| 1. |
- Devarakonda, Sravani, et al.
(author)
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Low-grade intestinal inflammation two decades after pelvic radiotherapy.
- 2023
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In: EBioMedicine. - 2352-3964. ; 94
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Journal article (peer-reviewed)abstract
- Radiotherapy is effective in the treatment of cancer but also causes damage to non-cancerous tissue. Pelvic radiotherapy may produce chronic and debilitating bowel symptoms, yet the underlying pathophysiology is still undefined. Most notably, although pelvic radiotherapy causes an acute intestinal inflammation there is no consensus on whether the late-phase pathophysiology contains an inflammatory component or not. To address this knowledge gap, we examined the potential presence of a chronic inflammation in mucosal biopsies from irradiated pelvic cancer survivors.We biopsied 24 cancer survivors two to 20 years after pelvic radiotherapy, and four non-irradiated controls. Using tandem mass tag (TMT) mass spectrometry and mRNA sequencing (mRNA-seq), we charted proteomic and transcriptomic profiles of the mucosal tissue previously exposed to a high or a low/no dose of radiation. Changes in the immune cell populations were determined with flow cytometry. The integrity of the protective mucus layers were determined by permeability analysis and 16S rRNA bacterial detection.942 proteins were differentially expressed in mucosa previously exposed to a high radiation dose compared to a low radiation dose. The data suggested a chronic low-grade inflammation with neutrophil activity, which was confirmed by mRNA-seq and flow cytometry and further supported by findings of a weakened mucus barrier with bacterial infiltration.Our results challenge the idea that pelvic radiotherapy causes an acute intestinal inflammation that either heals or turns fibrotic without progression to chronic inflammation. This provides a rationale for exploring novel strategies to mitigate chronic bowel symptoms in pelvic cancer survivors.This study was supported by the King Gustav V Jubilee Clinic Cancer Foundation (CB), The Adlerbertska Research Foundation (CB), The Swedish Cancer Society (GS), The Swedish State under the ALF agreement (GS and CB), Mary von Sydow's foundation (MA and VP).
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| 2. |
- Liang, Frank, et al.
(author)
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A Fraction of CD8+T Cells from Colorectal Liver Metastases Preferentially Repopulate Autologous Patient-Derived Xenograft Tumors as Tissue-Resident Memory T Cells
- 2022
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In: Cancers. - : MDPI AG. - 2072-6694. ; 14:12
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Journal article (peer-reviewed)abstract
- Simple Summary Treatment options for colorectal cancer (CRC) patients with liver metastases are often limited to liver surgery with or without chemotherapy. However, not all patients present operable colorectal liver metastases (CRLMs). Thus, alternative therapies that exploit the anti-tumor potential of tumor-infiltrating lymphocytes (TILs) are being evaluated. The establishment of markers connecting the phenotype to the function of tumor-reactive CD8+ TILs could aid diagnostic and therapeutic advances. In this regard, tissue-resident memory T cells (T-RM cells) could be a potential candidate for therapies targeting TILs. Putative tumor-reactive T-RM cells among CD8+ TILs likely co-express CD103 and CD39, since these markers indicate stable tumor residency and repeated response to antigens from the tumor environment, respectively. Our phenotypic and functional analyses of TILs in CRLM, with a specific focus on CD103+CD8+ T-RM cells, may guide the improvement of TIL-mediated CRC treatments. The diversity of T cells in the human liver may reflect the composition of TILs in CRLM. Our ex vivo characterization of CRLM vs. adjacent liver tissue detected CD103+CD39+CD8+ T-RM cells predominantly in CRLM, which prompted further assessments. These T-RM cells responded to cognate antigens in vitro. As functional activities of autologous TILs are central to the implementation of personalized cancer treatments, we applied a patient-derived xenograft (PDX) model to monitor TILs' capacity to control CRLM-derived tumors in vivo. We established PDX mice with CRLMs from two patients, and in vitro expansion of their respective TILs resulted in opposing CD4+ vs. CD8+ TIL ratios. These CRLMs also displayed mutated KRAS, which enabled trametinib-mediated inhibition of MEK. Regardless of the TIL subset ratio, persistent or transient control of CRLM-derived tumors of limited size by the transferred TILs was observed only after trametinib treatment. Of note, a portion of transferred TILs was observed as CD103+CD8+ T-RM cells that strictly accumulated within the autologous CRLM-derived tumor rather than in the spleen or blood. Thus, the predominance of CD103+CD39+CD8+ T-RM cells in CRLM relative to the adjacent liver and the propensity of CD103+CD8+ T-RM cells to repopulate the autologous tumor may identify these TILs as strategic targets for therapies against advanced CRC.
