Outcomes of proton therapy to infradiaphragmatic sites in pediatric patients with Hodgkin lymphoma.

BACKGROUND: Proton therapy (PT) has potential advantages in pediatric Hodgkin lymphoma (pHL). However, there are limited data on PT, specifically to infradiaphragmatic targets. We report on PT planning details, doses achieved to organs at risk (OARs), and clinical and toxicity outcomes for patients with pHL who received PT to infradiaphragmatic regions. METHODS: This is a retrospective study including patients treated between 2011 and 2022. Demographic and clinical factors were collected, and toxicity was reported using Common Terminology Criteria for Adverse Events (CTCAE) version 5.0. Dosimetric and clinical factors associated with key outcomes were assessed via Cox regression. Photon plans were generated for all patients, and the paired t-tests or Wilcoxon signed rank sum tests were used for dosimetric comparisons. RESULTS: Twenty-one patients comprising 22 PT courses were included. Median follow-up was 5.0 years, and mean age was 14.2 years. Median dose was 21 Gray equivalent (GyE) over 14 fractions. Top acute grade 1 (G1) toxicities included fatigue (59%) and anorexia (36%). Rates of acute G2 and G3+ toxicity were 18% and 0%, respectively. After PT, no local or marginal failures occurred. Five percent experienced disease progression, who were all successfully salvaged, and all patients were alive and disease-free at last follow-up. No secondary malignancies developed. Compared to photon radiotherapy, PT achieved significantly lower doses to the bowels, stomach, spleen, pancreatic tail, liver, kidneys, and pelvic bones. CONCLUSIONS: PT is well-tolerated and leads to excellent oncologic and toxicity outcomes with long-term follow-up. PT confers dosimetric advantages when compared to photons.

The transcription factor NR4A1 (Nur77) controls bone marrow differentiation and the survival of Ly6C- monocytes.

The transcription factors that regulate differentiation into the monocyte subset in bone marrow have not yet been identified. Here we found that the orphan nuclear receptor NR4A1 controlled the differentiation of Ly6C- monocytes. Ly6C- monocytes, which function in a surveillance role in circulation, were absent from Nr4a1-/- mice. Normal numbers of myeloid progenitor cells were present in Nr4a1-/- mice, which indicated that the defect occurred during later stages of monocyte development. The defect was cell intrinsic, as wild-type mice that received bone marrow from Nr4a1-/- mice developed fewer patrolling monocytes than did recipients of wild-type bone marrow. The Ly6C- monocytes remaining in the bone marrow of Nr4a1-/- mice were arrested in S phase of the cell cycle and underwent apoptosis. Thus, NR4A1 functions as a master regulator of the differentiation and survival of 'patrolling' Ly6C- monocytes.

NR4A1 (Nur77) deletion polarizes macrophages toward an inflammatory phenotype and increases atherosclerosis.

RATIONALE: NR4A1 (Nur77) is a nuclear receptor that is expressed in macrophages and within atherosclerotic lesions, yet its function in atherosclerosis is unknown. OBJECTIVE: Nur77 regulates the development of monocytes, particularly patrolling Ly6C(-) monocytes that may be involved in resolution of inflammation. We sought to determine how absence of nuclear receptor subfamily 4, group A, member 1 (NR4A1) in hematopoietic cells affected atherosclerosis development. METHODS AND RESULTS: Nur77(-/-) chimeric mice on a Ldlr(-/-) background showed a 3-fold increase in atherosclerosis development when fed a Western diet for 20 weeks, despite having a drastic reduction in Ly6C(-) patrolling monocytes. In a second model, mice deficient in both Nur77 and ApoE (ApoE(-/-)Nur77(-/-)) also showed increased atherosclerosis after 11 weeks of Western diet. Atherosclerosis was associated with a significant change in macrophage polarization toward a proinflammatory phenotype, with high expression of tumor necrosis factor-α and nitric oxide and low expression of Arginase-I. Moreover, we found increased expression of toll-like receptor 4 mRNA and protein in Nur77(-/-) macrophages as well as increased phosphorylation of the p65 subunit of NFκB. Inhibition of NFκB activity blocked excess activation of Nur77(-/-) macrophages. CONCLUSIONS: We conclude that the absence of Nur77 in monocytes and macrophages results in enhanced toll-like receptor signaling and polarization of macrophages toward a proinflammatory M1 phenotype. Despite having fewer monocytes, Nur77(-/-) mice developed significant atherosclerosis when fed a Western diet. These studies indicate that Nur77 is a novel target for modulating the inflammatory phenotype of monocytes and macrophages and may be important for regulation of atherogenesis.

