The efficacy and safety of neoadjuvant chemoradiotherapy combined with immunotherapy for locally advanced rectal cancer patients: a systematic review.

Background: Several studies have explored the effectiveness of PD-1/PD-L1 inhibitors combined with neoadjuvant chemoradiotherapy (nCRT) in the treatment of locally advanced rectal cancer(LARC), particularly in microsatellite stable(MSS) or mismatch repair proficient(pMMR) LARC patients. We undertook a single-arm systematic review to comprehensively evaluate the advantages and potential risks associated with the use of PD-1/PD-L1 inhibitors in conjunction with nCRT for patients diagnosed with locally advanced rectal cancer. Methods: The PubMed, Embase, Cochrane Library, ClinicalTrials.gov, ASCO and ESMO were searched for related studies. The main outcomes were pathologic complete response (pCR), major pathological response (MPR), anal preservation, and adverse effects (AEs). Results: Fourteen articles including 533 locally advanced rectal cancer (LARC) patients were analyzed. The pooled pCR, MPR, and anal preservation rates were 36%, 66% and 86%. Grade ≥3 adverse events occurred in 20%. Subgroup analysis showed that; dMMR/MSI-H had a pooled pCR (100%) and MPR (100%), pMMR/MSS had a pooled pCR (38%) and MPR (60%); the short-course radiotherapy and long-course radiotherapy had pooled pCR rates of 51% and 30%, respectively. The rates of pCR for the concurrent and sequential immuno-chemoradiotherapy subgroups at 30% and 40%, mirroring pCR rates for the PD-L1 and PD-1 inhibitor subgroups were 32% and 40%, respectively. Conclusion: In cases of locally advanced rectal cancer, PD-1/PD-L1 inhibitors combined with neoadjuvant chemoradiotherapy have shown promising response rates and acceptable toxicity profiles. PD-1/PD-L1 inhibitors combined with neoadjuvant chemoradiotherapy hence has a positive outcome even in MSS LARC patients. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/#myprospero, identifier CRD42023465380.

Case report: Durable complete response of a mucosal melanoma of the rectum after neoadjuvant immunotherapy with ipilimumab plus nivolumab.

Melanoma causes the majority of skin cancer-related deaths. Despite novel therapy options, metastatic melanoma still has a poor prognosis. Immune checkpoint inhibition (ICI) therapy has been shown to prolong overall survival in patients with advanced melanoma, but mucosal melanomas respond less favorably compared to melanomas of cutaneous origin. We report on a patient with a mucosal melanoma of the rectum diagnosed in June 2020. Since a surgical intervention in order to achieve a tumor-free situation would have required an amputation of the rectum, a neo-adjuvant systemic immunotherapy with ipilimumab and nivolumab was initiated. As restaging and colonoscopy after four doses of this combination immunotherapy showed a partial response, the patient decided against the pre-planned surgery and a maintenance therapy with nivolumab was started. Repeated colonoscopy showed a complete response after four doses of nivolumab. After ongoing ICI therapy with nivolumab and no evidence of tumor relapse, immunotherapy was stopped in July 2022 after nearly 2 years of continuous treatment. The patient remained tumor-free during further follow-up. Neo-adjuvant immunotherapy is getting more explored in advanced melanoma. By administering ICI therapy before surgical resection of an essentially operable tumor, a stronger and more diverse immunological response is supposed to be achieved. Our reported case demonstrates that this approach could also be effective in mucosal melanoma despite of its generally lower response to immunotherapy.

Case report: Decreased hemoglobin and multiple organ failure caused by ceftizoxime-induced immune hemolytic anemia in a Chinese patient with malignant rectal cancer.

