Centrosomal Protein 55 (CEP55) Drives Immune Exclusion and Resistance to Immune Checkpoint Inhibitors in Colorectal Cancer.

Colorectal cancer (CRC) currently ranks as the third most common cancer in the United States, and its incidence is on the rise, especially among younger individuals. Despite the remarkable success of immune checkpoint inhibitors (ICIs) in various cancers, most CRC patients fail to respond due to intrinsic resistance mechanisms. While microsatellite instability-high phenotypes serve as a reliable positive predictive biomarker for ICI treatment, the majority of CRC patients with microsatellite-stable (MSS) tumors remain ineligible for this therapeutic approach. In this study, we investigated the role of centrosomal protein 55 (CEP55) in shaping the tumor immune microenvironment in CRC. CEP55 is overexpressed in multiple cancer types and was shown to promote tumorigenesis by upregulating the PI3K/AKT pathway. Our data revealed that elevated CEP55 expression in CRC was associated with reduced T cell infiltration, contributing to immune exclusion. As CRC tumors progressed, CEP55 expression increased alongside sequential mutations in crucial driver genes (APC, KRAS, TP53, and SMAD4), indicating its involvement in tumor progression. CEP55 knockout significantly impaired tumor growth in vitro and in vivo, suggesting that CEP55 plays a crucial role in tumorigenesis. Furthermore, the CEP55 knockout increased CD8+ T cell infiltration and granzyme B production, indicating improved anti-tumor immunity. Additionally, we observed reduced regulatory T cell infiltration in CEP55 knockout tumors, suggesting diminished immune suppression. Most significantly, CEP55 knockout tumors demonstrated enhanced responsiveness to immune checkpoint inhibition in a clinically relevant orthotopic CRC model. Treatment with anti-PD1 significantly reduced tumor growth in CEP55 knockout tumors compared to control tumors, suggesting that inhibiting CEP55 could improve the efficacy of ICIs. Collectively, our study underscores the crucial role of CEP55 in driving immune exclusion and resistance to ICIs in CRC. Targeting CEP55 emerges as a promising therapeutic strategy to sensitize CRC to immune checkpoint inhibition, thereby improving survival outcomes for CRC patients.

Allogeneic tumor cell-derived extracellular vesicles stimulate CD8 T cell response in colorectal cancer.

Most colorectal cancer (CRC) patients present with a microsatellite-stable phenotype, rendering them resistant to immune checkpoint inhibitors (ICIs). Among the contributors to ICI resistance, tumor-derived extracellular vesicles (TEVs) have emerged as critical players. Previously we demonstrated that autologous transfer of TEVs without miR-424 can induce tumor antigen-specific immune responses in CRC models. Therefore, we postulated that allogeneic TEVs, modified to lack miR-424 and derived from an MC38 cells, could induce CD8+ T cell responses while restraining CT26 cell-based tumor. Here, we show that prophylactic administration of MC38 TEVs, without miR-424, showed a significant augmentation in CD8+ T-cells within CT26 tumors. This allogenic TEV effect was evident in CT26 tumors but not B16-F10 melanoma. Furthermore, we demonstrated the capacity of dendritic cells (DCs) to internalize TEVs, a possible mechanism to elicit immune response. Our investigation of autologously administered DCs, which had been exposed to modified TEVs, underscores their potential to dampen tumor growth while elevating CD8+ T cell levels vis-a-vis MC38 wild-type TEVs exposed to DCs. Notably, the modified TEVs were well tolerated and did not increase peripheral blood cytokine levels. Our findings underscore the potential of modified allogeneic TEVs without immune-suppressive factors to elicit robust T cell responses and limit tumor growth.

Allogeneic Tumor Cell-Derived Extracellular Vesicles Stimulate CD8 T Cell Response in Colorectal Cancer.

