bluuebunny/crossref_metadata_2025 huggingface dataset:
T cell receptor β-chain repertoire analysis to reveal potential predictive biomarker for the use of immune checkpoint blockade in patients with advanced solid tumors.
Authors: Li Ye , Jiao Shun Chang , Wu Liang Liang , Sun Sheng Jie , Wei Hua Feng , Liao Sheng You
Published: May 2019

e14152

Background: Immune checkpoint blockade (ICB) targeted to CTLA-4, PD-1, and PD-L1 can promote antitumor T cell immunity and clinical responses. Tracking the dynamic changes of the T Cell Receptor (TCR) Immune Repertoire can reflect the evolution of the immune system in tumor immunotherapy patients, predict the immune response to ICB, and provide high-value information for clinical monitoring of immune function status of cancer patients to make effective clinical decisions. The purpose of this study was to explore the correlation between the peripheral blood TCR repertoire dynamic changes and the immune response of patients with anti-PD-1/PD-L1, and to accurately guide the individualized treatment of immunotherapy. Methods: We sequenced the TCRβ genes from T cells contained in peripheral blood mononuclear cells (PBMCs) from 24 advanced solid tumors patients (PD/7, PR/4, SD/13) treated with immune checkpoint blockade. Peripheral blood samples were collected from baseline, 8-week, and 16-week after treatment for each patient. bioinformatic analysis was used to detect the diversity and clonality changes of TCRβ-CDR3 sequence before and after treatment with ICB in patients, and provide auxiliary technology for further guiding clinical tumor immunotherapy. Results: The analysis result of Chao index showed that the TCR-CDR3 diversity in the PR group was significantly increased after treatment than SD or PD group. (pvalue = 0.001), and the number of CDR3 aa clonotypes also increased significantly (pvalue = 0.012). A total of 61 Vβ gene fragments and 15 Jβ gene fragments were uniquely identified during immunotherapy . There was a significant difference in the number of Vβ gene fragments between baseline samples and after treatment in PR group (pvalue = 0.042). Conclusions: In this project, PBMC samples was used to evaluate the changes of TCR repertoire between before and after treatment with immune checkpoint blockade in advanced solid tumor patients. The immune function status was significantly changed before and after treatment. Patients with PR had significantly increased diversity in TCR repertoire than patients with SD or PD. The results provide T cell receptor β-chain repertoire sequencing and analysis has potential to assess whether immune checkpoint blockade promotes global T cell expansion and/or loss of T cell clonotype diversity, and to identify predictive biomarker for the use of immune checkpoint blockade.
Prospective cardiac function monitoring in immunotherapy-treated patients.
Authors: Aras Mandar A , Cuevas Lauren , Bibby Dwight , Fang Qizhi , Daud Adil , Schiller Nelson B , Tsai Katy K.
Published: May 2019

e14157

Background: Immune checkpoint inhibitors (ICI) demonstrate activity and long-term benefit in melanoma and other cancers. While certain immune-related toxicities have been well described, cardiac toxicities remain poorly characterized. We sought to characterize changes in cardiac function in advanced melanoma patients (pts) starting ICI treatment. Methods: A prospective, observational, single institution trial was performed to monitor cardiac function in pts receiving ICI. Eligible pts were adults with stage III/IV melanoma and ECOG 0-1, determined safe to exercise and without unstable cardiac symptoms. Pts received PD-1 monotherapy or combination ipilimumab/pembrolizumab. Rest and exercise stress cardiac testing (EKG, echocardiography) was conducted at baseline, at 3 months (mo), and at 6 mo. Patient-reported surveys were administered at baseline and at 6 mo. Results: Between Feb 2018 to Jan 2019, we prospectively identified 8 pts who completed baseline cardiac testing. As of Feb 2019, 6 pts completed 3-mo testing, and 4 completed 6-mo testing. Mean age was 57 years (range 34-68). 5 pts received pembrolizumab or nivolumab monotherapy, and 3 received ipilimumab/pembrolizumab. Resting LVOT-VTI decreased from 23.1±2.8 cm at baseline to 20.7±2.8cm at 6 mo (p = 0.047). Maximum HR at peak exercise decreased over time from 149.7±13 bpm at baseline to 114±20.5 bpm at 6 mo (p = 0.085) with concurrent decrease in maximum BP. Enrollment is ongoing. Conclusions: These data suggest measurable changes in left ventricular cardiac function at rest over time, in a prospective cohort of ICI-treated advanced melanoma pts. Further, we observe progressive loss of chronotropic and blood pressure response to exercise. These data have important implications for cancer survivorship in the era of immunotherapy. Updated data to be presented at the meeting include correlation with patient-reported symptoms. [Table: see text]
Host metabolic factors and prognosis in patients treated with immune checkpoint inhibitors for advanced malignancies.
Authors: Biello Federica , Audisio Marco , Genestroni Silvia , Borra Gloria , D'Avanzo Francesca , Lacidogna Gaetano , Sponghini Andrea Pietro , Rondonotti David , Forti Laura , Vignani Francesca , Barone Adesi Francesco , Di Maio Massimo , Sica Antonio , Gennari Alessandra
Published: May 2019