Background: Drug-induced immune hemolytic anemia (DIIHA) is a rare but serious condition, with an estimated incidence of one in 100,000 cases, associated with various antibiotics. This study reports on a case of ceftizoxime-induced hemolysis observed in a patient in China. Case description: A Chinese patient diagnosed with malignant rectal cancer underwent antimicrobial therapy after laparoscopic partial recto-sigmoid resection (L-Dixon). After receiving four doses of ceftizoxime, the patient developed symptoms including rash, itchy skin, and chest distress, followed by a rapid decline in hemoglobin levels, the presence of hemoglobin in the urine (hemoglobinuria), renal failure, and disseminated intravascular coagulation. Laboratory analysis revealed high-titer antibodies against ceftizoxime and red blood cells (RBCs) in the patient's serum, including immunoglobulin M (IgM) (1:128) antibodies and immunoglobulin G (IgG) (1:8) antibodies, with noted crossreactivity to ceftriaxone. Significant improvement in the patient's hemolytic symptoms was observed following immediate discontinuation of the drug, two plasma exchanges, and extensive RBC transfusion. Conclusion: This case, together with previous reports, underscores the importance of considering DIIHA in patients who exhibit unexplained decreases in hemoglobin levels following antibiotic therapy. A thorough examination of the patient's medical history can provide crucial insights for diagnosing DIIHA. The effective management of DIIHA includes immediate cessation of the implicated drug, plasma exchange, and transfusion support based on the identification of specific drug-dependent antibodies through serological testing.

Spatial transcriptomic analysis reveals local effects of intratumoral fusobacterial infection on DNA damage and immune signaling in rectal cancer.

Mucinous colorectal cancer (CRC) is a common histological subtype of colorectal adenocarcinoma, associated with a poor response to chemoradiotherapy. The commensal facultative anaerobes fusobacteria, have been associated with poor prognosis specifically in mesenchymal CRC. Interestingly, fusobacterial infection is especially prevalent in mucinous CRC. The objective of this study was therefore to increase our understanding of beneficial and detrimental effects of fusobacterial infection, by contrasting host cell signaling and immune responses in areas of high vs. low infection, using mucinous rectal cancer as a clinically relevant example. We employed spatial transcriptomic profiling of 106 regions of interest from 8 mucinous rectal cancer samples to study gene expression in the epithelial and immune segments across regions of high versus low fusobacterial infection. Fusobacteria high regions were associated with increased oxidative stress, DNA damage, and P53 signaling. Meanwhile regions of low fusobacterial prevalence were characterized by elevated JAK-STAT, Il-17, Il-1, chemokine and TNF signaling. Immune masks within fusobacterial high regions were characterized by elevated proportions of cytotoxic (CD8+) T cells (p = 0.037), natural killer (NK) cells (p < 0.001), B-cells (p < 0.001), and gamma delta T cells (p = 0.003). Meanwhile, fusobacteria low regions were associated with significantly greater M2 macrophage (p < 0.001), fibroblast (p < 0.001), pericyte (p = 0.002), and endothelial (p < 0.001) counts.

IFN-Type-I Response and Systemic Immunity in Rectal Adenocarcinoma Patients Treated with Conventional or Hypofractionated Neoadjuvant Radiotherapy.

The IFN-type-I pathway is involved in radiotherapy (RT)-mediated immune responses. Large RT fractions have been suggested to potently induce this pathway. Neoadjuvant hypofractionated short-course (scRT) and conventional long-course (lcRT) RT applied for the treatment of locally advanced rectal adenocarcinoma patients provides a unique model to address the immuno-stimulatory properties of RT on a systemic level. We prospectively analyzed the IFNß plasma levels and lymphocyte counts (LCs) of rectal adenocarcinoma patients before and after treatment with scRT (n = 22) and lcRT (n = 40). Flow cytometry was conducted to assess the effects on lymphocytic subpopulations in a subset of 20 patients. A statistically significant increase in the post-RT IFNß plasma levels was noted in patients undergoing scRT (p = 0.004). Improved pathological tumor regression was associated with elevated post-RT IFNß levels (p = 0.003). Although all patients experienced substantial lymphopenia after treatment, the post-RT LC of patients treated with scRT were significantly higher compared to lcRT (p = 0.001). Patients undergoing scRT displayed significantly lower percentages of regulatory CD4+/CD25+ T-cells after therapy (p = 0.02). scRT enables effective stimulation of the IFN-type-I pathway on a systemic level and confers decreased lymphocytic cytotoxicity and limited regulatory T-cell activation compared to lcRT, supporting its increasing role in immuno-RT trials.

Bioinformatics analysis of markers based on m6A related to prognosis combined with immune invasion of rectal adenocarcinoma.

