Colorectal cancer is the third most common cancer in the world with increasing incidence and mortality rates globally. Standard treatments for colorectal cancer have always been surgery, chemotherapy and radiotherapy which may be used in combination to treat patients. However, these treatments have many side effects due to their non-specificity and cytotoxicity toward any cells including normal cells that are growing and dividing. Furthermore, many patients succumb to relapse even after a series of treatments. Thus, it is crucial to have more alternative and effective treatments to treat CRC patients. Immunotherapy is one of the new alternatives in cancer treatment. The strategy is to utilize patients’ own immune systems in combating the cancer cells. Cancer immunotherapy overcomes the issue of specificity which is the major problem in chemotherapy and radiotherapy. The normal cells with no cancer antigens are not affected. The outcomes of some cancer immunotherapy have been astonishing in some cases, but some which rely on the status of patients’ own immune systems are not. Those patients who responded well to cancer immunotherapy have a better prognostic and better quality of life.

Regulatory T cells (Tregs), a subset of CD4+ or CD8+ T cells, play a pivotal role in regulating immune homeostasis. An increase in Tregs was reported in many tumors to be associated with immune suppression and evasion in cancer patients. Despite the importance of Tregs, the molecular signatures that contributed to their pathophysiological relevance remain poorly understood and controversial. In this study, we explored the gene expression profiles in Tregs derived from patients with colorectal cancer (CRC, n=15), colorectal polyps (P, n=15) and in healthy volunteers (N, n=15). Tregs were analyzed using CD4+CD25+CD127lowFoxP3+ antibody markers. Gene expression profiling analysis lead to the identification of 61 and 66 immune-related genes in Tregs derived from CRC and P patients, respectively but not in N-derived Treg samples. Of these, 30 genes were differentially expressed both in CRC- and P-derived Tregs when compared to N-derived Tregs. Most of the identified genes were involved in cytokine/chemokine mediators of inflammation, chemokine receptor, lymphocyte activation and T cell receptor (TCR) signaling pathways. This study highlights some of the molecular signatures that may affect Tregs expansion and possible suppression of function in cancer development. Our findings may provide a better understanding of the immunomodulatory nature of Tregs and could therefore open up new avenues in immunotherapy.

Colorectal cancer poses a major global health issue, profoundly affecting both mortality and morbidity rates across the world. A key obstacle in understanding the pathogenesis of colorectal cancer lies in its high inter-patient and spatial heterogeneity, making standard treatments ineffective. Commonly, the study on colorectal cancer relies on bulk RNA sequencing, offering an average gene expression profile for a heterogenous cell population. However, this approach obscures the heterogeneous characteristics of the cancer cells and hinders a comprehensive understanding of the complex interplay among different cell populations. Recently, the advent of single-cell RNA sequencing has been revolutionary, enabling researchers to analyze individual immune cells and overcome the limitations of bulk RNA sequencing. Through single-cell RNA sequencing, researchers have gained insights into the dynamic nature of the immune response to cancer and potential targets for immunotherapy. In this review, we discuss the technical aspects of single-cell RNA sequencing, the application of single-cell RNA sequencing in cancer immunology, and the potential of single-cell RNA in clinical settings. We believe that harnessing singlecell RNA sequencing in cancer research holds immense potential to drive the development of personalized immunotherapies, aiming to improve patient outcomes in colorectal cancer.

Objectives This study aimed to quantify how patient risk factors relate to COVID-19 severity across categories 1–5 in a prospective, hospital-based cohort. We hypothesised that greater severity would be associated with higher odds of intensive care unit (ICU) admission and in-hospital mortality. Secondary aims were to assess associations with age, viral variants, symptom clusters, lymphocyte count, fasting blood glucose and cytokine profiles. Design Prospective cohort study. Setting A secondary-care/tertiary-care hospital and linked community settings in Cheras, Kuala Lumpur, Malaysia. Participants This study was nested within the COVGEN project, a prospective COVID-19 cohort conducted at Hospital Canselor Tuanku Muhriz UKM (HCTM), Cheras Health Clinic and the Bandar Tun Razak COVID-19 Assessment Centre in Cheras, Kuala Lumpur, Malaysia, from 1 August 2021 to 31 October 2022. 2532 participants were enrolled at baseline. Eligible participants were Malaysian citizens aged 12–18 years (paediatric/adolescent) or ≥18 years who had reverse transcription-polymerase chain reaction–confirmed COVID-19 at recruitment and resided in Kuala Lumpur or Selangor. Patients who had a clinically unstable condition and those who declined participation (personally or via a next-of-kin or legal representative) were excluded. This analysis included 559 patients hospitalised at HCTM; after excluding five with incomplete questionnaires, 554 remained for analysis (413 admitted to general wards and 141 to ICUs). Categories 3–5 comprised hospitalised patients, whereas categories 1–2 included hospitalised individuals and a subset recruited from community settings. Primary and secondary outcome measures Primary outcomes included disease severity (categories 4–5 vs 1–3), ICU admission and in-hospital mortality. Secondary outcomes included associations with age strata, viral variant (delta vs omicron), symptom clusters, lymphocyte count, fasting blood glucose and cytokines: interferon gamma-inducible protein 10, interferon gamma, interleukins 8, 10, 2, 6 and 7 and tumour necrosis factor alpha. Results 141 of 554 (25.5%) patients required ICU care. Compared with milder categories, category 5 was associated with markedly higher odds of ICU admission (OR 204.50; 95% CI 37.54 to 1114.18; p<0.001) and death (OR 13.20, 95% CI 3.97 to 43.86; p55 versus <36 years for categories 3 (OR 4.575; 95% CI 1.472 to 14.218; p=0.009) and 5 (OR 18.003; 95% CI 1.139 to 284.601; p=0.040) and 36–55 versus <36 years for category 5 (OR 18.412; 95% CI 1.094 to 309.951; p=0.043). Delta accounted for 47.7% of infections compared with 17.9% for omicron, with a higher proportion of category 4 presentations among delta cases. Systemic, pulmonary/respiratory and gastrointestinal symptoms, lymphopenia, hyperglycaemia and elevated cytokines were each associated with increasing severity (p<0.05). Conclusions An increasing clinical severity category was strongly associated with ICU admission and mortality. Age, delta infection, specific symptom clusters, lymphopenia, hyperglycaemia and pro-inflammatory cytokines identified higher-risk patients, supporting risk-stratified management and prioritisation for enhanced monitoring.