Colorectal Cancer (CRC) is the second leading cause of cancer-related death in the United States. Most CRC patients present with a microsatellite stable (MSS) phenotype and are highly resistant to immunotherapies. Tumor extracellular vesicles (TEVs), secreted by tumor cells, can contribute to intrinsic resistance to immunotherapy in CRC. We previously showed that autologous TEVs without functional miR-424 induce anti-tumor immune responses. We hypothesized that allogeneic modified CRC-TEVs without miR-424 (mouse homolog miR-322) derived from an MC38 background would effectively stimulate CD8+ T cell response and limit CT26 tumor growth. Here we show that prophylactic administration of MC38 TEVs without functional miR-424 significantly increased CD8+ T cells in CT26 CRC tumors and limited tumor growth, not B16-F10 melanoma tumors. We further show that the depletion of CD4+ and CD8+ T cells abolished the protective effects of MC38 TEVs without functional miR-424. We further show that TEVs can be taken up by DCs in vitro, and subsequent prophylactic administration of autologous DCs exposed to MC38 TEVs without functional miR-424 suppressed tumor growth and increased CD8+ T cells compared to MC38 wild-type TEVs exposed to DCs, in Balb/c mice bearing CT26 tumors. Notably, the modified EVs were well tolerated and did not increase cytokine expression in peripheral blood. These findings suggest that allogeneic-modified CRC-EVs without immune suppressive miR-424 can induce antitumor CD8+ T cell responses and limit tumor growth in vivo.

Tumor-derived extracellular vesicles in the colorectal cancer immune environment and immunotherapy.

Despite significant advances in the screening, diagnosis, and treatment of colorectal cancer (CRC) immune checkpoint inhibitors (ICIs) continue to have limited utility outside of microsatellite-high disease. Given the durable response to immunotherapy seen across malignancies, increasing CRC response rates to ICI therapy is an active area of clinical research. An increasing body of work has demonstrated that tumor-derived extracellular vesicles (TEVs) are key modulators in tumor signaling and the determinants of the tumor microenvironment. Pre-clinical models have shown that TEVs are directly involved in antigen presentation and are involved in radiation-induced DNA damage signaling. Both direct and indirect modifications of these TEVs can alter CRC immunogenicity and ICI treatment response, making them attractive targets for potential therapeutic development. In addition, modified TEVs can be developed using several different mechanisms, with varied cargo including micro-RNAs and small peptide molecules. Recent work has shown strong pre-clinical evidence of injected modified TEV-induced ICI activity, with knockdown of the micro-RNA miR-424 in TEVs improving CRC immunogenicity and increasing anti-PD-1 activity in mouse models. Clinical trials are ongoing in the evaluation of modified TEVs in cancer therapy, but they appear to be a promising therapeutic target in CRC.

Tumor models to assess immune response and tumor-microbiome interactions in colorectal cancer.

Despite significant advances over the past 2 decades in preventive screening and therapy aimed at improving patient survival, colorectal cancer (CRC) remains the second most common cause of cancer death in the United States. The average 5-year survival rate of CRC patients with positive regional lymph nodes is only 40%, while less than 5% of patients with distant metastases survive beyond 5 years. There is a critical need to develop novel therapies that can improve overall survival in patients with poor prognoses, particularly since 60% of them are diagnosed at an advanced stage. Pertinently, immune checkpoint blockade therapy has dramatically changed how we treat CRC patients with microsatellite-instable high tumors. Furthermore, accumulating evidence shows that changes in gut microbiota are associated with the regulation of host antitumor immune response and cancer progression. Appropriate animal models are essential to deciphering the complex mechanisms of host antitumor immune response and tumor-gut microbiome metabolic interactions. Here, we discuss various mouse models of colorectal cancer that are developed to address key questions on tumor immune response and tumor-microbiota interactions. These CRC models will also serve as resourceful tools for effective preclinical studies.

ACKR4 in Tumor Cells Regulates Dendritic Cell Migration to Tumor-Draining Lymph Nodes and T-Cell Priming.

