Early Marker Of Response Research Articles

First, do no harm: the role of preclinical animal models in predicting adverse events in gene therapy clinical trials for Duchenne muscular dystrophy and X-Linked myotubular myopathy

The occurrence of severe adverse events (SAEs) in patients with Duchenne muscular dystrophy (DMD), X-linked myotubular myopathy (XLMTM), and other neuromuscular diseases treated with adeno-associated virus (AAV) constructs has prompted studies to improve the safety and efficacy of gene therapy. Physicians have weighed the medical tenet of “first, do no harm” against the perspective of patients with progressive life-threatening conditions who may accept greater risk. Regarding SAE pathogenesis, discussion has focused on total AAV exposure and patient mutations more likely to induce immunity, while stressing the limitations of animal models in predicting adverse events. Therapeutic strategies for reducing side effects have employed more myotropic AAV serotypes and efficient transgenes. Other recommendations include excluding certain DMD gene mutations associated with SAEs and substituting less immunogenic transgenes such as utrophin (DMD) and myotubularin-related protein (XLMTM). For the sake of preclinical studies, emphasis has been placed on outbred rodents and larger animals that better predict immunity. Here, the absence of side effects in canine DMD and XLMTM models might be explained partly by phenotypic differences between affected humans and dogs. Specifically, dystrophin- and myotubularin-deficient dogs exhibit milder lesions, including less muscle fat deposition and the absence of hepatopathy, respectively, which could lead to reduced immune responses to AAV constructs. To better predict future problems, thought should be given to tracking early subclinical markers of the innate immune response, especially complement activation. Regardless of steps taken to improve the predictive value of animal models for SAEs, some questions will only be answered through human clinical trials after carefully considering the risk-benefit ratio.

Albumin-To-Alkaline Phosphatase Ratio as a New Early Predictive Marker of Axillary Response in Breast Cancer Patients Undergoing Neoadjuvant Chemotherapy: A Pilot Study

Background and Objectives: The Albumin-to-Alkaline Phosphatase ratio (AAPR) is an easily applicable and cost-effective marker investigated as an outcome predictor in solid cancers. Preliminary evidence in breast cancer suggests that a low AAPR correlates with a poor response to neoadjuvant chemotherapy (NAC) in primary tumors. However, data regarding the axillary response are lacking. This study aims to evaluate whether the AAPR can predict the axillary response in initially nodal-positive (cN+) breast cancer patients undergoing NAC. Materials and Methods: Clinical and biochemical variables of cN+ breast cancer patients undergoing NAC were collected. Pre-NAC albumin and alkaline phosphatase serum values were utilized in the AAPR calculation. Fisher’s exact test was performed to identify differences between the two groups of patients (high and low AAPR according to the cut-off reported in the literature). The primary outcome was the nodal pathologic complete response (pCR) rate in the two groups of patients. Results: Nodal pCR was achieved in 20/45 (44.4%) patients. A total of 36/45 (80%) patients had an AAPR > 0.583. Among patient and tumor characteristics, the only statistically significant difference between the two groups was the axillary pCR between the low and high AAPR groups (p-value = 0.03, OR = 0.129, 95% CI = 0.00–0.835). Conclusions: This pilot study suggests that the pre-treatment AAPR might be an early predictor of axillary response in cN+ breast cancer patients undergoing NAC. This result justifies further investigation in larger prospective trials to validate this finding.

Tumour-informed liquid biopsies to monitor advanced melanoma patients under immune checkpoint inhibition

Immune checkpoint inhibitors (ICI) have significantly improved overall survival in melanoma patients. However, 60% experience severe adverse events and early response markers are lacking. Circulating tumour DNA (ctDNA) is a promising biomarker for treatment-response and recurrence detection. The prospective PET/LIT study included 104 patients with palliative combined or adjuvant ICI. Tumour-informed sequencing panels to monitor 30 patient-specific variants were designed and 321 liquid biopsies of 87 patients sequenced. Mean sequencing depth after deduplication using UMIs was 6000x and the error rate of UMI-corrected reads was 2.47×10−4. Variant allele fractions correlated with PET/CT MTV (rho=0.69), S100 (rho=0.72), and LDH (rho=0.54). A decrease of allele fractions between T1 and T2 was associated with improved PFS and OS in the palliative cohort (p = 0.008 and p < 0.001). ctDNA was detected in 76.9% of adjuvant patients with relapse (n = 10/13), while all patients without progression (n = 9) remained ctDNA negative. Tumour-informed liquid biopsies are a reliable tool for monitoring treatment response and early relapse in melanoma patients with ICI.

Cisplatin-Induced Metabolic Responses Measured with Raman Spectroscopy in Cancer Cells, Spheroids, and Canine-Derived Organoids.

