Tumor Repopulation Research Articles

A Caspase 3‐Hijacking Nanosystem Enhances Cancer Radiotherapy by Suppressing Tumor Repopulation

AbstractRadiotherapy is used in the treatment of ≈50% of patients with cancer. However, tumor repopulation is a major cause of treatment failure after radiotherapy. It is observed that apoptotic tumor following ionizing radiation (IR) accelerated the growth of surviving tumor cells. Here a Gasdermin E and Tannic acid‐based nanoassembly (GT) loaded with manganese tetroxide (Mn3O4) (termed as Mn3O4@GT) is developed to suppress tumor repopulation and improve the treatment outcome of radiotherapy. Mn3O4@GT enables an increase in the reactive oxygen species accumulation in tumor cells, enhancing radiotherapy‐mediated tumor killing. What's more, it can hijack activated caspase 3 to induce tumor pyroptosis, reversing apoptosis‐mediated tumor repopulation. In vivo results shows that Mn3O4@GT significantly reduced the IR induced tumor repopulation by 2.7 fold, resulting in 92% complete regression of tumors. In addition, Mn3O4@GT can sensitize tumors to anti‐PD‐L1 therapy by inducing immunogenic pyroptosis with 85% regression of distant tumors. The caspase 3‐hijacking nanosystem holds a great potential for improving the clinical benefits of radiotherapy.

Radiation therapy in the hypofractionation mode on the Halcyon elite linear accelerator: the first experience of St. Petersburg city clinical oncology dispensary

Stereotactic radiation therapy in the hypofractionation mode is a method in which high doses of ionizing radiation (more than 3 Gy per fraction) are delivered in a small number of fractions (from 2 to 5). Treatment and the process of preparation for it are carried out using special fixing devices, dosimetric planning techniques of high conformity, with control of the treatment position using x-ray imaging. which makes it possible to ensure the required irradiation accuracy. The advantages are a shorter course of treatment compared to standard fractionation, a high biologically effective dose, a reduced effect of tumor repopulation, greater efficiency under hypoxic conditions, and satisfactory results when irradiating radioresistant tumors. The article presents clinical cases of the use of hypofractionation modes on the Halcyon elite linear accelerator.

Dose-rate effects and tumor control probability in 177Lu-based targeted radionuclide therapy: a theoretical analysis

Objective.177Lu-based targeted radionuclide therapy (TRT) has become an important cancer treatment option in recent years, in particular in the treatment of advanced prostate cancer and metastasized neuroendocrine tumors. Although it is known from conventional radiotherapy that the temporal dynamics of the dose-rate can be of relevance for tumor cell survival, the analysis of TRT efficacy usually considers only the absorbed dose. Thus, the aim of this theoretical analysis is to shed light on the possible effects of the pattern of dose-rate in TRT on tumor control probability (TCP).Approach.For this purpose, TCP is studied numerically in a typical four-cycle treatment regime based on the mechanistic lethal-potentially lethal model and the Zaider-Minerbo model for TCP including repopulation of tumor cells.Main results.It is shown that the dose-rate pattern in TRT can have a substantial effect on TCP even though the absorbed dose in the tumor lesion is unchanged. These dose-rate effects are particularly evident when repair of potentially lethal lesions is slow.Significance.The results indicate that in some situations in the analysis of the efficacy of TRT it is necessary to consider the full dose-rate pattern instead of the absorbed dose alone. This can be highly relevant for optimization and further development of TRTs. In particular, it could be of relevancy in studying the efficacy of newly emerging treatment concepts that combine the use of TRT and drugs that inhibit DNA damage repair.

Chb-M', an Inhibitor of the RUNX Family Binding to DNA, Induces Apoptosis in p53-Mutated Non-Small Cell Lung Cancer and Inhibits Tumor Growth and Repopulation In Vivo.

Runt-related transcription factor (RUNX) proteins are considered to play various roles in cancer. Here, we evaluated the anticancer activity of Chb-M', a compound that specifically and covalently binds to the consensus sequence for RUNX family proteins, in p53-mutated non-small cell lung cancer cells. Chb-M' killed the cancer cells by inducing apoptosis. The compound showed an anticancer effect comparable to that of the clinically used drugs alectinib and ceritinib in vivo. Notably, Chb-M' extended the cancer-free survival of mice after ending treatment more effectively than did the other two drugs. The results presented here suggest that Chb-M' is an attractive candidate as an anticancer drug applicable to the treatment of non-small cell lung cancer and various other types of cancers.

