nCRT For Locally Advanced Rectal Cancer Research Articles

Characterization and proteomic analysis of plasma-derived small extracellular vesicles in locally advanced rectal cancer patients.

Neoadjuvant chemoradiotherapy (nCRT) stands as a pivotal therapeutic approach for locally advanced rectal cancer (LARC), yet the absence of a reliable biomarker to forecast its efficacy remains a challenge. Thus, this study aimed to assess whether the proteomic compositions of small extracellular vesicles (sEVs) might offer predictive insights into nCRT response among patients with LARC, while also delving into the proteomic alterations within sEVs post nCRT. Plasma samples were obtained from LARC patients both pre- and post-nCRT. Plasma-derived sEVs were isolated utilizing the TIO2-based method, followed by LC-MS/MS-based proteomic analysis. Subsequently, pathway enrichment analysis was performed to the Differentially Expressed Proteins (DEPs). Additionally, ROC curves were generated to evaluate the predictive potential of sEV proteins in determining nCRT response. Public databases were interrogated to identify sEV protein-associated genes that are correlated with the response to nCRT in LARC. A total of 16 patients were enrolled. Among them, 8 patients achieved a pathological complete response (good responders, GR), while the remaining 8 did not achieve a complete response (poor responders, PR). Our analysis of pretreatment plasma-derived sEVs revealed 67 significantly up-regulated DEPs and 9 significantly down-regulated DEPs. Notably, PROC (AUC: 0.922), F7 (AUC: 0.953) and AZU1 (AUC: 0.906) demonstrated high AUC values and significant differences (P value < 0.05) in discriminating between GR and PR patients. Furthermore, a signature consisting of 5 sEV protein-associated genes (S100A6, ENO1, MIF, PRDX6 and MYL6) was capable of predicting the response to nCRT, yielding an AUC of 0.621(95% CI: 0.454-0.788). Besides, this 5-sEV protein-associated gene signature enabled stratification of patients into low- and high-risk group, with the low-risk group demonstrating a longer overall survival in the testing set (P = 0.048). Moreover, our investigation identified 11 significantly up-regulated DEPs and 31 significantly down-regulated DEPs when comparing pre- and post-nCRT proteomic profiles. GO analysis unveiled enrichment in the regulation of phospholipase A2 activity. Differential expression of sEV proteins distinguishes between GR and PR patients and holds promise as predictive markers for nCRT response and prognosis in patients with LARC. Furthermore, our findings highlight substantial alterations in sEV protein composition following nCRT.

Factors associated with pathological complete remission after neoadjuvant chemoradiotherapy in locally advanced rectal cancer: a real-world clinical setting.

This study aims to identify factors associated with achieving a pathological complete remission (pCR) in patients with locally advanced rectal cancer (LARC) after neoadjuvant chemoradiotherapy (nCRT). We conducted a cohort analysis of 171 LARC patients who underwent curative resection post-nCRT at the First Affiliated Hospital of Guangxi Medical University between January 2015 and December 2021. The data encompassed clinical and pathological information. Univariate and binary logistic regression multivariate analyses were employed to examine the factors influencing pCR achievement after nCRT. Kappa value tests were utilized to compare clinical staging after nCRT with postoperative pathological staging. Postoperative histopathology revealed that of the 171 patients, 40 (23.4%) achieved TRG 0 grade (pCR group), while 131 (76.6%) did not achieve pCR, comprising 36 TRG1, 42 TRG2, and 53 TRG3 cases. Univariate analysis indicated that younger age (p=0.008), reduced tumor occupation of intestinal circumference (p =0.008), specific pathological types (p=0.011), and lower pre-nCRT CEA levels (p=0.003) correlated with pCR attainment. Multivariate analysis identified these factors as independent predictors of pCR: younger age (OR=0.946, p=0.004), smaller tumor occupation of intestinal circumference (OR=2.809, p=0.046), non-mucinous adenocarcinoma pathological type (OR=10.405, p=0.029), and lower pre-nCRT serum CEA levels (OR=2.463, p=0.031). Clinical re-staging post-nCRT compared to postoperative pathological staging showed inconsistent MRI T staging (Kappa=0.012, p=0.718, consistency rate: 35.1%) and marginally consistent MRI N staging (Kappa=0.205, p=0.001, consistency rate: 59.6%). LARC patients with younger age, presenting with smaller tumor circumferences in the intestinal lumen, lower pre-nCRT serum CEA levels, and non-mucinous adenocarcinoma are more likely to achieve pCR after nCRT. The study highlights the need for improved accuracy in clinical re-staging assessments after nCRT in LARC.

Abstract 1304: Employing widening funds to address inequity in cancer research - example of the STEPUPIORS collaborative rectal cancer project

