Differentially expressed messenger RNA/proteins can distinguish teratoma from necrosis in postchemotherapy retroperitoneal lymph node dissection tissue.

BACKGROUND: Before postchemotherapy retroperitoneal lymph node dissection (pcRPLND), in patients with metastasized germ cell tumors (GCTs), those harboring necrosis (NEC) cannot be distinguished from those who have teratoma (TER), resulting in relevant overtreatment, whereas microRNA-371a-3p may be predictive for viable GCT. The purpose of this study was to explore messenger RNA (mRNA) and proteins to distinguish TER from NEC in pcRPLND tissue. METHODS: The discovery cohort consisted in total of 48 patients, including 16 each with TER, viable GCT, and NEC. Representative areas were microdissected. A NanoString panel and proteomics were used to analyze 770 genes and >5000 proteins. The most significantly and differentially expressed combination of both parameters, mRNA and its associated protein, between TER and NEC was validated using immunohistochemistry (IHC) in an independent validation cohort comprising 66 patients who were not part of the discovery cohort. RESULTS: The authors observed that anterior gradient protein 2 homolog (AGR2) and keratin, type I cytoskeletal 19 (KRT19) were significantly differentially expressed in TER versus NEC in mRNA and protein analyses (proteomics). The technical validation using IHC was successful in the same patients. These proteins were further validated by IHC in the independent patient cohort and exhibited significantly higher levels in TER versus NEC (p < .0001; area under the curve, 1.0; sensitivity and specificity, 100% each). CONCLUSIONS: The current study demonstrated that KRT19 and AGR2 mRNA and protein are overexpressed in TER versus NEC in pcRPLND tissue and might serve as a future diagnostic target to detect TER, for instance, by functional imaging, to avoid overtreatment. PLAIN LANGUAGE SUMMARY: The proteins and the corresponding genes called AGR2 and KRT19 can differentiate between teratoma and necrosis in remaining tumor masses after chemotherapy in patients who have metastasized testicular cancer. This may be a way to improve presurgical diagnostics and to reduce the current overtreatment of patients with necrosis only, who could be treated sufficiently by surveillance.

Transcriptome analysis reveals upregulation of immune response pathways at the invasive tumour front of metastatic seminoma germ cell tumours.

BACKGROUND: Testicular germ cell tumours (TGCTs) have a high metastasis rate. However, the mechanisms related to their invasion, progression and metastasis are unclear. Therefore, we investigated gene expression changes that might be linked to metastasis in seminomatous testicular germ cell tumour (STGCT) patients. METHODS: Defined areas [invasive tumour front (TF) and tumour centre (TC)] of non-metastatic (with surveillance and recurrence-free follow-up >2 years) and metastatic STGCTs were collected separately using laser capture microdissection. The expression of 760 genes related to tumour progression and metastasis was analysed using nCounter technology and validated with quantitative real-time PCR and enzyme-linked immunosorbent assay. RESULTS: Distinct gene expression patterns were observed in metastatic and non-metastatic seminomas with respect to both the TF and TC. Comprehensive pathway analysis showed enrichment of genes related to tumour functions such as inflammation, angiogenesis and metabolism at the TF compared to the TC. Remarkably, prominent inflammatory and cancer-related pathways, such as interleukin-6 (IL-6) signalling, integrin signalling and nuclear factor-κB signalling, were significantly upregulated in the TF of metastatic vs non-metastatic tumours. CONCLUSIONS: IL-6 signalling was the most significantly upregulated pathway in metastatic vs non-metastatic tumours and therefore could constitute a therapeutic target for future personalised therapy. In addition, this is the first study showing intra- and inter-tumour heterogeneity in STGCT.

Prediction of Radioresistant Prostate Cancer Based on Differentially Expressed Proteins.