Abstract PO1-24-06: Prolactin Hormone-Induced Mammary Differentiation and Anti-tumorigenic Role in Breast Cancer

Abstract The premise of differentiation therapy in cancer is a strategy that aims at engaging-forward differentiation and cellular reprograming restricting the proliferative, tumor repopulation, stemness, EMT and metastatic capacities of tumor cells leading to the cessation of the aggressive tumor phenotype and offering the cancer patients improved survival for decades. Therefore, deciphering molecular mechanisms deriving differentiation in normal mammary and breast cancer cells may allow the discovery of innovative differentiation-based biomarkers and therapeutics in breast cancer. Lactation is an intricate process that results in the ability of the breast cells to produce a complex nutritious biological fluid to nurse the new-born. While the beneficial effects of breastfeeding to the infant is irrefutable, its effects on maternal health are less investigated especially in relation to breast cancer that still affects 1 in 8 women in high income countries and 1 in 20 globally. Importantly, epidemiological studies have linked breastfeeding to reduced risk of breast cancer by promoting terminal differentiation of the breast epithelial cells. While breastfeeding process is regulated by multiple hormones, growth factors, and transcriptional regulators, the lactation hormone prolactin (PRL) is known to be directly implicated in the hormonal control of breastfeeding. Extensive research including our own work has shown that PRL/PRL receptor (PRLR) pathway derives mammary gland development and importantly mammary epithelial cell terminal differentiation allowing successful lactation. In contrast the role of PRL in breast cancer is still to be fully defined. Importantly, we have accumulated compelling evidence using in-silico publicly available patient data sets and molecular data implicating this pathway as a clinically relevant pro-differentiation pathway driving differentiation and cellular reprograming suppressing stemness, EMT, metastasis and tumorigenesis in breast cancer. To better characterize PRL signaling mediating its pro-differentiation effects, we used large-scale unbiased proteomics analysis of PRLR/interacting proteins in breast cancer cells. Our analyses revealed several novel PRLR-interactors. We have further confirmed these interactions through co-immunoprecipitations/western blotting analyses in breast cancer cells representative the different breast cancer subtypes. We also examined their functional contributions to mammary differentiation and tumorigenesis using in vitro and in vivo assays. Moreover, we defined their potential clinical relevance using bioinformatics in silico databases of breast cancer. Together our study delineates novel PRL/PRLR-downstream signaling mediators important for mammary differentiation and tumorigenesis and may thus be useful as biomarkers and/or therapeutic targets in breast cancer. Citation Format: Dana Hamam, Suhad Ali. Prolactin Hormone-Induced Mammary Differentiation and Anti-tumorigenic Role in Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-24-06.

Pancreatic cancer cells hijack tumor suppressive microRNA-26a to promote radioresistance and potentiate tumor repopulation

Pancreatic cancer is one of the most lethal cancers with significant radioresistance and tumor repopulation after radiotherapy. As a type of short non-coding RNA that regulate various biological and pathological processes, miRNAs might play vital role in radioresistance. We found by miRNA sequencing that microRNA-26a (miR-26a) was upregulated in pancreatic cancer cells after radiation, and returned to normal state after a certain time. miR-26a was defined as a tumor suppressive miRNA by conventional tumor biology experiments. However, transient upregulation of miR-26a after radiation significantly promoted radioresistance, while stable overexpression inhibited radioresistance, highlighting the importance of molecular dynamic changes after treatment. Mechanically, transient upregulation of miR-26a promoted cell cycle arrest and DNA damage repair to promote radioresistance. Further experiments confirmed HMGA2 as the direct functional target, which is an oncogene but enhances radiosensitivity. Moreover, PTGS2 was also the target of miR-26a, which might potentiate tumor repopulation via delaying the synthesis of PGE2. Overall, this study revealed that transient upregulation of miR-26a after radiation promoted radioresistance and potentiated tumor repopulation, highlighting the importance of dynamic changes of molecules upon radiotherapy.