Abstract Introduction: Treatment of locally advanced rectal cancer (LARC) using neoadjuvant chemoradiotherapy (nCRT) improves local control and survival, but response varies. Patients treated at the Institute for Oncology and Radiology of Serbia (IORS) who undergo surgery upon complete response might be exposed to risks and poor quality of life. A consortium dedicated to redefining prognostic and predictive biomarkers in rectal cancer was formed within the framework of the STEPUPIORS Horizon Europe project. The current study evaluated whether employing collaborative widening funds might lead to sustainable strategies for cancer treatment, lowering inequity and steering societal impact, especially in countries with limited resources, using a pilot rectal cancer project. Patients and Methods: The analysis was performed in the period Oct 01, 2022, to Oct 01, 2023, with a Serbian coordinating institution and partners from Spain, Greece, and the Netherlands as expert twinning centers. RT was administered to 75 patients with LARC using volumetric modulated arc therapy-simultaneous integrated boost approach/concomitant chemotherapy (5FU/Leucovorin, week 1&amp;5) for the first time in Serbia. A comprehensive approach towards building human capacities on biobanking, multiomics analyses and project management was developed through intensive training and expert visits. Consensus consortium decisions were reached on regular partner meetings. Results: Patients with clinical complete response (16.0%) and initially distant-located tumors were enrolled in the watch and wait approach and are under follow up. Developed pipelines for multiomics analyses led to 4 high-quality papers on new predictive biomarkers to nCRT in LARC and the deposition of omics data in open-science repositories following FAIR data principles. A first Serbian cancer biobank was successfully installed. Fifteen IORS researchers (87% female), were certified in biobanking. A grant management office was established to provide and support the sustainability of future research projects. This led to 5 new grant applications and enrollment of 1 female project member in a Masters program for Management in the Health Care System. Conclusion: This study showed that even within the first 12 months, implementation of a 3-year collaborative grant dedicated to a specific research problem, might significantly change educational and treatment strategies enabling better patient care (OCEBM level of evidence 3). The results highlight that disparities in cancer outcomes and health equity might only be addressed by progressing beyond descriptions of said outcomes and taking specific action. Systematic, long-term strategic initiatives and collaborations among relevant stakeholders and ethical, legal and societal structures are needed in more countries. Acknowledgements: Horizon Europe Project STEPUPIORS (101079217). Citation Format: Milena Cavic, Ana Djuric, Mladen Marinkovic, Aleksandra Stanojevic, Miljana Tanic, Snezana Bjelogrlic, Jelena Spasic, Ana Damjanovic, Marija Djordjic Crnogorac, Ana Vuletic, Katarina Mirjacic Martinovic, Marko Radulovic, Suzana Stojanovic-Rundic, Ana Krivokuca, Radmila Jankovic, Sergi Castellvi-Bel, Jerome Zoidakis, Remond J. Fijneman. Employing widening funds to address inequity in cancer research - example of the STEPUPIORS collaborative rectal cancer project [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1304.

Abstract 5143: Molecular investigation of a polymorphic variant in JMJD1C that associates with chemoradiotherapy outcomes in locally advanced rectal and head and neck cancer

Abstract Background: Neoadjuvant chemoradiotherapy (nCRT) is standard therapy for locally advanced rectal cancer (LARC). While only a minority of patients are expected to have a complete response, all are at risk of toxicity. Currently no biomarkers reliably predict response to nCRT in LARC. Further, standard therapy in locally advanced head and neck squamous cell carcinoma (HNSCC) is adjuvant CRT. A major cause of poor outcomes in HNSCC is regional persistence or recurrence post-CRT. No biomarkers, apart from HPV, predict CRT response in HNSCC. CRT causes double-strand breaks (DSBs) and methylation of histones on lysine residues is known to affect chromatin, influencing DSB repair. We hypothesized that single nucleotide polymorphisms (SNP) in the histone 3 lysine 9 (H3K9) demethylase-encoding JMJD1C influenced CRT response. Methods: LARC patients (n=84) were divided into poor (PoR) and complete (CR) responders based on Neoadjuvant Rectal score. Lymphocyte DNA was sequenced, and LARC patient-derived cells were treated with DSB-inducing agents to assess viability. Sequencing was confirmed in an independent cohort of CRT-treated HNSCC patients (n=90) divided into: disease-free (n=30), recurrent disease (n=28), and never-disease-free (n=32). HNSCCs were in the larynx or oral cavity. Not all patients received HPV-testing; p16 expression was positive for 19/43 tumors. CRISPR/Cas9 editing of HPV-negative HNSCC lines (SCC9, Cal27) generated JMJD1C WT or SNP isogenic pairs. Bulk RNA-sequencing, colony survival, and immunofluorescence studies were performed. Results: In LARC patients, CRs were enriched for a coding, missense germline SNP in JMJD1C versus PoRs (discovery n=30, p&amp;lt;0.00001; validation n=54, p=0.001, Fisher’s Exact). When subjected to DSB, LARC patient-derived cells with the SNP displayed greater sensitivity than their WT counterparts (n=12, p&amp;lt;0.001, Mann-Whitney). In HNSCC, a correlation was identified between the same JMJD1C SNP and no recurrence (p=0.0003, Fisher’s Exact). HNSCC SNP cells showed changes in mRNA levels of DNA repair and immune response genes versus HNSCC WT cells (q-value&amp;lt;0.05). Upon irradiation, kinetics of foci resolution of the DNA damage response protein MDC1, specific to DSBs, were slower in SNP versus WT HNSCC cells (p&amp;lt;0.05, Mann Whitney). Further, there was reduced colocalization of BRCA1 with RAP80, a crucial factor for homology-directed DSB repair, in SNP versus WT HNSCC cells (p&amp;lt;0.01, Mann-Whitney).Targeted introduction of the SNP into HNSCC cell lines reduced viability and radiation response. Conclusion: Our study identifies a polymorphic coding variant in JMJD1C, as a predictive biomarker for CRT effectiveness in LARC and HNSCC, and defines a DNA repair mechanism by which JMJD1C influences response to irradiation. Citation Format: Adria Hasan, Elena V. Demidova, Philip Czyzewicz, Shreya M. Shah, Karthik Devarajan, Thomas J. Galloway, Margret B. Einarson, Barbara Burtness, Erica A. Golemis, Joshua E. Meyer, Sanjeevani Arora. Molecular investigation of a polymorphic variant in JMJD1C that associates with chemoradiotherapy outcomes in locally advanced rectal and head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5143.