Ex vivo assessment of drug response with conventional cell viability assays remains the standard practice for guiding initial therapeutic choices. However, such ensemble approaches fail to capture heterogeneities in treatment response and cannot identify early markers of response. Here, we leverage Raman spectroscopy (RS) as an accurate, low-cost, extraction-free, and label-free approach to track metabolic changes in cancer cells, spheroids, and organoids in response to cisplatin treatment. We identified 12 statistically significant metabolites in cells and 19 metabolites in spheroids and organoids as a function of depth. We show that the cisplatin treatment of 4T1 cells and spheroids results in a shift in metabolite levels; metabolites including nucleic acids such as DNA, 783 cm-1 with p = 0.00021 for cells; p = 0.02173 for spheroids, major amino acids such as threonine, 1338 cm-1 with p = 0.00045 for cells; p = 0.01022 for spheroids, proteins such as amide III, 1248 cm-1 with p = 0.00606 for cells; p = 0.00511 for spheroids serve as early predictors of response. Our RS findings were also applicable to canine-derived organoids, showing spatial variations in metabolic changes as a function of organoid depth in response to cisplatin. Further, the metabolic pathways such as tricarboxylic acid (TCA)/citric acid cycle and glyoxylate and dicarboxylate metabolism that drive drug response showed significant differences based on organoid depth, replicating the heterogeneous treatment response seen in solid tumors where there is a difference from the periphery to the tumor core. Our study showcases the versatility of RS as a predictive tool for treatment response applicable from cells to organotypic cultures, that has the potential to decrease animal burden and readout time for preclinical drug efficacy.

Peri-Tumoural Lipid Composition and Hypoxia for Early Immune Response to Neoadjuvant Chemotherapy in Breast Cancer.

The deregulation of monounsaturated, polyunsaturated, and saturated fatty acids (MUFAs, PUFAs, SFAs) from de novo synthesis and hypoxia are central metabolic features of breast tumour. Early response markers for neoadjuvant chemotherapy (NACT) are critical for stratified treatment for patients with breast cancer, and restoration of lipid metabolism and normoxia might precede observable structural change. In this study, we hypothesised that peri-tumoural lipid composition and hypoxia might be predictive and early response markers in patients with breast cancer undergoing NACT. Female patients with breast cancer were scanned on a 3T clinical MRI scanner at baseline and Cycle1, with acquisition of lipid composition maps of MUFAs, PUFAs, and SFAs, and hypoxia maps of effective transverse relaxation rate R2*. The percentage change in lipid composition and hypoxia at Cycle1 was calculated with reference to baseline. Tumour-associated macrophages were analysed based on immunostaining of CD163 from biopsy and resection, with the percentage change in the resected tumour calculated across the entire NACT. We found no significant difference in lipid composition and R2* between good and poor responders at baseline and Cycle1; however, the correlation between the percentage change in MUFAs and PUFAs against CD163 suggested the modulation in lipids with altered immune response might support the development of targeted therapies.

Diffusion-weighted magnetic resonance imaging as an early prognostic marker of chemoradiotherapy response in squamous cell carcinoma of the anus: An individual patient data meta-analysis

Background and purposeSquamous cell carcinoma of the anus (SCCA) can recur after chemoradiotherapy (CRT). Early prediction of treatment response is crucial for individualising treatment. Existing data on radiological biomarkers is limited and contradictory. We performed an individual patient data meta-analysis (IPM) of four prospective trials investigating whether diffusion-weighted (DW) magnetic resonance imaging (MRI) in weeks two to three of CRT predicts treatment failure in SCCA. Material and methodsIndividual patient data from four trials, including paired DW-MRI at baseline and during CRT, were combined into one dataset. The association between ADC volume histogram parameters and treatment failure (locoregional and any failure) was assessed using logistic regression. Pre-defined analysis included categorising patients into a change in the mean ADC of the delineated tumour volume above and below 20%. ResultsThe study found that among all included 142 patients, 11.3 % (n = 16) had a locoregional treatment failure. An ADC mean change of <20 % and >20 % resulted in a locoregional failure rate of 16.7 % and 8.0 %, respectively. However, no other ADC-based histogram parameter was associated with locoregional or any treatment failure. ConclusionsDW-MRI standard parameters, as an isolated biomarker, were not found to be associated with increased odds of treatment failure in SCCA in this IPM. Radiological biomarker investigations involve multiple steps and can result in heterogeneous data. In future, it is crucial to include radiological biomarkers in large prospective trials to minimize heterogeneity and maximize learning.

Single-cell Transcriptional Landscape of Temporal Neutrophil Response to Burn Wound in Larval Zebrafish.

Neutrophils accumulate early in tissue injury. However, the cellular and functional heterogeneity of neutrophils during homeostasis and in response to tissue damage remains unclear. In this study, we use larval zebrafish to understand neutrophil responses to thermal injury. Single-cell transcriptional mapping of myeloid cells during a 3-d time course in burn and control larvae revealed distinct neutrophil subsets and their cell-cell interactions with macrophages across time and conditions. The trajectory formed by three zebrafish neutrophil subsets resembles human neutrophil maturation, with varying transition patterns between conditions. Through ligand-receptor cell-cell interaction analysis, we found that neutrophils communicate more in burns in a pathway and temporal manner. Finally, we identified the correlation between zebrafish myeloid signatures and human burn severity, establishing GPR84+ neutrophils as a potential marker of early innate immune response in burns. This work builds a comparative single-cell transcriptomic framework to identify neutrophil markers of tissue damage using model organisms.