INTRODUCTION: Although relapses after radiotherapy are common in prostate cancer (PCA) patients, those with a high risk for radioresistance cannot be identified prior to treatment yet. Therefore, this proof-of-concept study was performed to compare protein expression profiles of patients with radio-recurrent PCA to patients treated with primary radical prostatectomy separated by Gleason risk groups. We hypothesized that radio-recurrent PCA have a similar protein expression as high-risk Gleason PCA. METHODS: Patient cohorts consisted of (i) 31 patients treated with salvage prostatectomy for locally recurrent PCA after primary radiotherapy and (ii) 94 patients treated with primary prostatectomy split into a Gleason high-risk (≥4 + 3; n = 42 [44.7%]) versus a low-risk group (≤3 + 4; n = 52 [55.3%]). Immunohistochemistry was performed using 15 antibodies with known association to radioresistance in PCA in vitro. ELISA was used for validation of selected markers in serum. RESULTS: Androgen receptor (AR) was overexpressed in most radio-recurrent PCA (89.7%) and in most primary high-risk Gleason PCA (87.8%; p = 0.851), while only 67.3% of the low-risk group showed an expression (p = 0.017). Considering the highest Gleason pattern in primary PCA, aldo-keto reductase family 1 member C3 (AKR1C3) was most similarly expressed by patients with radio-recurrent PCA and patients with Gleason patterns 4 and 5 (p = 0.827 and p = 0.893) compared to Gleason pattern 3 (p = 0.20). These findings were supported by ELISA. CONCLUSION: This is the first study to evaluate protein markers in order to predict radioresistance in PCA. Our results point to AR and AKR1C3 as the most promising markers that might help stratify patients for radiotherapy.

68 Ga-PSMA-PET/CT helps to select patients for salvage radical prostatectomy with local recurrence after primary radiotherapy for prostate cancer.

OBJECTIVE: To investigate the diagnostic performance of gallium-68 prostate-specific membrane antigen positron emission tomography/computed tomography (68 Ga-PSMA PET/CT) in patients with recurrent prostate cancer with regard to the presence of lymph node metastases (LNM) and local recurrences after primary radiotherapy. PATIENTS AND METHODS: We retrospectively reviewed 142 patients following salvage radical prostatectomy (sRP), 50 of which had a 68 Ga-PSMA PET/CT performed as a preoperative staging module. Predictive clinical parameters were analysed in a multivariate Cox regression analysis. Sensitivity, specificity, positive (PPV) and negative predictive values (NPV) and the accuracy of 68 Ga-PSMA PET/CT were analysed with regard to LNM and local recurrence. RESULTS: In all, 613 lymph nodes were resected in 40 patients and 23 lymph nodes had metastatic deposits in 14 patients. In all patients local recurrence could have been found with 68 Ga-PSMA PET/CT. Sensitivity, specificity, PPV and NPV and accuracy on a per lymph node basis were 34.78% (16.38-57.2%), 100% (99.38-100%), 100%, 97.52% (96.69-98.15%) and 97.55% (96.00-98.62%). For detecting local recurrence, the sensitivity and PPV were both 100% with an accuracy of 100% (92.89-100%). CONCLUSION: 68 Ga-PSMA PET/CT should be the standard imaging in biochemical recurrent prostate cancer. With this imaging module one detects first local recurrence and can detect locoregional and distant metastases more precisely than standard CT and bone scan.

Differential expression and in vivo secretion of the antimicrobial peptides psoriasin (S100A7), RNase 7, human beta-defensin-2 and -3 in healthy human skin.

Antimicrobial peptides (AMP) are key players in the skin's defense system. Previous observations suggest a site- and age-dependent expression of individual AMP. We investigated the expression and secretion patterns of four important AMP in a representative collective of healthy human skin samples. Levels of psoriasin, RNase 7 and hBD-3 expression - assessed by immunohistochemistry - varied between different body localisations. Older individuals expressed hBD-2 more frequently. No gender-related expression was observed. The in vivo secretion of psoriasin, measured in skin washing fluids using ELISA, was related to body localisation and age, whereas RNase 7 secretion showed no significant differences regarding these variables. HBD-2 and -3 secretion could not be detected. Our findings suggest the usage of control samples matching localisation and approximate age (in the case of hBD-2) for comparative immunohistochemical analysis. To avoid bias through great interindividual differences, sufficient large collectives should be used for in vivo secretion analyses.

MET-FISH Evaluation Algorithm: Proposal of a Simplified Method.