BACKGROUND: Colorectal cancer (CRC) is a common form of cancer, with rectal cancer accounting for approximately one-third of all cases. Among rectal cancers, 95% are classified as rectal adenocarcinoma (READ). Emerging evidence suggests that long noncoding RNAs (lncRNAs) play a significant role in the development and progression of various cancers. In our study, we aimed to identify differentially expressed lncRNAs potentially associated with m6A and establish a risk assessment model to predict clinical outcomes for READ patients. METHODS: The READ dataset from the TCGA database was utilized in this study to synergistically and logically integrate m6A and lncRNA, while employing bioinformatics technology for the identification of suitable biomarkers. A risk prediction model comprising m6A-associated lncRNAs was constructed to investigate the prognostic, diagnostic, and biological functional relevance of these m6A-related lncRNAs. RESULTS: Our research builds a composed of three related to m6A lncRNA rectal gland cancer prognosis model, and the model has been proved in the multi-dimensional can serve as the potential of the prognosis of rectal gland cancer biomarkers. Our study constructed a prognostic model of rectal adenocarcinoma consisting of three related m6A lncRNAs: linc00702, ac106900.1 and al583785.1. CONCLUSION: The model has been validated as a potential prognostic biomarker for rectal cancer in multiple dimensions, aiming to provide clinicians with an indicator to assess the duration of straight adenocarcinoma. This enables early detection of rectal cancer and offers a promising target for immunotherapy.

CD11c+ and IRF8+ cell densities in rectal cancer biopsies predict outcomes of neoadjuvant chemoradiotherapy.

Approximately 20% of locally advanced rectal cancer (LARC) patients treated preoperatively with chemoradiotherapy (CRT) achieve pathologically confirmed complete regression. However, there are no clinically implemented biomarkers measurable in biopsies that are predictive of tumor regression. Here, we conducted multiplexed immunophenotyping of rectal cancer diagnostic biopsies from 16 LARC patients treated preoperatively with CRT. We identified that patients with greater tumor regression had higher tumor infiltration of pan-T cells and IRF8+HLA-DR+ cells prior to CRT. High IRF8+HLA-DR+ cell density was further associated with prolonged disease-specific survival with 83% survival at 5 y compared to 28% in patients with low infiltration. Contrastingly, low CD11c+ myeloid cell infiltration prior to CRT was a putative biomarker associated with longer 3- and 5-y disease-free survival. The results demonstrate the potential use of rectal cancer diagnostic biopsies to measure IRF8+ HLA-DR+ cells as predictors of CRT-induced tumor regression and CD11c+ myeloid cells as predictors of LARC patient survival.

Tumor-infiltrating lymphocytes, PD-L1, and MMR-deficiency combined characterization may identify subgroups of rectal cancer patients who would benefit from immunotherapy.

INTRODUCTION: Mismatch repair deficiency, immunological fertility, and PD-L1 expression status are key histopathological and molecular features defining tumor responsiveness to immunotherapy and, eventually, prognosis. These were investigated in a series of locally advanced rectal cancer patients treated with postoperative chemotherapy and radiotherapy. MATERIALS AND METHODS: Tumor-infiltrating lymphocyte (TIL) density was assessed in hematoxylin-eosin tissue sections. PD-L1 expression and the expression of MMR proteins (MLH1, PSM2, MSH2, and MSH6) were assessed with immunohistochemistry. Their association with histopathological variables (node involvement and tumor budding) and prognosis was assessed. RESULTS: The TIL-density was significantly higher in the invading tumor front and was inversely related to tumor budding and directly with better overall survival (OS) and distant metastasis-free survival (DMFS) (p = 0.02 and 0.02, respectively). Cancer cell PD-L1 expression was related to high TIL-density (p < 0.01) but not to prognosis, although its overexpression defined a trend for poorer OS in patients with high TIL-density. High PD-L1 expression by stroma infiltrating immune cells was linked with better OS and DMFS (p = 0.007 and 0.001, respectively. MMR deficiency was recorded in 26.2 % of cases, and this was linked with higher TIL-density, but not with prognosis. CONCLUSIONS: Dense intratumoral lymphocytic infiltration relates to a better prognosis in rectal cancer, although it is also linked with PD-L1 expression that may adversely modulate the anti-tumor effects of TILs. This latter subgroup of patients (high TIL-density/high cancer cell PD-L1 expression) could be an additional target for anti-PD-1/PD-L1 immunotherapy, along with the established subgroup of MMR deficient patients.