Surviving the Storm: The Role of Poly- and Depolyploidization in Tissues and Tumors.

Polyploidization and depolyploidization are critical processes in the normal development and tissue homeostasis of diploid organisms. Recent investigations have revealed that polyaneuploid cancer cells (PACCs) exploit this ploidy variation as a survival strategy against anticancer treatment and for the repopulation of tumors. Unscheduled polyploidization and chromosomal instability in PACCs enhance malignancy and treatment resistance. However, their inability to undergo mitosis causes catastrophic cellular death in most PACCs. Adaptive ploid reversal mechanisms, such as multipolar mitosis, centrosome clustering, meiosis-like division, and amitosis, counteract this lethal outcome and drive cancer relapse. The purpose of this work is to focus on PACCs induced by cytotoxic therapy, highlighting the latest discoveries in ploidy dynamics in physiological and pathological contexts. Specifically, by emphasizing the role of "poly-depolyploidization" in tumor progression, the aim is to identify novel therapeutic targets or paradigms for combating diseases associated with aberrant ploidies.

Abstract 4264: The TWEAK-Fn14-NF-kB signaling axis enhances expression of integrin AVB3 in a post-chemotherapy tumor microenvironment

Abstract High grade serous ovarian cancer (HGSOC) patients initially respond to chemotherapy but a majority relapse with chemoresistant disease. Research supports the notion that tumor-initiating cells (TICs), a subpopulation of drug resistant stem-like tumor cells, are responsible for facilitating relapse and therapies targeting these fecund cells may prolong remission. The tumor microenvironment (TME) plays a crucial role in therapy resistance, and tumor relapse. Cytotoxic chemotherapies have been shown to modify the stroma of the TME which may favor the survival of TICs displaying specific adhesion proteins. We and others have shown that NF-kB transcription factors RelA and RelB respond to signals from the ovarian TME to promote proliferation, chemoresistance, and survival. Our results further show that RelA and RelB regulate expression of specific integrins. Recently we found that TNF-like inducer of apoptosis (TWEAK), a cytokine involved in tissue repair, and its only known receptor Fn14, are enriched following chemotherapy, leading to enhanced chemoresistance and survival of TICs. TWEAK also induces expression of specific integrins in ovarian TICs. Therefore, we hypothesize that NF-kB mediated integrin expression enables adhesion and survival of TICs to a chemotherapy modified extracellular matrix (ECM) to facilitate tumor repopulation and relapse. Using magnetic activated cell sorting (MACS) for CD117+ TICs, we found that a significant majority are enriched for integrin AVB3. Supporting a role for TWEAK-Fn14-NF-kB in mediating integrin expression we further show that integrin AVB3, but not AVB5 or A5B1, was significantly increased in CD117+ TICs in the presence of TWEAK and was dependent on Fn14 as demonstrated through knockdown experiments. RNA-sequencing analysis of tumor cells grown in spheroid conditions with RelA or RelB shRNA knockdown relative to control, show ITGAV is dependent on RelA while ITGB3 expression is dependent on RelB. Taken together, our findings suggest CD117+ TICs have enhanced expression of AVB3 and is further induced by TWEAK mediated NF-kB activity for adaptation to a stressful post-chemotherapy environment. Ongoing in vitro and in vivo studies are investigating the therapeutic benefit of targeting AVB3 or TWEAK in CD117+ TICs for prevention of adhesion and tumor repopulation following chemotherapy. These studies will uncover interactions between TICs and ECM through their unique expression of integrins and clarifies the role of NF-kB transcription factors RelA and RelB in regulating TIC survival in the post-chemotherapy TME. Citation Format: Omar Lujano Olazaba, Mikella Robinson, Cecilia Gallo, Mena Shammas, Katelyn Shelby, Luisjesus Cruz, Samuel F. Gilbert, Carrie D. House. The TWEAK-Fn14-NF-kB signaling axis enhances expression of integrin AVB3 in a post-chemotherapy tumor microenvironment [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 4264.

Understanding the impact of radiotherapy fractionation on overall survival in a large head and neck squamous cell carcinoma dataset: a comprehensive approach combining mechanistic and machine learning models.