Attention mechanism based multi-sequence MRI fusion improves prediction of response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer

BackgroundAccurate prediction of response to neoadjuvant chemoradiotherapy (nCRT) is very important for treatment plan decision in locally advanced rectal cancer (LARC). The aim of this study was to investigate whether self-attention mechanism based multi-sequence fusion strategy applied to multiparametric magnetic resonance imaging (MRI) based deep learning or hand-crafted radiomics model construction can improve prediction of response to nCRT in LARC.MethodsThis retrospective analysis enrolled 422 consecutive patients with LARC who received nCRT before surgery at two hospitals. All patients underwent multiparametric MRI scans with three imaging sequences. Tumor regression grade (TRG) was used to assess the response of nCRT based on the resected specimen. Patients were separated into 2 groups: poor responders (TRG 2, 3) versus good responders (TRG 0, 1). A self-attention mechanism, namely channel attention, was applied to fuse the three sequence information for deep learning and radiomics models construction. For comparison, other two models without channel attention were also constructed. All models were developed in the same hospital and validated in the other hospital.ResultsThe deep learning model with channel attention mechanism achieved area under the curves (AUCs) of 0.898 in the internal validation cohort and 0.873 in the external validation cohort, which was the best performed model in all cohorts. More importantly, both the deep learning and radiomics model that applied channel attention mechanism performed better than those without channel attention mechanism.ConclusionsThe self-attention mechanism based multi-sequence fusion strategy can improve prediction of response to nCRT in LARC.

MRI Tumor Regression Grade Combined with T2-Weighted Volumetry May Predict Histopathological Response in Locally Advanced Rectal Cancer following Neoadjuvant Chemoradiotherapy-A New Scoring System Proposal.

Modern studies focus on the discovery of innovative methods to improve the value of post-treatment magnetic resonance imaging (MRI) in the prediction of pathological responses to preoperative neoadjuvant chemoradiotherapy (nCRT) in locally advanced rectal cancer (LARC). The aim of this study was to assess the potential benefits of combining magnetic resonance tumor regression grade (mrTRG) with T2-weighted volumetry in the prediction of pathological responses to nCRT in LARC. This was a cohort study conducted on patients with histopathologically confirmed LARC in a period from 2020 to 2022. After histopathological verification, all patients underwent initial MRI studies, while the follow-up MRI was performed after nCRT. Tumor characteristics, MRI estimated tumor regression grade (mrTRG) and tumor volumetry were evaluated both initially and at follow-up. All patients were classified into responders and non-responders according to pathological tumor regression grade (pTRG) and mrTRG. A total of 71 patients, mostly male (66.2%) were included in the study. The median tumor volume reduction rate was significantly higher in nCRT-responders compared to non-responders (79.9% vs. 63.3%) (p = 0.003). Based on ROC analysis, optimal cut-off value for tumor volume reduction rate was determined with an area under the curve (AUC) value of 0.724 (p = 0.003). Using the tumor volume reduction rate ≥75% with the addition of response to nCRT according to mrTRG, a new scoring system for prediction of pTRG to preoperative nCRT in LARC was developed. Diagnostic performance of prediction score was tested and the sensitivity, PPV, specificity, and NPV were 81.8%, 56.3%, 71.4%, and 89.7%, respectively. The combination of mrTRG and T2-weighted volumetry increases the MRI-based prediction of pTRG to preoperative nCRT in LARC. The proposed scoring system could aid in distinguishing responders to nCRT, as these patients could benefit from organ-preserving treatment and a "watch and wait" strategy.

Local resection versus radical resection after neoadjuvant chemoradiotherapy for patients with locally advanced rectal cancer: a propensity-score matched cohort analysis

BackgroundWe aimed to address the shortage of evidence regarding the safety of the local resection approach by comparing long-term oncological outcomes between patients managed by local resection and those who underwent radical resection.MethodsThis was a propensity-score matched cohort analysis study that included patients of all ages diagnosed with locally advanced rectal cancer (LARC) who had received neoadjuvant chemoradiotherapy (nCRT) at the Fujian Medical University Union Hospital and Fujian Medical University Affiliated Zhangzhou Hospital, China, between Jan 10, 2011, to Dec 28, 2021. Partial patients with a significant downstage of the tumor were offered management with the local resection approach, and most of the rest were offered radical resection if eligible.FindingsOne thousand six hundred ninety-three patients underwent radical resection after nCRT, and another 60 patients performed local resection. The median follow-up times were 44.0 months (interquartile range = 4–107 months). After propensity-core matching (PSM), in the Kaplan–Meier curves, local resection (n = 56) or radical resection (n = 211) was not significantly associated with 1-, 3-, and 5-year cumulative incidence of overall survival (OS) (HR = 1.103, 95% CI: 0.372 ~ 3.266), disease-free survival (DFS) ((HR = 0.972, 95% CI: 0.401 ~ 2.359), local recurrence (HR = 1.044, 95% CI: 0.225 ~ 4.847), and distant metastasis (HR = 0.818, 95% CI: 0.280 ~ 2.387) (all log-rank P > 0.05). Similarly, multivariate Cox regression analysis indicates that local excision still was not an independent risk factor for OS (HR = 0.863, 95% CI: 0.267 ~ 2.785, P = 0.805) and DFS (HR = 0.885, 95% CI: 0.353 ~ 2.215, p = 0.794).ConclusionLocal resection can be a management option in selected patients with middle-low rectal cancer after nCRT for LARC and without loss of oncological safety at five years.