e14162

Background: It is well established that an altered host metabolism has an impact on cancer outcome, possibly mediated by several mechanisms, including hyperglicaemia, hyperinsulinemia and presence of chronic inflammation. The aim of our analysis was to evaluate the correlation between host metabolism and clinical outcome in patients with advanced melanoma, kidney and non-small cell lung cancer (NSCLC), treated with immune checkpoint inhibitors (anti-CTLA4, anti PD1 and anti PDL1). Methods: The relationship between presence of type 2 diabetes mellitus (DMII) at baseline and outcome was assessed in 187 patients treated with immune checkpoint inhibitors in two cancer centers. Progression Free Survival (PFS) and Overall Survival (OS) were calculated by Kaplan-Meier estimation; multivariate Cox analysis was performed according to age, gender, BMI (normal < 25 kg/m2, overweight 25-30 kg/m2, obese > 30 kg/m2), type of cancer and line of treatment. Results: One-hundred-sixty-eight patients were available for our analysis. Twenty-eight patients (17%) were diabetic at baseline. Median age was 65 (range 25-80); 83 patients were males (49%); 82 (48%) had advanced melanoma, 83 (49%) NSCLC and 3 (3%) kidney cancer. One-hundred-two (60%) patients had BMI < 25, 51 (30%) were overweight and 16 (10%) were obese. The first line of treatment was immunotherapy in 83 (49%) patients. By univariable analysis median PFS was 4.2 months in non diabetics vs 6.4 in diabetics patients (HR 0.95; 95%CI 0.58-1.58); median OS was 6.17 and 9.1 months, respectively (HR 1.00; 95%CI 0.58-1.75). At multivariable analysis, taking into account DMII, BMI, sex, age, line of treatment and type of cancer, we found that BMI ≤25 was associated with a two fold increase in risk of progression (PD) or death (p = 0.005), whereas patients who received immunotherapy as second or subsequent line had a two fold increase in risk of PD or death (p = 0.003). Conclusions: The results of our analysis show that in patients with advanced cancer treated with immune checkpoint inhibitors, the presence of DMII does not adversely affect the clinical outcome. Conversely, lower BMI was associated with a significantly worse PFS and OS, independently from type of cancer, age and gender. As expected, patients who received immunotherapy in later lines of treatment had a significantly shorter survival.
Incidence and characteristics of emergency department presentations during immune checkpoint inhibitor therapy.
Authors: Smith Daniel Arnold , Laukamp Kai , Campbell Melanie , Devita Robert , Nelson Ariel Ann , Hoimes Christopher J. , Ramaiya Nikhil H.
Published: May 2019

e14164

Background: Immune checkpoint inhibitors (ICIs) have emerged as a novel class of anticancer agents with unique response and toxicity profiles. Oncology patients undergoing ICI therapy can present acutely with cancer- or treatment-related complications, but knowledge of these acute clinical presentations is limited. The objective of this study was to investigate the features of emergency department (ED) presentations of patients undergoing ICI therapy. Methods: A retrospective chart review was performed of 1044 adult oncology patients at a single institution from 2010-2018 who underwent treatment with one or more ICI. The number of patient visits to the ED during and up to one month following ICI treatment was recorded, in addition to various clinical and demographic data. These data were compared based on stratification by number of ED visits (0 visits, 1 visit, or ≥2 visits) using Likelihood Ratio Chi-Square and Mann–Whitney U tests. Results: Mean age for the 1044 patients receiving ICI therapy was 64±13 years, with 57% males and 43% females. Primary cancer distribution included 42.0% lung, 24.2% melanoma, 6.9% head & neck, 5.1% kidney, 4.0% bladder, and 17.8% other malignancy. 83.4% of patients were treated with a single ICI, 14.9% with 2 ICIs, and 1.2% with 3-4 ICIs. 56.0% of patients had no ED visits during their treatment duration, 27.0% had 1 ED visit, and 17.0% had ≥2 ED visits. Patients with lung, kidney, and bladder cancer were more likely to present to the ED (p = < 0.001). Black ethnicity was the only demographic feature associated with more ED visits (p = 0.017). Patients receiving ≥2 ICIs or monotherapy with nivolumab, pembrolizumab, or atezolizumab more frequently presented to the ED compared to other ICIs (p = < 0.001). Patients with 1 or ≥2 ED visits had longer durations of ICI therapy (136±12 days and 216±15 days, respectively) compared to patients with no ED visits (127±8 days) (p = < 0.001). Patients with no ED visits also demonstrated better overall survival (p = < 0.001). Conclusions: More frequent ED visits during ICI therapy is statistically associated with several key clinical factors, including primary cancer type, ethnicity, specific ICI agent, ICI therapy duration, and overall survival.
An immune response-associated genomic model to predict early resistance to PD1/PD-L1 blockade among real-world TMB-high Chinese cancer patients.
Authors: Xu Qiang , Lu Shichun , Wang Guan , Wei Jinwang , Wu Angela , Dai Chun , Cai Xin , Luo Ying , Sun Wending
Published: May 2019

e14167

Background: TMB-high was used as a favorable biomarker to predict the response to anti-PD-1/PD-L1 therapy, but a substantial portion of the TMB-high patients suffered from early resistance with limited clinical benefits but potential severe adverse effects. Methods: Whole-exome sequencing were performed in more than 200 advanced cancer patients, who received anti-PD-1 therapy alone or in combination with chemotherapy and targeted therapy across 20 provinces of China, with quarterly follow-ups. The genomic profiles of the patients with progressive disease (PD) and complete remission (CR) outcomes were individually analyzed and the potential resistance and hypersensitive biomarkers were identified. These biomarkers were formulated into an immune response-associated genomic (IRAG) model based on XGBoost algorithm to predict the sensitivity to PD-1 blockage and early resistance among TMB-high patients. Results: Non-synonymous somatic mutations (CTNNB1, B2M, PD-L1 and HLA class I genes), somatic gene amplifications (MDM2/4, CDK4, CCND1/FGF3/FGF4/FGF19, FLT3 and MYC), chromosome 9p21 deletion, and loss of heterozygosity in chromosome 6 HLA-I region were identified as potential resistance biomarkers in the PD group. TMB-high, microsatellite instability (MSI), TMB-low with indel-proportion high, POLE loss-of-function mutations, Epstein-Barr virus infection and DNA mismatch repair-deficient germline and somatic mutations were identified as potential hypersensitivity biomarkers in the CR group. By combining all genomic resistance and hypersensitivity biomarkers, we developed a quantitative IRAG model and separated the patients into three groups: TMB-low, TMB-high/IRAG-low and TMB-high/IRAG-high. The PFS in TMB-high/IRAG-low group was characterized with early clinical resistance, similar to the TMB-low group (median: 3.8 vs. 4.2 months, p = 0.22). The reanalysis of the patient set with known clinical outcomes showed that the TMB-high/IRAG-high group had a significantly longer PFS than the other two groups. (median: 10 vs. 3.8, 4.2 months, p < 0.001). The TMB-high/IRAG-high group clearly showed higher chromosomal stability with fewer somatic copy number alterations. Conclusions: We developed a model based on one single genomic assay (whole-exome) to successfully identify those TMB-high patients who suffered from early resistance to PD-1 inhibitors. Larger cohort and independent analyses are necessary to verify the validity of these results.
Development of imjSCORE for early prediction of response to nivolumab among patients with advanced cancer.
Authors: Janicek Milos Josef , Cirstea Diana , Denis Gerald V , Lerner Adam
Published: May 2019

