TFEB drives mTORC1 hyperactivation and kidney disease in Tuberous Sclerosis Complex.

Tuberous Sclerosis Complex (TSC) is caused by TSC1 or TSC2 mutations, leading to hyperactivation of mechanistic target of rapamycin complex 1 (mTORC1) and lesions  in multiple organs including lung (lymphangioleiomyomatosis) and kidney (angiomyolipoma and renal cell carcinoma). Previously, we found that TFEB is constitutively active in TSC. Here, we generated two mouse models of TSC in which kidney pathology is the primary phenotype. Knockout of TFEB rescues kidney pathology and overall survival, indicating that TFEB is the primary driver of renal disease in TSC. Importantly, increased mTORC1 activity in the TSC2 knockout kidneys is normalized by TFEB knockout. In TSC2-deficient cells, Rheb knockdown or Rapamycin treatment paradoxically increases TFEB phosphorylation at the mTORC1-sites and relocalizes TFEB from nucleus to cytoplasm. In mice, Rapamycin treatment normalizes lysosomal gene expression, similar to TFEB knockout, suggesting that Rapamycin's benefit in TSC is TFEB-dependent. These results change the view of the mechanisms of mTORC1 hyperactivation in TSC and may lead to therapeutic avenues.

Ancestry-driven recalibration of tumor mutational burden and disparate clinical outcomes in response to immune checkpoint inhibitors.

The immune checkpoint inhibitor (ICI) pembrolizumab is US FDA approved for treatment of solid tumors with high tumor mutational burden (TMB-high; ≥10 variants/Mb). However, the extent to which TMB-high generalizes as an accurate biomarker in diverse patient populations is largely unknown. Using two clinical cohorts, we investigated the interplay between genetic ancestry, TMB, and tumor-only versus tumor-normal paired sequencing in solid tumors. TMB estimates from tumor-only panels substantially overclassified individuals into the clinically important TMB-high group due to germline contamination, and this bias was particularly pronounced in patients with Asian/African ancestry. Among patients with non-small cell lung cancer treated with ICIs, those misclassified as TMB-high from tumor-only panels did not associate with improved outcomes. TMB-high was significantly associated with improved outcomes only in European ancestries and merits validation in non-European ancestry populations. Ancestry-aware tumor-only TMB calibration and ancestry-diverse biomarker studies are critical to ensure that existing disparities are not exacerbated in precision medicine.

Variation in targetable genomic alterations in non-small cell lung cancer by genetic ancestry, sex, smoking history, and histology.

BACKGROUND: Genomic alterations in 8 genes are now the targets of FDA-approved therapeutics in non-small cell lung cancer (NSCLC), but their distribution according to genetic ancestry, sex, histology, and smoking is not well established. METHODS: Using multi-institutional genetic testing data from GENIE, we characterize the distribution of targetable genomic alterations in 8 genes among 8675 patients with NSCLC (discovery cohort: DFCI, N = 3115; validation cohort: Duke, Memorial Sloan Kettering Cancer Center, Vanderbilt, N = 5560). For the discovery cohort, we impute genetic ancestry from tumor-only sequencing and identify differences in the frequency of targetable alterations across ancestral groups, smoking pack-years, and histologic subtypes. RESULTS: We identified variation in the prevalence of KRASG12C, sensitizing EGFR mutations, MET alterations, ALK, and ROS1 fusions according to the number of smoking pack-years. A novel method for computing continental (African, Asian, European) and Ashkenazi Jewish ancestries from panel sequencing enables quantitative analysis of the correlation between ancestry and mutation rates. This analysis identifies a correlation between Asian ancestry and EGFR mutations and an anti-correlation between Asian ancestry and KRASG12C mutation. It uncovers 2.7-fold enrichment for MET exon 14 skipping mutations and amplifications in patients of Ashkenazi Jewish ancestry. Among never/light smokers, targetable alterations in LUAD are significantly enriched in those with Asian (80%) versus African (49%) and European (55%) ancestry. Finally, we show that 5% of patients with squamous cell carcinoma (LUSC) and 17% of patients with large cell carcinoma (LCLC) harbor targetable alterations. CONCLUSIONS: Among patients with NSCLC, there was significant variability in the prevalence of targetable genomic alterations according to genetic ancestry, histology, and smoking. Patients with LUSC and LCLC have 5% rates of targetable alterations supporting consideration for sequencing in those subtypes.