Response assessment of locally advanced rectal cancer after neoadjuvant chemoradiotherapy: Is apparent diffusion coefficient useful on 3T magnetic resonance imaging?

To assess the diagnostic value of apparent diffusion coefficient (ADC) on 3T device for the prediction of tumoral response to neoadjuvant chemoradiotherapy (nCRT) and for the response assessment after nCRT in patients with locally advanced rectal cancer (LARC), using pathology as a reference. Forty-one patients affected by LARC undergoing 3.0T MRI before and after nCRT were retrospectively selected. After the conventional acquisition of high resolution T2-weighted sequences, diffusion-weighted MRI (DW-MRI) was performed using a spin-echo echo-planar sequence with multiple b values (150, 500, 1000, 1500 s/mm2 ). Fitted ADC values were calculated for each rectal lesion before and after nCRT by drawing a hand-made region of interest (ROI) around the tumour outline. All patients underwent surgery and pathological staging (classified according to tumour regression grading [TRG] and to tumour and node [TN]) represented the reference standard. Pretreatment ADC value (pre-ADC), ADC value obtained after nCRT (post-ADC) and the difference between post-ADC and pre-ADC (ΔADC) were correlated with both the TRG classes and the TN staging system in each patient. The ADC values obtained in the post nCRT examination and the ΔADC were statistically related both to TRG (p=0.0004; p=0.0126, respectively) and TN (p=0.0484; p=0.0673, respectively) stages at histopathology. On the contrary, the pre-ADC was not related either to the TRG classes or to the lesion TN staging system (p > 0.05). 3 T DW-MRI using ADC value can be useful to assess the efficacy of nCRT in LARC; in fact, post-ADC and ΔADC values improve MR capability to evaluate tumour response.

What genes predict response to neoadjuvant chemoradiation in locally advanced rectal cancer? A systematic review based on gene expression profiling.

200 Background: Curative management of locally advanced rectal cancer (LARC) involves neoadjuvant chemoradiation therapy (nCRT) followed by radical surgery. Pathological complete response (pCR) is seen in 15-27% of patients. The response to nCRT may be influenced by genetic variations. Gene signatures therefore may serve as biomarkers for predicting response or resistance to nCRT. We conducted a systematic review on genetic biomarkers for response prediction to nCRT in LARC and identify common genes or ontology pathways. Methods: A systematic search of electronic databases (MEDLINE, PubMed and EMBASE) was performed for relevant recent studies. Studies were screened as per the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines with search terms “rectal neoplasm”, “rectal cancer”, “gene”, “gene expression”, “gene expression profile”. The relevant studies were selected for full-text review. Studies that reported gene expression profile to predict response to nCRT were included. Results: Twelve of 367 studies screened were included. Most studies used microarray or real-time reverse transcription PCR. Six of these assessed for gene predictors of pCR with 2 studies reporting correlation between certain genes and pCR: thymidylate synthetase; gene set (including LMNB2, MLF2, DDX1, EIF4A1, FOXO3A, ILF3 and SEMA3C); 19 genes including (NPSR1, HYAL1, FGFBP1, PLLP and MALL). Common gene ontology pathways identified included DNA break repair via homologous recombination and apoptotic pathways. Conclusions: Multiple genes were identified as prospective biomarkers for nCRT in LARC patients. Further studies are required to validate these identified genes to personalize treatment for LARC patients.

Evaluation of Neutrophil-Lymphocyte Ratio and Platelet-Lymphocyte Ratio on Predicting Responsiveness to Neoadjuvant Chemoradiotherapy in Locally Advanced Rectal Cancer Patients.

Objective Neutrophil-lymphocyte ratio (NLR) and Platelet-lymphocyte ratio (PLR) have been proposed as prognostic biomarkers in multiple cancers. However, the implications of NLR and PLR in the responsiveness to neoadjuvant chemoradiotherapy (nCRT) remain to be clarified in locally advanced rectal cancer (LARC) patients. This retrospective study investigated the prognostic value of NLR and PLR in nCRT responsiveness of LARC patients. Methods A total number of 86 patients diagnosed with LARC and treated with nCRT and total mesorectal excision were retrospectively followed from 2013 to 2016. Receiver operating characteristic (ROC) curve was used to determine the cutoff values of NLR and PLR, and the patients were divided into NLR elevation and NLR decrease groups, or PLR elevation and PLR decrease groups. The correlation between NLR and PLR changes, and clinicopathological factors were analyzed. The relationship between NLR and PLR changes and the curative responsiveness towards nCRT were further evaluated. Results NLR and PLR changes after nCRT were significantly correlated with the distance of tumors to the anus and BMI (body mass index) (P < 0.05). The clinical remission rate of patients with NLR reduction was 72.09% (31/43), which was significantly higher than that in patients with NLR increment (22/43, 51.16%). There was no significant difference in the clinic remission rate between the patients with PLR reduction and those with PLR increment (P > 0.05). However, the pathological responsiveness rate was significantly higher in patients with PLR reduction (21/43, 48.84%) when compared to the ones with PLR increment (9/43, 20.9%) (P = 0.036). Conclusion Our data indicate that in LARC patients with nCRT, the reduction of NLR and the reduction of PLR could serves as predictors for the clinic remission rate and pathological responsiveness rate, respectively. The combination of NLR and PLR changes may be employed as a simple and effective prognostic parameter to predict the treatment outcome of nCRT in LARC.

