Aging limits stemness and tumorigenesis in the lung by reprogramming iron homeostasis.

Aging is associated with a decline in the number and fitness of adult stem cells 1-4 . Aging-associated loss of stemness is posited to suppress tumorigenesis 5,6 , but this hypothesis has not been tested in vivo . Here, using physiologically aged autochthonous genetically engineered mouse models and primary cells 7,8 , we demonstrate aging suppresses lung cancer initiation and progression by degrading stemness of the alveolar cell of origin. This phenotype is underpinned by aging-associated induction of the transcription factor NUPR1 and its downstream target lipocalin-2 in the cell of origin in mice and humans, leading to a functional iron insufficiency in the aged cells. Genetic inactivation of the NUPR1-lipocalin-2 axis or iron supplementation rescue stemness and promote tumorigenic potential of aged alveolar cells. Conversely, targeting the NUPR1- lipocalin-2 axis is detrimental to young alveolar cells via induction of ferroptosis. We find that aging-associated DNA hypomethylation at specific enhancer sites associates with elevated NUPR1 expression, which is recapitulated in young alveolar cells by inhibition of DNA methylation. We uncover that aging drives a functional iron insufficiency, which leads to loss of stemness and tumorigenesis, but promotes resistance to ferroptosis. These findings have significant implications for the therapeutic modulation of cellular iron homeostasis in regenerative medicine and in cancer prevention. Furthermore, our findings are consistent with a model whereby most human cancers initiate in young individuals, revealing a critical window for such cancer prevention efforts.

Tumoral Intraductal Neoplasms of the Bile Ducts Comprise Morphologically and Genetically Distinct Entities.

CONTEXT.­: Tumoral (grossly visible) intraductal neoplasms of the bile ducts are still being characterized. OBJECTIVE.­: To investigate their morphologic, immunohistochemical, and molecular features. DESIGN.­: Forty-one cases were classified as gastric-, intestinal-, pancreatobiliary-type intraductal papillary neoplasm (IPN), intraductal oncocytic papillary neoplasm (IOPN), or intraductal tubulopapillary neoplasm (ITPN) on the basis of histology. All neoplasms were subjected to targeted next-generation sequencing. RESULTS.­: The mean age at diagnosis was 69 years (42-81 years); male to female ratio was 1.3. Most neoplasms (n = 23, 56%) were extrahepatic/large (mean size, 4.6 cm). The majority (n = 32, 78%) contained high-grade dysplasia, and 68% (n = 28) revealed invasion. All gastric-type IPNs (n = 9) and most ITPNs/IOPNs showed consistent colabeling for CK7/MUC6, which was less common among others (P = .004). Intestinal-type IPNs (n = 5) showed higher rates of CK20 expression than others (P < .001). Overall, the most commonly mutated genes included TP53 and APC, while copy number variants affected ELF3 and CDKN2A/B. All gastric-type IPNs contained an alteration affecting the Wnt signaling pathway; 7 of 9 (78%) showed aberrations in the MAPK pathway. Mutations in APC and KRAS were common in gastric-type IPNs as compared with others (P = .01 for both). SMAD4 was more frequently mutated in intestinal-type IPNs (P = .02). Pancreatobiliary-type IPNs (n = 14) exhibited frequent alterations in tumor suppressor genes including TP53, CDKN2A/B, and ARID2 (P = .04, P = .01 and P = .002, respectively). Of 6 IOPNs analyzed, 3 (50%) revealed ATP1B1-PRKACB fusion. ITPNs (n = 6) showed relatively few recurrent genetic aberrations. Follow-up information was available for 38 patients (median, 58.5 months). The ratio of disease-related deaths was higher for the cases with invasion (56% versus 10%). CONCLUSIONS.­: Tumoral intraductal neoplasms of the bile ducts, similar to their counterparts in the pancreas, are morphologically and genetically heterogeneous.

Integrated digital pathology at scale: A solution for clinical diagnostics and cancer research at a large academic medical center.