CNS bridging radiotherapy achieves rapid cytoreduction before CAR T-cell therapy for aggressive B-cell lymphomas.

ABSTRACT: Chimeric antigen receptor (CAR) T-cell therapy (CART) for central nervous system lymphoma (CNSL) is a promising strategy, yet responses are frequently not durable. Bridging radiotherapy (BRT) is used for extracranial lymphoma in which it can improve CART outcomes through cytoreduction of high-risk lesions. We hypothesized that BRT would achieve similar, significant cytoreduction before CART for CNSL (CNS-BRT). We identified patients with CNSL with non-Hodgkin B-cell lymphoma who received CNS-BRT before commercial CART. Cytoreduction from CNS-BRT was calculated as change in lesion size before CART. Twelve patients received CNS-BRT, and the median follow-up among survivors is 11.8 months (interquartile range, 8.5-21.9). Ten patients had CNSL (9 secondary, 1 primary) and 2 patients had epidural disease (evaluable for toxicity). All 10 patients with CNSL had progressive disease at the time of CNS-BRT. Of 12 patients, 1 experienced grade ≥3 cytokine release syndrome, and 3 of 12 patients experienced grade ≥3 immune effector cell-associated neurotoxicity syndrome. CNS-BRT achieved a 74.0% (95% confidence interval, 62.0-86.0) mean reduction in lesion size from baseline (P = .014) at a median of 12 days from BRT completion and before CART infusion. Best CNS response included 8 complete responses, 1 partial response, and 1 progressive disease. Three patients experienced CNS relapse outside the BRT field. Preliminary data suggest CNS-BRT achieves rapid cytoreduction and is associated with a favorable CNS response and safety profile. These data support further study of BRT as a bridging modality for CNSL CART.

Metabolic tumor volume response after bridging therapy determines chimeric antigen receptor T-cell outcomes in large B cell lymphoma.

PURPOSE: Greater disease burden is a well-established predictor of poorer outcomes following chimeric antigen receptor T-cell therapy (CART). While bridging therapy (BT) is widely used between leukapheresis and CAR T infusion, limited data has evaluated the impact of BT on CART outcomes. In this study, we hypothesized that the quantitative dynamics of radiomic cytoreduction during bridging are prognostic. PATIENTS AND METHODS: Patients with large B-cell lymphoma (LBCL) treated with CD19-CART from 2016-2022 were included. Metabolic tumor volume (MTV) was determined for all patients on pre-leukapheresis PET and on post-BT/pre-infusion PET in those who received BT. Patients were stratified into 'High' and 'Low' disease burden using an MTV cutpoint of 65.4cc established by maximally selected log-rank statistic for progression free survival (PFS). RESULTS: Of 191 patients treated with CART, 144 (75%) received BT. In the BT cohort, 56% had any reduction in MTV post-BT. On multivariate analysis, MTV trajectory across the bridging period remained significantly associated with PFS (p<0.001), however notably patients with improved MTV (High->Low) had equivalent PFS compared to those with initially and persistently low MTV (Low->Low) (HR for High->Low MTV: 2.74, CI: 0.82-9.18). There was a reduction in any Grade ICANS in the High->Low MTV cohort as compared to High->High (13 vs. 41%, p=0.05). CONCLUSIONS: This is the first study to use radiomics to quantify disease burden pre- and post-BT in a large real world LBCL cohort. We demonstrate that effective BT can enable initially high-disease burden patients to achieve post-CART outcomes comparable to low-disease burden patients.

Conventional and novel [18F]FDG PET/CT features as predictors of CAR-T cell therapy outcome in large B-cell lymphoma.

