PRMT inhibition induces a viral mimicry response in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype with the worst prognosis and few effective therapies. Here we identified MS023, an inhibitor of type I protein arginine methyltransferases (PRMTs), which has antitumor growth activity in TNBC. Pathway analysis of TNBC cell lines indicates that the activation of interferon responses before and after MS023 treatment is a functional biomarker and determinant of response, and these observations extend to a panel of human-derived organoids. Inhibition of type I PRMT triggers an interferon response through the antiviral defense pathway with the induction of double-stranded RNA, which is derived, at least in part, from inverted repeat Alu elements. Together, our results represent a shift in understanding the antitumor mechanism of type I PRMT inhibitors and provide a rationale and biomarker approach for the clinical development of type I PRMT inhibitors.

AP-1 activity is a major barrier of human somatic cell reprogramming.

Human induced pluripotent stem cells (iPSCs) technology has been widely applied to cell regeneration and disease modeling. However, most mechanism of somatic reprogramming is studied on mouse system, which is not always generic in human. Consequently, the generation of human iPSCs remains inefficient. Here, we map the chromatin accessibility dynamics during the induction of human iPSCs from urine cells. Comparing to the mouse system, we found that the closing of somatic loci is much slower in human. Moreover, a conserved AP-1 motif is highly enriched among the closed loci. The introduction of AP-1 repressor, JDP2, enhances human reprogramming and facilitates the reactivation of pluripotent genes. However, ESRRB, KDM2B and SALL4, several known pluripotent factors promoting mouse somatic reprogramming fail to enhance human iPSC generation. Mechanistically, we reveal that JDP2 promotes the closing of somatic loci enriching AP-1 motifs to enhance human reprogramming. Furthermore, JDP2 can rescue reprogramming deficiency without MYC or KLF4. These results indicate AP-1 activity is a major barrier to prevent chromatin remodeling during somatic cell reprogramming.

The roles of epigenetics in cancer progression and metastasis.

Cancer metastasis remains a major clinical challenge for cancer treatment. It is therefore crucial to understand how cancer cells establish and maintain their metastatic traits. However, metastasis-specific genetic mutations have not been identified in most exome or genome sequencing studies. Emerging evidence suggests that key steps of metastasis are controlled by reversible epigenetic mechanisms, which can be targeted to prevent and treat the metastatic disease. A variety of epigenetic mechanisms were identified to regulate metastasis, including the well-studied DNA methylation and histone modifications. In the past few years, large scale chromatin structure alterations including reprogramming of the enhancers and chromatin accessibility to the transcription factors were shown to be potential driving force of cancer metastasis. To dissect the molecular mechanisms and functional output of these epigenetic changes, it is critical to use advanced techniques and alternative animal models for interdisciplinary and translational research on this topic. Here we summarize our current understanding of epigenetic aberrations in cancer progression and metastasis, and their implications in developing new effective metastasis-specific therapies.

Assessment of Manufacturing Related Deficiencies for Modified Release Tablet in Abbreviated New Drug Applications.

Generic drugs play an important role in public health. However, the first review cycle approval rate for Abbreviated New Drug Applications (ANDAs) is generally low. To identify if the drug product (DP) manufacturing related deficiencies are the potential root causes of low first review cycle approval of the modified release (MR) tablet ANDAs, we collected and analyzed the review recommendations from each review discipline and the DP manufacturing (process and facility) related deficiencies for original MR tablet ANDAs submitted between FY17 and FY19. We identified 193 original MR tablet ANDAs. The analysis showed that 12% of the applications were approved in first review cycle, while 88% received complete responses (CR). Of the 169 CR applications, 91% were found inadequate for multiple review disciplines. A total of 1345 DP manufacturing process-related deficiencies were issued to 184 ANDAs during the first review cycle. We have identified common deficiencies across ANDAs based on DP manufacturing process categories. The top deficiencies cited reasons include facilities are out of compliance/not ready to commercialize/not ready for inspection; critical process parameter (CPP) ranges are not proposed/proposed CPP ranges are too wide and/or not supported by studied range and no in-process controls (IPCs) are proposed/proposed IPCs acceptance criteria (limits) are too wide and/or not supported by observed data etc. Avoiding the common DP manufacturing deficiencies may reduce the need for issuing DP manufacturing related deficiencies in information requests (IRs), discipline review letters (DRLs), and CRs for MR tablet ANDAs.

KDM5 histone demethylases repress immune response via suppression of STING.