OBJECTIVE: Broad adoption of digital pathology (DP) is still lacking, and examples for DP connecting diagnostic, research, and educational use cases are missing. We blueprint a holistic DP solution at a large academic medical center ubiquitously integrated into clinical workflows; researchapplications including molecular, genetic, and tissue databases; and educational processes. MATERIALS AND METHODS: We built a vendor-agnostic, integrated viewer for reviewing, annotating, sharing, and quality assurance of digital slides in a clinical or research context. It is the first homegrown viewer cleared by New York State provisional approval in 2020 for primary diagnosis and remote sign-out during the COVID-19 (coronavirus disease 2019) pandemic. We further introduce an interconnected Honest Broker for BioInformatics Technology (HoBBIT) to systematically compile and share large-scale DP research datasets including anonymized images, redacted pathology reports, and clinical data of patients with consent. RESULTS: The solution has been operationally used over 3 years by 926 pathologists and researchers evaluating 288 903 digital slides. A total of 51% of these were reviewed within 1 month after scanning. Seamless integration of the viewer into 4 hospital systems clearly increases the adoption of DP. HoBBIT directly impacts the translation of knowledge in pathology into effective new health measures, including artificial intelligence-driven detection models for prostate cancer, basal cell carcinoma, and breast cancer metastases, developed and validated on thousands of cases. CONCLUSIONS: We highlight major challenges and lessons learned when going digital to provide orientation for other pathologists. Building interconnected solutions will not only increase adoption of DP, but also facilitate next-generation computational pathology at scale for enhanced cancer research.

Tipifarnib Inhibits HRAS-Driven Dedifferentiated Thyroid Cancers.

Of the three RAS oncoproteins, only HRAS is delocalized and inactivated by farnesyltransferase inhibitors (FTI), an approach yet to be exploited clinically. In this study, we treat mice bearing Hras-driven poorly differentiated and anaplastic thyroid cancers (Tpo-Cre/HrasG12V/p53flox/flox ) with the FTI tipifarnib. Treatment caused sustained tumor regression and increased survival; however, early and late resistance was observed. Adaptive reactivation of RAS-MAPK signaling was abrogated in vitro by selective RTK (i.e., EGFR, FGFR) inhibitors, but responses were ineffective in vivo, whereas combination of tipifarnib with the MEK inhibitor AZD6244 improved outcomes. A subset of tumor-bearing mice treated with tipifarnib developed acquired resistance. Whole-exome sequencing of resistant tumors identified a Nf1 nonsense mutation and an activating mutation in Gnas at high allelic frequency, supporting the on-target effects of the drug. Cell lines modified with these genetic lesions recapitulated tipifarnib resistance in vivo This study demonstrates the feasibility of targeting Ras membrane association in cancers in vivo and predicts combination therapies that confer additional benefit.Significance: Tipifarnib effectively inhibits oncogenic HRAS-driven tumorigenesis and abrogating adaptive signaling improves responses. NF1 and GNAS mutations drive acquired resistance to Hras inhibition, supporting the on-target effects of the drug. Cancer Res; 78(16); 4642-57. ©2018 AACR.

PIK3CA mutations rarely demonstrate genotypic intratumoral heterogeneity and are selected for in breast cancer progression.