Treating head and neck squamous cell carcinomas (HNSCC), especially human papillomavirus negative (HPV-) and locally advanced cases, remains difficult. Our previous analyses of radiotherapy-only HNSCC clinical trials data using mechanistically-motivated models of tumor repopulation and killing by radiotherapy predicted that hyperfractionation with twice-daily fractions, or hypofractionation involving increased doses/fraction and reduced treatment durations, both improve tumor control and reduce late normal tissue toxicity, compared with standard protocols using 35×2 Gy. Here we further investigated the validity of these conclusions by analyzing a large modern dataset on 3,346 HNSCC radiotherapy patients from the University Health Network in Toronto, Canada, where 42.5% of patients were also treated with chemotherapy. We used a two-step approach that combines mechanistic modeling concepts with state-of-the-art machine learning, beginning with Random Survival Forests (RSF) for an exploratory analysis and followed by Causal Survival Forests (CSF) for a focused causal analysis. The mechanistic concept of biologically effective dose (BED) was implemented for the standard dose-independent (DI) tumor repopulation model, our alternative dose-dependent (DD) repopulation model, and a simple model with no repopulation (BEDsimp). These BED variants were included in the RSF model, along with age, stage, HPV status and other relevant variables, to predict patient overall survival (OS) and cause-specific mortality (deaths from the index cancer, other cancers or other causes). Model interpretation using Shapley Additive Explanations (SHAP) values and correlation matrices showed that high values of BEDDD or BEDDI, but not BEDsimp, were associated with decreased patient mortality. Targeted causal inference analyses were then performed using CSF to estimate the causal effect of each BED variant on OS. They revealed that high BEDDD (>61.8 Gy) or BEDDI (>57.6 Gy), but not BEDsimp, increased patient restricted mean survival time (RMST) by 0.5-1.0 years and increased survival probability (SP) by 5-15% several years after treatment. In addition to population-level averages, CSF generated individual-level causal effect estimates for each patient, facilitating personalized medicine. These findings are generally consistent with those of our previous mechanistic modeling, implying the potential benefits of altered radiotherapy fractionation schemes (e.g. 25×2.4 Gy, 20×2.75 Gy, 18×3.0 Gy) which increase BEDDD and BEDDI and counteract tumor repopulation more effectively than standard fractionation. Such regimens may represent potentially useful hypofractionated options for treating HNSCC.

Opportunities in Cancer Therapies: Deciphering the Role of Cancer Stem Cells in Tumour Repopulation.

Tumour repopulation during treatment is a well acknowledged yet still challenging aspect of cancer management. The latest research results show clear evidence towards the existence of cancer stem cells (CSCs) that are responsible for tumour repopulation, dissemination, and distant metastases in most solid cancers. Cancer stem cell quiescence and the loss of asymmetrical division are two powerful mechanisms behind repopulation. Another important aspect in the context of cancer stem cells is cell plasticity, which was shown to be triggered during fractionated radiotherapy, leading to cell dedifferentiation and thus reactivation of stem-like properties. Repopulation during treatment is not limited to radiotherapy, as there is clinical proof for repopulation mechanisms to be activated through other conventional treatment techniques, such as chemotherapy. The dynamic nature of stem-like cancer cells often elicits resistance to treatment by escaping drug-induced cell death. The aims of this scoping review are (1) to describe the main mechanisms used by cancer stem cells to initiate tumour repopulation during therapy; (2) to present clinical evidence for tumour repopulation during radio- and chemotherapy; (3) to illustrate current trends in the identification of CSCs using specific imaging techniques; and (4) to highlight novel technologies that show potential in the eradication of CSCs.

ALDH2 dysfunction and alcohol cooperate in cancer stem cell enrichment.