Relapse and toxicity limit the effectiveness of chimeric antigen receptor T-cell (CAR-T) therapy for large B-cell lymphoma (LBCL), yet biomarkers that predict outcomes and toxicity are lacking. We examined radiomic features extracted from pre-CAR-T 18F-fluorodeoxyglucose positron emission tomography/computed tomography ([18F]FDG PET/CT) scans (n = 341) of 180 patients (121 male; median age, 66 years). Three conventional (maximum standardized uptake value [SUVmax], metabolic tumor volume [MTV], total lesion glycolysis [TLG]) and 116 novel radiomic features were assessed, along with inflammatory markers, toxicities, and outcomes. At both pre-apheresis and pre-infusion time points, conventional PET features of disease correlated with elevated inflammatory markers. At pre-infusion, MTV was associated with grade ≥ 2 cytokine release syndrome (odds ratio [OR] for 100 mL increase: 1.08 [95% confidence interval (CI), 1.01-1.20], P = 0.031), and SUVmax was associated with failure to achieve complete response (CR) (OR 1.72 [95% CI, 1.24-2.43], P < 0.001). Higher pre-apheresis and pre-infusion MTV values were associated with shorter progression-free survival (PFS) (HR for 10-unit increase: 1.11 [95% CI, 1.05-1.17], P < 0.001; 1.04 [95% CI, 1.02-1.07], P < 0.001) and shorter overall survival (HR for 100-unit increase: 1.14 [95% CI, 1.07-1.21], P < 0.001; 1.04 [95% CI, 1.02-1.06], P < 0.001). A combined MTV and LDH measure stratified patients into high and low PFS risk groups. Multiple pre-infusion novel radiomic features were associated with CR. These quantitative conventional [18F]FDG PET/CT features obtained before CAR-T cell infusion, which were correlated with inflammation markers, may provide prognostic biomarkers for CAR-T therapy efficacy and toxicity. The use of conventional and novel radiomic features may thus help identify high-risk patients for earlier interventions.

Characterization of Central Nervous System Clinico-Genomic Outcomes in ALK-Positive Non-Small Cell Lung Cancer Patients with Brain Metastases Treated with Alectinib.

INTRODUCTION: Highly effective brain-penetrant ALK-targeted tyrosine kinase inhibitors (TKIs) have been developed for the management of NSCLC patients with brain metastases (BM). Local therapy (LT) such as SRS or therapeutic craniotomy is increasingly being deferred for such patients. Herein we report detailed patient- and lesion-level intracranial outcomes and co-mutational genomic profiles from a cohort of NSCLC patients with BM treated with alectinib, with or without LT. METHODS: We retrospectively reviewed ALK fusion-positive NSCLC patients with BMs who received alectinib at the diagnosis of BM from 1/2012 and 5/2021. Outcome variables included intracranial progression-free survival (iPFS), overall survival (OS), duration of TKI therapy, and CNS response rates. Genomic characteristics from tumor specimens were assessed with MSK-IMPACT, a next-generation sequencing (NGS)-based genomic profiling assay. RESULTS: A total of 38 patients with 114 CNS lesions were included. Twelve of these patients also received contemporaneous LT (SRS, WBRT, or surgical resection). Maximal BM diameter in the TKI + LT group was greater (p < 0.003) but despite this difference, iPFS (TKI only, HR 1.21, 95 % CI 0.51-2.89; p = 0.66) and OS (TKI only, HR 5.99, 95 % CI 0.77-46.6; p = 0.052) were similar between groups and trended towards more favorable outcomes with the addition of LT. SMARCA4 co-alterations were associated with inferior OS (HR 8.76, 1.74-44.2; p = 0.009). CONCLUSIONS: Our study demonstrated that patients with ALK fusion-positive NSCLC treated with TKI + LT had larger BM and higher likelihood of pre-treatment neurologic symptoms. Despite these differences, iPFS was similar between groups. Results should be interpreted with caution as our study was limited by an underpowered sample size. SMARCA4 co-alterations were associated with inferior OS and these findings warrant further investigation.

Bridging Radiation Rapidly and Effectively Cytoreduces High-Risk Relapsed/Refractory Aggressive B Cell Lymphomas Prior to Chimeric Antigen Receptor T Cell Therapy.