PIK3CA gene mutations are the most common activating mutations in human breast cancer. Its association with hormone receptor-positive breast cancer makes it a prime target for clinical therapeutic advances to maintain anti-estrogen responsiveness. In anticipation of this therapeutic approach, we have evaluated intratumoral heterogeneity in primary breast cancers with regard to PIK3CA mutation status. In addition, we have assessed for the presence of the mutation in paired pre-invasive breast cancer and metastases. To assess for intratumoral heterogeneity, separate tumor blocks from primary breast cancers (n = 63) were genotyped for PIK3CA mutations. Available paired tissue samples from breast tumors known to harbor mutations underwent massARRAY genotyping (n = 70) to identify PIK3CA and AKT1(E17K) mutations. Cores were macro-dissected from matched tissue, including normal breast, benign lymph nodes (LN), ductal carcinoma in situ, regional LN metastases, and distant metastases. Matched samples underwent genetic fingerprinting by multiple SNP genotyping to confirm genetic identity. Intratumoral heterogeneity is minimal with a concordance rate of 95.2% between two different blocks from primary breast cancers. Complete concordance of PIK3CA mutations is noted between primary breast cancer and DCIS. PIK3CA mutations in primary breast cancer are detected in matched regional LNs (91.7%) and distant metastases (100%). Mutation detection by massARRAY genotyping is sensitive but may be affected by sample quality. Intratumoral heterogeneity as measured by PIK3CA genotype is rare; PIK3CA mutations occur early and are selected for in breast cancer progression. HapMap analysis is an essential control for paired sample analysis. This data is clinically important, particularly, for the design of therapies targeting the PI3K/AKT pathway, as it offers confidence that the detection of PIK3CA mutations in the invasive primary tumor will accurately reflect breast cancer biology.

PIK3CA mutation associates with improved outcome in breast cancer.

PURPOSE: In breast cancer, somatic mutations in the PIK3CA gene are common. The prognostic implication of these activating mutations remains uncertain as moderately sized studies have yielded variable outcomes. Our aim was to determine the prognostic implications of PIK3CA mutations in breast cancer. EXPERIMENTAL DESIGN: Archival formalin-fixed paraffin-embedded primary breast tumors, from 590 patients selected for known vital status with a median follow-up of 12.8 years and a tumor >1 cm, were genotyped for PIK3CA mutations. Mutation rates and associations between mutation site and clinicopathologic characteristics were assessed. Progression-free survival, overall survival, and breast cancer-specific survival were examined using Kaplan-Meier or competing risk methodology. RESULTS: PIK3CA mutation is identified in 32.5% of breast cancers. PIK3CA mutation significantly associates with older age at diagnosis, hormone receptor positivity, HER2 negativity, lower tumor grade and stage, and lymph node negativity. Patients with PIK3CA mutated tumors have significant improvement in overall survival (P = 0.03) and breast cancer-specific survival (P = 0.004). Analysis for PIK3CA mutation site-specific associations reveals that the H1047R kinase domain mutation highly associates with node negativity (P = 0.007), whereas helical domain hotspot mutations associate with older age at diagnosis (P = 0.004). CONCLUSION: This study defines the positive prognostic significance of PIK3CA mutations. This work is clinically relevant, as it will significantly affect the design of clinical trials planned for phosphatidylinositol 3-kinase-targeted therapy. Future work may define a population of older age breast cancer patients in whom therapy can be minimized.

NF-kappaB regulates androgen receptor expression and prostate cancer growth.

Prostate cancers that progress during androgen-deprivation therapy often overexpress the androgen receptor (AR) and depend on AR signaling for growth. In most cases, increased AR expression occurs without gene amplification and may be due to altered transcriptional regulation. The transcription factor nuclear factor (NF)-kappaB, which is implicated in tumorigenesis, functions as an important downstream substrate of mitogen-activated protein kinase, phosphatidylinositol 3-kinase, AKT, and protein kinase C and plays a role in other cancer-associated signaling pathways. NF-kappaB is an important determinant of prostate cancer clinical biology, and therefore we investigated its role in the regulation of AR expression. We found that NF-kappaB expression in prostate cancer cells significantly increased AR mRNA and protein levels, AR transactivation activity, serum prostate-specific antigen levels, and cell proliferation. NF-kappaB inhibitors decrease AR expression levels, prostate-specific antigen secretion, and proliferation of prostate cancer cells in vitro. Furthermore, inhibitors of NF-kappaB demonstrated anti-tumor activity in androgen deprivation-resistant prostate cancer xenografts. In addition, levels of both NF-kappaB and AR were strongly correlated in human prostate cancer. Our data suggest that NF-kappaB can regulate AR expression in prostate cancer and that NF-kappaB inhibitors may have therapeutic potential.