MET amplifications (METamp) occur in 5% of NSCLC and represent in most case mechanisms of resistance to ALK and/or EGFR-targeted therapies. METamp detection can be performed using different techniques, although Fluorescence In-Situ Hybridization (FISH) remains the gold-standard, especially in the context of subclonality. To date current evaluation algorithms of MET amplifications are time consuming. Aim of the study was to identify a faster, equally reliable diagnostic algorithm for the detection of METamp, which is currently classified in negativity and low/intermediate/high-level amplification. N=497 NSCLC cases with available MET-FISH data had been selected. The results based on the first evaluated 20 cells had been re-calculated and compared with the definitive results based on 60 cells. For n=464 (93.4%) identical results had been obtained when counting 20 cells instead of 60 cells. Thirty-three cases (5.6%) showed a discrepancy, leading to an incorrect upgrade to a higher diagnostic category (n=25) and to an incorrect downgrade (n=8). We propose a simplified, yet equally reliable MET FISH-algorithm: after accurate screening of the whole tumor slide, twenty tumor cells have to be evaluated and results calculated: If the result is negative, or if all criteria of high-level METamp are fulfilled, the case can be signed out as such. All other cases should be considered as equivocal and additional 40 cells have to be counted. Given that, reliable results can be obtained by counting 20 cells only and an "equivocal" category for cases that need further investigation have been clearly defined.

Psoriasin: key molecule of the cutaneous barrier?

Psoriasin (S100 A7) was discovered two decades ago as a protein abundantly expressed in psoriatic keratinocytes. Even though much scientific research has been carried out on the characterization of psoriasin, only recent studies point to an important role of psoriasin as an antimicrobial and immunomodulatory protein in skin and other epithelia. In this review, we provide an overview of the major findings in psoriasin research and discuss novel studies highlighting the role of psoriasin as an important effector molecule of the cutaneous barrier.

The combination of microRNA-371a-3p and 375-5p can distinguish viable germ cell tumor and teratoma from necrosis in postchemotherapy retroperitoneal lymph node dissection specimens.

BACKGROUND: To identify a combination of microRNAs (miRNA) to differentiate between viable tumor (V) or teratoma (T) and necrosis/fibrosis (N) in pcRPLND specimens of metastatic germ cell tumor (GCT) patients with residual masses ≥1 cm after chemotherapy. Biomarker guided therapy could reduce overtreatment with pcRPLND in patients with only N. METHODS: We selected 48 metastatic GCT patients who had undergone pcRPLND. V, pure T and N was shown in the resected tissue of 16 patients, respectively. Of these areas total RNA was isolated and miRNA expression was analyzed for miR-371a-3p, 375-3p, and 375-5p using qPCR. ROC analysis was performed for each miRNA and for all combinations in order to determine the discriminatory capacity of V and T vs. N. RESULTS: On comparing V vs. N miR-371a-3p achieved the highest fold change (FC) of 31.1 (P=0.023) while for T vs. N miR-375-5p performed best (FC 64.2; P<0.001). Likewise, the most accurate AUC for V was 0.75 using miR-371a-3p, for T 0.80 using miR-375-5p. Combining the best performing miRNAs for V and T resulted in an AUC of 0.94 with a sensitivity of 93.75, specificity of 93.75, PPV of 96.8 and NPV of 83.3. CONCLUSIONS: By combining miR-371a-3p and miR-375-5p in pcRPLND tissue samples V and T could be distinguished from necrosis/fibrosis with great accuracy. This combination of miRNAs might serve as new biomarker in the future, in order to spare miRNA-negative patients from pcRPLND. However, further studies analyzing patient's serum are needed to confirm the clinical impact of these biomarkers.

Differential expression of antimicrobial peptides in margins of chronic wounds.

Skin wounds usually heal without major infections, although the loss of the mechanical epithelial barrier exposes the tissue to various bacteria. One reason may be the expression of antimicrobial peptides (AMP) of which some [human beta-defensins (hBD) and LL-37] were recently shown to support additionally certain steps of wound healing. There are no studies which have compared expression patterns of different classes of AMP in chronic wounds. The aim of our study was therefore to analyse the expression profile of hBD-2, hBD-3, LL-37, psoriasin and RNase 7 by immunohistochemistry from defined wound margins of chronic venous ulcers. We detected a strong induction of psoriasin and hBD-2 in chronic wounds in comparison with healthy skin. Except for stratum corneum, no expression of RNase 7 and LL-37 was detected in the epidermis while expression of hBD-3 was heterogeneous. Bacterial swabs identified Staphylococcus aureus and additional bacterial populations, but no association between colonization and AMP expression was found. The differential expression of AMP is noteworthy considering the high bacterial load of chronic ulcers. Clinically, supplementation of AMP with the capability to enhance wound healing besides restricting bacterial overgrowth could present a physiological support for treatment of disturbed wound healing.