e14169

Background: Cross-sectional Imaging (CI) plays an essential role in early assessment of patients on immune checkpoint inhibitors (ICIs), yet often remains inconclusive in the setting of advanced disease, prior cancer treatment, toxicity, and cancer unrelated findings. Best timing of scans is also uncertain and may affect interpretation. We propose here a robust, reproducible imjScore based on measurable tumor response on the first two re-staging scans and their ratios that may predict treatment impact. Our approach incorporates the prognostic aspect of early imaging, time lag of ICIs effect, and alternative measures of treatment impact. Methods: Retrospective review of CI studies was performed in 92 patients with advanced stage solid tumors treated with Nivolumab during 6/2015-8/2018 at Boston Medical Center utilizing standard clinical protocols. Time on therapy (TOTx) was used as a surrogate marker of therapy impact. Modified imRECIST score utilizing Σ of 4 discrete cancer lesions at baseline (< 1 month before Nivolumab), 1st, and 2nd CI since initiation of Nivolumab were adjusted for TOTx. Our imjSCORE averages "predicted days on therapy" calculated from best fit function of tumor change/day [%] at each time point vs. TOTx and ratios between 1st and 2nd scan. Results: Median TOTx was 70 days (range from 1 to 1029). Differences in imRECIST scores derived from two early restaging CI studies adjusted for TOTx were able to identify with high statistical significance two groups of "SHORT" vs "LONG" sojourn on Nivolumab based on Median of 70 days. Early progression curve was associated with shorter benefit of Nivolumab; subset with pseudo-progression curve showed still significantly shorter sojourn on Nivolumab compared to initial regression or stable score curves (P < .002). Conclusions: The development of imjScore may differentiate with high likelihood patients with short versus long sojourn on Nivolumab. Our CI-based evaluation using the imjSCORE suggests a clear benefit from early imaging with a response curve and standardized point analysis of the 1st and 2nd post-treatment scans. If validated with other ICIs, this scoring tool may allow early prediction of ICIs treatment response and clinical outcome. Further correlation of imjSCORE with clinical biomarkers may be useful to build predictive nomograms of "short" versus "long" treatment impact.
Recurrent and de-novo autoimmune hemolytic anemia in patients treated with immunotherapy for advanced cancer.
Authors: Jain Natasha A. , Rogers Kerry Anne , Wang Tzu-Fei , Zhao Songzhu , Wei Lai , Otterson Gregory Alan , Owen Dwight Hall
Published: May 2019

e14170

Background: Immunotherapy (IO) has brought dramatic clinical benefit to cancer patients. Serious immune-related adverse events (irAE) have been reported, however, hematological toxicities such as autoimmune hemolytic anemia (AIHA) remains poorly characterized. Here we performed a single institution retrospective study to determine the incidence and risk factors for AIHA and if prior positive direct anti-globulin test (DAT) or anti-RBC alloantibodies (alloAb) on type and screen predicts subsequent AIHA in patients treated with IO. Methods: This is a retrospective review of patients treated with IO between November 2012 and February 2017 at the Ohio State University. Patients who received IO either on clinical trial or as standard of care were included. The electronic medical records were reviewed for patients who developed AIHA. Overall survival (OS) was calculated from the date of initiation of IO to death from any cause or date or last follow-up. Medians and 95% CIs were estimated using Kaplan-Meier method and differences compared using the Log-Rank test. The association between irAE and DAT or alloAb testing was studied using Fisher’s exact test. Results: In 1,097 patients treated with IO, we identified 179 (16.6%) patients who had a DAT or alloAb testing at any time, 126 (11.5%) had this done prior to receiving IO. Of the 179 tested 10 (5.6%) had positive alloAb and 7 (3.9%) had a positive DAT. During treatment, two patients with prior history of AIHA experienced recurrence of AIHA during treatment. Three patients had positive DAT after IO, one was diagnosed as a delayed hemolytic transfusion reaction, and the patient required transfusion every 2 weeks since DAT positivity. The other two with positive DAT were not diagnosed with AIHA and did not require intervention. Patients with positive baseline alloAb (n = 6) had higher rate of irAE than patients without (n = 120) (13.3% vs 2.1%, p = 0.03), including 2 thyroid dysfunction, 2 skin rash, 2 myalgia, 2 hemolytic anemia. Patients with a positive DAT at baseline did not have a higher rate of irAEs (p = 0.09). Median OS for the entire cohort was 7.6 (5.8-9.9) months. No association was found between baseline positive DAT or alloAb and OS (p = 0.1748 and p = 0.3164 respectively). Conclusions: The incidence of hemolytic anemia in patients treated with IO was low. Presence of alloAb at baseline may predict irAE during IO treatment as patients with positive alloAb at baseline had a higher rate of irAE than those without.
A novel noninvasive biomarker based on peripheral PD-1posi CD8 T-cell receptor repertoire correlated with clinical outcomes to immunotherapy in non-small cell lung cancer.
Authors: Han Jie-Fei , Wang Zhijie , Bai Hua , Chen Si , Wang Yuqi , Duan Jianchun , Wang Jie
Published: May 2019