A combination of stromal PD-L1 and tumoral nuclear ß-catenin expression as an indicator of colorectal carcinoma progression and resistance to chemoradiotherapy in locally advanced rectal carcinoma.

Programmed cell death-1 (PD-1) and its ligand (PD-L1) are significant mediators of immune suppression in the tumor microenvironment. We focused on the immunological impact of PD-1/PD-L1 signaling during tumor progression in colorectal carcinoma (CRC) and its association with resistance to neoadjuvant chemoradiotherapy (NCRT) in locally advanced rectal carcinoma (LAd-RC). Histopathological and immunohistochemical analyses of 100 CRC cases (including 34 RC) without NCRT and 109 NCRT-treated LAd-RC cases were performed. Membranous tumoral PD-L1 expression was identified in 9 of 100 (9%) CRC cases, including 1 of 34 (2.9%) RC cases, but PD-L1 immunopositivity was not associated with any clinicopathological factors, with the exception of deficient mismatch repair (dMMR) status. In contrast, stromal PD-L1+ immune cells, which frequently exhibited coexpression of PD-1 and CD8 markers, were significantly correlated with tumor vessel invasion, nuclear ß-catenin+ tumor budding cancer stem cell (CSC)-like features, and unfavorable prognosis. In the LAd-RC cases, stromal CD8+ (but not PD-L1+) immune cell infiltration in pretreatment-biopsied samples was significantly and positively associated with therapeutic efficacy. After NCRT, tumoral PD-L1 expression was observed in only 2 of 83 (2.4%) tumors, independent of dMMR status, whereas high stromal PD-L1+ and tumoral nuclear ß-catenin positivity were significantly linked to a poor response to NCRT and high tumor budding features. In addition, high stromal PD-L1 immunoreactivity was significantly associated with poorer overall survival. In conclusion, a combination of stromal PD-L1+ immune cells and nuclear ß-catenin+ tumor budding may contribute to tumor progression in CRC and resistance to NCRT in LAd-RC, through formation of niche-like lesions that exhibit immune resistance and CSC properties.

The Prognostic Value of FoxP3+ Tumour-Infiltrating Lymphocytes in Rectal Cancer Depends on Immune Phenotypes Defined by CD8+ Cytotoxic T Cell Density.

Tumour-infiltrating FoxP3+ regulatory T cells have been identified as both positive and negative prognostic factors in colorectal cancer (CRC) and rectal cancer (RC). In this study we investigated whether immune phenotypes, defined by CD8+ cytotoxic T cell density, may influence the prognostic association of FoxP3+ T cell densities in RC. Tissue microarrays from 154 rectal cancer resections were immunohistochemically double stained for CD8 and FoxP3. CD8+ and FoxP3+ cell densities were measured in the stromal and intraepithelial compartment. Stromal FoxP3+ cell densities were not associated with 10-year overall survival (OS). In the "immune-desert" phenotype, defined by very low stromal CD8+ cell density, a high density of stromal FoxP3+ T cells displayed a tendency towards an association with decreased 10-year OS (p = 0.179). In "inflamed" tumours, defined by high intraepithelial CD8+ T cell infiltration, the opposite was the case and high stromal FoxP3+ T cell densities were a positive prognostic factor (p = 0.048). Additionally, patients with an increased FoxP3/CD8 cell density ratio demonstrated a strong trend towards decreased 10-year OS (p = 0.066). These contrasting findings suggest functional heterogeneity within the group of FoxP3+ T cells. They are consistent with experimental studies which reported suppressive and non-suppressive populations of FoxP3+ T cells in CRC. Furthermore, our study demonstrates that CD8 immunohistochemistry may act as an instrument to identify tumours infiltrated by possibly functionally differing FoxP3+ T cell subtypes.

Relationship between stromal regulatory T cells and the response to neoadjuvant chemotherapy for locally advanced rectal cancer.