Facial nerve paralysis caused by a T-cell lymphoma.

Facial nerve paralysis due to the infiltration by a lymphoma is rare and the prognosis remains poor. If perineural spread and meningeosis are suspected, quick interdisciplinary diagnostic work-up is recommended. It should include magnetic resonance imaging, biopsy of the lesion, bone marrow biopsy, and lumbar puncture. Therapy should be initiated immediately.

Surgical resection of symptomatic brain metastases improves the clinical status and facilitates further treatment.

BACKGROUND: Brain metastases (BM) frequently cause focal neurological deficits leading to a reduced Karnofsky performance score (KPS). Since KPS is routinely used to guide the choice of adjuvant therapy, we hypothesized that improving KPS by surgical resection may improve the chance for adjuvant treatment and ultimately result in better survival. We therefore analyzed the course of a large cohort undergoing resection of symptomatic brain metastases in the context of further treatment and clinical outcome. PATIENTS AND METHODS: In a bi-centric retrospective analysis we retrieved baseline, clinical, and treatment-related parameters of patients operated on BM between 2010 and 2019. Survival was calculated using Kaplan-Meier estimates; prognostic factors for survival were analyzed by Log-rank test and Cox proportional hazards. RESULTS: We included 750 patients with a median age of 61 (19-87) years. The functional status was significantly improved by surgical resection, with a median preoperative (KPS) of 80 (10-100) increasing to 90 (0-100) after surgery (P < .0001). Moreover, surgery improved the RTOG recursive partitioning analysis (RPA) class from III to I/II in 82 patients. Postoperative local radiotherapy and systemic treatment were associated with significantly longer survival (P < .0001 for each). Systemic treatment was provided significantly more frequently in patients with a fair postoperative clinical status (KPS ≥ 70; P < .0001). The postoperative clinical status, postoperative radiotherapy, systemic treatment, controlled systemic disease and < 4 BM were independent predictors for survival. CONCLUSION: The resection of symptomatic BM may restore clinical status, so enhancing the likelihood of receiving adjuvant treatment, and therefore leading to improved overall survival.

Timing of Development of Symptomatic Brain Metastases from Non-Small Cell Lung Cancer: Impact on Symptoms, Treatment, and Survival in the Era of Molecular Treatments.

OBJECTIVE: We attempted to analyze whether early presentation with brain metastases (BM) represents a poor prognostic factor in patients with non-small cell lung cancer (NSCLC), which should guide the treatment team towards less intensified therapy. PATIENTS AND METHODS: In a retrospective bi-centric analysis, we identified patients receiving surgical treatment for NSCLC BM. We collected demographic-, tumor-, and treatment-related parameters and analyzed their influence on further survival. RESULTS: We included 377 patients. Development of BM was precocious in 99 (26.3%), synchronous in 152 (40.3%), and metachronous in 126 (33.4%) patients. The groups were comparable in terms of age (p = 0.76) and number of metastases (p = 0.11), and histology (p = 0.1); however, mutational status significantly differed (p = 0.002). The precocious group showed the worst clinical status as assessed by Karnofsky performance score (KPS) upon presentation (p < 0.0001). Resection followed by postoperative radiotherapy was the predominant treatment modality for precocious BM, while in syn- and metachronous BM surgical and radio-surgical treatment was balanced. Overall survival (OS) did not differ between the groups (p = 0.76). A good postoperative clinical status (KPS ≥ 70) and the application of any kind of adjuvant systemic therapy were independent predictive factors for OS. CONCLUSION: Early BM presentation was not associated with worse OS in NSCLC BM patients.

ATM activity in T cells is critical for immune surveillance of lymphoma in vivo.