e14174

Background: Although advances on exploration of biomarkers personalized the immune checkpoint blockades (ICBs) delivery, non-invasive blood-based approach remainsan intriguing area for further investigation.This study investigated the feasibility of peripheral isolated PD-1posiCD8T cell receptor (TCR) repertoire profiling in predicting the clinical outcomes of ICBs treatment for non-small cell lung cancer (NSCLC) patients. Methods: The study comprised two independent cohorts (A and B), ultimately recruiting total 40 from 51 patients with stage IIIB-IV NSCLCs who received anti-PD-1/PD-L1 therapy between March 14, 2017 and May 2, 2018. Peripheral blood samples of pre- and post-ICBs (the timepoint of first imaging evaluation) were prospectively collected, and PD-1posiCD8 T cells were isolated by flow cytometry for TCR sequencing. The diversity and clonality of the TCR repertoire were calculated for biomarker profiling. Cohort A (n = 25) was used as a training set for discovery of TCR diversity in the prediction of response to ICBs, and cohort B (n = 15) as a validation set. Progression-free survival (PFS) and response to ICBs treatment were correlated with TCR diversity and clonality. Results: In cohort A, patients with high pre-ICBs PD-1posiCD8 TCR diversity had longer PFS than those with low diversity (6.5 vs. 2.6 months, p = .045), which were substantially validated in cohort B. By using the incorporated set, pre-ICBs PD-1posiCD8 TCR diversity exhibited an optimal Youden's index of 0.81 with a sensitivity of 0.87 and a specificity of 0.94 in clarifying clinical response of ICBs (PR+SD vs. PD). Patients with increased PD-1posiCD8 TCR clonality demonstrated significantly improved PFS (7.3 vs. 2.7 months, p = .005) compared to those with decreased TCR clonality. Interestingly, two patients with initial pseudo-PD exhibited similar change of TCR clonality and expansion of dominant TCR clones with those with PR, but not PD. Conclusions: Peripheral PD-1posiCD8 TCR repertoire sequencing might be a promising non-invasive approach for selecting patients who could benefit from ICBs treatment, which guaranteed the further investigation and validation by larger cohorts.
Correlation of CD28 expression on CD8+ T cells with prognosis in advanced lung adenocarcinomas and squamous cell carcinomas.
Authors: Liu Chao , Jing Wang , Zhu Hui , Yu Jinming
Published: May 2019

e14185

Background: Programmed death-1 (PD-1)-targeted therapies that treat non-small cell lung cancer (NSCLC) to salvage exhausted CD8 T cells depends on CD28 costimulatory pathway. However, the prognostic values of CD8+CD28+ T and CD8+CD28- T in advanced NSCLC and the impact of histological type on them remains unclear. Methods: Of 232 advanced NSCLC patients, 101 treatment-naïve individuals were eligible and included in our study. Flow cytometry was used to evaluate CD8+CD28+ T, CD8+CD28- T, CD4+CD25hi T, B, natural killer, γδT, and natural killer T cells in patients’ peripheral blood. Results: Multivariate analysis showed that increased levels of CD8+CD28+ T cells independently predicted favorable overall survival (OS) [hazard ratio (HR): 0.59, 95% confidence interval (CI): 0.28-0.93, P = 0.034] and progression-free survival (PFS) (HR: 0.60, 95%CI: 0.27-0.98, P = 0.033) in ADs but not in SCCs. In addition, high levels of CD8+CD28- T cells independently predicted unfavorable OS (HR: 1.57, 95%CI: 1.13-3.34, P = 0.039) and PFS (HR: 2.08, 95%CI: 1.02-4.23, P = 0.043) in SCCs but not in ADs. ADs had higher levels of CD4+CD25hi T and CD8+CD28- T, and lower NK (all P < 0.05) cell counts than SCCs. Conclusions: The prognostic values of peripheral CD8+CD28+ and CD8+CD28- T cells in advanced NSCLC patients differ depending on histological type. These results could help refine the CD28-dependent PD-1-targeted therapies for advanced NSCLC with ADs and SCCs.
Comprehensive analysis of lymphocyte loss kinetics in patients treated with focal fractionated radiation therapy.
Authors: Yalamanchali Anirudh , Jin Jian-Yue , Zhang Hong , Grossman Stuart A. , Hobbs Robert F , Ellsworth Susannah G.
Published: May 2019