BACKGROUND: In addition to the direct power of anticancer drugs, the effectiveness of anticancer therapy depends on the host immune function. The present study investigated whether or not the reduction rate and histological response of preoperative chemotherapy were related to the immune microenvironment surrounding a primary tumor of the rectum. METHODS: Sixty-five patients received preoperative chemotherapy followed by resection from 2012 to 2014; all of these patients were retrospectively analyzed. CD3, CD8, and FoxP3 were immunohistochemically examined as markers for T lymphocytes, cytotoxic T lymphocytes, and regulatory T lymphocytes (Treg), respectively. The correlation between the tumor-infiltrating lymphocyte composition and the tumor reduction rate and histological response to neoadjuvant chemotherapy was investigated. RESULTS: The average tumor reduction rate was 41.5% ± 18.8%. According to RECIST, 47 patients (72.3%) achieved a partial response (PR), and 1 patient (1.5%) achieved a complete response (CR). Eight patients (12.3%) showed a grade 2 histological response, and 2 (3.1%) showed a grade 3 response. A multivariate analysis demonstrated that a low Treg infiltration in stromal cell areas was significantly associated with the achievement of a PR or CR [odds ratio (OR) 7.69; 95% confidence interval (CI) 1.96-33.33; p < 0.01] and a histological grade 2 or 3 response (OR 11.11; 95% CI 1.37-98.04; p = 0.02). CONCLUSION: A low Treg infiltration in the stromal cell areas may be a marker of a good response to neoadjuvant chemotherapy in patients with locally advanced rectal cancer.

Squamous cell carcinoma of the rectum in an HIV-1 patient on antiretroviral therapy.

Primary squamous cell carcinoma of the rectum is a very rare malignancy in clinical practice. There are only a few case-studies related to SCC rectum in HIV-1 infected persons in the literature. We report here a case in which the patient, in spite of receiving combination antiretroviral therapy and optimum virological control, suffered from SC C rectum. He was treated with chemoradiation but he relapsed soon and now receiving palliative treatment with Cisplatin and Fluorouracil. There are no case-reports on this issue from India and probably this is the first case-report on SCC rectum in HIV-1 infection adequately treated with cART. Since this is very aggressive disease, outcome is poor.

Potential role of tumor-infiltrating T-, B-lymphocytes, tumor-associated macrophages and IgA-secreting plasma cells in long-term survival in the rectal adenocarcinoma patients.

AIMS: Many studies investigated the associations between the role of immune cells of rectal cancer microenvironment and survival during the first 5 years post-surgery. This is problematic as this disease has the potential to progress even after 5 years after relapse and infiltrating immune cells could play key roles. Therefore, this retrospective study investigates expression and roles of tumor-infiltrating T-lymphocytes (TIL-T), tumor-infiltrating B-lymphocytes (TILB), IgA+ plasma cells (IgA+ PC) and tumor-associated macrophages (TAM) in patients with or without progression over 5 years survival with rectal adenocarcinoma. MAIN METHODS: Here we used immunohistochemical staining of CD3, CD20, IgA, CD68 positive cells and its detection in rectal cancer stroma. Data was analyzed using Mann Whitney U test, ROC, survival and Cox's regression analysis. KEY FINDINGS: The number of TIL-T (p = 0.0276), TIL-B (p < 0.0001) and IgA+ PC (p = 0.015) immune cells was significantly higher in rectal cancer stroma of patients with favorable outcome. Univariate Cox's regression analysis revealed a predictive role of TIL-T (HR = 0.482; 95% CI, 0.303 to 0.704; p < 0.0001), TIL-B (HR = 0.301; 95% CI, 0.198 to 0.481; p < 0.0001) and IgA+-PC (HR = 0.488; 95% CI, 0.322 to 0.741; p < 0.0001). Multivariate Cox's regression analysis showed prognostic role of TIL-B (HR = 0.940; 95% CI, 0.914 to 0.968; p < 0.0001) and IgA+-PC (HR = 0.985; 95% CI, 0.975 to 0.996; p = 0.006) play role in long time survival. SIGNIFICANCE: CD20+ TIL-B and IgA+ cells have significant associations with long -term survival of patients with rectal cancer, with potential therapeutic intervention in cancer immunotherapy.

