Efficacy and Safety of Dose-Escalated Alectinib in Patients With Metastatic ALK-Positive NSCLC and Central Nervous System Relapse on Standard-Dose Alectinib.

Introduction: Central nervous system (CNS) metastases remain a common challenge in patients with ALK-positive NSCLC. We previously reported reinduction of CNS responses using dose-intensified alectinib in two patients with CNS progression on standard-dose alectinib. Nevertheless, this strategy has not been assessed in larger cohorts. Methods: Patients were eligible for this retrospective study if they had metastatic ALK-positive NSCLC with CNS relapse on alectinib 600 mg twice daily dosing and subsequently received escalated dosing (900 mg twice daily) of alectinib. CNS efficacy was assessed per the modified Response Evaluation Criteria in Solid Tumors version 1.1. Results: Among 27 patients, median duration of dose-escalated alectinib was 7.7 months (95% confidence interval [CI]: 4.8-10.9), with median overall time-to-progression (TTP) of 7.1 months (95% CI: 4.4-9.6). Among 25 CNS response-assessable patients, CNS objective response rate was 12.0% (95% CI: 2.5-31.2) and CNS disease control rate was 92.0% (95% CI: 74.0-99.0), with median CNS duration of disease control of 5.3 months (95% CI: 3.4-8.3) and median CNS TTP of 7.1 months (95% CI: 4.4-9.6). Among four patients with measurable CNS disease at baseline, three experienced a best intracranial response of stable disease and one experienced intracranial partial response with CNS TTP ranging from 4.1 to 7.7 months. No patient required drug discontinuation due to treatment-related adverse event or experienced grade 3 or higher treatment-related adverse events. Conclusions: Dose-intensified alectinib was found to have tolerability and activity in patients with ALK-positive NSCLC who experienced CNS relapse on standard-dose alectinib and represents one clinically viable strategy for this population.

Lorlatinib and capmatinib in a ROS1-rearranged NSCLC with MET-driven resistance: tumor response and evolution.

Acquired drug resistance remains a major problem across oncogene-addicted cancers. Elucidation of mechanisms of resistance can inform rational treatment strategies for patients relapsing on targeted therapies while offering insights into tumor evolution. Here, we report acquired MET amplification as a resistance driver in a ROS1-rearranged lung adenocarcinoma after sequential treatment with ROS1 inhibitors. Subsequent combination therapy with lorlatinib plus capmatinib, a MET-selective inhibitor, induced intracranial and extracranial tumor response. At relapse, sequencing of the resistant tumor revealed a MET D1246N mutation and loss of MET amplification. We performed integrated molecular analyses of serial tumor and plasma samples, unveiling dynamic alterations in the ROS1 fusion driver and MET bypass axis at genomic and protein levels and the emergence of polyclonal resistance. This case illustrates the complexity of longitudinal tumor evolution with sequential targeted therapies, highlighting challenges embedded in the current precision oncology paradigm and the importance of developing approaches that prevent resistance.

Efficacy and Tolerability of ALK/MET Combinations in Patients With ALK-Rearranged Lung Cancer With Acquired MET Amplification: A Retrospective Analysis.

Introduction: MET amplification is a potentially actionable resistance mechanism in ALK-rearranged (ALK+) lung cancer. Studies describing treatment outcomes of this molecular subgroup are lacking. Methods: We assembled a cohort of patients with ALK+ lung cancer and acquired MET amplification (identified by tissue or plasma) who received regimens targeting both ALK and MET. Efficacy and safety were assessed using the Response Evaluation Criteria in Solid Tumors version 1.1 and Common Terminology Criteria for Adverse Events version 4.03, respectively. Results: A total of 12 patients were included in the series. MET amplification was detected after a median of 1.5 (range 1-5) lines of therapy. Four distinct regimens were implemented to address MET amplification: crizotinib (n = 2), lorlatinib plus crizotinib (n = 6), alectinib plus capmatinib (n = 3), and alectinib plus crizotinib (n = 1). Partial responses were observed in five (42%) of 12 patients, including patients who received crizotinib (n = one of two), lorlatinib plus crizotinib (n = three of six), and alectinib plus capmatinib (n = one of three). Primary progression was observed in four patients (33%). Grades 1 to 2 peripheral edema, occurring in seven (58%) patients, was found with both crizotinib and capmatinib. One patient required dose reduction of capmatinib plus alectinib for persistent grade 2 edema. Across the regimens, one patient discontinued therapy for toxicity, specifically neurocognitive toxicity from lorlatinib plus crizotinib. At progression on ALK+ MET therapy, potential resistance mechanisms included MET copy number changes and ALK kinase domain mutations. Conclusions: Combined ALK and MET inhibition is associated with moderate antitumor activity in patients with ALK+ NSCLC with concurrent MET amplification. Prospective studies are indicated to confirm activity and identify individuals most likely to benefit from the treatment.