The proximal DNA damage response kinase ATM is frequently inactivated in human malignancies. Germline mutations in the ATM gene cause Ataxia-telangiectasia (A-T), characterized by cerebellar ataxia and cancer predisposition. Whether ATM deficiency impacts on tumor initiation or also on the maintenance of the malignant state is unclear. Here, we show that Atm reactivation in initially Atm-deficient B- and T cell lymphomas induces tumor regression. We further find a reduced T cell abundance in B cell lymphomas from Atm-defective mice and A-T patients. Using T cell-specific Atm-knockout models, as well as allogeneic transplantation experiments, we pinpoint impaired immune surveillance as a contributor to cancer predisposition and development. Moreover, we demonstrate that Atm-deficient T cells display impaired proliferation capacity upon stimulation, due to replication stress. Altogether, our data indicate that T cell-specific restoration of ATM activity or allogeneic hematopoietic stem cell transplantation may prevent lymphomagenesis in A-T patients.

AATF suppresses apoptosis, promotes proliferation and is critical for Kras-driven lung cancer.

A fundamental principle in malignant tranformation is the ability of cancer cells to escape the naturally occurring cell-intrinsic responses to DNA damage. Tumors progress despite the accumulation of DNA lesions. However, the underlying mechanisms of this tolerance to genotoxic stress are still poorly characterized. Here, we show that replication stress occurs in Kras-driven murine lung adenocarcinomas, as well as in proliferating murine embryonic and adult tissues. We identify the transcriptional regulator AATF/CHE-1 as a key molecule to sustain proliferative tissues and tumor progression in parts by inhibiting p53-driven apoptosis in vivo. In an autochthonous Kras-driven lung adenocarcinoma model, deletion of Aatf delayed lung cancer formation predominantly in a p53-dependent manner. Moreover, targeting Aatf in existing tumors through a dual recombinase strategy caused a halt in tumor progression. Taken together, these data suggest that AATF may serve as a drug target to treat KRAS-driven malignancies.

The Cdkn1aSUPER Mouse as a Tool to Study p53-Mediated Tumor Suppression.

Cdkn1a, which encodes p21, functions as a major route for p53-mediated cell-cycle arrest. However, the consequence of Cdkn1a gene dosage on tumor suppression has not been systematically investigated. Here, we employed BAC transgenesis to generate a Cdkn1aSUPER mouse, which harbors an additional Cdkn1a allele within its natural genomic context. We show that these mice display enhanced cell-cycle arrest and reduced apoptosis in response to genotoxic stress. Furthermore, using a chemically induced skin cancer model and an autochthonous Kras-driven lung adenocarcinoma model, we show that Cdkn1aSUPER mice display a cancer protection phenotype that is indistinguishable from that observed in Tp53SUPER animals. Moreover, we demonstrate that Tp53 and Cdkn1a cooperate in mediating cancer resistance, using a chemically induced fibrosarcoma model. Overall, our Cdkn1aSUPER allele enabled us to assess the contribution of Cdkn1a to Tp53-mediated tumor suppression.

KRAS mutation in papillary fibroelastoma: a true cardiac neoplasm?

Primary cardiac tumours are rare and mostly benign lesions. Recent publications report that cardiac papillary fibroelastomas are the most common benign primary heart tumour, outnumbering myxomas. However, there is no consensus about their aetiology. We investigated the molecular profile of these tumours using next generation sequencing in a cohort of 16 cases. Eleven of 14 (79%) analysable tumours showed mutations of the KRAS oncogene. Our results provide unambiguous evidence that a significant proportion of these lesions are genuine neoplastic tumours caused by an oncogenic driver mutation.

Comparison of Blood Collection Tubes from Three Different Manufacturers for the Collection of Cell-Free DNA for Liquid Biopsy Mutation Testing.

The improvement in sensitive techniques has allowed the detection of tumor-specific aberrations in circulating tumor (ct) DNA. Amplification-refractory mutation system PCR has been used for the analysis of ctDNA to detect resistance-causing mutations in the epidermal growth factor receptor gene (eg, EGFR T790M) in lung cancer patients. So far, Streck tubes have been widely used as standard blood collection tubes. Here, we compared blood collection tubes manufactured by Streck with tubes from Roche and Qiagen regarding their utility in stabilizing ctDNA in plasma samples. Venous blood from healthy donors was collected in tubes from Streck, Roche, and Qiagen. Samples were spiked with artificially fragmented EGFR T790M-mutated DNA and stored for different periods of time or spiked with different DNA amounts before plasma preparation. Extracted ctDNA was analyzed by amplification-refractory mutation system PCR. Mutant DNA, spiked into blood samples from healthy donors at quantities of 1 or 3 ng, was still reliably detectable in all samples after 7 days. EGFR T790M could be detected when spiking was performed with an amount of artificial ctDNA of 0.5 ng when tubes from Roche and Qiagen were used. Blood collection tubes from Roche and Qiagen are highly suitable for ctDNA stabilization and subsequent liquid biopsy testing. Even low ctDNA concentrations allow the detection of somatic mutations.