Peripheral blood CD45RO+T cells is a predictor of the effectiveness of neoadjuvant chemoradiotherapy in locally advanced rectal cancer.

ABSTRACT: To investigate the relationship between the changes in circulating CD45RO+T lymphocyte subsets following neoadjuvant therapy for rectal cancer in patients with locally advanced rectal cancer.The clinicopathological data of 185 patients with rectal cancer who received neoadjuvant therapy in the General Surgery Department of Beijing Chaoyang Hospital affiliated to Capital Medical University from June 2015 to June 2017 were analyzed. Venous blood samples were collected 1 week before neoadjuvant therapy and 1 week before surgery, and the expression of CD45RO+T was detected by flow cytometry. The receiver operating characteristic curve analysis was used to determine the optimal cut-off point of CD45RO+ratio. Log-rank test and multivariate Cox regression were used to analyze the overall survival rate (OS) and disease-free survival rate (DFS) associated with CD45RO+ratio.Circulating CD45RO+ratio of 1.07 was determined as the optimal cut-off point and CD45RO+ratio-high was associated with lower tumor regression grade grading (P = .031), T stage (P = .001), and tumor node metastasis (TNM) stage (P = .012). The 3-year DFS and OS rate in the CD45RO+ratio-high group was significantly higher than that in the CD45RO+ratio-low group (89.2% vs 60.1%, P<.001; 94.4% vs 73.2%, P<.001). The multivariate Cox analysis revealed that elevated CD45RO+ratio was an independent factor for better DFS (OR, 0.339; 95% CI, 0.153-0.752; P = .008) and OS (OR, 0.244; 95% CI,0.082-0.726; P = .011).Circulating CD45RO+ratio could predict the tumor regression grade of neoadjuvant therapy for rectal cancer, as well as long-term prognosis. These findings could be used to stratify patients and develop alternative strategies for adjuvant therapy.

Prognostic value of tumour microenvironment-related genes by TCGA database in rectal cancer.

Rectal cancer is a common malignant tumour and the progression is highly affected by the tumour microenvironment (TME). This study intended to assess the relationship between TME and prognosis, and explore prognostic genes of rectal cancer. The gene expression profile of rectal cancer was obtained from TCGA and immune/stromal scores were calculated by Estimation of Stromal and Immune cells in Malignant Tumors using Expression data (ESTIMATE) algorithm. The correlation between immune/stromal scores and survival time as well as clinical characteristics were evaluated. Differentially expressed genes (DEGs) were identified according to the stromal/immune scores, and the functional enrichment analyses were conducted to explore functions and pathways of DEGs. The survival analyses were conducted to clarify the DEGs with prognostic value, and the protein-protein interaction (PPI) network was performed to explore the interrelation of prognostic DEGs. Finally, we validated prognostic DEGs using data from the Gene Expression Omnibus (GEO) database by PrognoScan, and we verified these genes at the protein levels using the Human Protein Atlas (HPA) databases. We downloaded gene expression profiles of 83 rectal cancer patients from The Cancer Genome Atlas (TCGA) database. The Kaplan-Meier plot demonstrated that low-immune score was associated with worse clinical outcome (P = .034), metastasis (M1 vs. M0, P = .031) and lymphatic invasion (+ vs. -, P < .001). A total of 540 genes were screened as DEGs with 539 up-regulated genes and 1 down-regulated gene. In addition, 60 DEGs were identified associated with overall survival. Functional enrichment analyses and PPI networks showed that the DEGs are mainly participated in immune process, and cytokine-cytokine receptor interaction. Finally, 19 prognostic genes were verified by GSE17536 and GSE17537 from GEO, and five genes (ADAM23, ARHGAP20, ICOS, IRF4, MMRN1) were significantly different in tumour tissues compared with normal tissues at the protein level. In summary, our study demonstrated the associations between TME and prognosis as well as clinical characteristics of rectal cancer. Moreover, we explored and verified microenvironment-related genes, which may be the potential key prognostic genes of rectal cancer. Further clinical samples and functional studies are needed to validate this finding.

The circulating soluble form of the CD40 costimulatory immune checkpoint receptor and liver metastasis risk in rectal cancer.