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| 3. |
- Liang, Frank, et al.
(author)
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Antigen Presenting Cells from Tumor and Colon of Colorectal Cancer Patients Are Distinct in Activation and Functional Status, but Comparably Responsive to Activated T Cells.
- 2021
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In: Cancers. - : MDPI AG. - 2072-6694. ; 13:20
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Journal article (peer-reviewed)abstract
- Although mouse models of CRC treatments have demonstrated robust immune activation, it remains unclear to what extent CRC patients' APCs and TILs interact to fuel or quench treatment-induced immune responses. Our ex vivo characterization of tumor and adjacent colon cell suspensions suggest that contrasting environments in these tissues promoted inversed expression of T cell co-stimulatory CD80, and co-inhibitory programmed death (PD)-ligand1 (PD-L1) on intratumoral vs. colonic APCs. While putative tumor-specific CD103+CD39+CD8+ TILs expressed lower CD69 (early activation marker) and higher PD-1 (extended activation/exhaustion marker) than colonic counterparts, the latter had instead higher CD69 and lower PD-1 levels. Functional comparisons showed that intratumoral APCs were inferior to colonic APCs regarding protein uptake and upregulation of CD80 and PD-L1 after protein degradation. Our attempt to model CRC treatment-induced T cell activation in vitro showed less interferon (IFN)-γ production by TILs than colonic T cells. In this model, we also measured APCs' CD80 and PD-L1 expression in response to activated co-residing T cells. These markers were comparable in the two tissues, despite higher IFN- γ exposure for colonic APCs. Thus, APCs within distinct intratumoral and colonic milieus showed different activation and functional status, but were similarly responsive to signals from induced T cell activation.
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| 4. |
- Liang, Frank, et al.
(author)
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Cryopreservation of Whole Tumor Biopsies from Rectal Cancer Patients Enable Phenotypic and In Vitro Functional Evaluation of Tumor-Infiltrating T Cells
- 2021
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In: Cancers. - : MDPI AG. - 2072-6694. ; 13:10
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Journal article (peer-reviewed)abstract
- Simple Summary Colorectal cancer (CRC) remains the third most common malignancy. Tumor-infiltrating lymphocytes (TILs) have emerged as correlates to CRC patient outcome after treatment. The pro- or anti-tumor responses of TILs are usually assessed in cell suspensions of fresh tumors that were surgically removed a few hours earlier. We propose a platform for concurrent enumeration and in vitro functional evaluation of TILs in cryopreserved tumor biopsies, offering the benefit of postponing tumor processing and analyses of TILs in cell suspensions until clinical post-treatment responses are established. Our platform is practical considering the inconsistent time when patient samples become available for research purposes and can be readily utilized by other laboratories. With a fresh portion of tumor biopsies as benchmark, we validated the recovery of viable TILs capable of interferon (IFN)-gamma responses in the cryopreserved portion of same biopsies. Ultimately, this platform could provide sufficient information on TILs, to also predict patient outcome after CRC treatments. TILs comprise functionally distinct conventional and unconventional T cell subsets and their role in responses to CRC treatments is poorly understood. We explored recovery of viable TILs from cryopreserved tumor biopsies of (chemo)-radiated patients with rectal cancer to establish a platform for retrospective TIL analyses of frozen tumors from pre-selected study cohorts. Frequencies of TIL subsets and their capacity to mount IFN-gamma responses in cell suspensions of fresh vs. cryopreserved portions of the same tumor biopsies were determined for platform validation. The percentages and proportions of CD4+ TILs and CD8+ cytotoxic T lymphocytes (CTLs) among total TILs were not affected by cryopreservation. While recovery of unconventional gamma delta T cells and mucosal-associated invariant T cells (MAIT cells) was stable after cryopreservation, the regulatory T cells (Tregs) were reduced, but in sufficient yields for quantification. IFN-gamma production by in vitro-stimulated CD4+ TILs, CTLs, gamma delta T cells, and MAIT cells were proportionally similar in fresh and cryopreserved tumor portions, albeit the latter displayed lower levels. Thus, the proposed platform intended for TIL analyses on cryopreserved tumor biobank biopsies holds promises for studies linking the quantity and quality of TIL subsets with specific clinical outcome after CRC treatment.
