognize cancer cells expressing a particular surface antigen. CART cell therapy has been astonishingly successful at eradicating certain B cell malignancies, but relapse is common, and efficacy is lacking in many cancers. Gene editing of CART cells is being investigated to enhance efficacy and safety and to develop off-the-shelf products. Currently, genome engineering tools used to modify CART cells include zinc finger nucleases, transposons, TALENs, and CRISPR-Cas9. CRISPR-Cas9 uses a trans-activating (tracrRNA): CRISPR RNA (crRNA) duplex to trigger imprecise DNA repair through targeted double stranded breaks, causing indels and often resulting in loss of protein function. Gene-edited CART cells have entered the clinic to provide an allogeneic cell source (TALEN TCRα knockout), safer treatment (CRISPR-Cas9 GM-CSF knockout), and resistance to exhaustion (CRISPR-Cas9 PD-1 knockout). CRISPR-Cas9 PD-1 knockout (PD-1k/o) CART cells were well-tolerated in a first-in-human clinical trial. However, clinically tested CRISPR-Cas9-edited CART cells showed only modest loss of function (~25%) of PD-1 upon infusion. Additionally, off-target editing has been observed in the clinic and remains a concern. We hypothesized that using next-generation CRISPR-Cas12a systems will result in enhanced editing efficiency and precision. CRISPR-Cas12a has a smaller protein component than CRISPR-Cas9, uses a single crRNA without a tracrRNA for simplified delivery and leaves staggered 5' overhangs. These properties, along with lower intrinsic off-target activity than Cas9, render Cas12a a powerful gene editing tool. First, we compared the knockout efficiency of Cas9 and Cas12a in three therapeutically relevant genetic targets in T cells by delivering ribonucleoprotein complexes containing the crRNA and Cas protein of interest. We showed that Cas12a more effectively knocked out CD3, GM-CSF, and PD-1 expression compared to Cas9 (Figure 1A), demonstrating the potential of Cas12a in further genetically editing T cell therapies. We then used electroporation with Cas9 and Cas12a to generate PD-1k/o in lentivirally transduced CD19-targeted CART (CART19) cells with the aim of making exhaustion-resistant CART19 cells through CRISPR gene editing. CART19 and PD-1k/o CART19 cells were repeatedly stimulated with CD19+ NALM6 target cells for one week, and exhaustion marker expression was measured over time with flow cytometry. The expression of CTLA4, TIM3, and LAG3 were similar between CART19 groups, but PD-1 expression was lower in Cas9 PD-1k/o CART19 cells and almost completely eradicated in Cas12a PD-1k/o CART19 cells compared to wildtype or mock shocked CART19 cells (Figure 1B). We then compared the functionality of wildtype, mock shocked, and Cas9 or Cas12a PD-1k/o CART19 cells in vitro to ensure that neither the electroporation process nor PD-1 knockout impaired CART19 cell antitumor activity. Over a range of effector-to-target ratios and with repeated stimulation with target cells, cytotoxicity was comparable across all CART19 cell groups (Figure 1C). All CART19 cell groups demonstrated robust proliferation in response to both nonspecific and antigen-specific stimulation and over one week of repeated antigen stimulation with NALM6 target cells (Figure 1D). We also confirmed that all CART19 cell groups demonstrated strong degranulation and cytokine production in response to nonspecific and antigen-specific stimulation, regardless of electroporation or PD-1 knockout (Figure 1E). In summary, our data demonstrate that Cas12a can be used as a gene editing tool to efficiently knock out therapeutically relevant genes in CART19 cell therapy. Additionally, Cas12a demonstrated improved knockout efficiency over Cas9 in three different genomic targets. PD-1 knockout via Cas9 or Cas12a reduced PD-1 expression on the CART19 cell surface, and PD-1 expression was almost completely ablated with Cas12a gene editing. Electroporation and PD-1 knockout did not impact the effector functions of the CART19 cells, including cytotoxicity, degranulation, cytokine secretion, or proliferation. In vivo studies assessing the antitumor efficacy and CART19 cell persistence are ongoing. Overall, Cas12a is a promising, efficient method of gene knockout to enhance the safety and efficacy of CART cells. Disclosures Sakemura: Humanigen: Patents & Royalties. Cox:Humanigen: Patents & Royalties. Kenderian:MorphoSys: Research Funding; Sunesis: Research Funding; Tolero: Research Funding; BMS: Research Funding; Juno: Research Funding; Gilead: Research Funding; Kite: Research Funding; Novartis: Patents & Royalties, Research Funding; Torque: Consultancy; Humanigen: Consultancy, Patents & Royalties, Research Funding; Mettaforge: Patents & Royalties; Lentigen: Research Funding.

Abstract Improvements in treatments and prognosis in glioma patients has led patients the opportunity for pregnancy despite their diagnosis. In this retrospective case series of 13 patients evaluated at Mayo Clinic, we identified patients who were pregnant at or after glioma diagnosis from 1998-2019. Clinical information was obtained through the electronic medical record. Tumor grade, treatments, peripartum complications, and pediatric outcomes were analyzed. Median follow-up for pregnant patients was 13.5 years (range, 0-24). Of the 13 patients, 2 patients received temozolomide and radiation during pregnancy for glioblastoma and had no peripartum complications. Median survival time in these patients after further treatments was 23.5 months (range, 23.3-23.8). Two children from these pregnancies had no fetal malformations or pediatric developmental delays with median follow-up of 8.8 years (range, 4.7-12.8). Three patients received radiation during pregnancy for high grade gliomas and had no fetal malformations or peripartum complications. Eight patients had one or multiple pregnancies with median 2 (range, 1-3) children at or after diagnosis with 14 children total. Eleven of the 14 children had no fetal malformations or peripartum complications and were conceived without reproductive assistive technologies. Two children, who were identical twins, were born with a placental malformation but otherwise had normal pediatric development. One child was born pre-term and had long-term developmental delays.