Cyclic GMP-AMP (cGAMP) synthase (cGAS) stimulator of interferon genes (STING) senses pathogen-derived or abnormal self-DNA in the cytosol and triggers an innate immune defense against microbial infection and cancer. STING agonists induce both innate and adaptive immune responses and are a new class of cancer immunotherapy agents tested in multiple clinical trials. However, STING is commonly silenced in cancer cells via unclear mechanisms, limiting the application of these agonists. Here, we report that the expression of STING is epigenetically suppressed by the histone H3K4 lysine demethylases KDM5B and KDM5C and is activated by the opposing H3K4 methyltransferases. The induction of STING expression by KDM5 blockade triggered a robust interferon response in a cytosolic DNA-dependent manner in breast cancer cells. This response resulted in resistance to infection by DNA and RNA viruses. In human tumors, KDM5B expression is inversely associated with STING expression in multiple cancer types, with the level of intratumoral CD8+ T cells, and with patient survival in cancers with a high level of cytosolic DNA, such as human papilloma virus (HPV)-positive head and neck cancer. These results demonstrate a novel epigenetic regulatory pathway of immune response and suggest that KDM5 demethylases are potential targets for antipathogen treatment and anticancer immunotherapy.

Metformin Antagonizes Cancer Cell Proliferation by Suppressing Mitochondrial-Dependent Biosynthesis.

Metformin is a biguanide widely prescribed to treat Type II diabetes that has gained interest as an antineoplastic agent. Recent work suggests that metformin directly antagonizes cancer cell growth through its actions on complex I of the mitochondrial electron transport chain (ETC). However, the mechanisms by which metformin arrests cancer cell proliferation remain poorly defined. Here we demonstrate that the metabolic checkpoint kinases AMP-activated protein kinase (AMPK) and LKB1 are not required for the antiproliferative effects of metformin. Rather, metformin inhibits cancer cell proliferation by suppressing mitochondrial-dependent biosynthetic activity. We show that in vitro metformin decreases the flow of glucose- and glutamine-derived metabolic intermediates into the Tricarboxylic Acid (TCA) cycle, leading to reduced citrate production and de novo lipid biosynthesis. Tumor cells lacking functional mitochondria maintain lipid biosynthesis in the presence of metformin via glutamine-dependent reductive carboxylation, and display reduced sensitivity to metformin-induced proliferative arrest. Our data indicate that metformin inhibits cancer cell proliferation by suppressing the production of mitochondrial-dependent metabolic intermediates required for cell growth, and that metabolic adaptations that bypass mitochondrial-dependent biosynthesis may provide a mechanism of tumor cell resistance to biguanide activity.

Comments Regarding "Solubility determination and thermodynamic model analysis of L-α-glyceryl phosphorylcholine in different organic solvents of 278.15-323.15 K".

A polemic is given regarding several of the calculated curve-fit parameters that Zhou and coworkers reported in their published paper. The calculated curve-fit parameters for the Combined Nearly Ideal Binary Solvent/Redlich-Kister, Jouyban-Acree-van't Hoff, Sun and modified Apelblat models were found to give mole fraction solubilities that exceeded unity. Our analysis also found that the mean relative absolute percent deviations provided by the authors were significantly underestimated.

Validation of the Burn Survivor Fear-Avoidance Questionnaire and Its Association With Pain Intensity, Catastrophizing, and Disability.