e14191

Background: Radiation (RT)-induced lymphopenia (RIL) is associated with worse survival in patients with solid tumors as well as lower response rates to checkpoint inhibitors. This analysis aimed to model the kinetics of lymphocyte depletion during RT to assist in predicting RIL risk. Methods: This registry-based study included 419 patients who received either total body irradiation (TBI; n = 30), stereotactic body RT (SBRT; n = 73), or conventional chemoradiation (CRT; n = 316). For each patient, serial absolute lymphocyte counts (ALCs) were plotted against RT fraction number. The initial 3 weeks of treatment for conventionally irradiated patients and the entirety of treatment for SBRT and TBI patients were fit to exponential decay in the form ALC(x) = ae-bx. From those fits, percent per fraction loss in ALC (PFLAC) was calculated as PFLAC = (1 – e-b)*100, and multivariable regression was performed to find its significant predictors. Results: Curves were well fitted by exponential decay for all RT techniques (median linearized R2 0.98, 0.93, and 0.97 for patients treated with TBI, SBRT, and CRT, respectively). In CRT patients, apparent ALC loss rate slowed after week 3, potentially due to lymphocyte repopulation or other factors. TBI and SBRT patients completed RT before the end of the exponential decay phase, and their ALC loss rates remained unchanged throughout RT. Initial PFLAC varied significantly with treatment technique. Mean PFLAC was 35.5%, 24.3%, and 10.77% for patients treated with TBI, SBRT, and CRT, respectively (p < 0.001). Significant predictors of PFLAC varied by site and included field size, dose per fraction, mean spleen dose, chemotherapy backbone, and age. In pancreas cancer patients, gemcitabine was associated with a higher PFLAC (mean = 10.7) than 5-FU (mean = 8.3) after adjustment for covariates (p < 0.001). Finally, total % ALC loss during RT was highly correlated with PFLAC (p < 0.001). Conclusions: Lymphocyte depletion kinetics during the initial phase of fractionated RT are characterized by pure exponential decay. Initial PFLAC is strongly correlated with RT planning parameters and predicts total % ALC loss. The highest ALC loss rates were associated with RT-only regimens, implying that concurrent chemotherapy is not solely responsible for lymphopenia in patients receiving CRT. This work may also assist in selecting patients for adaptive RT approaches to mitigate RIL risk.
Precision probiotic therapy enhances immune checkpoint therapy efficacy in melanoma bearing mice.
Authors: Frankel Arthur E. , Honda Kenya , Roberts Bruce , Szabady Rose , Reddy Amit , Lightcap Johnny , McClellan Steve , Deshmukh Sachin Kumar , Koh Andrew Y.
Published: May 2019

e14195

Background: Immune checkpoint inhibitor therapy, ICT, achieves remissions in melanoma patients but factors modulating response are not well defined. Our group (Frankel et al. Neoplasia 2017) and others have identified specific gut microbiota associated with improved ICT response. Recently, we identified specific gut microbiota that induce adaptive immune responses and potentiate ICT (Tanoue et al. Nature 2019). In this study, we determined whether this predefined consortia of gut microbiota augment ICT efficacy in melanoma bearing mice. Methods: Mice (C57BL/6, 6-8wk old, female, Jackson, n = 4-12 mice) received ± antibiotic water (penicillin G 1500U/mL + streptomycin 2mg/mL) for 6 d to deplete gut microbiota. Mice were then inoculated with 105 B16F10 melanoma cells SQ. At d 4, 8, 12 post-tumor inoculation, 0.2 mg anti-mCTLA4 + anti-mPD1 antibodies (Bio X Cell) were administered IP. Precision probiotic therapies included Vedanta Bioscience VE800 (Tanoue et al., Nature 2019), VE804 (same as VE800 without R. lactatiformans and F. ulcerans), VE411 (four Clostridial firmicutes) (Narushima, 2014), and Lactobacillus acidophilus (ATCC 4356) probiotics were given via gavage (1x109 cfu) starting day +1 after tumor inoculation and 3xwkly. Loss of survival was defined as death or tumor diameters ≥ 2 cm. Tumor growth inhibition, TGI = (1- mean treated tumor volume/mean control tumor volume) x 100%. Tumor mononuclear cells were isolated for flow cytometry for murine CD4, CD8, and CD11c. Results: TGI in mice with intact gut microbiota and treated with ICT was 84 ± 4% (SEM). Pre-treatment antibiotics reduced TGI to 38 ± 11%. Groups treated with Vedanta VE800, VE804, and VE411 exhibited TGIs of 77 ± 9, 61 ± 8, and 69%, respectively, whereas treatment with Lactobacillus acidophilus achieved TGIs 57%. VE800 treated mice had significantly increased length of survival compared to mice treated with antibiotics (p = 0.0008, log-rank test). Length of survival was not significantly different between groups with intact gut microbiota and those pretreated with antibiotics and dosed with VE800 (p = 0.52, log-rank test). ICT increased tumor CD4 cells to 11% from 2% and CD8 cells to 9% from 1%., however pre-treatment with antibiotics reduced CD4 cells to 4% and CD8 cells to 1%. Conclusions: Defined consortia of gut microbiota facilitate ICT efficacy. These preclinical studies lay the foundation for optimizing the host response to ICT.
Correlation of inflammation-related markers with MDSC and IL-17, and use as prognostic indicators in patients with advanced gastric and colorectal cancers.
Authors: Shibata Masahiko , Ujiie Daisuke , Ashizawa Mai , Kikuchi Tomohiro , Okayama Hirokazu , Fujita Shoutarou , Sakamoto Wataru , Nakajima Takahiro , Endo Hisahito , Saito Motonobu , Momma Tomoyuki , Shimura Tatsuo , Kono Koji , Takenoshita Seiichi
Published: May 2019

e14204

Background: Although a causal relationship for inflammation and immunity of cancer is more widely accepted today, the precise cell mechanisms mediating this relationship have not been elucidated. Accumulating evidence suggests that myeloid-derived suppressor cells (MDSC), may contribute to the negative regulation of immune responses during cancer and inflammation. IL-17 is a pro-inflammatory cytokine that is primarily secreted by T helper (Th)17 cells and we have reported that IL-17 correlates with immunosuppressive conditions in patients with cancer. Methods: PBMC (peripheral blood mononuclear cells) were harvested from 106 patients including 43 with gastric and 63 with colorectal cancer. PBMC were stimulated with PHA (phytohemagglutinin) and the production of IL-17 was measured by ELISA. MDSC were detected by flow cytometry (CD11b+,CD14-,CD33+). The levels of CRP (C-reacting protein) and NLR (neutrophil to lymphocyte ratio) were used as inflammatory markers. Results: Both of MDSC and IL-17 production were increased in patients with advanced stages, and correlated with each other, inflammatory markers and immune suppression. The patients were divided with average levels of MDSC and IL-17 production and the prognosis were analyzed with Kaplan-Meier method. The overall survival of patients with high MDSC or high IL-17 production were significantly worse than those with low MDSC or low IL-17 production, respectively, in patients with stages III and IV, although the differences were not significant in patients with stages I and II. Conclusions: Thus these inflammatory markers are closely related with systemic inflammation involving IL-17 and with immunosuppression driven by MDSC, and are effective prognostic indicators in patients with stages III and IV gastric and colorectal cancer.
Microsatellite instability assessment of FFPE tumor samples through targeted next-generation sequencing.
Authors: Wong-Ho Elaine
Published: May 2019