Greater tumor burden before CD19-targeted chimeric antigen receptor T cell (CAR-T) therapy predicts lower complete response rate and shorter overall survival (OS) in patients with aggressive non-Hodgkin lymphoma (NHL). Recent patterns of failure studies have identified lesion characteristics, including size, standard uptake value (SUV), and extranodal location, as associated with post-CAR-T therapy failure. Here we analyzed the effect of bridging radiation-containing treatment (BRT) on pre-CAR-T therapy lesion- and patient-level characteristics and post-CAR-T therapy outcomes, including patterns of failure. Consecutive NHL patients who received radiation therapy from 30 days before leukapheresis until CAR T cell infusion were reviewed. Metabolic tumor volume (MTV) was contoured with a threshold SUV of 4. The first post-CAR-T therapy failures were categorized as preexisting/new/mixed with respect to pre-CAR-T therapy disease and in-field/marginal/distant with respect to BRT. Forty-one patients with diffuse large B cell lymphoma (DLBCL; n = 33), mantle cell lymphoma (n = 7), or Burkitt lymphoma (n = 1) were identified. BRT significantly improved established high-risk parameters of post-CAR-T therapy progression, including in-field median MTV (45.5 cc to .2 cc; P < .001), maximum SUV (18.1 to 4.4; P < .001), diameter (5.5 cm to 3.2 cm; P < .001), and lactate dehydrogenase (LDH; 312 to 232; P = .025). DLBCL patients with lower LDH levels post-BRT had improved progression-free survival (PFS; P = .001). In DLBCL, first failures were new in 7 of 19 patients, preexisting in 5 of 19, and mixed in 7 of 19; with respect to BRT, 4 of 19 were in-field and 4 of 19 were marginal. Post-CAR-T therapy survival was similar in patients with initially low MTV and those with newly low MTV post-BRT using a statistically determined threshold of 16 cc (PFS, 26 months versus 31 months; OS unreached for both). BRT produced significant cytoreductions in diameter, SUV, MTV, and LDH, all predictors of poor post-CAR-T therapy outcomes. Similar PFS and OS in patients with initially low MTV and those who achieved newly low MTV after BRT suggest that BRT may "convert" poor-risk patients to better risk. In the future, the response to BRT may allow for risk stratification and individualization of bridging strategies.

Trends in Use of and Medicare Spending on Short-Course Radiotherapy for Lymphomas From 2015 to 2019.

This cross-sectional study uses Centers for Medicare & Medicaid Services payment data to examine use of short-course radiotherapy from 2015 to 2019 among Medicare beneficiaries with indolent lymphoma.

Outcomes With Local Therapy and Tyrosine Kinase Inhibition in Patients With ALK/ROS1/RET-Rearranged Lung Cancers.

PURPOSE: Local therapy prolongs progression-free survival in patients with oligometastatic non-small-cell lung cancers treated with chemotherapy. We previously reported that local therapy also prolongs survival and time to next therapy in patients on tyrosine kinase inhibitors (TKIs) for EGFR-mutant lung adenocarcinomas. Here, we investigate the role of local therapy in patients progressing on TKIs for ALK/ROS1/RET-rearranged lung adenocarcinomas. MATERIALS AND METHODS: Patients with advanced ALK/ROS/RET-rearranged lung adenocarcinomas who underwent radiation, surgery, or percutaneous thermal ablation from 2012 to 2020 for progression on an ALK/ROS1/RET TKI were included. Progression patterns were identified. Times from local therapy to progression, next therapy, and death were measured. RESULTS: Sixty-one patients with ALK (n = 37), ROS1 (n = 12), and RET (n = 12) fusions were identified. Patients received radiotherapy (92%), surgery (13%), and percutaneous thermal ablation (8%). Local therapy was administered for solitary/oligoprogressive (94%) or polyprogressive (6%) disease. For most patients (85%), local therapy addressed all progressing sites. The median times from any local therapy to subsequent progression and next systemic therapy were 6.8 months (95% CI, 5.1 to 8.1) and 10 months (95% CI, 8.4 to 15.3), respectively. Third or greater local therapy was associated with shorter time to progression and next therapy than first/second local therapies (hazard ratio, 4.97; P < .001 and hazard ratio, 2.48; P < .001). The median overall survival from first local therapy was 34 months (95% CI, 26 to not reached). CONCLUSION: Local therapy for progression on ALK, ROS1, or RET TKIs is associated with clinically meaningful time on continued TKI therapy beyond progression, especially earlier in the course of disease.