Identification of proteomic markers for prediction of the response to 5-Fluorouracil based neoadjuvant chemoradiotherapy in locally advanced rectal cancer patients

BackgroundNeoadjuvant chemoradiotherapy (nCRT) prior to surgery is the standard treatment for patients with locally advanced rectal cancer (LARC), while parts of them show poor therapeutic response accompanied by therapy adverse effects. Predictive biomarkers for nCRT response could facilitate the guidance on treatment decisions but are still insufficient until now, which limits the clinical applications of nCRT in LARC patients.MethodsIn our study, 37 formalin-fixed paraffin-embedded tumor biopsies were obtained from patients with LARC before receiving 5-fluorouracil based nCRT. Proteomics analyses were conducted to identify the differentially expressed proteins (DEPs) between total responders (TR) and poor responders (PR). The DEPs were validated via ROC plotter web tool and their predictive performance was evaluated by receiver operating characteristic analysis. Functional enrichment analyses were performed to further explore the potential mechanisms underlying nCRT response.ResultsAmong 3,998 total proteins, 91 DEPs between TR and PR were screened out. HSPA4, NIPSNAP1, and SPTB all with areas under the curve (AUC) ~ 0.8 in the internal discovery cohort were independently validated by the external mRNA datasets (AUC ~ 0.7), and their protein levels were linearly correlated with the graded responses to nCRT in the internal cohort. The combination of HSPA4 and SPTB could distinctly discriminate the TR and PR groups (AUC = 0.980, p < 0.0001). Moreover, multiple combinations of the three proteins realized increased specificity and/or sensitivity, while achieving favorable predictive value when moderate responders were introduced into the ROC analysis. Pathways including DNA damage repair, cell cycle, and epithelial mesenchymal transition were involved in nCRT response according to the enrichment analysis results.ConclusionsHSPA4, SPTB and NIPSNAP1 in tumor biopsies and/or their optional combinations might be potential predictive markers for nCRT response in patients with LARC. The DEPs and their related functions have implications for the potential mechanisms of treatment response to nCRT in patients with LARC.

MRI Radiomics Model Predicts Pathologic Complete Response of Rectal Cancer Following Chemoradiotherapy.

Background Preoperative assessment of pathologic complete response (pCR) in locally advanced rectal cancer (LARC) after neoadjuvant chemoradiotherapy (nCRT) is increasingly needed for organ preservation, but large-scale validation of an MRI radiomics model remains lacking. Purpose To evaluate radiomics models based on T2-weighted imaging and diffusion-weighted MRI for predicting pCR after nCRT in LARC and compare their performance with visual assessment by radiologists. Materials and Methods This retrospective study included patients with LARC (clinical stage T3 or higher, positive nodal status, or both) who underwent post-nCRT MRI and elective resection between January 2009 and December 2018. Surgical histopathologic analysis was the reference standard for pCR. Radiomic features were extracted from the volume of interest on T2-weighted images and apparent diffusion coefficient (ADC) maps from post-nCRT MRI to generate three models: T2 weighted, ADC, and both T2 weighted and ADC (merged). Radiomics signatures were generated using the least absolute shrinkage and selection operator with tenfold cross-validation. Three experienced radiologists independently rated tumor regression grades at MRI and compared these with the radiomics models' diagnostic outcomes. Areas under the curve (AUCs) of the radiomics models and pooled readers were compared by using the DeLong method. Results Among 898 patients, 189 (21%) achieved pCR. The patients were chronologically divided into training (n = 592; mean age ± standard deviation, 59 years ± 12; 388 men) and test (n = 306; mean age, 59 years ± 12; 190 men) sets. The radiomics signatures of the T2-weighted, ADC, and merged models demonstrated AUCs of 0.82, 0.79, and 0.82, respectively, with no evidence of a difference found between the T2-weighted and merged models (P = .49), while the ADC model performed worse than the merged model (P = .02). The T2-weighted model had higher classification performance (AUC, 0.82 vs 0.74 [P = .009]) and sensitivity (80.0% vs 15.6% [P < .001]), but lower specificity (68.4% vs 98.6% [P < .001]) than the pooled performance of the three radiologists. Conclusion An MRI-based radiomics model showed better classification performance than experienced radiologists for diagnosing pathologic complete response in patients with locally advanced rectal cancer after neoadjuvant chemoradiotherapy. © RSNA, 2022 Online supplemental material is available for this article. See also the editorial by Taylor in this issue.

THUNDER 2: THeragnostic Utilities for Neoplastic DisEases of the Rectum by MRI guided radiotherapy