e14213

Background: Immunotherapies to treat malignancies have shown great promise, however, only a fraction of cancer patients respond to immunotherapy, making it imperative to identify biomarkers that predict response. Cancer-associated instabilities at microsatellite locations throughout the genome have been shown to be predictive of response to immunotherapy treatment. Here we determine the accuracy of determining the MSI status through targeted sequencing using Ion Torrent sequencing technology. Methods: We have identified optimal chemistry and developed a novel algorithm to assess MSI status of samples using a large number of markers on the Ion GeneStudio S5 Series sequencer. The diverse marker set includes monomers that vary in length between 10 BP and 40 BP in addition to di-and tri-nucleotide STR markers. Each sample is assigned an MSI score based on features that measure MSI response of markers in the assay. Results: We evaluated performance of the MSI markers in the context of a comprehensive targeted next generation sequencing assay with the MSI algorithm. A large set of FFPE tumor samples and MSI controls were evaluated. The resulting scores were in concordance with results from capillary electrophoresis studies. Conclusions: In summary, a next-generation sequencing based assay using multiple markers was developed to assign MSI status to FFPE tumor samples. The accuracy of the assay was validated using an orthogonal test. This allows us to use Ion Torrent sequencing technology to identify changes in complex repeat regions of the genome as part of a comprehensive genomic profiling approach.
Modified ketogenic diet in solid tumors: A Veteran Affairs Pittsburgh Healthcare System case series.
Authors: Tan Jocelyn Lai , Carrick Jennifer , Passero Vida Almario , Shields Jenna , Liman Andrew Dede , Rai Hema , Harrold Lauri
Published: May 2019

e14217

Background: Cancers prefer glucose due to mitochondrial dysfunction. Ketogenic diets fuel normal cells while starving cancer. Sparse data exists on ketogenic diets in solid tumors. Methods: Studied 5 advanced solid tumor patients on a modified ketogenic diet plus standard of care chemotherapy. Daily carbohydrate limited to 20-40 grams, no sweets or starch. Histories and clinical courses are presented. Results: Case 1: 70 year old male, medically inoperable Stage IIIA poorly differentiated squamous cell lung cancer and non-hodgkins lymphoma. Treated with radiation and paclitaxel/carboplatin. After 5 months, lung cancer relapsed in a neck mass, treated with chemoradiation plus the diet. At 46 months, scans and blood counts were normal. Case 2: 42 year old, 417 lb. male diabetic stage IV colon cancer, spread to lungs and liver. Progressed on 5FU/bevacizumab/oxaliplatin, irinotecan, tipiracil/trifluridine, regorafenib but responded to 5FU/irinotecan rechallenge plus liver chemoembolization. Dieted, mild ketosis but lost 64 lbs, stable disease at 6.5 years. Case 3: 42 year old male, Stage III BRAF V600 E melanoma, Vemurafenib intolerant. Bulky axillary nodes resected plus diet for 3 years. Complete remission at 7 yrs 9 months. Case 4: 57 year old male, medically inoperable liver cancer, normal AFP values and normal scans after 5 months of sorafenib. AFP later increased to 690 ng/ml. Advised to change treatment but he refused and started the ketogenic diet with the sorafenib. After 5 months, AFP improved to 445 ng/ml. Sorafenib and diet interrupted for 3 months, AFP rose to 1017. At 7 yrs, 4 months, back on sorafenib and diet, AFP 1520 mg/ml, negative scans. Case 5: 49 year old male, aggressive digital papillary skin adenocarcinoma, multi-organ metastases. Had side-effects with many chemotherapies (paclitaxel/bevacizumab/carboplatin, sorafenib, gemcitabine, oxaliplatin/5FU, cyclophosphamide) Stable disease on diet despite multiple therapy changes. At 34 months, he was off the diet, tumors grew plus new brain lesions (resected). Alive on atezolizumab at 36 months. Conclusions: Ketogenic diets in human solid tumors seem well tolerated and may improve response and survival after standard therapy.
Spatially-resolved, multiplexed quantification of protein and mRNA distribution and abundance in colorectal cancer tumor microenvironment with multiple modalities of NanoString GeoMx Digital Spatial Profiler.
Authors: Demirkan Gokhan , Merritt Chris , Hinerfeld Douglas , Ong Giang , Sorg Kristina , Balasundaram Gayathri , Dunaway Dwayne , Geiss Gary K , Beechem Joseph M
Published: May 2019