Spectrum of Mechanisms of Resistance to Crizotinib and Lorlatinib in ROS1 Fusion-Positive Lung Cancer.

PURPOSE: Current standard initial therapy for advanced, ROS proto-oncogene 1, receptor tyrosine kinase fusion (ROS1)-positive (ROS1+) non-small cell lung cancer (NSCLC) is crizotinib or entrectinib. Lorlatinib, a next-generation anaplastic lymphoma kinase/ROS1 inhibitor, recently demonstrated efficacy in ROS1+ NSCLC, including in crizotinib-pretreated patients. However, mechanisms of lorlatinib resistance in ROS1+ disease remain poorly understood. Here, we assessed mechanisms of resistance to crizotinib and lorlatinib. EXPERIMENTAL DESIGN: Biopsies from patients with ROS1 + NSCLC progressing on crizotinib or lorlatinib were profiled by genetic sequencing. RESULTS: From 55 patients, 47 post-crizotinib and 32 post-lorlatinib biopsies were assessed. Among 42 post-crizotinib and 28 post-lorlatinib biopsies analyzed at distinct timepoints, ROS1 mutations were identified in 38% and 46%, respectively. ROS1 G2032R was the most commonly occurring mutation in approximately one third of cases. Additional ROS1 mutations included D2033N (2.4%) and S1986F (2.4%) post-crizotinib and L2086F (3.6%), G2032R/L2086F (3.6%), G2032R/S1986F/L2086F (3.6%), and S1986F/L2000V (3.6%) post-lorlatinib. Structural modeling predicted ROS1L2086F causes steric interference to lorlatinib, crizotinib, and entrectinib, while it may accommodate cabozantinib. In Ba/F3 models, ROS1L2086F, ROS1G2032R/L2086F, and ROS1S1986F/G2032R/L2086F were refractory to lorlatinib but sensitive to cabozantinib. A patient with disease progression on crizotinib and lorlatinib and ROS1 L2086F received cabozantinib for nearly 11 months with disease control. Among lorlatinib-resistant biopsies, we also identified MET amplification (4%), KRAS G12C (4%), KRAS amplification (4%), NRAS mutation (4%), and MAP2K1 mutation (4%). CONCLUSIONS: ROS1 mutations mediate resistance to crizotinib and lorlatinib in more than one third of cases, underscoring the importance of developing next-generation ROS1 inhibitors with potency against these mutations, including G2032R and L2086F. Continued efforts are needed to elucidate ROS1-independent resistance mechanisms.

Treatment with Next-Generation ALK Inhibitors Fuels Plasma ALK Mutation Diversity.

PURPOSE: Acquired resistance to next-generation ALK tyrosine kinase inhibitors (TKIs) is often driven by secondary ALK mutations. Here, we investigated utility of plasma genotyping for identifying ALK resistance mutations at relapse on next-generation ALK TKIs. EXPERIMENTAL DESIGN: We analyzed 106 plasma specimens from 84 patients with advanced ALK-positive lung cancer treated with second- and third-generation ALK TKIs using a commercially available next-generation sequencing (NGS) platform (Guardant360). Tumor biopsies from TKI-resistant lesions underwent targeted NGS to identify ALK mutations. RESULTS: By genotyping plasma, we detected an ALK mutation in 46 (66%) of 70 patients relapsing on a second-generation ALK TKI. When post-alectinib plasma and tumor specimens were compared, there was no difference in frequency of ALK mutations (67% vs. 63%), but plasma specimens were more likely to harbor ≥2 ALK mutations (24% vs. 2%, P = 0.004). Among 29 patients relapsing on lorlatinib, plasma genotyping detected an ALK mutation in 22 (76%), including 14 (48%) with ≥2 ALK mutations. The most frequent combinations of ALK mutations were G1202R/L1196M and D1203N/1171N. Detection of ≥2 ALK mutations was significantly more common in patients relapsing on lorlatinib compared with second-generation ALK TKIs (48% vs. 23%, P = 0.017). Among 15 patients who received lorlatinib after a second-generation TKI, serial plasma analysis demonstrated that eight (53%) acquired ≥1 new ALK mutations on lorlatinib. CONCLUSIONS: ALK resistance mutations increase with each successive generation of ALK TKI and may be underestimated by tumor genotyping. Sequential treatment with increasingly potent ALK TKIs may promote acquisition of ALK resistance mutations leading to treatment-refractory compound ALK mutations.