Prevalence of pathogenic germline cancer risk variants in testicular cancer patients: Identifying high risk groups.

BACKGROUND: Germline studies in testicular cancer have focused on unselected populations but so far have not led to recommendations for testicular cancer screening. OBJECTIVE: Herein, we hypothesized that men with testicular cancer and an additional risk factor for hereditary cancer predisposition carry a higher rate of pathogenic variants than men with testicular cancer without another risk factor. METHODS AND RESULTS: 187 patients with a personal history of testicular cancer underwent germline testing via Invitae. Patients were divided into low-risk and high-risk patients. Low-risk patients (n=83) had testicular cancer as their only primary malignancy without a family history of testicular cancer. High-risk patients (n=104) had additional primary malignancies and/or a family history of testicular cancer. 23.1% of patients harbored pathogenic germline variants with 19.6% carrying actionable variants. Among low-risk patients, 13.5% carried pathogenic variants versus 29.9% in the high-risk cohort. Of patients with a family history of non-testicular cancers and a personal history of additional primary malignancies, 32% harbored pathogenic variants. CONCLUSION: High-risk patients are twice as likely to harbor pathogenic variants compared to low-risk patients. Importantly, patients with a family history of cancer and other primary malignancies represent a subset of patients that may benefit from genetic evaluation.

CDH1 germline variants are enriched in patients with colorectal cancer, gastric cancer, and breast cancer.

BACKGROUND AND AIMS: CDH1 germline variants have been linked to heritability in diffuse gastric (DGC) and lobular breast cancer (LBC). Studies have not yet assessed whether CDH1 is a cancer-susceptibility gene in other cancers. Herein, we mapped the landscape of pathogenic and likely pathogenic (P/LP) germline variants in CDH1 across various cancers and ethnicities. METHODS: We evaluated CDH1 germline P/LP variants in 212,944 patients at one CLIA-certified laboratory (Invitae) and described their frequency in 7 cancer types. We screened for CDH1 variant enrichment in each cancer relative to a cancer-free population from The Genome Aggregation Database version 3 (gnomADv3). RESULTS: CDH1 P/LP variants were identified in 141 patients, most commonly in patients with DGC (27/408, 6.6%) followed by colorectal signet-ring cell cancer (CSRCC; 3/79, 3.8%), gastric cancer (56/2756, 2%), and LBC (22/6809, 0.3%). CDH1 P/LP variants were enriched in specific ethnic populations with breast cancer, gastric cancer, CRC, LBC, DGC, and CSRCC compared to matched ethnicities from gnomADv3. CONCLUSION: We report for the first time the prevalence of P/LP CDH1 variants across several cancers and show significant enrichment in CDH1 P/LP variants for patients with CSRCC, DGC, and LBC across various ethnicities. Future prospective studies are warranted to validate these findings.

TSC2 regulates lysosome biogenesis via a non-canonical RAGC and TFEB-dependent mechanism.

Tuberous Sclerosis Complex (TSC) is caused by TSC1 or TSC2 mutations, resulting in hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1). Transcription factor EB (TFEB), a master regulator of lysosome biogenesis, is negatively regulated by mTORC1 through a RAG GTPase-dependent phosphorylation. Here we show that lysosomal biogenesis is increased in TSC-associated renal tumors, pulmonary lymphangioleiomyomatosis, kidneys from Tsc2+/- mice, and TSC1/2-deficient cells via a TFEB-dependent mechanism. Interestingly, in TSC1/2-deficient cells, TFEB is hypo-phosphorylated at mTORC1-dependent sites, indicating that mTORC1 is unable to phosphorylate TFEB in the absence of the TSC1/2 complex. Importantly, overexpression of folliculin (FLCN), a GTPase activating protein for RAGC, increases TFEB phosphorylation at the mTORC1 sites in TSC2-deficient cells. Overexpression of constitutively active RAGC is sufficient to relocalize TFEB to the cytoplasm. These findings establish the TSC proteins as critical regulators of lysosomal biogenesis via TFEB and RAGC and identify TFEB as a driver of the proliferation of TSC2-deficient cells.