BACKGROUND: In colorectal cancer, the inflamed tumour microenvironment with its angiogenic activities is immune- tolerant and incites progression to liver metastasis. We hypothesised that angiogenic and inflammatory factors in serum samples from patients with non-metastatic rectal cancer could inform on liver metastasis risk. METHODS: We measured 84 angiogenic and inflammatory markers in serum sampled at the time of diagnosis within the population-based cohort of 122 stage I-III patients. In a stepwise manner, the statistically strongest proteins associated with time to development of liver metastasis were analysed in the corresponding serum samples from 273 stage II-III rectal cancer patients in three independent cohorts. RESULTS: We identified the soluble form of the costimulatory immune checkpoint receptor cluster of differentiation molecule 40 (sCD40) as a marker of liver metastasis risk across all patient cohorts-the higher the sCD40 level, the shorter time to liver metastasis. In patients receiving neoadjuvant treatment, the sCD40 value remained an independent variable associated with progression to liver metastasis along with the local treatment response. Of note, serum sCD40 was not associated with progression to lung metastasis. CONCLUSIONS: Circulating sCD40 is a marker of liver metastasis risk in rectal cancer and may be developed for use in clinical practice.

Neoadjuvant chemoradiation alters biomarkers of anticancer immunotherapy responses in locally advanced rectal cancer.

BACKGROUND: Neoadjuvant chemoradiation therapy (CRT) is a widely used preoperative treatment strategy for locally advanced rectal cancer (LARC). However, a few studies have evaluated the molecular changes caused by neoadjuvant CRT in these cancer tissues. Here, we aimed to investigate changes in immunotherapy-related immunogenic effects in response to preoperative CRT in LARC. METHODS: We analyzed 60 pairs of human LARC tissues before and after irradiation from three independent LARC cohorts, including a LARC patient RNA sequencing (RNA-seq) dataset from our cohort and GSE15781 and GSE94104 datasets. RESULTS: Gene ontology analysis showed that preoperative CRT significantly enriched the immune response in LARC tissues. Moreover, gene set enrichment analysis revealed six significantly enriched Kyoto Encyclopedia of Genes and Genomes pathways associated with downregulated genes, including mismatch repair (MMR) genes, in LARC tissues after CRT in all three cohorts. Radiation also induced apoptosis and downregulated various MMR system-related genes in three colorectal cancer cells. One patient with LARC showed a change in microsatellite instability (MSI) status after CRT, as demonstrated by the loss of MMR protein and PCR for MSI. Moreover, CRT significantly increased tumor mutational burden in LARC tissues. CIBERSORT analysis revealed that the proportions of M2 macrophages and CD8 T cells were significantly increased after CRT in both the RNA-seq dataset and GSE94104. Notably, preoperative CRT increased various immune biomarker scores, such as the interferon-γ signature, the cytolytic activity and the immune signature. CONCLUSIONS: Taken together, our findings demonstrated that neoadjuvant CRT modulated the immune-related characteristics of LARC, suggesting that neoadjuvant CRT may enhance the responsiveness of LARC to immunotherapy.

Differential and longitudinal immune gene patterns associated with reprogrammed microenvironment and viral mimicry in response to neoadjuvant radiotherapy in rectal cancer.