Integrated Radiology, Pathology, and Pharmacy Program to Accelerate Access to Osimertinib.

PURPOSE: Targeted therapy yields superior outcomes relative to genotype-agnostic therapy for patients with epidermal growth factor receptor (EGFR)-mutant lung cancer. Workflows that facilitate timely detection of EGFR mutations and early dispensation of osimertinib can improve management of this disease. METHODS: We developed an Integrated Radiology, Pathology, and Pharmacy Program to minimize delays in initiating osimertinib. The intervention consisted of parallel workflows coupling interventional radiology, surgical pathology, and analysis of nucleic acids from frozen tissue with early pharmacy engagement. We compared time to EGFR testing results and time to treatment for participating patients with those of historical cohorts. RESULTS: Between January 2020 and December 2021, 222 patients participated in the intervention. The median turnaround time from biopsy to EGFR results was 1 workday. Forty-nine (22%) tumors harbored EGFR exon 19 deletions or EGFR L858R. Thirty-one (63%) patients were prescribed osimertinib via the intervention. The median interval between osimertinib prescription and osimertinib dispensation was 3 days; dispensation occurred within 48 hours for 42% of patients. The median interval between biopsy and osimertinib dispensation was 5 days. Three patients received osimertinib within 24 hours of EGFR results. Compared with patients with EGFR-mutant non-small-cell lung cancer who were diagnosed through routine workflows, the intervention led to a significant reduction in median time between biopsy and EGFR results (1 v 7 days; P < .01) and median time to treatment initiation (5 v 23 days; P < .01). CONCLUSION: Combining radiology and pathology workflows with early parallel pharmacy engagement leads to a significant reduction in time to initiating osimertinib. Multidisciplinary integration programs are essential to maximize clinical utility of rapid testing.

Community pharmacists' counseling regarding nicotine replacement therapy: A secret shopper study.

BACKGROUND: Nicotine replacement therapy (NRT) is a safe and effective non-prescription tobacco cessation treatment. While most community-based pharmacists periodically provide patient education regarding NRT, there is a gap in real-world evidence assessing the counseling provided. OBJECTIVES: To assess community pharmacist counseling regarding NRT in a real-world setting. METHODS: A cross-sectional secret shopper audit was conducted to collect data regarding NRT counseling from 120 community pharmacist encounters. Seventeen trained college of pharmacy students presented to community pharmacies using a standardized script asking about 1 of 3 common NRT products (patch, gum, and lozenge). Pharmacies were randomly selected from a list of all community pharmacies open to the public in Bexar County, Texas. A standardized assessment form was used to document product availability, counseling length, whether or not the 7 counseling points and 6 assessment questions that could help guide the pharmacist's counseling regarding NRT products were provided without prompting, and potential inaccuracy of any recommendations and counseling points. Descriptive statistics were calculated, and analysis of variance and Fisher's exact test were used to test for variation across site type and time of day. RESULTS: NRT was available for purchase without speaking to pharmacy staff in 99 of 120 (83%) pharmacies. The mean length of counseling was 136 (standard deviation = 91) seconds. The most common points discussed were recommended strength (72%), tapering schedule (58%), and assessment of the daily number of cigarettes smoked (56%). Forty-one (34%) pharmacists provided one or more potentially inaccurate counseling points, the most common being inaccurate tapering schedule (provided during 31 (26%) encounters). Only 15% of pharmacists referred auditors for additional help or recommended a follow-up. CONCLUSION: NRT was commonly accessible in community pharmacies outside of the pharmacy area. Opportunities for pharmacists to provide more complete and accurate information to better assist patients with safe and effective smoking cessation were identified.