Clozapine treatment and astrocyte activity in treatment resistant schizophrenia: A proton magnetic resonance spectroscopy study

Clozapine is the only antipsychotic approved for treating treatment-resistant schizophrenia (TRS), characterized by persistent positive symptoms despite adequate antipsychotic treatment. Unfortunately, clozapine demonstrates clinical efficacy in only ~30–60 % of patients with TRS (clozapine-responders; ClzR+), while the remaining ~40–70 % are left with no pharmacological recourse for improvement (clozapine-resistant; ClzR−). Mechanism(s) underlying clozapine's superior efficacy remain unclear. However, in vitro evidence suggests clozapine may mitigate glutamatergic dysregulations observed in TRS, by modulating astrocyte activity in ClzR+, but not ClzR−. A factor that if proven correct, may help the assessment of treatment response and development of more effective antipsychotics. To explore the presence of clozapine-astrocyte interaction and clinical improvement, we used 3 T proton-magnetic resonance spectroscopy to quantify levels of myo-Inositol, surrogate biomarker of astrocyte activity, in regions related to schizophrenia neurobiology: Dorsal-anterior-cingulate-cortex (dACC), left-dorsolateral-prefrontal-cortex (left-DLPFC), and left-striatum (left-striatum) of 157 participants (ClzR− = 30; ClzR+ = 37; responders = 38; controls = 52). Clozapine treatment was assessed using clozapine to norclozapine plasma levels, 11–12 h after last clozapine dose. Measures for symptom severity (i.e., Positive and Negative Symptoms Scale) and cognition (i.e., Mini-Mental State Examination) were also recorded. Higher levels of myo-Inositol were observed in TRS groups versus responders and controls (dACC (p < 0.001); left-striatum (p = 0.036); left-DLPFC (p = 0.023)). In ClzR+, but not ClzR−, clozapine to norclozapine ratios were positively associated with myo-Inositol levels (dACC (p = 0.004); left-DLPFC (p < 0.001)), and lower positive symptom severity (p < 0.001). Our results support growing in vitro evidence of clozapine-astrocyte interaction in clozapine-responders. Further research may determine the viability of clozapine-astrocyte interactions as an early marker of clozapine response.

NURS-21. EXPLORATION OF CORRELATIVE STUDIES IN EARLY PHASE TRIALS FOR PEDIATRIC AND YOUNG ADULT CENTRAL NERVOUS SYSTEM (CNS) TUMORS AND THE POTENTIAL TRANSLATION TO CLINICAL CARE

Abstract BACKGROUND Correlative studies are common in early phase clinical trials. In addition to primary aims establishing maximum tolerated dose or efficacy, supplementary information from biospecimens is frequently collected with patient consent. The Pediatric Neuro-Oncology Consortium (PNOC) has included several correlative studies with implications for understanding patterns in the development of tumors, response to treatment, and prognosis. A needs assessment survey of PNOC institutions by the Advanced Practice Provider and Investigational Nurse Committee (APPNIC) highlighted that nurses and advanced practice providers (APPs) desire further education regarding correlative studies. METHODS Blood, stool, cerebrospinal fluid (CSF) and hair strands are examples of correlative biospecimens incorporated in PNOC protocols. We will share a review of the literature and implications for practice. RESULTS Blood, drawn in conjunction with clinical labs, is commonly analyzed for circulating tumor DNA to conduct molecular profiling in blood compared to tissue, explore less invasive means of molecular evolution of tumor somatic mutations, and identify early markers of tumor response or resistance. Stool is collected to utilize the gut microbiome as a lens into treatment response in CNS tumors via the gut-brain axis. Stool samples are compared to normal controls and longitudinally within patients during treatment. CSF sampling provides less invasive disease monitoring, tumor molecular characterization, markers of tumor response or resistance, and drug pharmacokinetics. Hair sampling can be used to further quantify drug levels for pharmacokinetics and assess treatment compliance. The robustness of these correlative samples collected at various timepoints (diagnosis, pre-surgery and post-surgery, post-radiation and/or with recurrence) informs on prognosis, response to treatment; and ultimately, interventions to improve outcomes for pediatric and young adult CNS tumors. CONCLUSION Our assessment identified a need for APP and nursing-focused education on biospecimen collection to inform on emerging clinical and research advances while improving protocol compliance.

Intraoperative Oxygen Consumption and Postoperative Immune Response in Colorectal Oncological Surgery: A Prospective, Monocentric Pilot Study.