According to the Fear-avoidance (FA) model, FA beliefs can lead to disability due to avoidance of activities expected to result in pain or further injury. Extensive research on the relationship of FA, pain, catastrophizing, and disability has been generated with patients suffering from chronic neck and back pain, but little research has been conducted with burn survivors. To address this need, the Burn Survivor FA Questionnaire (BSFAQ) was developed (Langlois J, Vincent-Toskin, S, Duchesne, P et al. Fear-avoidance beliefs and behaviors of burn survivors: A mixed-methods approach. Burns 2021;47:175-89.) but has not been validated. Thus, the primary objective of this study was to investigate the construct validity of the BSFAQ among burn survivors. The secondary objective was to examine the relationship between FA and 1) pain intensity and 2) catastrophizing at baseline, 3 months and 6 months postburn, and 3) disability among burn survivors at 6 months postburn. A prospective mixed-methods approach was used to examine the construct validity by comparing the quantitative scores of the BSFAQ to independently performed qualitative interviews of burn survivors (n = 31) that explored their lived experiences, to determine if the BSFAQ discriminated those who had, from those who did not have FA beliefs. Data for the secondary objective, scores of burn survivors (n = 51) pain intensity (numeric rating scale), catastrophizing (pain catastrophizing scale), and disability (Burn Specific Health Scale-brief), were collected through a retrospective chart review. For the primary objective, Wilcoxon rank sum test results showed a statistically significant difference (P = .015) between the BSFAQ scores of participants who were identified from the qualitative interviews as fear avoidant compared to those who were identified as non fear avoidant, with a receiver operating characteristic curve indicating that the BSFAQ correctly predicted FA 82.4% of the time. For the secondary objective, Spearman correlation test results showed a moderate correlation between FA and 1) pain at baseline (r = .466, P = .002), 2) catastrophizing thoughts over time (r = .557, P = .000; r = .470, P = .00; r = .559, P = .002 respectively at each time point), and 3) disability at 6 months postburn (r = -.643, P = .000). These results support that the BSFAQ is able to discriminate which burn survivors are experiencing FA beliefs. It is also consistent with the FA model since burn survivors who express FA are more likely to report higher levels of pain early during their recovery that correlates with persistently elevated catastrophizing thoughts and ultimately results in higher self-reported disability. The BSFAQ demonstrates construct validity and is able to correctly predict fear-avoidant burn survivors; however, additional research is required to further examine the BSFAQ's clinimetric properties.

Instrumented insoles for assessment of gait in patients with vestibular schwannoma.

Background: Imbalance and gait disturbances are common in patients with vestibular schwannoma (VS) and can result in significant morbidity. Current methods for quantitative gait analysis are cumbersome and difficult to implement. Here, we use custom-engineered instrumented insoles to evaluate the gait of patients diagnosed with VS. Methods: Twenty patients with VS were recruited from otology, neurosurgery, and radiation oncology clinics at a tertiary referral center. Functional gait assessment (FGA), 2-minute walk test (2MWT), and uneven surface walk test (USWT) were performed. Custom-engineered instrumented insoles, equipped with an 8-cell force sensitive resistor (FSR) and a 9-degree-of-freedom inertial measurement unit (IMU), were used to collect stride-by-stride spatiotemporal gait parameters, from which mean values and coefficients of variation (CV) were determined for each patient. Results: FGA scores were significantly correlated with gait metrics obtained from the 2MWT and USWT, including stride length, stride velocity, normalized stride length, normalized stride velocity, stride length CV, and stride velocity CV. Tumor diameter was negatively associated with stride time and swing time on the 2MWT; no such association existed between tumor diameter and FGA or DHI. Conclusions: Instrumented insoles may unveil associations between VS tumor size and gait dysfunction that cannot be captured by standardized clinical assessments and self-reported questionnaires.

Mode-locked femtosecond all-normal all-PM Yb-doped fiber laser using a nonlinear amplifying loop mirror.

We report on a new design for a passively mode locked fibre laser employing all normal dispersion polarisation maintaining fibres operating at 1 µm. The laser produces linearly polarized, linearly chirped pulses that can be recompressed down to 344 fs. Compared to previous laser designs the cavity is mode-locked using a nonlinear amplifying fibre loop mirror that provides an additional degree of freedom allowing easy control over the pulse parameters. This is a robust laser design with excellent reliability and lifetime.

Human WDR5 promotes breast cancer growth and metastasis via KMT2-independent translation regulation.

Metastatic breast cancer remains a major cause of cancer-related deaths in women, and there are few effective therapies against this advanced disease. Emerging evidence suggests that key steps of tumor progression and metastasis are controlled by reversible epigenetic mechanisms. Using an in vivo genetic screen, we identified WDR5 as an actionable epigenetic regulator that is required for metastatic progression in models of triple-negative breast cancer. We found that knockdown of WDR5 in breast cancer cells independently impaired their tumorigenic as well as metastatic capabilities. Mechanistically, WDR5 promotes cell growth by increasing ribosomal gene expression and translation efficiency in a KMT2-independent manner. Consistently, pharmacological inhibition or degradation of WDR5 impedes cellular translation rate and the clonogenic ability of breast cancer cells. Furthermore, a combination of WDR5 targeting with mTOR inhibitors leads to potent suppression of translation and proliferation of breast cancer cells. These results reveal novel therapeutic strategies to treat metastatic breast cancer.

CECR2 drives breast cancer metastasis by promoting NF-κB signaling and macrophage-mediated immune suppression.