BackgroundNeoadjuvant chemoradiation therapy (nCRT) is the standard treatment modality in locally advanced rectal cancer (LARC). Since response to radiotherapy (RT) is dose dependent in rectal cancer, dose escalation may lead to higher complete response rates. The possibility to predict patients who will achieve complete response (CR) is fundamental. Recently, an early tumour regression index (ERI) was introduced to predict pathological CR (pCR) after nCRT in LARC patients.The primary endpoints will be the increase of CR rate and the evaluation of feasibility of delta radiomics-based predictive MRI guided Radiotherapy (MRgRT) model.MethodsPatients affected by LARC cT2-3, N0-2 or cT4 for anal sphincter involvement N0-2a, M0 without high risk features will be enrolled in the trial. Neoadjuvant CRT will be administered using MRgRT. The initial RT treatment will consist in delivering 55 Gy in 25 fractions on Gross Tumor Volume (GTV) plus the corresponding mesorectum and 45 Gy in 25 fractions on the drainage nodes. Chemotherapy with 5-fluoracil (5-FU) or oral capecitabine will be administered continuously.A 0.35 Tesla MRI will be acquired at simulation and every day during MRgRT. At fraction 10, ERI will be calculated: if ERI will be inferior than 13.1, the patient will continue the original treatment; if ERI will be higher than 13.1 the treatment plan will be reoptimized, intensifying the dose to the residual tumor at the 11th fraction to reach 60.1 Gy.At the end of nCRT instrumental examinations are to be performed in order to restage patients. In case of stable disease or progression, the patient will undergo surgery. In case of major or complete clinical response, conservative approaches may be chosen. Patients will be followed up to evaluate toxicity and quality of life.The number of cases to be enrolled will be 63: all the patients will be treated at Fondazione Policlinico Universitario A. Gemelli IRCCS in Rome.DiscussionThis clinical trial investigates the impact of RT dose escalation in poor responder LARC patients identified using ERI, with the aim of increasing the probability of CR and consequently an organ preservation benefit in this group of patients.Trial registrationClinicalTrials.gov Identifier: NCT04815694 (25/03/2021).

Pre-Existing Tumoral B Cell Infiltration and Impaired Genome Maintenance Correlate with Response to Chemoradiotherapy in Locally Advanced Rectal Cancer.

Locally advanced rectal cancer (LARC) remains a medical challenge. Reliable biomarkers to predict which patients will significantly respond to neoadjuvant chemoradiotherapy (nCRT) have not been identified. We evaluated baseline genomic and transcriptomic features to detect differences that may help predict response to nCRT. Eligible LARC patients received nCRT (3D-LCRT 50.4 Gy plus capecitabine 825 mg/m/bid), preceded by three cycles of CAPOX in high systemic-relapse risk tumors, and subsequent surgery. Frozen tumor biopsies at diagnosis were sequenced using a colorectal cancer panel. Transcriptomic data was used for pathway and cell deconvolution inferential algorithms, coupled with immunohistochemical validation. Clinical and molecular data were analyzed according to nCRT outcome. Pathways related to DNA repair and proliferation (p < 0.005), and co-occurrence of RAS and TP53 mutations (p = 0.001) were associated with poor response. Enrichment of expression signatures related to enhanced immune response, particularly B cells and interferon signaling (p < 0.005), was detected in good responders. Immunohistochemical analysis of CD20 cells validated the association of good response with B cell infiltration (p = 0.047). Findings indicate that the presence of B cells is associated with successful tumor regression following nCRT in LARC. The prevalence of simultaneous RAS and TP53 mutations along with a proficient DNA repair system that may counteract chemoradio-induced DNA damage was associated with poor response.

Germline and Somatic Pharmacogenomics to Refine Rectal Cancer Patients Selection for Neo-Adjuvant Chemoradiotherapy

Neoadjuvant chemoradiotherapy (nCRT) followed by radical surgery is the standard of care for patients with Locally Advanced Rectal Cancer (LARC). Current selection for nCRT is based on clinical criteria regardless of any molecular marker. Pharmacogenomics may be a useful strategy to personalize and optimize nCRT in LARC. This review aims to summarize the most recent and relevant findings about the role of germline and somatic pharmacogenomics in the prediction of nCRT outcome in patients with LARC, discussing the state of the art of their application in the clinical practice. A systematic literature search of the PubMed database was completed to identify relevant English-language papers published up to January 2020. The chemotherapeutic backbone of nCRT is represented by fluoropyrimidines, mainly metabolized by DPD (Dihydro-Pyrimidine Dehydrogenase, DPYD). The clinical impact of testing DPYD*2A, DPYD*13, c.2846A > T and c.1236G > A-HapB3 before a fluoropyrimidines administration to increase treatment safety is widely acknowledged. Other relevant target genes are TYMS (Thymidylate Synthase) and MTHFR (Methylene-Tetrahydro-Folate Reductase), whose polymorphisms were mainly studied as potential markers of treatment efficacy in LARC. A pivotal role of a TYMS polymorphism in the gene promoter region (rs34743033) was reported and was pioneeringly used to guide nCRT treatment in a phase II study. The pharmacogenomic analysis of other pathways mostly involved in the cellular response to radiation damage, as the DNA repair and the activation of the inflammatory cascade, provided less consistent results. A high rate of somatic mutation in genes belonging to PI3K (Phosphatidyl-Inositol 3-Kinase) and MAPK (Mitogen-Activated Protein Kinase) pathways, as BRAF (V-raf murine sarcoma viral oncogene homolog B1), KRAS (Kirsten Rat Sarcoma viral oncogene homolog), NRAS (Neuroblastoma RAS viral (v-ras) oncogene homolog), PIK3CA (Phosphatidyl-Inositol-4,5-bisphosphate 3-Kinase, Catalytic Subunit Alpha), as well as TP53 (Tumor Protein 53) was reported in LARC. Their pharmacogenomic role, already defined in colorectal cancer, is under investigation in LARC with promising results concerning specific somatic mutations in KRAS and TP53, as predictors of tumor response and prognosis. The availability of circulating tumor DNA in plasma may also represent an opportunity to monitor somatic mutations in course of therapy.