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| 5. |
- Rezapour, Azar, et al.
(author)
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A type I interferon footprint in pre-operative biopsies is an independent biomarker that in combination with CD8(+) T cell quantification can improve the prediction of response to neoadjuvant treatment of rectal adenocarcinoma
- 2023
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In: OncoImmunology. - 2162-402X. ; 12:1
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Journal article (peer-reviewed)abstract
- Tailored treatment for patients with rectal cancer requires clinically available markers to predict their response to neoadjuvant treatment. The quantity of tumor-infiltrating lymphocytes (TILs) in pre-operative tumor biopsies has been suggested to predict a favorable response, but opposing results exist. A biopsy-adapted Immunoscore (ISB) based on TILs has recently emerged as a promising predictor of tumor regression and prognosis in (colo)rectal cancer. We aimed to refine the ISB for prediction of response using multiplex immunofluorescence (mIF) on pre-operative rectal cancer biopsies. We combined the distribution and density of conventional T cell subsets and ?dT cells with a type I Interferon (IFN)-driven response assessed using Myxovirus resistance protein A (MxA) expression. We found that pathological complete response (pCR) following neoadjuvant treatment was associated with type I IFN. Stratification of patients according to the density of CD8(+) in the entire tumor tissue and MxA(+) cells in tumor stroma, where equal weight was assigned to both parameters, resulted in improved predictive quality compared to the ISB. This novel stratification approach using these two independent parameters in pre-operative biopsies could potentially aid in identifying patients with a good chance of achieving a pCR following neoadjuvant treatment.
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| 6. |
- Rezapour, Azar
(author)
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Immune Cell Profiling of Colorectal Cancers: Unravelling the Connection to Treatment Responses
- 2023
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Doctoral thesis (other academic/artistic)abstract
- Understanding the interplay between tumor cells and the immune system holds the key to the development of more effective treatments for cancer patients. Therefore, we have emphasized our efforts on deciphering the intricacies of these interactions to pave for more effective therapeutic strategies. We observed that different microenvironments in tumor and adjacent colon tissues influence immune cell behavior differently. Notably, cells known as antigen-presenting cells within the tumor show different activation and functional status compared to those in the colon. Despite this, they react similarly when exposed to signals from activated T cells, immune cells responsible for attacking cancer cells. The challenge remains in predicting how patients with rectal cancer will respond to treatments. Several studies have highlighted the potential of using tumor-infiltrating lymphocytes (TILs) as markers to predict treatment outcomes. An emerging predictive tool, the biopsy-adapted Immunoscore (ISB), assesses TILs to forecast tumor regression. To enhance its accuracy, we used a method incorporating multiplex immunofluorescence. By focusing on certain T cell subsets and the expression of Myxovirus resistance protein A (MxA), a component indicative of anti-tumoral inflammation, this refined method provided a better predictive framework. Specifically, the presence of MxA+ cells in the tumor stroma, combined with the density of CD8+ T cells, offered improved predictions regarding patient with a complete response to neoadjuvant treatment i.e., no detectable rectal cancer after the chemoradiotherapy. However, while TIL density and the presence of MxA-expressing cells provided insights regarding complete responders, our approach did not distinguish non-responders from treatment responders. A more nuanced approach revealed that non-responders had a higher proportion of a particular T-cell subset, CD8+CD103+39- present in the tumor but not in in paired rectal tissue prior to treatment. In addition, the non-responders exhibited a lower expression of PD-1 on TILs compared to responders, indicating an inadequate tumor microenvironment for immune cell activation. However, when T cells were stimulated in vitro, their responses were overall similar, regardless of how the tumor responded to the neoadjuvant treatment in the patient. In conclusion, while tools like the ISB offer predictive insights, refining these with multiparametric immune activity screens of the pre-operative biopsy can provide a more accurate picture of the patient’s ensuing responses to neoadjuvant treatment. This has significant implications for patient stratification and could possibly lead to the development of more personalized treatment regimens of rectal cancers.
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