Psychosocial needs of young breast cancer survivors in Mexico City, Mexico.

OBJECTIVE: Young breast cancer survivors in Mexico face distinct psychosocial challenges that have not been characterized. This study aims to describe the psychosocial needs of young breast cancer survivors in Mexico at 5 or more years of survivorship, identifying areas of focus for early interventions. METHODS: Breast cancer patients diagnosed at age 40 or prior with 5 or more years since diagnosis were invited to participate in one-on-one 30-60 minute semi-structured audio-recorded interviews at the Instituto Nacional de Cancerología in Mexico City. Transcripts were coded using thematic analysis with NVivo software. RESULTS: 25 women participated. Five major phenomena emerged from analysis: (1) minimization of fertility concerns; (2) persistence of body image disturbance over time; (3) barriers to employment during survivorship; (4) impact on family relationships and social networks; & (5) unmet psychological care and informational needs. CONCLUSIONS: Early interventions with a focus on fertility loss education, access to reconstructive surgery and body image support, guidance during return-to-work, assistance with childcare, integration of psychological care and the fulfillment of informational needs could ameliorate long-term psychological and social distress for young breast cancer survivors in Mexico.

Improved Overall Survival and Locoregional Disease Control With Concurrent PD-1 Pathway Inhibitors and Stereotactic Radiosurgery for Lung Cancer Patients With Brain Metastases.

PURPOSE: Despite the emerging role of programmed cell death-1 (PD-1) pathway inhibitors for patients with advanced lung cancer, a paucity of data are available on the activity of these agents among patients with brain metastases. We investigated the outcomes of PD-1 pathway inhibitors and stereotactic radiosurgery (SRS) for the treatment of patients with brain metastases from lung cancer. METHODS AND MATERIALS: We retrospectively reviewed the medical records of non-small-cell lung cancer patients with brain metastases consecutively treated with PD-1 pathway inhibitors and SRS at our institution from 2012 to 2017. Overall survival (OS), distant brain failure (DBF), and local control (LC) were assessed using Kaplan-Meier estimates and Cox regression models. RESULTS: We identified 37 patients treated with SRS to 85 lesions (90.6% intact and 9.4% resected) and a median total of 7 doses of PD-1 pathway inhibitors (83.8% nivolumab, 10.8% atezolizumab, 5.4% pembrolizumab). Most lesions were treated with 18 Gy in a single fraction (n = 61; 71.8%). Patients treated with concurrent SRS and PD-1 pathway inhibitors had longer OS and reduced rates of DBF compared with patients treated with SRS before or after PD-1 pathway inhibitor therapy (1-year OS, 87.3% vs 70.0% vs 0%, P = .008; 1-year DBF, 38.5% vs 65.8% vs 100%, P = .042). LC was favorable among lesions treated with SRS concurrent with or after PD-1 pathway inhibitor therapy compared with before PD-1 pathway inhibitor therapy (1-year LC, 100% vs 72.3%, P = .016). Three lesions transiently enlarged after SRS and then had partially or completely resolved on follow-up imaging. Four patients required steroids for SRS-associated toxicity. No patient experienced grade ≥ 4 toxicity. CONCLUSIONS: Concurrent treatment with SRS and PD-1 pathway inhibitors was associated with favorable OS and locoregional disease control. This combination of therapy was well tolerated and merits further evaluation in larger cohorts in a prospective setting.

Safety of Combined PD-1 Pathway Inhibition and Intracranial Radiation Therapy in Non-Small Cell Lung Cancer.