e14218

Background: Characterization of the spatial distribution and abundance of proteins and mRNAs with morphological context within tissues enables a better understanding of biological systems in many research areas, including immunology and oncology. However, it has proven difficult to perform such studies in a highly multiplexed manner. To address this unmet need, we have developed a novel optical-barcode based microscope and tissue-sampling platform designed to simultaneously analyze hundreds of proteins or mRNAs on a single FFPE section from distinct tissue spatial regions. Methods: Using colorectal cancer FFPE sections we spatially resolve protein and mRNA expression of immune-oncology targets and present a series of modalities and associated applications for the GeoMx Digital Spatial Profiler platform. Results: Microsatellite stable (MSS) or instable (MSI) colorectal tumors will be characterized to evaluate active and suppressive immune mechanisms in both immune dense regions and tumor versus stroma by utilizing segment profiling. Regions from the invasive margins and tumor centers will be investigated with contour profiling to define different immunosuppressive and activated immune phenotypes. Evaluation of tumor versus stroma will also identify pathways related to each compartment that are different between MSI and MSS tumors. These techniques can be used to discover drug mechanism of action or immune activation status, as well as to facilitate prediction of treatment response and disease progression or investigation of specific rare cell populations’ molecular profiles. Conclusions: This study will demonstrate multiplexed pathway-level protein and gene expression analysis from discrete regions within a colorectal tumor (tumor center and immune invasive margin), enabling systematic interrogation of immune activity in FFPE samples. Further studies could eventually lead to the identification of unique localized immune characteristics to guide combination therapeutic approaches.
Applying machine learning strategy for microsatellite status detection in plasma sample type.
Authors: Zhao Lili , Han Wenbo , Zhou Wei , Fang Lu , Hong Yuanyuan , Tian Chen , Chen Weizhi , He Ji
Published: May 2019

e14219

Background: Microsatellite(MS) status is an important biomarker in cancer diagnosis . Most of available detection algorithms were based on figuring out number of validated allele types from control and cases, then determine the MS status by identify the difference. That would be suitable for tissue like samples due to their clear and sufficient tumor nucleic acid content. But for liquid biopsy such as plasma, lower ctDNA concentration and higher background ratio induced from laboratory and sequencing procedure make it necessary to develop novo method for plasma MS status detection. Methods: We developed an machine learning based approach to detect the MS status by using NGS sequencing data from plasma sample. 98 blood cell , 12 tumor tissue samples with PCR confirmed MSS status and 87 plasma samples from 134 patients were used to build normal baseline, and check the sequencing stability and population polymorphism of 207 candidate SSR locus in our panel. 97 locus then passed and were chosen as original biomarkers for model construction. For each coming sample, the allele probability distribution were obtained for each locus. Followed by calculating the Kullback–Leibler Divergence (KLD) value between target and baseline samples for each locus, the KLD value was used as input for feature selection and machine learning model fitting. Finally a leave one out strategy was used for model performance evaluation. Results: We obtained 17 most relevant SSR locus with suitable for MS status analysis of plasma sample. By using a SVM model fitted with 49 real patients sample data (21 MSI and 28 MSS) , The MS status prediction of plasma sample was of high sensitivity and specificity which were 90.47% and 96.42% respectively. Conclusions: Our results suggest that our MSI calculation model by using KLD value and SVM method is appropriate for the plasma sample. Further large-scale prospective studies are needed to validate our model.
MDNA109: Effect of an interleukin-2 superkine on CD8 T-cell properties in the tumor microenvironment.
Authors: Rafei Moutih , Fidai Shafique , Merchant Rosemina , Merchant Fahar
Published: May 2019

e14220

Background: Proleukin (a cysteine-modified variant of interleukin (IL)-2) is the only common-γc cytokine approved for the treatment of metastatic melanoma and renal cell carcinoma. Its therapeutic use is however hampered by its short half-life and severe toxicity. At low doses, Proleukin administration leads to preferential activation of regulatory T cells (Tregs) due to their expression of the high affinity trimeric IL-2 receptor (IL-2R) consisting of CD25, CD122 and CD132 instead of cytotoxic CD8 T cells expressing the intermediate affinity IL-2R (CD122 and CD132). Methods: To bypass these limitations, we evaluated engineered variants of IL-2 (MDNA109 superkine) exhibiting enhanced affinity towards CD122 including their long-acting and bispecific superkine fusions. This approach allows preferential activation of CD8 T cells while displaying reduced adverse effects in vivo. Results: Both Biacore analysis and signaling studies on human peripheral blood mononuclear cells confirmed enhanced binding of MDNA109 to CD122 and STAT5 activation, respectively. When tested in vivo, MDNA109 co-administration with the immune-checkpoint blockers anti-programmed cell death (PD)-1 or anti-cytotoxic T-Lymphocyte-Associated Protein (CTLA)4 cured mice with pre-established MC38 or CT26 colon cancers respectively. In addition, bi-weekly administration of the long-acting MDNA109-Fc fusion led to similar therapeutic outcomes in the B16F10 model when compared to MDNA109 administered daily. Finally, in vitro and in-vivo results using (a) MDNA109 muteins with completely impaired CD25 binding activity and (b) novel bispecific superkine fusions consisting of MDNA109 and a dual IL-4/IL-13 super-antagonist capable of selective CD8 T-cell activation while mitigating the suppressive functions of myeloid-derived suppressor cells and tumor-associated macrophages will be presented. Conclusions: Altogether, these data demonstrate that both MDNA109 and MDNA109-Fc are therapeutically superior to Proleukin. Furthermore, use of MDNA109-based strategies will not only ensure complete abrogation of adverse effects, but will in addition eliminate immunosuppression caused by both Tregs and myeloid cells.
An umbrella for the cytokine storm: Enabling precision detection of CRS in CAR-T therapy.
Authors: Quan Qimin
Published: May 2019