3.0\xa0T MRI IVIM-DWI for predicting the efficacy of neoadjuvant chemoradiation for locally advanced rectal cancer

PurposeThe purpose of this study was to determine the diagnostic performance of intravoxel incoherent motion (IVIM) on assessing response to neoadjuvant chemoradiation (nCRT) in patients with Locally Advanced Rectal Cancer (LARC).Methods50 patients with rectal cancer who underwent magnetic resonance (MR) imaging before and after nCRT, the values of pre-nCRT and post-nCRT IVIM-DWI parameters apparent diffusion coefficient (ADC), diffusion coefficient (D), false diffusion coefficient (D*), and perfusion fraction (f), together with the percentage changes (∆% parametric value) induced by nCRT were calculated. According to the patient's response to nCRT, the patients were divided into pathological complete response (pCR) and non-pCR groups, Good Response (GR) group and Poor Response (PR) group, and the above values were compared between different groups. Univariate and multiple logistic regression analysis were done to investigate the relation between different parameters and patient nCRT. Draw ROC curve according to sensitivity and specificity, and compare its diagnostic efficacy.ResultsThere were no significant differences in the baseline data of 50 patients. After nCRT, the ADC and D values for LARC increased significantly (all p < 0.05). The pCR group (n = 9) had higher preD*, pref, postD*, ∆%ADC and ∆%D values than the non-pCR group (n = 41) (all p < 0.05). The GR group (n = 17) exhibited higher post D, ∆%ADC and ∆%D values than the PR group (n = 33) (all p < 0.05). From the results of Logistic regression analysis found that ∆%ADC and ∆%D were significantly correlated with patients' response to nCRT. Based on ROC analysis, ∆%D had a higher area under the curve value than ∆%ADC (p = 0.009) in discriminating the pCR from non-pCR groups.ConclusionsIVIM-DWI technology may be helpful in identifying the pCR and GR patients to nCRT for LARC.

MRI Radiomics for Prediction of Tumor Response and Downstaging in Rectal Cancer Patients after Preoperative Chemoradiation

PurposeThis study aimed to investigate radiomic features extracted from magnetic resonance imaging (MRI) scans performed before and after neoadjuvant chemoradiotherapy (nCRT) in predicting response of locally advanced rectal cancer (LARC). Methods and MaterialsThirty-nine patients who underwent nCRT for LARC were included, with 294 radiomic features extracted from MRI that was performed before (pre-CRT) and 6 to 8 weeks after completing nCRT (post-CRT). Based on tumor regression grade (TRG), 26 patients were classified as having a histopathologic good response (GR; TRG 0-1) and 13 as non-GR (TRG 2-3). Tumor downstaging (T-downstaging) occurred in 25 patients. Univariate analyses were performed to assess potential radiomic and delta-radiomic predictors for TRG in pathologic complete response (pCR) versus non-pCR, GR versus non-GR, and T-downstaging. The support vector machine-based multivariate model was used to select the best predictors for TRG and T-downstaging. ResultsWe identified 13 predictive features for pCR versus non-pCR, 14 for GR versus non-GR, and 16 for T-downstaging. Pre-CRT gray-level run length matrix nonuniformity, pre-CRT neighborhood intensity difference matrix (NIDM) texture strength, and post-CRT NIDM busyness predicted all 3 treatment responses. The best predictor for GR versus non-GR was pre-CRT global minimum combined with clinical N stage in the multivariate analysis. The best predictor for T-downstaging was the combination of pre-CRT gray-level co-occurrence matrix correlation, NIDM-texture strength, and gray-level co-occurrence matrix variance. The pre-CRT, post-CRT, and delta radiomic-based models had no significant difference in predicting all 3 responses. ConclusionsPre-CRT MRI, post-CRT MRI, and delta radiomic-based models have the potential to predict tumor response after nCRT in LARC. These data, if validated in larger cohorts, can provide important predictive information to aid in clinical decision making.

College of American Pathologists Tumor Regression Grading System for Long-Term Outcome and Adjuvant Chemotherapy in Patients with Locally Advanced Rectal Cancer: Result from a Multicentre, Observational, Cohort Study

Background: NCCN's Rectal Cancer Guideline Panel recommends the American Joint Committee on Cancer and the College of American Pathologists tumor regression grading (AJCC/CAP TRG) system to evaluate pathologic response to neoadjuvant chemoradiotherapy (nCRT) for locally advanced rectal cancer (LARC). Yet, the clinical significance of AJCC/CAP TRG system has not been fully defined. Methods: This was a retrospectively designed, multicentre and prospectively maintained cohort study. Of which, LARC cases from one institution formed discovery cohort to determine the relationship between AJCC/CAP TRG system and overall survival (OS), disease free survival (DFS), local recurrence-free survival (LRFS), and distant metastasis-free survival (DMFS), as well as the risk of disease progression (death, disease relapse, local recurrence, and distant metastasis). Moreover, patients from other institutions formed the validation cohort. Kaplan-Meier survival functions were computed and the hazard ratios calculated, with the Cox regression hazards model. Findings: The discovery cohort (940 cases) found and validation cohort (2156 cases) further confirmed that the inferior AJCC/CAP TRG categories were closely correlated with an unfavorable OS, DFS, LRFS and DMFS, as well as a higher risk of disease progression (all P < 0·05). Significantly, the pairwise comparison revealed that any 2 of the 4 TRG categories had a distinguished survival (OS, DFS, LRFS and DMFS) ratio and risk of disease progression. In contrast, the similar findings were not observed at the ypT and ypN stage categories patients, as some two consecutive ypT and ypN categories patients had the comparable survival (OS, DFS, LRFS and DMFS) ratio and risk of disease progression. Importantly, after the propensity score matching, Kaplan-Meier analysis showed that the AJCC/CAP TRG0 category patients treated with or without adjuvant chemotherapy had a similar OS, DFS, LRFS and DMFS. Conversely, for the AJCC/CAP TRG1-3 categories cases, adjuvant chemotherapy treatment significantly improved the 3-year OS (90·2% vs. 84·6%, P < 0·001). Receiver operating characteristic and multivariate analysis demonstrated AJCC/CAP TRG system was an independent prognostic surrogate in discovery cohort and validation cohort (all P < 0·05). Interpretation: Based on one of the largest dataset, AJCC/CAP TRG system, a useful prognostic surrogate, appears ideal for further strategizing therapy after nCRT for LARC. Funding: National Natural Science Foundation of China, International Center for Genetic Engineering and Biotechnology grant. Declaration of Interest: All authors declare no competing interests. Ethical Approval: The Clinical Ethics Review Committee at the Sixth Affiliated Hospital of Sun Yat-sen University approved this study.