Surgical resection is the key treatment for colorectal cancer, but the extent of surgical trauma has been implied as a key factor for the oncologic outcome. The immune stress response to surgical trauma generates a cascade of immunological events implying neutrophils' perioperative change generating NETosis, N killer decrease, and platelets' activation that may influence postoperative surgical outcome, tumor cell growth, and future oncogenesis. The present study aimed to investigate the correlation between intraoperative oxygen consumption (VO2) and the dynamic variation of neutrophils, lymphocytes, and platelets in the perioperative period to identify an intraoperative tool that could predict the postoperative immune response. Twenty-six colorectal oncological surgical patients were enrolled in an observational, prospective, monocentric study, over 18 months. Serum neutrophils, lymphocytes, and thrombocytes values were collected in the preoperative period and on the third postoperative day, oxygen consumption was measured and recorded every 15 min during surgery using indirect calorimetry. We compared oxygen consumption measurements registered 30 min after induction of anesthesia (VO2a) and the first value registered after abdominal wall closure (VO2b) to perioperative variation of absolute neutrophils (VNC), lymphocytes (VLC), and platelets (VPC) count. Our results proved a significant correlation between VO2 variation and neutrophils' perioperative dynamic assessed by VNC (correlation coefficient = 0.547, p < 0.01, 95% confidence interval (CI) =0.175, 0.783). We also noticed a correlation between VPC and VO2 (correlation coefficient = -0.603, p < 0.01, 95% CI = -0.815, -0.248). No correlation could be shown between VO2 and VLC variation (p = 0.39). In conclusion, intraoperative VO2 variation measured by indirect calorimetry correlates well with perioperative neutrophils and platelets count dynamic variations and can be used as an early prognosis marker of postoperative immune response and surgical outcome in colorectal oncological surgery.

Abstract PO2-14-06: CIRCULATING TUMOR DNA DYNAMICS IN THE FIRST TREATMENT LINE OF THE SONIA TRIAL

Abstract Background and objective: CDK4/6 inhibitors (CDK4/6i) improve the outcome of patients with ER+/HER2- advanced breast cancer when added to endocrine therapy in the first line or second line of treatment. The recently reported SONIA trial shows no difference in progression free survival after two lines of treatment (PFS2), overall survival (OS) or quality of life benefit of CDK4/6i in first-line compared to second-line, while adverse events are substantially higher when CDK4/6i is added in the first-line. However, a subgroup of patients shows early progression on endocrine mono-treatment and not all patients respond equally well to CDK4/6i. This highlights the need for early response markers during the first weeks of treatment to reduce unnecessary toxicity while providing the most optimal care. In this SONIA side study, we aim to investigate whether we could identify patients who will progress rapidly during first line of therapy by using circulating tumor DNA (ctDNA) dynamics. Methods: Blood samples were obtained from patients included in the SONIA trial in both the first line CDK4/6i + aromatase inhibitor (AI) combination arm (arm A) and the first line AI monotherapy arm (arm B) at three time points: before start, after two weeks and after three months of first line treatment. Cell free DNA (cfDNA) was isolated from all samples and subsequently baseline cfDNA was sequenced with the Oncomine™ Breast cfDNA Assay v2 (Thermofisher), a focused NGS panel of 10 genes. Dedicated dPCR assays were used to track an identified driver mutation over time. Results: Blood samples of 332 patients (206 in arm A and 126 in arm B) before start of first line treatment and after two weeks of treatment were obtained. At baseline, in 142 of the 332 (42.7%) patients at least one somatic mutation was detected. In 30 patients two or more mutations were identified, in which case the mutation with the highest variant allele frequency (VAF) was followed over time. The most frequently mutated genes were PIK3CA (102 patients (30.7% of the total included cohort)), TP53 (32 patients (9.6%)), SF3B1 (8 patients (2.4%) and ESR1 (6 patients (1.8%)). The median observed VAF at baseline was 4.1% (IQR: 1.5%-13.1%). Of the 142 patients with detectable ctDNA at baseline, 141 patients and 100 patients, respectively, had blood samples taken after 2 weeks and after 3 months of treatment. After two weeks, the VAF of the tracked mutation was increased in 15.2% (14/92) of the patients in arm A and in 18.4% (9/49) of the patients in arm B. At the same time 40.2% (37/92) of the patients in arm A and 34.7% (17/49) of the patients in arm B had cleared ctDNA. No difference was observed between the arms for both an increase in VAF (P=0.63, Chi-square test) and ctDNA clearance (P=0.52, Chi-square test). After 3 months an increased VAF of the tracked mutation compared to baseline was observed in 6.3% (4/64) of the patients in arm A and 8.3% (3/36) of the patients in arm B. At this timepoint 59.4% (38/64) of the patients in arm A and 61.1% (22/36) of the patients in arm B had cleared ctDNA. No difference was observed between the arms for both ctDNA clearance (P=0.86, Chi-square test) and increase in VAF (P=0.70, Chi-square test) after three months. Conclusions: ctDNA was detected at baseline in almost half of the patients in the SONIA study. Compared to baseline, a decreasing number of patients had an elevation of the VAF of the tracked mutation and an increasing number of patients showed ctDNA clearance from 2 weeks to 3 months after start of treatment. There was no difference between the two arms observed regarding ctDNA dynamics. Interestingly, irrespective of treatment arm, about 1/3rd of patients remained ctDNA-positive even after 3 months of treatment. Associations of these ctDNA data with the clinical outcome data are currently being evaluated and will be presented at the meeting. Citation Format: Elisabeth M Jongbloed, Stefania Stella, Lotte van Bergen, Corine Beaufort, Jean Helmijr, Vanja de Weerd, Frans Erdkamp, Joan Heijns, Yvonne Kamm, Anne-Marie van Riel, Quirine van Rossum-Schornagel, Maurice Jansen, Inge RHM Konings, Gabe Sonke, John WM Martens, Agnes Jager, Saskia M Wilting. CIRCULATING TUMOR DNA DYNAMICS IN THE FIRST TREATMENT LINE OF THE SONIA TRIAL [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-14-06.