The alcohol metabolite acetaldehyde is a potent human carcinogen linked to esophageal squamous cell carcinoma (ESCC) initiation and development. Aldehyde dehydrogenase 2 (ALDH2) is the primary enzyme that detoxifies acetaldehyde in the mitochondria. Acetaldehyde accumulation causes genotoxic stress in cells expressing the dysfunctional ALDH2E487K dominant negative mutant protein linked to ALDH2*2, the single nucleotide polymorphism highly prevalent among East Asians. Heterozygous ALDH2*2 increases the risk for the development of ESCC and other alcohol-related cancers. Despite its prevalence and link to malignant transformation, how ALDH2 dysfunction influences ESCC pathobiology is incompletely understood. Herein, we characterize how ESCC and preneoplastic cells respond to alcohol exposure using cell lines, three-dimensional organoids and xenograft models. We find that alcohol exposure and ALDH2*2 cooperate to increase putative ESCC cancer stem cells with high CD44 expression (CD44H cells) linked to tumor initiation, repopulation and therapy resistance. Concurrently, ALHD2*2 augmented alcohol-induced reactive oxygen species and DNA damage to promote apoptosis in the non-CD44H cell population. Pharmacological activation of ALDH2 by Alda-1 inhibits this phenotype, suggesting that acetaldehyde is the primary driver of these changes. Additionally, we find that Aldh2 dysfunction affects the response to cisplatin, a chemotherapeutic commonly used for the treatment of ESCC. Aldh2 dysfunction facilitated enrichment of CD44H cells following cisplatin-induced oxidative stress and cell death in murine organoids, highlighting a potential mechanism driving cisplatin resistance. Together, these data provide evidence that ALDH2 dysfunction accelerates ESCC pathogenesis through enrichment of CD44H cells in response to genotoxic stressors such as environmental carcinogens and chemotherapeutic agents.

SURG-31. DEFINING THE NEW POSSIBILITIES FOR CESIUM BRACHYTHERAPY IN PATIENTS WITH PERIVENTRICULAR GBM

Abstract INTRODUCTION Re-irradiation of patients with external beam radiotherapy (EBRT) for recurrent glioblastoma (GBM) continues to present a therapeutic dilemma due to a high risk of radiation induced necrosis (RN). In contrast, brachytherapy represents a safer, more conformal radiation option aimed at optimizing local control while minimizing the risk of brain tissue damage. Immediate intra-operative implantation with brachytherapy prevents tumor repopulation and offers patients two procedures in one setting. However, due to the risk of seed migration, surgical resection and brachytherapy in periventricular regions was deemed unachievable. In heavily pretreated periventricular GBM, where further EBRT is complicated by RN, a barrier between the ventricular lining and tumor cavity is needed to prevent seed migration. We present two patients where such a barrier allowed successful implantation of brachytherapy to improve disease control. METHODS Two patients, previously treated with surgery and EBRT, presented with recurrent GBM abutting the ventricles. Following GTR of the tumors, a barrier was created by patching the ventricle with Gelfoam to prevent seed migration. Intraoperative brachytherapy implantation of suture stranded Cs-131 seeds (Isoray) for patient 1 and collagen embedded Cs-131 (GammaTile) for patient 2 was performed. A total of 20 Cs-131 seeds on strings threaded through Surgicel, and 6 GammaTiles (24 Cs-131 seeds), were used implanted in patients 1 and 2, respectively. Surgicel covered the implanted seeds and the resection cavity filled with Adherus. RESULTS Post-operative CT scan demonstrated all 20 Cs-131 seeds and 24 Cs-131 seeds in the surgical cavities of patients 1 and 2, respectively, and had serial post operative imaging with MRI scans showing Cs-131 seeds layering surgical cavity, as late as 6 months in total follow up. To date, both patients have had durable local control and no seed migration.