Factors Associated With Developing Neurocognitive Adverse Events in Patients Receiving Lorlatinib After Progression on Other Targeted Therapies.

INTRODUCTION: The safety profile of lorlatinib includes neurocognitive adverse events (NAEs). Baseline factors associated with developing NAEs remain poorly characterized. METHODS: Records from patients who received lorlatinib through prospective studies at Massachusetts General Hospital (MGH, n = 124) or the phase 1/2 B7461001 (NCT01970865; n = 248) study were reviewed to identify potential associations between comorbidities, baseline medications, and NAEs. RESULTS: Most patients experienced a NAE (MGH: 60%, B7461001: 49%). Cognitive effects occurred in 40% and 29% of patients in the MGH and B7461001 cohorts, respectively. Brain metastases (p = 0.008), brain radiation (p = 0.033), psychiatric illness (p = 0.008), psychiatric medications (p < 0.001), antiepileptics (p < 0.001), and stimulants (p = 0.026) were associated with developing cognitive effects in B7461001. Mood effects occurred in 36% and 23% of patients in the MGH and B7461001 cohorts, respectively. In the MGH cohort, psychiatric illness (p = 0.02) and stimulants (p = 0.01) were associated with developing mood effects whereas brain surgery (p = 0.020), psychiatric medications (p < 0.001), benzodiazepines (p = 0.002), and sedatives (p = 0.034) were associated with developing mood effects in B7461001. Psychotic effects were infrequent (MGH: 3%, B7461001: 9%) and were associated with brain surgery in the MGH cohort (p = 0.001) and age in B7461001 (p = 0.014). Speech effects were observed in 23% and 11% of patients in the MGH and B7461001 cohorts, respectively. Brain radiation (p = 0.012) and antiepileptics (p < 0.001) were associated with speech effects in B7461001. Dose reductions were implemented for 52% and 18% of patients with NAEs in MGH and B7461001 cohorts, respectively, with mitigating effect. CONCLUSIONS: Neurocognitive effects from lorlatinib are common. Lorlatinib-related NAEs may be influenced by multiple factors, including brain metastases, brain radiation, psychiatric illness, and use of neurotropic medications.

Analysis of lorlatinib analogs reveals a roadmap for targeting diverse compound resistance mutations in ALK-positive lung cancer.

Lorlatinib is currently the most advanced, potent and selective anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitor for the treatment of ALK-positive non-small cell lung cancer in the clinic; however, diverse compound ALK mutations driving therapy resistance emerge. Here, we determine the spectrum of lorlatinib-resistant compound ALK mutations in patients, following treatment with lorlatinib, the majority of which involve ALK G1202R or I1171N/S/T. We further identify structurally diverse lorlatinib analogs that harbor differential selective profiles against G1202R versus I1171N/S/T compound ALK mutations. Structural analysis revealed increased potency against compound mutations through improved inhibition of either G1202R or I1171N/S/T mutant kinases. Overall, we propose a classification of heterogenous ALK compound mutations enabling the development of distinct therapeutic strategies for precision targeting following sequential tyrosine kinase inhibitors.

Response to Immune Checkpoint Inhibition as Monotherapy or in Combination With Chemotherapy in Metastatic ROS1-Rearranged Lung Cancers.