Molecular Profiling to Understand Treatment Resistance and Response in Glioblastoma

INTRODUCTION: Patients with glioblastoma experience a wide variation in response to standard treatment, with nearly 30% experiencing tumour progression during treatment, and nearly 6% surviving more than 5 years. To date, there are few non-invasive clinical biomarkers to predict response to firstline treatment. Chemical exchange saturation transfer (CEST) MRI may have the potential to fill this gap. CEST MRI is sensitive to treatment-induced changes and changes in tumor metabolism1. Our team has obtained CEST data for patients before, during and after standard chemoradiation treatment, and found that CEST provides markers of early response and can identify early, standard and late progressors before treatment initiation.2 METHODS: Patients (n = 104) with primary, IDH wild-type glioblastoma were imaged with CEST-MRI at multiple time points throughout standard chemoradiation treatment. DNA and RNA were co-extracted from matched normal and tumour pairs and processed for whole genome sequencing and gene expression analysis using Nanostring. Clinical variables such as age, extent of resection, sex, ECOG status and MGMT promoter methylated were also collected. A survival analysis was conducted using the Kaplan-Meier method with log-rank tests. Univariate and multi-variate hazard ratios for clinical variables were calculated by fitting Cox Proportional Hazards Models. RESULTS: Early progressors reported a median progression-free survival (PFS) of 142 days compared to 296.5 in standard and 832 days in late progressors (p &lt; 0.0001). Early progressors also harbored distinct and statistically significant differences in gene expression and genomic alterations, namely in DNA damage repair, glucose transport and arginine metabolism pathways. A gene signature was prognostic of PFS and overall survival in a Cox proportional hazards model. CONCLUSIONS: This data may serve as a radiogenomic biomarker to assess treatment response within early phases of treatment and allow for a personalized treatment plan.

CD4+ T cell activation distinguishes response to anti-PD-L1+anti-CTLA4 therapy from anti-PD-L1 monotherapy

Cancer patients often receive a combination of antibodies targeting programmed death-ligand 1 (PD-L1) and cytotoxic T lymphocyte antigen-4 (CTLA4). We conducted a window-of-opportunity study in head and neck squamous cell carcinoma (HNSCC) to examine the contribution of anti-CTLA4 to anti-PD-L1 therapy. Single-cell profiling of on- versus pre-treatment biopsies identified Tcell expansion as an early response marker. In tumors, anti-PD-L1 triggered the expansion of mostly CD8+ Tcells, whereas combination therapy expanded both CD4+ and CD8+ Tcells. Such CD4+ Tcells exhibited an activated T helper 1 (Th1) phenotype. CD4+ and CD8+ Tcells co-localized with and were surrounded by dendritic cells expressing Tcell homing factors or antibody-producing plasma cells. Tcell receptor tracing suggests that anti-CTLA4, but not anti-PD-L1, triggers the trafficking of CD4+ naive/central-memory Tcells from tumor-draining lymph nodes (tdLNs), via blood, to the tumor wherein Tcells acquire a Th1 phenotype. Thus, CD4+ Tcell activation and recruitment from tdLNs are hallmarks of early response to anti-PD-L1 plus anti-CTLA4 in HNSCC.

Spatial heterogeneity of peri-tumoural lipid composition in postmenopausal patients with oestrogen receptor positive breast cancer