Autocrine regulation of tumor cell repopulation by Hsp70-HMGB1 alarmin complex

BackgroundCancer recurrence is regulated by a variety of factors, among which is the material of dying tumor cells; it is suggested that remaining after anti-cancer therapy tumor cells receive a signal from proteins called damage-associated molecular patterns (DAMPs), one of which is heat shock protein 70 (Hsp70).MethodsTwo models of tumor repopulation were employed, based on minimal population of cancer cells and application of conditioned medium (CM). To deplete the CMs of Hsp70 affinity chromatography on ATP-agarose and immunoprecipitation were used. Cell proliferation and the dynamics of cell growth were measured using MTT assay and xCELLigence technology; cell growth markers were estimated using qPCR and with the aid of ELISA for prostaglandin E detection. Immunoprecipitation followed by mass-spectrometry was employed to identify Hsp70-binding proteins and protein-protein interaction assays were developed to reveal the above protein complexes.ResultsIt was found that CM of dying tumor cells contains tumor regrowth-initiating factors and the removal of one of them, Hsp70, caused a reduction in the relapse-activating capacity. The pull out of Hsp70 alone using ATP-agarose had no effect on repopulation, while the immunodepletion of Hsp70 dramatically reduced its repopulation activity. Using proteomic and immunochemical approaches, we showed that Hsp70 in conditioned medium binds and binds another abundant alarmin, the High Mobility Group B1 (HMGB1) protein; the complex is formed in tumor cells treated with anti-cancer drugs, persists in the cytosol and is further released from dying tumor cells. Recurrence-activating power of Hsp70-HMGB1 complex was proved by the enhanced expression of proliferation markers, Ki67, Aurka and MCM-10 as well as by increase of prostaglandin E production and autophagy activation. Accordingly, dissociating the complex with Hsp70 chaperone inhibitors significantly inhibited the pro-growth effects of the above complex, in both in vitro and in vivo tumor relapse models.ConclusionsThese data led us to suggest that the abundance of the Hsp70-HMGB1 complex in the extracellular matrix may serve as a novel marker of relapse state in cancer patients, while specific targeting of the complex may be promising in the treatment of cancers with a high risk of recurrence.

Prostate Cancer Stem Cells: Biology and Treatment Implications.

Stem cells differentiate into mature organ/tissue-specific cells at a steady pace under normal conditions, but their growth can be accelerated during the process of tissue healing or in the context of certain diseases. It is postulated that the proliferation and growth of carcinomas are sustained by the presence of a vital cellular compartment resembling stem cells residing in normal tissues: 'stem-like cancer cells' or cancer stem cells (CSCs). Mutations in prostate stem cells can lead to the formation of prostate cancer. Prostate CSCs (PCSCs) have been identified and partially characterized. These express surface markers include CD44, CD133, integrin α2β1, and pluripotency factors like OCT4, NANOG, and SOX2. Several signaling pathways are also over-activated, including Notch, PTEN/Akt/PI3K, RAS-RAF-MEK-ERK and HH. Moreover, PCSCs appear to induce resistance to radiotherapy and chemotherapy, while their presence has been linked to aggressive cancer behavior and higher relapse rates. The development of treatment policies to target PCSCs in tumors is appealing as radiotherapy and chemotherapy, through cancer cell killing, trigger tumor repopulation via activated stem cells. Thus, blocking this reactive stem cell mobilization may facilitate a positive outcome through cytotoxic treatment.

Predicting tumor repopulation through the gene panel derived from radiation resistant colorectal cancer cells

BackgroundTumor cells with the capability of radiation resistance can escape the fate of cell death after radiotherapy, serving as the main cause of treatment failure. Repopulation of tumors after radiotherapy is dominated by this group of residual cells, which greatly reduce the sensitivity of recurrent tumors to the therapy, resulting in poor clinical outcomes. Therefore, revealing the mechanism of radiation resistant cells participating in tumor repopulation is of vital importance for cancer patients to obtain a better prognosis.MethodsCo-expressed genes were searched by using genetic data of radiation resistant cells (from GEO database) and TCGA colorectal cancer. Univariate and multivariate Cox regression analysis were performed to define the most significant co-expressed genes for establishing prognostic indicator. Logistic analysis, WGCNA analysis, and other types of tumors were included to verify the predictive ability of the indicator. RT-qPCR was carried out to test expression level of key genes in colorectal cancer cell lines. Colongenic assay was utilized to test the radio-sensitivity and repopulation ability of key gene knockdown cells.ResultsPrognostic indicator based on TCGA colorectal cancer patients containing four key radiation resistance genes (LGR5, KCNN4, TNS4, CENPH) was established. The indicator was shown to be significantly correlated with the prognosis of colorectal cancer patients undergoing radiotherapy, and also had an acceptable predictive effect in the other five types of cancer. RT-qPCR showed that expression level of key genes was basically consistent with the radiation resistance level of colorectal cancer cells. The clonogenic ability of all key gene knockdown cells decreased after radiation treatment compared with the control groups.ConclusionsOur data suggest that LGR5, KCNN4, TNS4 and CENPH are correlated with radiation sensitivity of colorectal cancer cells, and the indicator composed by them can reflect the prognosis of colorectal cancer patients undergoing radiation therapy. Our data provide an evidence of radiation resistant tumor cells involved in tumor repopulation, and give patients undergoing radiotherapy an approving prognostic indicator with regard to tumor progression.