Colorectal cancer (CRC) is one of the most common malignancies in both morbidity and mortality. Immune checkpoint blockade (ICB) treatments have been successful in a portion of mismatch repair-deficient (dMMR) CRC patients but have failed in mismatch repair-proficient (pMMR) CRC patients. Atypical Chemokine Receptor 4 (ACKR4) is implicated in regulating dendritic cell (DC) migration. However, the roles of ACKR4 in CRC development and anti-tumor immunoregulation are not known. By analyzing human CRC tissues, transgenic animals, and genetically modified CRC cells lines, our study revealed an important function of ACKR4 in maintaining CRC immune response. Loss of ACKR4 in CRC is associated with poor immune infiltration in the tumor microenvironment. More importantly, loss of ACKR4 in CRC tumor cells, rather than stromal cells, restrains the DC migration and antigen presentation to the tumor-draining lymph nodes (TdLNs). Moreover, tumors with ACKR4 knockdown become less sensitive to immune checkpoint blockade. Finally, we identified that microRNA miR-552 negatively regulates ACKR4 expression in human CRC. Taken together, our studies identified a novel and crucial mechanism for the maintenance of the DC-mediated T-cell priming in the TdLNs. These new findings demonstrate a novel mechanism leading to immunosuppression and ICB treatment resistance in CRC.

Tumor-Secreted Extracellular Vesicles Regulate T-Cell Costimulation and Can Be Manipulated To Induce Tumor-Specific T-Cell Responses.

BACKGROUND & AIMS: Colorectal cancer is a major cause of cancer-related deaths worldwide. Immune checkpoint blockade therapies are effective in 30%-60% of the microsatellite instable-high subtype. Unfortunately, most patients with colorectal cancer (>85%) have microsatellite stable tumors that do not respond. In this study, we aimed to decipher the underlying tumor-intrinsic mechanisms critical for improving immunotherapy in colorectal cancer. METHODS: We used human and mouse tumor samples, cell lines, human colorectal cancer organoids, and various syngeneic orthotopic mouse models of late-stage colorectal cancer to define the effects of tumor cell-secreted extracellular vesicles (EVs) on antitumor immune response. RESULTS: Our analyses of human colorectal cancer immune profiles and tumor-immune cell interactions showed that tumor-secreted EVs containing microRNA miR-424 suppressed the CD28-CD80/86 costimulatory pathway in tumor-infiltrating T cells and dendritic cells, leading to immune checkpoint blockade resistance. Modified tumor-secreted EVs with miR-424 knocked down enhanced T-cell-mediated antitumor immune response in colorectal cancer tumor models and increased the immune checkpoint blockade response. Intravenous injections of modified tumor-secreted EVs induced tumor antigen-specific immune responses and boosted the immune checkpoint blockade efficacy in colorectal cancer models that mimic aggressively progressing, late-stage disease. CONCLUSIONS: Collectively, we show a critical role for tumor-secreted EVs in antitumor immune regulation and immunotherapy response, which could be developed as a novel treatment for immune checkpoint blockade-resistant colorectal cancer.

Parasexual recombination enables Aspergillus fumigatus to persist in cystic fibrosis.

Aspergillus fumigatus is a saprobic fungus that causes a range of pulmonary diseases, some of which are characterised by fungal persistence such as is observed in cystic fibrosis (CF) patients. Creation of genetic variation is critical for A. fumigatus to adapt to the lung environment, but biofilm formation, especially in CF patients, may preclude mutational supply in A. fumigatus due to its confinement to the hyphal morphotype. We tested our hypothesis that genetic variation is created through parasexual recombination in chronic biofilms by phenotypic and genetic analysis of A. fumigatus isolates cultured from different origins. As diploids are the hallmark of parasex, we screened 799 A. fumigatus isolates obtained from patients with CF, chronic pulmonary lung disease and acute invasive aspergillosis, and from the environment for spore size. Benomyl sensitivity, nuclear content measurements through fluorescence-activated cell sorting and scanning electron microscopy were used to confirm the diploid state of large size spores. Whole genome sequencing was used to characterise diploid-associated genetic variation. We identified 11 diploids in isolates recovered from six of 11 (55%) CF patients and from one of 24 (4%) chronic aspergillosis patients, but not in 368 isolates from patients with acute Aspergillus infection and the environment. Diploid formation was associated with accumulation of mutations and variable haploid offspring including a voriconazole-resistant isolate. Parasexual recombination allows A. fumigatus to adapt and persist in CF patients, and plays a role in azole resistance development. Our findings are highly significant for understanding the genetics and biology of A. fumigatus in the human lung.