Metastasis is the major cause of cancer-related deaths due to the lack of effective therapies. Emerging evidence suggests that certain epigenetic and transcriptional regulators drive cancer metastasis and could be targeted for metastasis treatment. To identify epigenetic regulators of breast cancer metastasis, we profiled the transcriptomes of matched pairs of primary breast tumors and metastases from human patients. We found that distant metastases are more immune inert with increased M2 macrophages compared to their matched primary tumors. The acetyl-lysine reader, cat eye syndrome chromosome region candidate 2 (CECR2), was the top up-regulated epigenetic regulator in metastases associated with an increased abundance of M2 macrophages and worse metastasis-free survival. CECR2 was required for breast cancer metastasis in multiple mouse models, with more profound effect in the immunocompetent setting. Mechanistically, the nuclear factor κB (NF-κB) family member v-rel avian reticuloendotheliosis viral oncogene homolog A (RELA) recruits CECR2 to increase chromatin accessibility and activate the expression of their target genes. These target genes include multiple metastasis-promoting genes, such as TNC, MMP2, and VEGFA, and cytokine genes CSF1 and CXCL1, which are critical for immunosuppression at metastatic sites. Consistent with these results, pharmacological inhibition of CECR2 bromodomain impeded NF-κB-mediated immune suppression by macrophages and inhibited breast cancer metastasis. These results reveal that targeting CECR2 may be a strategy to treat metastatic breast cancer.

Nutrigenetic reprogramming of oxidative stress.

Retinal disorders such as retinitis pigmentosa, age-related retinal degeneration, oxygen-induced retinopathy, and ischemia-reperfusion injury cause debilitating and irreversible vision loss. While the exact mechanisms underlying these conditions remain unclear, there has been a growing body of evidence demonstrating the pathological contributions of oxidative stress across different cell types within the eye. Nuclear factor erythroid-2-related factor (Nrf2), a transcriptional activator of antioxidative genes, and its regulator Kelch-like ECH-associated protein 1 (Keap1) have emerged as promising therapeutic targets. The purpose of this review is to understand the protective role of the Nrf2-Keap1 pathway in different retinal tissues and shed light on the complex mechanisms underlying these processes. In the photoreceptors, we highlight that Nrf2 preserves their survival and function by maintaining oxidation homeostasis. In the retinal pigment epithelium, Nrf2 similarly plays a critical role in oxidative stabilization but also maintains mitochondrial motility and autophagy-related lipid metabolic processes. In endothelial cells, Nrf2 seems to promote proper vascularization and revascularization through concurrent activation of antioxidative and angiogenic factors as well as inhibition of inflammatory cytokines. Finally, Nrf2 protects retinal ganglion cells against apoptotic cell death. Importantly, we show that Nrf2-mediated protection of the various retinal tissues corresponds to a preservation of functional vision. Altogether, this review underscores the potential of the Nrf2-Keap1 pathway as a powerful tool against retinal degeneration. Key insights into this elegant oxidative defense mechanism may ultimately pave the path toward a universal therapy for various inherited and environmental retinal disorders.

TFEB-mediated endolysosomal activity controls human hematopoietic stem cell fate.

It is critical to understand how human quiescent long-term hematopoietic stem cells (LT-HSCs) sense demand from daily and stress-mediated cues and then transition into bioenergetically active progeny to differentiate and meet these cellular needs. However, the demand-adapted regulatory circuits of these early steps of hematopoiesis are largely unknown. Here we show that lysosomes, sophisticated nutrient-sensing and signaling centers, are regulated dichotomously by transcription factor EB (TFEB) and MYC to balance catabolic and anabolic processes required for activating LT-HSCs and guiding their lineage fate. TFEB-mediated induction of the endolysosomal pathway causes membrane receptor degradation, limiting LT-HSC metabolic and mitogenic activation, promoting quiescence and self-renewal, and governing erythroid-myeloid commitment. In contrast, MYC engages biosynthetic processes while repressing lysosomal catabolism, driving LT-HSC activation. Our study identifies TFEB-mediated control of lysosomal activity as a central regulatory hub for proper and coordinated stem cell fate determination.

Photochemistry in photonic crystal fiber nanoreactors.