Hyperthermia Reduces Irradiation-Induced Tumor Repopulation in an In Vivo Pancreatic Carcinoma Model.

Pancreatic cancer has a poor prognosis due to its aggressive nature and ability to metastasize at anearly stage. Currently, its management is still a challenge because this neoplasm is resistant to conventional treatment approaches, among which is chemo-radiotherapy (CRT), due to the abundant stromal compartment involved in the mechanism of hypoxia. Hyperthermia, among other effects, counteracts hypoxia by promoting blood perfusion and thereby can enhance the therapeutic effect of radiotherapy (RT). Therefore, the establishment of integrated treatments would be a promising strategy for the management of pancreatic carcinoma. Here, the effects of joint radiotherapy/hyperthermia (RT/HT) on optimized chick embryo chorioallantoic membrane (CAM) pancreatic tumor models are investigated. This model enables a thorough assessment of the tumor-arresting effect of the combined approach as well as the quantitative evaluation of hypoxia and cell cycle-associated mechanisms by both gene expression analysis and histology. The analysis of the lower CAM allows to investigate the variation of the metastatic behaviors of the cancer cells associated with the treatments. Overall, this study provides a potentially effective combined strategy for the non-invasive management of pancreatic carcinoma.

Cancer Cells Enter an Adaptive Persistence to Survive Radiotherapy and Repopulate Tumor.

Repopulation of residual tumor cells impedes curative radiotherapy, yet the mechanism is not fully understood. It is recently appreciated that cancer cells adopt a transient persistence to survive the stress of chemo- or targeted therapy and facilitate eventual relapse. Here, it is shown that cancer cells likewise enter a "radiation-tolerant persister" (RTP) state to evade radiation pressure in vitro and in vivo. RTP cells are characterized by enlarged cell size with complex karyotype, activated type I interferon pathway and two gene patterns represented by CST3 and SNCG. RTP cells have the potential to regenerate progenies via viral budding-like division, and type I interferon-mediated antiviral signaling impaired progeny production. Depleting CST3 or SNCG does not attenuate the formation of RTP cells, but can suppress RTP cells budding with impaired tumor repopulation. Interestingly, progeny cells produced by RTP cells actively lose their aberrant chromosomal fragments and gradually recover back to a chromosomal constitution similar to their unirradiated parental cells. Collectively, this study reveals a novel mechanism of tumor repopulation, i.e., cancer cell populations employ a reversible radiation-persistence by poly- and de-polyploidization to survive radiotherapy and repopulate the tumor, providing a new therapeutic concept to improve outcome of patients receiving radiotherapy.

Subclonal evolution and expansion of spatially distinct THY1-positive cells is associated with recurrence in glioblastoma

PurposeGlioblastoma(GBM) is a lethal disease characterized by inevitable recurrence. Here we investigate the molecular pathways mediating resistance, with the goal of identifying novel therapeutic opportunities. Experimental designWe developed a longitudinal in vivo recurrence model utilizing patient-derived explants to produce paired specimens(pre- and post-recurrence) following temozolomide(TMZ) and radiation(IR). These specimens were evaluated for treatment response and to identify gene expression pathways driving treatment resistance. Findings were clinically validated using spatial transcriptomics of human GBMs. ResultsThese studies reveal in replicate cohorts, a gene expression profile characterized by upregulation of mesenchymal and stem-like genes at recurrence. Analyses of clinical databases revealed significant association of this transcriptional profile with worse overall survival and upregulation at recurrence. Notably, gene expression analyses identified upregulation of TGFβ signaling, and more than one-hundred-fold increase in THY1 levels at recurrence. Furthermore, THY1-positive cells represented <10% of cells in treatment-naïve tumors, compared to 75-96% in recurrent tumors. We then isolated THY1-positive cells from treatment-naïve patient samples and determined that they were inherently resistant to chemoradiation in orthotopic models. Additionally, using image-guided biopsies from treatment-naïve human GBM, we conducted spatial transcriptomic analyses. This revealed rare THY1+ regions characterized by mesenchymal/stem-like gene expression, analogous to our recurrent mouse model, which co-localized with macrophages within the perivascular niche. We then inhibited TGFBRI activity in vivo which decreased mesenchymal/stem-like protein levels, including THY1, and restored sensitivity to TMZ/IR in recurrent tumors. ConclusionsThese findings reveal that GBM recurrence may result from tumor repopulation by pre-existing, therapy-resistant, THY1-positive, mesenchymal cells within the perivascular niche.