e14223

Background: Cytokine release syndrome (CRS), a systemic inflammatory response observed with monoclonal antibody drugs and adoptive T cell treatments, has become a major issue for CAR-T therapy. CRS can present as a mild reaction requiring minimally invasive supportive care up to a severe systemic response resulting in patient death. Monitoring this response during these therapeutic treatments is non-trivial due the wide range of biomarker concentrations, small sample volumes, and long assay times. Current analytical methods are unable to address these needs, limiting the precision of CAR-T therapy and effective management of its side effects. Methods: Emerging studies in this area have focused in establishing a panel of predictive biomarkers to manage dosing and early interventions, among them, IFNγ, IL6, TNFα, MIP1 have shown predicative powers in pediatric patients. Nevertheless, a significant improvement (100x) on the detection sensitivity is required to predict the CRS response with currently available methods. In addition, CRS-associated biomarkers including CRP and ferritin vary from 10ng/mL-10mg/mL while other predictive biomarkers (eg, IL6, IFNγ, etc.) vary from 1pg/mL-100ng/mL. At present, no analytical tool, known to us, can provide this large dynamic range ( > 9 logs), with the requisite lower limit of detection, in a rapid single test to predict and differentiate low, medium or high grade responses. Results: We present the NanoMosaic platform, the technology that has requisite sensitivity and breadth of dynamic range to enable precision detection based on precise quantitation of CRS-relevant biomarkers. NanoMosaic technology is enabled by single molecule nanoneedle sensors that are densely integrated on a silicon chip and manufactured with a CMOS-compatible process. Absolute quantitation is achieved by imaging the spectrum of nanoneedles, corresponding directly to the number of molecules. Conclusions: Direct comparison of different protein biomarkers with orders of magnitude concentration variations becomes possible in one platform and small sample sizes. We envision NanoMosaic technology will not only drive biomarker discovery, but also enable precise dosing management for CAR-T therapy.
Collagen affinity improves efficacy and safety of antibody and cytokine cancer immunotherapy.
Authors: Ishihara Ako , Ishihara Jun , Hubbell Jeffrey A , Swartz Melody A
Published: May 2019

e14232

Background: Cancer immunotherapy with immune checkpoint inhibitors (CPI) and interleukin (IL)-2 have demonstrated clinical antitumor efficacy but is frequently accompanied with severe side-effects caused by excessive and systemic immune system activation. Methods: We addressed this need by targeting both the CPI antibodies anti-CTLA4 (aCTLA4) + anti-PD-L1 (aPD-L1) and the cytokine IL-2 to tumors via conjugation (for the antibodies) or recombinant fusion (for the cytokine) to a collagen-binding domain (CBD) derived from the blood protein von Willebrand factor (VWF) A3 domain. This approach harnesses the exposure of tumor stroma collagen to blood components due to the leakiness of the tumor vasculature. Results: We show that intravenously (i.v.) administered CBD protein accumulated in tumors in an orthotopic MMTV-PyMT breast tumor model. CBD conjugation or fusion decreased the systemic toxicity of both aCTLA4+aPD-L1 combination therapy and IL-2. CBD-conjugation to CPI abolished T cell infiltration into the liver and eliminated hepatotoxicity as well as decreased systemic cytokine release in the blood serum. CBD-fusion to IL-2 ameliorated capillary leak syndrome and pulmonary edema. These side-effects have been reported in the clinic. CBD-CPI and CBD-IL-2 significantly suppressed tumor growth compared to their unmodified forms in B16F10 melanoma, CT26 colon carcinoma and MMTV-PyMT slow growing breast cancer models, and both CBD-CPI and CBD-IL-2 increased tumor-infiltrating CD8+ T cells; increases in the ratio of effector CD8+T cells to T regulatory cells were observed. In an orthotopic breast tumor model, combination treatment with CPI and IL-2 eradicated tumors in 9/13 animals with the CBD-modified drugs, whereas it did so in only 1/13 animals with the unmodified drugs. Conclusions: Our data suggest that the A3 domain of VWF can be used to engineer immunotherapies (e.g. CPI and cytokine) with high translational promise as systemically-administered tumor targeting drugs with improved safety and efficacy compared to their native forms.
Discovery of a novel Toll-like Receptor 7 agonist for systemic immunotherapy of cancer.
Authors: Appleman James Richard , Webber Stephen Evan
Published: May 2019

e14246

Background: While ICPIs (immune checkpoint inhibitors) have fundamentally changed the practice of cancer therapy for tumors arising from many different tissues, ways to increase both response rate and durability are critically needed. On theoretical grounds, combining stimulators of innate immunity with activators of adaptive immunity should lead to better outcomes. We have therefore created a series of novel Toll-like receptor 7 (TLR7) agonists that are intended to be combined with activators of adaptive immunity for the treatment of cancer. We have previously reported a series of unusual properties that must be met for successful drug candidates targeting TLR7 intended for chronic and systemic administration for the treatment of cancer. Methods: Key assays in our testing cascade include: cellular reporter assays and hPBMC assays to evaluate selectivity and potency; characterization in cynomolgus monkeys: appropriate PK with active TLR7 agonist, efficient oral delivery into systemic circulation with prodrugs of the TLR7 agonist, and targeted pharmacodynamic response at relevant oral dose (gating for candidate selection); antitumor activity alone and in combination with other immunotherapeutics in syngeneic rodent tumor models. Results: Appropriate potency at and specificity for TLR7 was achieved early in this discovery program. Furthermore, the PK profile after IV administration met our previously defined criteria. However, oral absorption of candidate prodrugs of two lead molecules was limited. In one class of prodrugs this was due to insufficient lipophilicity; in another class this was a consequence of MDR1 activity. Next-generation candidates have been synthesized; these candidates are expected to have substantially better oral delivery in vivo based upon results in predictive in vitro models. Testing in advanced animal models is in progress. Conclusions: From our original starting point - a relatively weak TLR7 agonist with no oral bioavailability - we have invented a novel series of molecules that are designed to be dosed QOD continuously over a 24-month period to appropriately engage innate immunity at a level that is well-tolerated by the patient while increasing treatment response rate and durability.