We report the use of a liquid-filled hollow-core photonic crystal fiber (PCF) as a highly controlled photochemical reactor. Hollow-core PCFs have several major advantages over conventional sample cells: the sample volume per optical path length is very small (2.8 nL cm(-1) in the fiber used), long optical path lengths are possible as a result of very low intrinsic waveguide loss, and furthermore the light travels in a diffractionless single mode with a constant transverse intensity profile. As a proof of principle, the (very low) quantum yield of the photochemical conversion of vitamin B(12), cyanocobalamin (CNCbl) to hydroxocobalamin ([H(2)OCbl](+)) in aqueous solution was measured for several pH values from 2.5 to 7.5. The dynamics of the actively induced reaction were monitored in real-time by broadband absorption spectroscopy. The PCF nanoreactor required ten thousand times less sample volume compared to conventional techniques. Furthermore, the enhanced sensitivity and optical pump intensity implied that even systems with very small quantum yields can be measured very quickly--in our experiments one thousand times faster than in a conventional cuvette.

Specific chromatin landscapes and transcription factors couple breast cancer subtype with metastatic relapse to lung or brain.

BACKGROUND: Few somatic mutations have been linked to breast cancer metastasis, whereas transcriptomic differences among primary tumors correlate with incidence of metastasis, especially to the lungs and brain. However, the epigenomic alterations and transcription factors (TFs) which underlie these alterations remain unclear. METHODS: To identify these, we performed RNA-seq, Chromatin Immunoprecipitation and sequencing (ChIP-seq) and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) of the MDA-MB-231 cell line and its brain (BrM2) and lung (LM2) metastatic sub-populations. We incorporated ATAC-seq data from TCGA to assess metastatic open chromatin signatures, and gene expression data from human metastatic datasets to nominate transcription factor biomarkers. RESULTS: Our integrated epigenomic analyses found that lung and brain metastatic cells exhibit both shared and distinctive signatures of active chromatin. Notably, metastatic sub-populations exhibit increased activation of both promoters and enhancers. We also integrated these data with chromosome conformation capture coupled with ChIP-seq (HiChIP) derived enhancer-promoter interactions to predict enhancer-controlled pathway alterations. We found that enhancer changes are associated with endothelial cell migration in LM2, and negative regulation of epithelial cell proliferation in BrM2. Promoter changes are associated with vasculature development in LM2 and homophilic cell adhesion in BrM2. Using ATAC-seq, we identified a metastasis open-chromatin signature that is elevated in basal-like and HER2-enriched breast cancer subtypes and associates with worse prognosis in human samples. We further uncovered TFs associated with the open chromatin landscapes of metastatic cells and whose expression correlates with risk for metastasis. While some of these TFs are associated with primary breast tumor subtypes, others more specifically correlate with lung or brain metastasis. CONCLUSIONS: We identify distinctive epigenomic properties of breast cancer cells that metastasize to the lung and brain. We also demonstrate that signatures of active chromatin sites are partially linked to human breast cancer subtypes with poor prognosis, and that specific TFs can independently distinguish lung and brain relapse.

Methionine Metabolism Shapes T Helper Cell Responses through Regulation of Epigenetic Reprogramming.

Epigenetic modifications on DNA and histones regulate gene expression by modulating chromatin accessibility to transcription machinery. Here we identify methionine as a key nutrient affecting epigenetic reprogramming in CD4+ T helper (Th) cells. Using metabolomics, we showed that methionine is rapidly taken up by activated T cells and serves as the major substrate for biosynthesis of the universal methyl donor S-adenosyl-L-methionine (SAM). Methionine was required to maintain intracellular SAM pools in T cells. Methionine restriction reduced histone H3K4 methylation (H3K4me3) at the promoter regions of key genes involved in Th17 cell proliferation and cytokine production. Applied to the mouse model of multiple sclerosis (experimental autoimmune encephalomyelitis), dietary methionine restriction reduced the expansion of pathogenic Th17 cells in vivo, leading to reduced T cell-mediated neuroinflammation and disease onset. Our data identify methionine as a key nutritional factor shaping Th cell proliferation and function in part through regulation of histone methylation.

Gefitinib inhibition of drug resistance to doxorubicin by inactivating ABCG2 in thyroid cancer cell lines.