XBP1‐elicited environment by chemotherapy potentiates repopulation of tongue cancer cells by enhancing miR‐22/lncRNA/KAT6B‐dependent NF‐κB signalling

Tumour repopulation initiated by residual tumour cells in response to cytotoxic therapy has been described clinically and biologically, but the mechanisms are unclear. Here, we aimed to investigate the mechanisms for the tumour-promoting effect in dying cells and for tumour repopulation in surviving tongue cancer cells. Tumour repopulation in vitro and in vivo was represented by luciferase activities. The differentially expressed cytokines in the conditioned medium (CM) were identified using a cytokine array. Gain or loss of function was investigated using inhibitors, neutralising antibodies, shRNAs and ectopic overexpression strategies. We found that dying tumour cells undergoing cytotoxic therapy increase the growth of living tongue cancer cells in vitro and in vivo. Dying tumour cells create amphiregulin (AREG)- and basicfibroblast growth factor (bFGF)-based extracellular environments via cytotoxic treatment-induced endoplasmic reticulum stress. This environment stimulates growth by activating lysine acetyltransferase 6B (KAT6B)-dependent nuclear factor-kappa B (NF-κB) signalling in living tumour cells. As direct targets of NF-κB, miR-22 targets KAT6B to repress its expression, but long noncoding RNAs (lncRNAs) (XLOC_003973 and XLOC_010383) counter the effect of miR-22 to enhance KAT6B expression. Moreover, we detected increased AREG and bFGF protein levels in the blood of tongue cancer patients with X-box binding protein-1 (XBP1) activation in tumours under cytotoxic therapy and found that XBP1 activation is associated with poor prognosis of patients. We also detected activation of miR-22/lncRNA/KAT6B/NF-κB signalling in recurrent cancers compared to paired primary tongue cancers. We identified the molecular mechanisms of cell death-induced tumour repopulation in tongue cancer. Such insights provide new avenues to identify predictive biomarkers and effective strategies to address cancer progression.

A comparative and prospective study of two different radiation fractionation schedules with concurrent chemotherapy in locally advanced head-and-neck squamous cell carcinoma

Background: Accelerated fractionation radiotherapy has radiobiological advantage of preventing accelerated tumor repopulation and logistic advantage of treating more patients than conventionally fractionated radiotherapy because of its relatively shorter treatment duration. Aims and Objectives: In this study, we compared accelerated fractionation with conventionally fractionated radiotherapy in terms of tumor response and acute toxicities for the treatment of locally advanced head-and-neck carcinomas. Materials and Methods: Patients with Stage III and IVA carcinoma of head-and-neck region were randomized into two groups. The study group patients received accelerated radiotherapy to a total dose of 66Gy in 33 fractions, 2Gy/fraction, 6 fractions/week over a time period of 5.5 weeks. Control group received conventionally fractionated radiotherapy to same total dose and fraction size but 5 fractions/week, over a time period of 6.5 weeks. Both groups received concurrent weekly Cisplatin. All patients were followed up weekly for treatment related acute toxicity during the treatment and then at every month for 6 months after completion of treatment. Results: About 26.6% patients of study arm achieved complete response in comparison to 25.6% of control arm, but the difference was not statistically significant (P=0.957). Although statistically not significant, higher grade of skin toxicity (60%vs.35%, P=0.179) and xerostomia (46% vs. 29%, P=0.155) was also numerically higher in accelerated fractionation. Conclusion: For locally advanced head-and-neck carcinoma, accelerated fractionation radiotherapy with concurrent chemotherapy can be considered as an acceptable and effective alternative of conventionally fractionated concurrent chemoradiotherapy in terms of treatment response and acute toxicity profile.