Acquired Resistance to Immune Checkpoint Blockade Therapies.

Immune checkpoint blockade therapy (ICBT) has revolutionized the treatment and management of numerous cancers, yet a substantial proportion of patients who initially respond to ICBT subsequently develop resistance. Comprehensive genomic analysis of samples from recent clinical trials and pre-clinical investigation in mouse models of cancer provide insight into how tumors evade ICBT after an initial response to treatment. Here, we summarize our current knowledge on the development of acquired ICBT resistance, by examining the mechanisms related to tumor-intrinsic properties, T-cell function, and tumor-immune cell interactions. We discuss current and future management of ICBT resistance, and consider crucial questions remaining in this field of acquired resistance to immune checkpoint blockade therapies.

Epidemiological and histopathological characteristics of head and neck cancers in Bhutan from 2011 to 2017: a retrospective descriptive study.

BACKGROUND: Head and neck cancers are among the commonest cancers in the developing world. Personal habits, such as the use of tobacco, betel nut and alcohol are strongly associated with the development of head and neck cancers at certain sites. Therefore, they are among the preventable cancers. In Bhutan, there has not yet been a study conducted on head and neck cancers. OBJECTIVE: To describe baseline epidemiological and histopathological characteristics of head and neck cancers in Bhutan. METHODS: This is a 7-year descriptive study of all cases of head and neck cancers presented at the Jigme Dorji Wangchuk National Referral Hospital from 2011 to 2017. The data were collected from the hospital's medical records section, histopathology records, patient referral unit and some treatment centres in India. Prior approval was sought from the Research and Ethics Board for Health, the Ministry of Health and the hospital management. RESULTS: There were a total of 515 cases of head and neck cancers from 2011 to 2017. The crude incidence rate was 10 per 100,000 and the overall age adjusted rate was 12.3 (95% CI 9.5-15.1) per 100,000 population. The prevalence during this 7-year period was 69.1 per 100,000 population. The commonest cancers are thyroid, oral cavity, hypopharyngeal, laryngeal and nasopharyngeal cancer in decreasing order. Head and neck cancers are more common in males than females in the majority of sites except thyroid, salivary gland and sinonasal malignancies. Thyroid cancers and nasopharyngeal cancers are found to affect younger age groups. Tashigang (48) followed by Paro (43) recorded the highest number of cases. Squamous cell carcinoma is the commonest histopathology type in almost all the cases, while papillary carcinoma is the commonest among thyroid cancers. Personal habits, such as smoking, chewing tobacco, betel nut and alcohol consumption, were found to be more common among patients suffering from oral cavity, laryngeal, hypopharyngeal and oropharyngeal cancers. CONCLUSION: Head and neck cancers are the third most common cancer in Bhutan after stomach cancer and cervical cancer. Thyroid, oral cavity and hypopharynx are the top three anatomical sites for head and neck cancers in Bhutan. The current epidemiological and histopathological profile of head and neck cancers will form a baseline of information and basis for further research on head and neck cancers in Bhutan.

Chemotherapy but Not the Tumor Draining Lymph Nodes Determine the Immunotherapy Response in Secondary Tumors.

Immunotherapies are used as adjuvant therapies for cancers. However, knowledge of how traditional cancer treatments affect immunotherapies is limited. Using mouse models, we demonstrate that tumor-draining lymph nodes (TdLNs) are critical for tumor antigen-specific T cell response. However, removing TdLNs concurrently with established primary tumors did not affect the immune checkpoint blockade (ICB) response on localized secondary tumor due to immunotolerance in TdLNs and distribution of antigen-specific T cells in peripheral lymphatic organs. Notably, treatment response improved with sequential administration of 5-fluorouracil (5-FU) and ICB compared with concurrent administration of ICB with 5-FU. Immune profiling revealed that using 5-FU as induction treatment increased tumor visibility to immune cells, decreased immunosuppressive cells in the tumor microenvironment, and limited chemotherapy-induced T cell depletion. We show that the effect of traditional cytotoxic treatment, not TdLNs, influences immunotherapy response in localized secondary tumors. We postulate essential considerations for successful immunotherapy strategies in clinical conditions.