Deregulation of lipid composition in adipose tissue adjacent to breast tumour is observed in ex vivo and animal models. Novel non-invasive magnetic resonance imaging (MRI) allows rapid lipid mapping of the human whole breast. We set out to elucidate the spatial heterogeneity of peri-tumoural lipid composition in postmenopausal patients with oestrogen receptor positive (ER +) breast cancer. Thirteen participants (mean age, 62 ± [SD] 6 years) with ER + breast cancer and 13 age-matched postmenopausal healthy controls were scanned on MRI. The number of double bonds in triglycerides was computed from MRI images to derive lipid composition maps of monounsaturated, polyunsaturated, and saturated fatty acids (MUFA, PUFA, SFA). The spatial heterogeneity measures (mean, median, skewness, entropy and kurtosis) of lipid composition in the peri-tumoural region and the whole breast of participants and in the whole breast of controls were computed. The Ki-67 proliferative activity marker and CD163 antibody on tumour-associated macrophages were assessed histologically. Mann Whitney U or Wilcoxon tests and Spearman’s coefficients were used to assess group differences and correlations, respectively. For comparison against the whole breast in participants, peri-tumoural MUFA had a lower mean (median (IQR), 0.40 (0.02), p < .001), lower median (0.42 (0.02), p < .001), a negative skewness with lower magnitude (− 1.65 (0.77), p = .001), higher entropy (4.35 (0.64), p = .007) and lower kurtosis (5.13 (3.99), p = .001). Peri-tumoural PUFA had a lower mean (p < .001), lower median (p < .001), a positive skewness with higher magnitude (p = .005) and lower entropy (p = .002). Peri-tumoural SFA had a higher mean (p < .001), higher median (p < .001), a positive skewness with lower magnitude (p < .001) and lower entropy (p = .012). For comparison against the whole breast in controls, peri-tumoural MUFA had a negative skewness with lower magnitude (p = .01) and lower kurtosis (p = .009), however there was no difference in PUFA or SFA. CD163 moderately correlated with peri-tumoural MUFA skewness (rs = − .64), PUFA entropy (rs = .63) and SFA skewness (rs = .59). There was a lower MUFA and PUFA while a higher SFA, and a higher heterogeneity of MUFA while a lower heterogeneity of PUFA and SFA, in the peri-tumoural region in comparison with the whole breast tissue. The degree of lipid deregulation was associated with inflammation as indicated by CD163 antibody on macrophages, serving as potential marker for early diagnosis and response to therapy.

P576 Fecal calprotectin could predict clinical and endoscopic response to ustekinumab therapy

Abstract Background Ustekinumab (UST) is one of the most used biologic drugs for the treatment of Crohn’s Disease (CD) and Ulcerative Colitis (UC). However, not all patients achieve clinical remission and even less mucosal healing. There is currently scarce knowledge about the early markers of therapeutic response in this setting, with particular regard to mucosal healing. The aim of this multicenter study was to evaluate the role of fecal calprotectin (FC) as early predictor of mucosal healing. Methods We enrolled consecutive patients with CD or UC treated with UST for at least 48 weeks at four referral centers in the last 5 years. Therapeutic response was evaluated in terms of clinical remission (defined as a Partial Mayo Score &amp;lt;2 for patients with UC and as Harvey Bradshaw Index &amp;lt;5 for patients with CD, without concomitant corticosteroid therapy) and mucosal healing (defined as a Mayo Endoscopic Score &amp;lt;2 for UC and as the disappearance of ulcers for CD). We collected clinical and demographic data, as well as fecal calprotectin (FC) and C-Reactive Protein (CRP) levels at baseline and at week 24. Results Among 196 patients enrolled (73 [37.2%] women), 167 were affected by CD, whereas 29 by UC. Globally, 128 (65.3%) patients achieved clinical remission and 83 (42.3%) endoscopic remission at 48 weeks of treatment. There were no group differences in age, sex, BMI, baseline clinical and endoscopic indexes, or baseline CRP levels between patients achieving or not either clinical or endoscopic remission. Baseline calprotectin levels were lower in patients who achieved endoscopic remission (median [interquartile range] 391 [103, 1050] versus 825 [317, 1800] µg/mL,p=0.002). At week 24, lower calprotectin levels were observed in both the groups who achieved clinical (160 [50, 426] versus 439 [146, 1000] µg/mL,p&amp;lt;0.0001) and endoscopic remission (114 [43, 265] versus 392 [139, 912] µg/mL,p&amp;lt;0.0001) compared with non-responders. The effects of week-24 calprotectin levels on clinical remission (β= -0.003,p&amp;lt;0.0001) and endoscopic remission (β= -0.004,p&amp;lt;0.0001) remained significant in multivariable models adjusted for age, sex, and BMI. A cut-point of 268 µg/mL was identified as the optimal threshold of week-24 calprotectin for predicting clinical remission (sensitivity 65%, specificity 67%, ROC AUC 0.689), while a cut-point of 309 µg/mL was identified to predict endoscopic remission (sensitivity 79%, specificity 59%, ROC AUC 0.712). Conclusion Our data suggest that FC assessment after 24 weeks of treatment with UST could represent a promising early marker of response, even in terms of mucosal healing, in both CD and UC.

An Exploratory Study of Early Immune Response Markers for Pembrolizumab in Urothelial Tract Cancer

Background: This prospective pilot study explored the potential of the innate immune system’s response to cancer-related immuno-stimulants as a predictive biomarker for Immune Checkpoint Inhibitor (ICI) effectiveness, using pembrolizumab-treated metastatic urothelial tract cancer (mUTC) patients as the study population. Methods: We included ten mUTC patients and assessed their innate immune responses before the first and second pembrolizumab cycles with the TruCulture® immunoassay. We also executed survival analysis and compared cytokine release. Results: R848-induced IFNα and HKCA-induced IL-10 values decreased in patients with disease progression (n = 7), while these values increased in non-progressing patients (n = 3), denoting a significant difference (p = 0.00192 and p = 0.00343, respectively). Further, an increased R848-induced IFNα response correlated with extended survival (log-rank p-value of 0.048). Conclusion: Our small study identified distinct immune response patterns following pembrolizumab’s first cycle in mUTC patients, hypothesizing the potential of an increased R848-induced IFNα response for improved survival outcomes. Further confirmatory studies are in progress.