INTRODUCTION: ROS1 fusions are oncogenic drivers in 1% to 3% of NSCLCs. The activity of immune checkpoint inhibitor (ICI) monotherapy or in combination with chemotherapy (chemotherapy with ICI [chemo-ICI]) in these tumors and their immunophenotype have not been systematically described. METHODS: In this multi-institutional retrospective study, tumor programmed death-ligand 1 (PD-L1) expression and tumor mutational burden (TMB) were evaluated in patients with ROS1-rearranged NSCLC. Time-to-treatment discontinuation (TTD) and objective response rate (ORR) (Response Evaluation Criteria in Solid Tumors [RECIST] version 1.1) were calculated for patients treated with ICI or chemo-ICI in the metastatic setting. RESULTS: A total of 184 patients were identified. Among 146 assessable cases, PD-L1 expression was less than 1% in 60 (41%), 1% to 49% in 35 (24%), and greater than or equal to 50% in 51 tumors (35%). Of 100 (92%) TMB-assessable tumors, 92 had less than 10 mutations per megabase. TMB was significantly lower for ROS1-rearranged tumors (n = 97) compared with tumors with EGFR (n = 1250) or KRAS alterations (n = 1653) and all other NSCLC tumors (n = 2753) evaluated with Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (median TMB = 2.6 versus 3.5, 7.0, and 6.1 mutations per megabase, p < 0.001). Among patients treated with ICI, median TTD was 2.1 months (95% confidence interval [CI]: 1.0-4.2 mo; n = 28) and ORR 13% (2 of 16 RECIST-assessable; 95% CI: 2%-38%). Among patients treated with chemo-ICI, median TTD was 10 months (95% CI: 4.7-14.1 mo; n = 11) and ORR 83% (5 of 6 RECIST-assessable; 95% CI: 36%-100%). There was no difference in PD-L1 expression (p = 0.91) or TMB (p = 0.83) between responders and nonresponders. CONCLUSIONS: Most ROS1-rearranged NSCLCs have low PD-L1 expression and TMB. The activity of ICI in these tumors is modest. In contrast, chemo-ICI can achieve meaningful activity.

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.

A Phase 2 Study of Capmatinib in Patients With MET-Altered Lung Cancer Previously Treated With a MET Inhibitor.

INTRODUCTION: Capmatinib is approved for MET exon 14-altered NSCLC on the basis of activity in targeted therapy-naive patients. We conducted a phase 2 study to assess the efficacy of capmatinib in patients previously treated with a MET inhibitor. METHODS: Patients with advanced NSCLC harboring MET amplification or MET exon 14 skipping alterations received capmatinib 400 mg twice daily. The primary end point was the objective response rate. Secondary end points included progression-free survival, disease control rate (DCR), intracranial response rate, and overall survival. Circulating tumor DNA was analyzed to identify capmatinib resistance mechanisms. RESULTS: A total of 20 patients were enrolled between May 2016 and November 2019, including 15 patients with MET skipping alterations and five patients with MET amplification. All patients had received crizotinib; three had also received other MET-directed therapies. The median interval between crizotinib and capmatinib was 22 days (range: 4-374). Two patients (10%) achieved an objective response to capmatinib and 14 had stable disease, yielding a DCR of 80%. Among five patients who discontinued crizotinib for intolerance, the DCR was 83%, including two patients with the best tumor shrinkage of -25% and -28%. Intracranial DCR among four patients with measurable brain metastases was 100%, with no observed intracranial objective responses. Overall, the median progression-free survival and overall survival were 5.5 (95% confidence interval: 1.3-11.0) and 11.3 (95% confidence interval: 5.5-not reached) months, respectively. MET D1228 and Y1230 mutations and MAPK alterations were recurrently detected in postcrizotinib, precapmatinib plasma. New and persistent MET mutations and MAPK pathway alterations were detected in plasma at progression on capmatinib. CONCLUSIONS: Capmatinib has modest activity in crizotinib-pretreated MET-altered NSCLC, potentially owing to overlapping resistance mechanisms.

Coronavirus Disease 2019 Infection in a Patient Population with Lung Cancer: Incidence, Presentation, and Alternative Diagnostic Considerations.