BACKGROUND: Rectal cancers show a highly varied response to neoadjuvant radiotherapy/chemoradiation (RT/CRT) and the impact of the tumor immune microenvironment on this response is poorly understood. Current clinical tumor regression grading systems attempt to measure radiotherapy response but are subject to interobserver variation. An unbiased and unique histopathological quantification method (change in tumor cell density (ΔTCD)) may improve classification of RT/CRT response. Furthermore, immune gene expression profiling (GEP) may identify differences in expression levels of genes relevant to different radiotherapy responses: (1) at baseline between poor and good responders, and (2) longitudinally from preradiotherapy to postradiotherapy samples. Overall, this may inform novel therapeutic RT/CRT combination strategies in rectal cancer. METHODS: We generated GEPs for 53 patients from biopsies taken prior to preoperative radiotherapy. TCD was used to assess rectal tumor response to neoadjuvant RT/CRT and ΔTCD was subjected to k-means clustering to classify patients into different response categories. Differential gene expression analysis was performed using statistical analysis of microarrays, pathway enrichment analysis and immune cell type analysis using single sample gene set enrichment analysis. Immunohistochemistry was performed to validate specific results. The results were validated using 220 pretreatment samples from publicly available datasets at metalevel of pathway and survival analyses. RESULTS: ΔTCD scores ranged from 12.4% to -47.7% and stratified patients into three response categories. At baseline, 40 genes were significantly upregulated in poor (n=12) versus good responders (n=21), including myeloid and stromal cell genes. Of several pathways showing significant enrichment at baseline in poor responders, epithelial to mesenchymal transition, coagulation, complement activation and apical junction pathways were validated in external cohorts. Unlike poor responders, good responders showed longitudinal (preradiotherapy vs postradiotherapy samples) upregulation of 198 immune genes, reflecting an increased T-cell-inflamed GEP, type-I interferon and macrophage populations. Longitudinal pathway analysis suggested viral-like pathogen responses occurred in post-treatment resected samples compared with pretreatment biopsies in good responders. CONCLUSION: This study suggests potentially druggable immune targets in poor responders at baseline and indicates that tumors with a good RT/CRT response reprogrammed from immune "cold" towards an immunologically "hot" phenotype on treatment with radiotherapy.

Combination of lymphocyte count and albumin concentration as a new prognostic biomarker for rectal cancer.

Although numerous studies have highlighted the prognostic values of various inflammation-related markers, clinical significance remains to be elucidated. The prognostic values of inflammation-related biomarkers for rectal cancer were investigated in this study. A total of 448 patients with stage II/III rectal cancer undergoing curative resection were enrolled from the discovery cohort (n = 240) and validation cohort (n = 208). We comprehensively compared the prognostic values of 11 inflammation-related markers-derived from neutrophil, lymphocyte, platelet, monocyte, albumin, and C-reactive protein for overall survival (OS) and recurrence-free survival (RFS). Among 11 inflammation-related markers, only "lymphocyte × albumin (LA)" was significantly associated with both OS and RFS in the discovery cohort (P = 0.007 and 0.015, respectively). Multivariate analysis indicated that low LA was significantly associated with poor OS (hazard ratio [HR] 2.19, 95% confidence interval [CI] 1.09-4.58, P = 0.025), and poor RFS (HR 1.61, 95% CI 1.01-2.80, P = 0.048). Furthermore, using the discovery cohort, we confirmed that low LA was significantly associated with poor OS (HR 2.89, 95% CI 1.42-6.00, P = 0.002), and poor RFS (HR 1.79, 95% CI 1.04-2.95, P = 0.034). LA can be a novel prognostic biomarker for stage II/III rectal cancer.

Chr20q Amplification Defines a Distinct Molecular Subtype of Microsatellite Stable Colorectal Cancer.

Colorectal cancer is the third leading cause of cancer-related death in the United States. About 15% of colorectal cancers are associated with microsatellite instability (MSI) due to loss of function in the DNA mismatch repair pathway. This subgroup of patients has better survival rates and is more sensitive to immunotherapy. However, it remains unclear whether microsatellite stable (MSS) patients with colorectal cancer can be further stratified into subgroups with differential clinical characteristics. In this study, we analyzed The Cancer Genome Atlas data and found that Chr20q amplification is the most frequent copy number alteration that occurs specifically in colon (46%) and rectum (61%) cancer and is mutually exclusive with MSI. Importantly, MSS patients with Chr20q amplification (MSS-A) were associated with better recurrence-free survival compared with MSS patients without Chr20q amplification (MSS-N; P = 0.03). MSS-A tumors were associated with high level of chromosome instability and low immune infiltrations. In addition, MSS-A and MSS-N tumors were associated with somatic mutations in different driver genes, with high frequencies of mutated TP53 in MSS-A and mutated KRAS and BRAF in MSS-N. Our results suggest that MSS-A and MSS-N represent two subtypes of MSS colorectal cancer, and such stratification may be used to improve therapeutic treatment in an individualized manner. SIGNIFICANCE: This study shows that chromosome 20q amplification occurs predominately in microsatellite-stable colorectal cancer and defines a distinct subtype with good prognosis, high chromosomal instability, distinct mutation profiles, and low immune infiltrations.