INTRODUCTION: Intracranial metastases are a common cause of morbidity and mortality in patients with advanced NSCLC, and are frequently managed with radiation therapy (RT). The safety of cranial RT in the setting of treatment with immune checkpoint inhibitors (ICIs) has not been established. METHODS: We identified patients with advanced NSCLC with brain metastases who received cranial RT and were treated with or without programmed cell death 1/programmed death ligand 1 inhibitors between August 2013 and September 2016. RT-related adverse events (AEs) were retrospectively evaluated and analyzed according to ICI treatment status, cranial RT type, and timing of RT with respect to ICI. RESULTS: Of 163 patients, 50 (31%) received ICIs, whereas 113 (69%) were ICI naive. Overall, 94 (58%), 28 (17%), and 101 (62%) patients received stereotactic radiosurgery, partial brain irradiation, and/or whole brain RT, respectively. Fifty percent of patients received more than one radiation course. We observed no significant difference in rates of all-grade AEs and grade 3 or higher AEs between the ICI-naive and ICI-treated patients across different cranial RT types (grade ≥3 AEs in 8% of ICI-naive patients versus in 9% of ICI-treated patients for stereotactic radiosurgery [p = 1.00] and in 8% of ICI-naive patients versus in 10% of ICI-treated patients for whole brain RT [p = 0.71]). Additionally, there was no difference in AE rates on the basis of timing of ICI administration with respect to RT. CONCLUSIONS: Treatment with an ICI and cranial RT was not associated with a significant increase in RT-related AEs, suggesting that use of programmed cell death 1/programmed death ligand 1 inhibitors in patients receiving cranial RT may have an acceptable safety profile. Nonetheless, additional studies are needed to validate this approach.

Tracking the Evolution of Resistance to ALK Tyrosine Kinase Inhibitors through Longitudinal Analysis of Circulating Tumor DNA.

PURPOSE: ALK rearrangements predict for sensitivity to ALK tyrosine kinase inhibitors (TKIs). However, responses to ALK TKIs are generally short-lived. Serial molecular analysis is an informative strategy for identifying genetic mediators of resistance. Although multiple studies support the clinical benefits of repeat tissue sampling, the clinical utility of longitudinal circulating tumor DNA analysis has not been established in ALK-positive lung cancer. METHODS: Using a 566-gene hybrid-capture next-generation sequencing (NGS) assay, we performed longitudinal analysis of plasma specimens from 22 ALK-positive patients with acquired resistance to ALK TKIs to track the evolution of resistance during treatment. To determine tissue-plasma concordance, we compared plasma findings to results of repeat biopsies. RESULTS: At progression, we detected an ALK fusion in plasma from 19 (86%) of 22 patients, and identified ALK resistance mutations in plasma specimens from 11 (50%) patients. There was 100% agreement between tissue- and plasma-detected ALK fusions. Among 16 cases where contemporaneous plasma and tissue specimens were available, we observed 100% concordance between ALK mutation calls. ALK mutations emerged and disappeared during treatment with sequential ALK TKIs, suggesting that plasma mutation profiles were dependent on the specific TKI administered. ALK G1202R, the most frequent plasma mutation detected after progression on a second-generation TKI, was consistently suppressed during treatment with lorlatinib. CONCLUSIONS: Plasma genotyping by NGS is an effective method for detecting ALK fusions and ALK mutations in patients progressing on ALK TKIs. The correlation between plasma ALK mutations and response to distinct ALK TKIs highlights the potential for plasma analysis to guide selection of ALK-directed therapies.

Sequential ALK Inhibitors Can Select for Lorlatinib-Resistant Compound ALK Mutations in ALK-Positive Lung Cancer.