INTRODUCTION: Lung cancer is associated with severe coronavirus disease 2019 (COVID-19) infections. Symptom overlap between COVID-19 and lung cancer may complicate diagnostic evaluation. We aimed to investigate the incidence, symptoms, differential diagnosis, and outcomes of COVID-19 in patients with lung cancer. METHODS: To determine an at-risk population for COVID-19, we retrospectively identified patients with lung cancer receiving longitudinal care within a single institution in the 12 months (April 1, 2019 to March 31, 2020) immediately preceding the COVID-19 pandemic, including an "active therapy population" treated within the last 60 days of this period. Among patients subsequently referred for COVID-19 testing, we compared symptoms, laboratory values, radiographic findings, and outcomes of positive versus negative patients. RESULTS: Between April 1, 2019 and March 31, 2020, a total of 696 patients received longitudinal care, including 406 (58%) in the active therapy population. Among 55 patients referred for COVID-19 testing, 24 (44%) were positive for COVID-19, representing a cumulative incidence of 3.4% (longitudinal population) and 1.5% (active therapy population). Compared with patients who were COVID-19 negative, those who were COVID-19 positive were more likely to have a supplemental oxygen requirement (11% versus 54%, p = 0.005) and to have typical COVID-19 pneumonia imaging findings (5 versus 56%, p = 0.001). Otherwise, there were no marked differences in presenting symptoms. Among patients who were COVID-19 negative, alternative etiologies included treatment-related toxicity (26%), atypical pneumonia (22%), and disease progression (22%). A total of 16 patients positive for COVID-19 (67%) required hospitalization, and seven (29%) died from COVID-related complications. CONCLUSIONS: COVID-19 was infrequent in this lung cancer population, but these patients experienced high rates of morbidity and mortality. Oncologists should maintain a low threshold for COVID-19 testing in patients with lung cancer presenting with acute symptoms.

Small cell transformation of ROS1 fusion-positive lung cancer resistant to ROS1 inhibition.

Histologic transformation from non-small cell to small cell lung cancer has been reported as a resistance mechanism to targeted therapy in EGFR-mutant and ALK fusion-positive lung cancers. Whether small cell transformation occurs in other oncogene-driven lung cancers remains unknown. Here we analyzed the genomic landscape of two pre-mortem and 11 post-mortem metastatic tumors collected from an advanced, ROS1 fusion-positive lung cancer patient, who had received sequential ROS1 inhibitors. Evidence of small cell transformation was observed in all metastatic sites at autopsy, with inactivation of RB1 and TP53, and loss of ROS1 fusion expression. Whole-exome sequencing revealed minimal mutational and copy number heterogeneity, suggestive of "hard" clonal sweep. Patient-derived models generated from autopsy retained features consistent with small cell lung cancer and demonstrated resistance to ROS1 inhibitors. This case supports small cell transformation as a recurring resistance mechanism, and underscores the importance of elucidating its biology to expand therapeutic opportunities.

Imaging Features and Patterns of Metastasis in Non-Small Cell Lung Cancer with RET Rearrangements.

Rearranged during transfection proto-oncogene (RET) fusions represent a potentially targetable oncogenic driver in non-small cell lung cancer (NSCLC). Imaging features and metastatic patterns of advanced RET fusion-positive (RET+) NSCLC are not well established. Our goal was to compare the imaging features and patterns of metastases in RET+, ALK+ and ROS1+ NSCLC. Patients with RET+, ALK+, or ROS1+ NSCLC seen at our institution between January 2014 and December 2018 with available pre-treatment imaging were identified. The clinicopathologic features, imaging characteristics, and the distribution of metastases were reviewed and compared. We identified 215 patients with NSCLC harboring RET, ALK, or ROS1 gene fusion (RET = 32; ALK = 116; ROS1 = 67). Patients with RET+ NSCLC were older at presentation compared to ALK+ and ROS1+ patients (median age: RET = 64 years; ALK = 51 years, p < 0.001; ROS = 54 years, p = 0.042) and had a higher frequency of neuroendocrine histology (RET = 12%; ALK = 2%, p = 0.025; ROS1 = 0%, p = 0.010). Primary tumors in RET+ patients were more likely to be peripheral (RET = 69%; ALK = 47%, p = 0.029; ROS1 = 36%, p = 0.003), whereas lobar location, size, and density were comparable across the three groups. RET+ NSCLC was associated with a higher frequency of brain metastases at diagnosis compared to ROS1+ NSCLC (RET = 32%, ROS1 = 10%; p = 0.039. Metastatic patterns were otherwise similar across the three molecular subgroups, with high incidences of lymphangitic carcinomatosis, pleural metastases, and sclerotic bone metastases. RET+ NSCLC shares several distinct radiologic features and metastatic spread with ALK+ and ROS1+ NSCLC. These features may suggest the presence of RET fusions and help identify patients who may benefit from further molecular genotyping.

MET Alterations Are a Recurring and Actionable Resistance Mechanism in ALK-Positive Lung Cancer.