OBJECTIVE: To investigate the regulation of the breast cancer resistance protein ABCG2/BRCP1 drug transporter by epidermal growth factor receptor (EGFR) kinase activity, and to determine whether gefitinib, an EGFR small molecule inhibitor, will modulate the effects of doxorubicin hydrochloride by inhibiting its extrusion from thyroid cancer cells. DESIGN: Extrusion assays using flow cytometry analysis were used to determine the ability of thyroid cancer cells to extrude the chemotherapy drug, doxorubicin, via the ABCG2 drug transporter in the presence or absence of gefitinib. Immunofluorescence was employed to determine the cellular expression of ABCG2. The ABCG2 expression in ARO and WRO cell lines was analyzed by Western blot analysis. Inactivation of EGFR kinase by gefitinib was analyzed by Western blot analysis and immunofluorescence. A terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay was performed to demonstrate ABCG2-mediated apoptosis in the presence of doxorubicin. Colony formation assays were performed to determine the effect of gefitinib on thyroid cancer cell survival in response to gefitinib, doxorubicin, or the combination of both drugs. RESULTS: Inhibition of EGFR kinase activity by gefitinib causes the translocation of the ABCG2 drug transporter away from the plasma membrane, resulting in a concomitant decrease in doxorubicin extrusion in thyroid cancer cell lines. Both ARO and WRO demonstrated differential ABCG2 expression, whereas both were sensitized to doxorubicin-induced apoptosis on ABCG2 knockdown with short interfering RNA. The addition of gefitinib increases doxorubicin-induced cell death in thyroid cancer cells as measured by colony formation assay. CONCLUSIONS: Epidermal growth factor receptor regulates the function of the drug transporter ABCG2/BCRP1 and correlates with ABCG2 protein expression levels. Inactivation of the EGFR kinase by gefitinib potentiates the cytotoxic effect of doxorubicin in thyroid cancer, most likely by decreasing the ability of the cell to extrude doxorubicin. The expression of ABCG2 may explain in part the ineffectiveness of doxorubicin as a single modality treatment for anaplastic thyroid cancer or for treatment of metastatic follicular thyroid cancer. Use of this combination treatment of gefitinib and doxorubicin may be a promising therapy for anaplastic thyroid and metastatic follicular thyroid cancer and needs to be investigated further.

Ubiquitination of tumor suppressor PML regulates prometastatic and immunosuppressive tumor microenvironment.

The tumor microenvironment plays an important role in tumor growth and metastasis. However, the mechanism by which tumor cells regulate the cell and non-cell constituents of surrounding stroma remains incompletely understood. Promyelocytic leukemia (PML) is a pleiotropic tumor suppressor, but its role in tumor microenvironment regulation is poorly characterized. PML is frequently downregulated in many cancer types, including lung cancer. Here, we identify a PML ubiquitination pathway that is mediated by WD repeat 4-containing cullin-RING ubiquitin ligase 4 (CRL4WDR4). Clinically, this PML degradation pathway is hyperactivated in lung cancer and correlates with poor prognosis. The WDR4/PML axis induces a set of cell-surface or secreted factors, including CD73, urokinase-type plasminogen activator receptor (uPAR), and serum amyloid A2 (SAA2), which elicit paracrine effects to stimulate migration, invasion, and metastasis in multiple lung cancer models. In xenograft and genetically engineered mouse models, the WDR4/PML axis elevates intratumoral Tregs and M2-like macrophages and reduces CD8+ T cells to promote lung tumor growth. These immunosuppressive effects were all reversed by CD73 blockade. Our study identifies WDR4 as an oncoprotein that negatively regulates PML via ubiquitination to promote lung cancer progression by fostering an immunosuppressive and prometastatic tumor microenvironment, suggesting the potential of immune-modulatory approaches for treating lung cancer with aberrant PML degradation.

EGFR regulates the side population in head and neck squamous cell carcinoma.

OBJECTIVE: To identify the presence of side population (SP) cells in established head and neck squamous carcinoma cell (HNSCC) lines and to determine the role of EGFR in the regulation of the side population of these cells. METHODS: SP cells were identified using flow cytometry analysis by the ability of these cells to extrude the Hoechst 33342 dye via the drug transporter BCRP1/ABCG2. Effect of EGFR on the side population was determined also by difference in Hoechst extrusion and by immunofluorescence. Immunohistochemical staining was performed to show the presence of the BCRP1/ABCG2 transporter and the phosphorylated form of EGFR in HNSCC tissue. RESULTS: SP cells are present in HNSCC cell lines. With the Hoechst 33342 extrusion assay, SP cells were found to comprise an average of 0.69% of the UMSCC10B cells and 0.91% of HN12 cells. Addition of the EGF ligand increased the SP population while inactivation of the EGFR kinase by Iressa significantly decreased SP. CONCLUSION: In established head and neck squamous cell carcinoma cell lines, SP cells were found using methods that determine expression and function of the drug transporter BCRP1/ABCG2. Activation of EGFR, a gene implicated in tumorigenesis in HNSCC leads to increased SP, and conversely, inhibition of EGFR leads to decrease in SP. This finding could help explain the role of EGFR in regulating cancer stem cells and thus tumorigenesis in HNSCC.