The cornerstone of treatment for advanced ALK-positive lung cancer is sequential therapy with increasingly potent and selective ALK inhibitors. The third-generation ALK inhibitor lorlatinib has demonstrated clinical activity in patients who failed previous ALK inhibitors. To define the spectrum of ALK mutations that confer lorlatinib resistance, we performed accelerated mutagenesis screening of Ba/F3 cells expressing EML4-ALK. Under comparable conditions, N-ethyl-N-nitrosourea (ENU) mutagenesis generated numerous crizotinib-resistant but no lorlatinib-resistant clones harboring single ALK mutations. In similar screens with EML4-ALK containing single ALK resistance mutations, numerous lorlatinib-resistant clones emerged harboring compound ALK mutations. To determine the clinical relevance of these mutations, we analyzed repeat biopsies from lorlatinib-resistant patients. Seven of 20 samples (35%) harbored compound ALK mutations, including two identified in the ENU screen. Whole-exome sequencing in three cases confirmed the stepwise accumulation of ALK mutations during sequential treatment. These results suggest that sequential ALK inhibitors can foster the emergence of compound ALK mutations, identification of which is critical to informing drug design and developing effective therapeutic strategies.Significance: Treatment with sequential first-, second-, and third-generation ALK inhibitors can select for compound ALK mutations that confer high-level resistance to ALK-targeted therapies. A more efficacious long-term strategy may be up-front treatment with a third-generation ALK inhibitor to prevent the emergence of on-target resistance. Cancer Discov; 8(6); 714-29. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 663.

Patterns of Metastatic Spread and Mechanisms of Resistance to Crizotinib in ROS1-Positive Non-Small-Cell Lung Cancer.

PURPOSE: The ROS1 tyrosine kinase is activated through ROS1 gene rearrangements in 1-2% of non-small cell lung cancer (NSCLC), conferring sensitivity to treatment with the ALK/ROS1/MET inhibitor crizotinib. Currently, insights into patterns of metastatic spread and mechanisms of crizotinib resistance among ROS1-positive patients are limited. PATIENTS AND METHODS: We reviewed clinical and radiographic imaging data of patients with ROS1- and ALK-positive NSCLC in order to compare patterns of metastatic spread at initial metastatic diagnosis. To determine molecular mechanisms of crizotinib resistance, we also analyzed repeat biopsies from a cohort of ROS1-positive patients progressing on crizotinib. RESULTS: We identified 39 and 196 patients with advanced ROS1- and ALK-positive NSCLC, respectively. ROS1-positive patients had significantly lower rates of extrathoracic metastases (ROS1 59.0%, ALK 83.2%, P=0.002), including lower rates of brain metastases (ROS1 19.4%, ALK 39.1%; P = 0.033), at initial metastatic diagnosis. Despite similar overall survival between ALK- and ROS1-positive patients treated with crizotinib (median 3.0 versus 2.5 years, respectively; P=0.786), ROS1-positive patients also had a significantly lower cumulative incidence of brain metastases (34% vs. 73% at 5 years; P<0.0001). Additionally, we identified 16 patients who underwent a total of 17 repeat biopsies following progression on crizotinib. ROS1 resistance mutations were identified in 53% of specimens, including 9/14 (64%) non-brain metastasis specimens. ROS1 mutations included: G2032R (41%), D2033N (6%), and S1986F (6%). CONCLUSIONS: Compared to ALK rearrangements, ROS1 rearrangements are associated with lower rates of extrathoracic metastases, including fewer brain metastases, at initial metastatic diagnosis. ROS1 resistance mutations, particularly G2032R, appear to be the predominant mechanism of resistance to crizotinib, underscoring the need to develop novel ROS1 inhibitors with activity against these resistant mutants.

Regulatory T cells occupy an isolated niche in the intestine that is antigen independent.

Regulatory T cells (Tregs) are CD4(+) T cells that maintain immune homeostasis and prevent autoimmunity. Like all CD4(+) T cells, Tregs require antigen-specific signals via T cell receptor-major histocompatibility complex class II (TCR-MHCII) interactions for their development. However, the requirement for MHCII in Treg homeostasis in tissues such as intestinal lamina propria (LP) is unknown. We examined LP Treg homeostasis in a transgenic mouse model that lacks peripheral TCR-MHCII interactions and generation of extrathymic Tregs (iTregs). Thymically generated Tregs entered the LP of weanlings and proliferated independently of MHCII to fill the compartment. The adult LP was a closed niche; new thymic Tregs were excluded, and Tregs in parabiotic pairs were LP resident. The isolated LP niche was interleukin-2 (IL-2) independent but dependent on commensal bacteria. Thus, an LP Treg niche can be filled, isolated, and maintained independently of antigen signals and iTregs. This niche may represent a tissue-specific mechanism for maintaining immune tolerance.