Targeting a non-oncogene addiction to the ATR/CHK1 axis for the treatment of small cell lung cancer.

Small cell lung cancer (SCLC) is a difficult to treat subtype of lung cancer. One of the hallmarks of SCLC is its almost uniform chemotherapy sensitivity. However, chemotherapy response is typically transient and patients frequently succumb to SCLC within a year following diagnosis. We performed a transcriptome analysis of the major human lung cancer entities. We show a significant overexpression of genes involved in the DNA damage response, specifically in SCLC. Particularly CHEK1, which encodes for the cell cycle checkpoint kinase CHK1, is significantly overexpressed in SCLC, compared to lung adenocarcinoma. In line with uncontrolled cell cycle progression in SCLC, we find that CDC25A, B and C mRNAs are expressed at significantly higher levels in SCLC, compared to lung adenocarcinoma. We next profiled the efficacy of compounds targeting CHK1 and ATR. Both, ATR- and CHK1 inhibitors induce genotoxic damage and apoptosis in human and murine SCLC cell lines, but not in lung adenocarcinoma cells. We further demonstrate that murine SCLC tumors were highly sensitive to ATR- and CHK1 inhibitors, while Kras G12D -driven murine lung adenocarcinomas were resistant against these compounds and displayed continued growth under therapy. Altogether, our data indicate that SCLC displays an actionable dependence on ATR/CHK1-mediated cell cycle checkpoints.

Ercc1 Deficiency Promotes Tumorigenesis and Increases Cisplatin Sensitivity in a Tp53 Context-Specific Manner.

KRAS-mutant lung adenocarcinoma is among the most common cancer entities and, in advanced stages, typically displays poor prognosis due to acquired resistance against chemotherapy, which is still largely based on cisplatin-containing combination regimens. Mechanisms of cisplatin resistance have been extensively investigated, and ERCC1 has emerged as a key player due to its central role in the repair of cisplatin-induced DNA lesions. However, clinical data have not unequivocally confirmed ERCC1 status as a predictor of the response to cisplatin treatment. Therefore, we employed an autochthonous mouse model of Kras-driven lung adenocarcinoma resembling human lung adenocarcinoma to investigate the role of Ercc1 in the response to cisplatin treatment. Our data show that Ercc1 deficiency in Tp53-deficient murine lung adenocarcinoma induces a more aggressive tumor phenotype that displays enhanced sensitivity to cisplatin treatment. Furthermore, tumors that relapsed after cisplatin treatment in our model develop a robust etoposide sensitivity that is independent of the Ercc1 status and depends solely on previous cisplatin exposure. Our results provide a solid rationale for further investigation of the possibility of preselection of lung adenocarcinoma patients according to the functional ERCC1- and mutational TP53 status, where functionally ERCC1-incompetent patients might benefit from sequential cisplatin and etoposide chemotherapy. IMPLICATIONS: This study provides a solid rationale for the stratification of lung adenocarcinoma patients according to the functional ERCC1- and mutational TP53 status, where functionally ERCC1-incompetent patients could benefit from sequential cisplatin and etoposide chemotherapy. Mol Cancer Res; 14(11); 1110-23. ©2016 AACR.

Genotype/Phenotype correlations in patients with hereditary breast cancer.

Of all breast cancer cases, 5-10% can be attributed to germline mutations, and the high-susceptibility genes BRCA1 and BRCA2 account for about 25-28% of these cases. For the remainder, several genes of moderate and low penetrance have been discovered. Histopathologic characteristics have been studied in small cohorts, but for most of the known non-BRCA1/2-associated hereditary breast cancers, the histologic and immunohistochemical phenotypes are not yet identified. Particularly BRCA1 tumors are associated with a distinct morphology and immunohistochemical characteristics that differ from sporadic breast cancer of age-matched controls. The recognition of features characteristic of these mutations can be helpful to identify patients likely to carry a germline mutation and to assess which gene should be screened for first, in families with a high occurrence of breast and ovarian cancer.