CDKN2A Alterations and Response to Immunotherapy in Solid Tumors.

PURPOSE: Immune checkpoint inhibitors (ICI) have shown clinical benefit in many types of metastatic cancers with only a few predictive biomarkers identified so far. CDKN2A is commonly altered in human cancers, but prior studies have provided conflicting evidence regarding the association between CDKN2A genomic alterations (GA) and response to ICIs. Herein, we examined the impact of loss-of-function CDKN2A alterations on response and survival in patients treated with ICIs. EXPERIMENTAL DESIGN: We studied the association between loss-of-function CDKN2A alterations and the response to ICIs in two independent cohorts of six different cancer types. Seven hundred and eighty-nine patients treated at Dana-Farber Cancer Institute (DFCI; Boston, MA) and 1,250 patients treated at Memorial Sloan Kettering Cancer Center (MSKCC; New York, NY) were included in the final analysis. Patients' tumors were sequenced using Oncopanel or MSK-IMPACT. RNA sequencing data from The Cancer Genome Atlas and IMvigor210 were used to investigate differences in the tumor microenvironment. RESULTS: In the DFCI cohort, CDKN2A GAs were associated with poor response and survival in patients with urothelial carcinoma treated with ICIs, but not those treated with platinum-based therapy. Similarly, CDKN2A GAs were associated with worse outcomes in the MSKCC urothelial carcinoma cohort treated with ICIs. There was no association of CDKN2A status with ICI treatment outcome in five other cancers: esophagogastric, head and neck, non-small cell lung, renal cell carcinoma, and melanoma. Immuno-inflammatory pathways were significantly reduced in expression in CDKN2A-altered tumors. CONCLUSIONS: Our data show that CDKN2A GAs were associated with reduced benefit from ICI therapy in urothelial carcinoma as well as changes in the tumor-immune microenvironment.

Trans-ethnic variation in germline variants of patients with renal cell carcinoma.

Prior studies of the renal cell carcinoma (RCC) germline landscape investigated predominantly patients of European ancestry. We examine the frequency of germline pathogenic and likely pathogenic (P/LP) variants in 1,829 patients with RCC from various ancestries. Overall, P/LP variants are found in 17% of patients, among whom 10.3% harbor one or more clinically actionable variants with potential preventive or therapeutic utility. Patients of African ancestry with RCC harbor significantly more P/LP variants in FH compared to patients of non-African ancestry with RCC and African controls from the Genome Aggregation Database (gnomAD). Patients of non-African ancestry have significantly more P/LP variants in CHEK2 compared to patients of African ancestry with RCC and non-Finnish Europeans controls. Non-Africans with RCC have more actionable variants compared to Africans with RCC. This work helps understand the underlying biological differences in RCC between Africans and non-Africans and paves the way to more comprehensive genomic characterization of underrepresented populations.

Therapeutic Targeting of DGKA-Mediated Macropinocytosis Leads to Phospholipid Reprogramming in Tuberous Sclerosis Complex.

Lymphangioleiomyomatosis is a rare destructive lung disease affecting primarily women and is the primary lung manifestation of tuberous sclerosis complex (TSC). In lymphangioleiomyomatosis, biallelic loss of TSC1/2 leads to hyperactivation of mTORC1 and inhibition of autophagy. To determine how the metabolic vulnerabilities of TSC2-deficient cells can be targeted, we performed a high-throughput screen utilizing the "Repurposing" library at the Broad Institute of MIT and Harvard (Cambridge, MA), with or without the autophagy inhibitor chloroquine. Ritanserin, an inhibitor of diacylglycerol kinase alpha (DGKA), was identified as a selective inhibitor of proliferation of Tsc2-/- mouse embryonic fibroblasts (MEF), with no impact on Tsc2+/+ MEFs. DGKA is a lipid kinase that metabolizes diacylglycerol to phosphatidic acid, a key component of plasma membranes. Phosphatidic acid levels were increased 5-fold in Tsc2-/- MEFs compared with Tsc2+/+ MEFs, and treatment of Tsc2-/- MEFs with ritanserin led to depletion of phosphatidic acid as well as rewiring of phospholipid metabolism. Macropinocytosis is known to be upregulated in TSC2-deficient cells. Ritanserin decreased macropinocytic uptake of albumin, limited the number of lysosomes, and reduced lysosomal activity in Tsc2-/- MEFs. In a mouse model of TSC, ritanserin treatment decreased cyst frequency and volume, and in a mouse model of lymphangioleiomyomatosis, genetic downregulation of DGKA prevented alveolar destruction and airspace enlargement. Collectively, these data indicate that DGKA supports macropinocytosis in TSC2-deficient cells to maintain phospholipid homeostasis and promote proliferation. Targeting macropinocytosis with ritanserin may represent a novel therapeutic approach for the treatment of TSC and lymphangioleiomyomatosis. SIGNIFICANCE: This study identifies macropinocytosis and phospholipid metabolism as novel mechanisms of metabolic homeostasis in mTORC1-hyperactive cells and suggest ritanserin as a novel therapeutic strategy for use in mTORC1-hyperactive tumors, including pancreatic cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/8/2086/F1.large.jpg.