Breast Multiparametric MRI for Prediction of Neoadjuvant Chemotherapy Response in Breast Cancer: The BMMR2 Challenge.

Purpose To describe the design, conduct, and results of the Breast Multiparametric MRI for prediction of neoadjuvant chemotherapy Response (BMMR2) challenge. Materials and Methods The BMMR2 computational challenge opened on May 28, 2021, and closed on December 21, 2021. The goal of the challenge was to identify image-based markers derived from multiparametric breast MRI, including diffusion-weighted imaging (DWI) and dynamic contrast-enhanced (DCE) MRI, along with clinical data for predicting pathologic complete response (pCR) following neoadjuvant treatment. Data included 573 breast MRI studies from 191 women (mean age [±SD], 48.9 years ± 10.56) in the I-SPY 2/American College of Radiology Imaging Network (ACRIN) 6698 trial (ClinicalTrials.gov: NCT01042379). The challenge cohort was split into training (60%) and test (40%) sets, with teams blinded to test set pCR outcomes. Prediction performance was evaluated by area under the receiver operating characteristic curve (AUC) and compared with the benchmark established from the ACRIN 6698 primary analysis. Results Eight teams submitted final predictions. Entries from three teams had point estimators of AUC that were higher than the benchmark performance (AUC, 0.782 [95% CI: 0.670, 0.893], with AUCs of 0.803 [95% CI: 0.702, 0.904], 0.838 [95% CI: 0.748, 0.928], and 0.840 [95% CI: 0.748, 0.932]). A variety of approaches were used, ranging from extraction of individual features to deep learning and artificial intelligence methods, incorporating DCE and DWI alone or in combination. Conclusion The BMMR2 challenge identified several models with high predictive performance, which may further expand the value of multiparametric breast MRI as an early marker of treatment response. Clinical trial registration no. NCT01042379 Keywords: MRI, Breast, Tumor Response Supplemental material is available for this article. © RSNA, 2024.

SGLT2 inhibitor dapagliflozin protects the kidney in a murine model of Balkan nephropathy.

Proximal tubular uptake of aristolochic acid (AA) forms aristolactam (AL)-DNA adducts, which cause a p53/p21-mediated DNA damage response and acute tubular injury. Recurrent AA exposure causes kidney function loss and fibrosis in humans (Balkan endemic nephropathy) and mice and is a model of (acute kidney injury) AKI to chronic kidney disease (CKD) transition. Inhibitors of the proximal tubule sodium-glucose transporter SGLT2 can protect against CKD progression, but their effect on AA-induced kidney injury remains unknown. C57BL/6J mice (15-wk-old) were administered vehicle or AA every 3 days for 3 wk (10 and 3 mg/kg ip in females and males, respectively). Dapagliflozin (dapa, 0.01 g/kg diet) or vehicle was initiated 7 days prior to AA injections. All dapa effects were sex independent, including a robust glycosuria. Dapa lowered urinary kidney-injury molecule 1 (KIM-1) and albumin (both normalized to creatinine) after the last AA injection and kidney mRNA expression of early DNA damage response markers (p53 and p21) 3 wk later at the study end. Dapa also attenuated AA-induced increases in plasma creatinine as well as AA-induced up-regulation of renal pro-senescence, pro-inflammatory and pro-fibrotic genes, and kidney collagen staining. When assessed 1 day after a single AA injection, dapa pretreatment attenuated AL-DNA adduct formation by 10 and 20% in kidney and liver, respectively, associated with reduced p21 expression. Initiating dapa application after the last AA injection also improved kidney outcome but in a less robust manner. In conclusion, the first evidence is presented that pretreatment with an SGLT2 inhibitor can attenuate the AA-induced DNA damage response and subsequent nephropathy.NEW & NOTEWORTHY Recurrent exposure to aristolochic acid (AA) causes kidney function loss and fibrosis in mice and in humans, e.g., in the form of the endemic Balkan nephropathy. Inhibitors of the proximal tubule sodium-glucose transporter SGLT2 can protect against CKD progression, but their effect on AA-induced kidney injury remains unknown. Here we provide the first evidence in a murine model that pretreatment with an SGLT2 inhibitor can attenuate the AA-induced DNA damage response and subsequent nephropathy.