PURPOSE: Most ALK-positive lung cancers will develop ALK-independent resistance after treatment with next-generation ALK inhibitors. MET amplification has been described in patients progressing on ALK inhibitors, but frequency of this event has not been comprehensively assessed. EXPERIMENTAL DESIGN: We performed FISH and/or next-generation sequencing on 207 posttreatment tissue (n = 101) or plasma (n = 106) specimens from patients with ALK-positive lung cancer to detect MET genetic alterations. We evaluated ALK inhibitor sensitivity in cell lines with MET alterations and assessed antitumor activity of ALK/MET blockade in ALK-positive cell lines and 2 patients with MET-driven resistance. RESULTS: MET amplification was detected in 15% of tumor biopsies from patients relapsing on next-generation ALK inhibitors, including 12% and 22% of biopsies from patients progressing on second-generation inhibitors or lorlatinib, respectively. Patients treated with a second-generation ALK inhibitor in the first-line setting were more likely to develop MET amplification than those who had received next-generation ALK inhibitors after crizotinib (P = 0.019). Two tumor specimens harbored an identical ST7-MET rearrangement, one of which had concurrent MET amplification. Expressing ST7-MET in the sensitive H3122 ALK-positive cell line induced resistance to ALK inhibitors that was reversed with dual ALK/MET inhibition. MET inhibition resensitized a patient-derived cell line harboring both ST7-MET and MET amplification to ALK inhibitors. Two patients with ALK-positive lung cancer and acquired MET alterations achieved rapid responses to ALK/MET combination therapy. CONCLUSIONS: Treatment with next-generation ALK inhibitors, particularly in the first-line setting, may lead to MET-driven resistance. Patients with acquired MET alterations may derive clinical benefit from therapies that target both ALK and MET.

Machine Learning Methods to Predict Lung Cancer Survival Using the Veterans Affairs Research Precision Oncology Data Commons.

We completed a pilot study to guide the development of the VA Research Precision Oncology Data Commons infrastructure as a collaboration platform with the greater research community. Our results using a small subset of patients from the VA's Precision Oncology Program demonstrate the feasibility of our data sharing platform to build predictive models for lung cancer survival using machine learning, as well as highlight the potential of target genome sequencing data.

Diet-induced obesity precipitates kidney dysfunction and alters inflammatory mediators in mice treated with Shiga Toxin 2.

Shiga Toxin (Stx)-producing E. coli (STEC) continue to be a prominent cause of foodborne outbreaks of hemorrhagic colitis worldwide, and can result in life-threatening diseases, including hemolytic uremic syndrome (HUS), in susceptible individuals. Obesity-associated immune dysfunction has been shown to be a risk factor for infectious diseases, although few studies have addressed the role of obesity in foodborne diseases. We hypothesized that obesity may affect the development of HUS through an alteration of immune responses and kidney function. We combined diet-induced obese (DIO) and HUS mouse models to look for differences in disease outcome between DIO and wild-type (WT) male and female C57 B l/6 mice. Following multiple intraperitoneal injections with endotoxin-free saline or sublethal doses of purified Stx2, we examined DIO and WT mice for signs of HUS development. DIO mice receiving Stx2 injections lost more body weight, and had significantly higher (p < 0.001) BUN, serum creatinine, and neutrophil counts compared to WT mice or DIO mice receiving saline injections. Lymphocyte counts were significantly (p < 0.05) lower in Stx2-treated obese mice compared to WT mice or saline-treated DIO mice. In addition to increased Stx2-induced kidney dysfunction, DIO mouse kidneys also had significantly increased expression of IL-1α, IL-1ß, IL-6, TNF-α, MCP-1, and KC RNA compared to saline controls (p < 0.05). Serum cytokine levels of IL-6 and KC were also significantly higher in Stx2-treated mice compared to saline controls, but there were no significant differences between the WT and DIO mice. WT and DIO mice treated with Stx2 exhibited significantly higher degrees of kidney tubular dilation and necrosis as well as some signs of tissue repair/regeneration, but did not appear to progress to the full pathology typically associated with human HUS. Although the combined obesity/HUS mouse model did not manifest into HUS symptoms and pathogenesis, these data demonstrate that obesity alters kidney function, inflammatory cells and cytokine production in response to Stx2, and may play a role in HUS severity in a susceptible model of infection.