CtDNA As a Potential Prognostic Marker for Locally Advanced Rectal Cancer Patients Receiving Neoadjuvant Chemo-Radiation Therapy on Disease-Free Survival (DFS)

Neoadjuvant chemo-radiation therapy (nCRT) plus surgery is the standard treatment for patients with locally advanced rectal cancer (LARC). However, how to predict and select patients who may achieve clinical complete response is still an unsolved issue. We conducted a pilot study to evaluate the potential role of ctDNA as a biomarker to predict treatment outcome. We found that ctDNA clearance was related to TRG level in previous studies. Now we try to explore using ctDNA as a prognostic factor and improve risk stratification in locally advanced rectal cancer (LARC). In this study, we recruited 119 patients with LARC receiving nCRT. 595 serial plasma samples were collected at d0, d15, d25 of radiotherapy as well before and 7 days post-surgery. The level of ctDNA was calculated by dynamic monitoring the mutant allele frequency of somatic mutations in plasma. Plasma and tissue samples were subjected to targeted-NGS using a 422 cancer-related genes panel. We followed up patients with concomitant CT until disease progression or death. Serial samples’ data analysis showed detection of pre-treatment mutations after completion of nCRT was significantly associated with worse disease-free survival (DFS) (P<0.05). Patients with ctDNA mutations clearance during nCRT significantly have a better DFS compare with patients with ctDNA non-clearance (P=0.02). The presence of TP53 mutations is predictive of treatment outcome and shows a statistically significant worse DFS (2 years) (54.3% VS 87.0%, P=0.0005). Predictive model based on support vector machine was developed for prediction of pCR achieving a mean AUC of 0.85 assessed by repeated cross validation. Through tracking clonal extinction, persistence and emergence, patients were grouped into four evolutionary subtypes with distinct TRG and DFS. Our data showed the prognostic value of ctDNA on DFS. In patients treated with nCRT, the presence of ctDNA after completion of nCRT was associated with an inferior DFS (P=0.02). Dynamic monitoring of ctDNA can be used to predict TRG and prognosis in LARC patients receiving nCRT. ctDNA sequencing depicts the evolutionary trajectories of sensitive and resistant clones during nCRT in LARC. CtDNA could potentially be used to guide patient selection for nCRT.

CtDNA as a potential prognostic marker for locally advanced rectal cancer patients with ‘watch and wait’ approach.

3544 Background: ‘Watch and Wait’ policy has currently led to growing interest for organ-preservation after neoadjuvant chemoradiation (nCRT) to improve quality of life. However, how to predict and select patients who may achieve clinical complete response is still an unsolved issue. We conducted a pilot study to evaluate the potential role of ctDNA as a biomarker to predict treatment outcome and improve risk stratification in locally advanced rectal cancer (LARC). Methods: In this study, we recruited 119 patients with LARC receiving nCRT. 595 serial plasma samples were collected at d0, d15, d25 of radiotherapy as well before and 7 days post surgery. The level of ctDNA was calculated by dynamic monitoring the mutant allele frequency of somatic mutations in plasma. Plasma and tissue samples were subjected to targeted-NGS using a 422 cancer-related genes panel. We followed up patients with concomitant CT until disease progression or death. Results: Detected mutation of TP53 and APC gene in pre-treatment samples was negatively correlated with patients’ response to nCRT. Alterations in homologous recombination and adherens junction pathways were associated with a better response (P &lt; 0.05). Detection of pre-treatment mutations in any time points during nCRT was significantly (P = 0.03) decreased from TRG3 to TRG0 group (33%, 29%, 22% and 4%, respectively); while detection of acquired mutations showed an opposite trend (P = 0.04). A predictive model based on support vector machine was developed for prediction of pCR achieving a mean AUC of 0.85 assessed by repeated cross validation. Further, detection of pre-treatment mutations after completion of nCRT was significantly associated with worse disease-free survival (DFS) (P &lt; 0.05). Through tracking clonal extinction, persistence and emergence, patients were grouped into four evolutionary subtypes with distinct TRG and DFS. Conclusions: Our data showed the prognostic value of ctDNA on DFS. Dynamic monitoring of ctDNA can be used to predict TRG and prognosis in LARC patients receiving nCRT. ctDNA sequencing depicts the evolutionary trajectories of sensitive and resistant clones during nCRT in LARC. CtDNA could potentially be used to guide patient selection for W&amp;W strategy.