The chicken chorioallantoic membrane as a low-cost, high-throughput model for cancer imaging

Mouse models are invaluable tools for radiotracer development and validation. They are, however, expensive, low throughput, and are constrained by animal welfare considerations. Here, we assessed the chicken chorioallantoic membrane (CAM) as an alternative to mice for preclinical cancer imaging studies. NCI-H460 FLuc cells grown in Matrigel on the CAM formed vascularized tumors of reproducible size without compromising embryo viability. By designing a simple method for vessel cannulation it was possible to perform dynamic PET imaging in ovo, producing high tumor-to-background signal for both 18F-2-fluoro-2-deoxy-D-glucose (18F-FDG) and (4S)-4-(3-18F-fluoropropyl)-L-glutamate (18F-FSPG). The pattern of 18F-FDG tumor uptake were similar in ovo and in vivo, although tumor-associated radioactivity was higher in the CAM-grown tumors over the 60 min imaging time course. Additionally, 18F-FSPG provided an early marker of both treatment response to external beam radiotherapy and target inhibition in ovo. Overall, the CAM provided a low-cost alternative to tumor xenograft mouse models which may broaden access to PET and SPECT imaging and have utility across multiple applications.

Early Peripheral Blood Blast Clearance As a Prognostic Marker for Early Treatment Response in Pediatric Acute Myeloid Leukemia

Background and Significance The cure rate of pediatric acute myeloid leukemia (AML) has improved significantly; however, relapses still occur in up to 30% of the total patient population in high-income countries (HICs), and in an even higher percentage of patients in low- and middle-income countries (LMICs). Early treatment response, cytogenetics and minimal residual disease (MRD) are well-known prognostic factors for treatment outcome; however, sophisticated techniques for identifying prognostic factors are often lacking in LMICs, and are sometimes unsuccessful in HICs. Early treatment response assessed by a bone marrow (BM) aspirate on day 15 or day 22 (D22) (depending on the applicable protocol) after initiation of chemotherapy is a known prognostic factor for outcome in patients with pediatric AML, but does not allow modifications of the first induction chemotherapy course (Creutzig 2014, Abrahamsson 2011, Kern 2003). In both pediatric acute lymphoid leukemia (ALL) and adult AML, early peripheral blood blast clearance (PBBC) is a known early indicator of treatment response (Manabe 2008, Arellano 2012), whereas to the best of our knowledge, this has not yet been reported in pediatric AML. In this study, we evaluated the significance of time to complete PBBC, and rate of PBBC after initiation of chemotherapy, in relation to disease status in BM D22, as measure of early treatment response that has prognostic significance. Study Design and Methods Pediatric patients with AML, aged 0-18 years, that were treated according to the NOPHO-DBH AML-2012 protocol were included. The first 5 days of induction therapy consisted of etoposide, followed by 7 days of low-dose cytarabine and mitoxantrone or liposomal daunorubicin. Peripheral blood (PB) was assessed by morphology at the time of diagnosis, daily in the first week, and thrice weekly until D22 or until occurrence of complete clearance of blasts. BM D22 was assessed by morphology and flow cytometry. A receiver operating characteristic (ROC) analysis was performed to evaluate the predictive power of days from starting chemotherapy to complete PBBC. The correlation between time to complete PBBC and BM D22 results was analyzed using a logistic regression model. Multivariable logistic regression models were applied for the association after adjusting for effects of other covariates. Rate of PBBC was defined as the percentage of the absolute PB blast count on the day of diagnosis that cleared with each day of therapy, on average, until D22 or the day of complete PBBC. Results In total, 319 patients were included, out of whom 241 were eligible for analysis, and 78 not eligible due to missing or incomplete data. Day 9 after initiation of chemotherapy was identified as the most discriminating cut-off point for predicting BM D22 response. Patients were categorized into early PBBC (≤9 days; n=159) and delayed PBBC (&amp;gt;9 days; n=82) groups. Early PBBC was associated with having &amp;lt;5% AML cells in BM D22 as assessed by both morphology (OR 3.11; P=0.005) and flow cytometry (OR 2.73; P=0.041). Early PBBC also showed a much lower likelihood of having &amp;gt;15% AML cells in BM D22 by morphology (OR 0.08; P=0.002). None of the early PBBC patients had &amp;gt;15% AML cells in BM D22 by flow cytometry. For 217 patients (selection due to missing data [n=24]), the rate of PBBC could be calculated, and a discriminative cut-off value of 11% was identified. Patients were categorized into high rate of PBBC (&amp;gt;11%; n=132) and low rate of PBBC (≤11%; n=85). High rate of PBBC was associated with having &amp;lt;5% AML cells in BM D22 by flow cytometry (OR 4.30; P=0.007) and negative MRD status (OR 2.02; P=0.004), while also showing a strong association with a lower likelihood of &amp;gt;15% AML cells in BM D22 (OR 0.11; P=0.007) by morphology. None of the high rate PBBC patients had &amp;gt;15% AML cells in BM D22 by flow cytometry. Conclusion Both early complete PBBC and rate of PBBC after initiation of chemotherapy holds promise as prognostic marker for clinical outcome in pediatric AML, as it correlates significantly with BM D22 response. These findings may offer an early and easily accessible prognostic factor, particularly beneficial for LMICs, as well as for patients with unsuccessful MRD analyses.