The Use of Social Media to Increase the Impact of Health Research: Systematic Review.

BACKGROUND: Academics in all disciplines increasingly use social media to share their publications on the internet, reaching out to different audiences. In the last few years, specific indicators of social media impact have been developed (eg, Altmetrics), to complement traditional bibliometric indicators (eg, citation count and h-index). In health research, it is unclear whether social media impact also translates into research impact. OBJECTIVE: The primary aim of this study was to systematically review the literature on the impact of using social media on the dissemination of health research. The secondary aim was to assess the correlation between Altmetrics and traditional citation-based metrics. METHODS: We conducted a systematic review to identify studies that evaluated the use of social media to disseminate research published in health-related journals. We specifically looked at studies that described experimental or correlational studies linking the use of social media with outcomes related to bibliometrics. We searched the Medical Literature Analysis and Retrieval System Online (MEDLINE), Excerpta Medica dataBASE (EMBASE), and Cumulative Index to Nursing and Allied Health Literature (CINAHL) databases using a predefined search strategy (International Prospective Register of Systematic Reviews: CRD42017057709). We conducted independent and duplicate study selection and data extraction. Given the heterogeneity of the included studies, we summarized the findings through a narrative synthesis. RESULTS: Of a total of 18,624 retrieved citations, we included 51 studies: 7 (14%) impact studies (answering the primary aim) and 44 (86%) correlational studies (answering the secondary aim). Impact studies reported mixed results with several limitations, including the use of interventions of inappropriately low intensity and short duration. The majority of correlational studies suggested a positive association between traditional bibliometrics and social media metrics (eg, number of mentions) in health research. CONCLUSIONS: We have identified suggestive yet inconclusive evidence on the impact of using social media to increase the number of citations in health research. Further studies with better design are needed to assess the causal link between social media impact and bibliometrics.

Coronavirus Goes Viral: Quantifying the COVID-19 Misinformation Epidemic on Twitter.

Background Since the beginning of the coronavirus disease 2019 (COVID-19) epidemic, misinformation has been spreading uninhibited over traditional and social media at a rapid pace. We sought to analyze the magnitude of misinformation that is being spread on Twitter (Twitter, Inc., San Francisco, CA) regarding the coronavirus epidemic.  Materials and methods We conducted a search on Twitter using 14 different trending hashtags and keywords related to the COVID-19 epidemic. We then summarized and assessed individual tweets for misinformation in comparison to verified and peer-reviewed resources. Descriptive statistics were used to compare terms and hashtags, and to identify individual tweets and account characteristics. Results The study included 673 tweets. Most tweets were posted by informal individuals/groups (66%), and 129 (19.2%) belonged to verified Twitter accounts. The majority of included tweets contained serious content (91.2%); 548 tweets (81.4%) included genuine information pertaining to the COVID-19 epidemic. Around 70% of the tweets tackled medical/public health information, while the others were pertaining to sociopolitical and financial factors. In total, 153 tweets (24.8%) included misinformation, and 107 (17.4%) included unverifiable information regarding the COVID-19 epidemic. The rate of misinformation was higher among informal individual/group accounts (33.8%, p: <0.001). Tweets from unverified Twitter accounts contained more misinformation (31.0% vs 12.6% for verified accounts, p: <0.001). Tweets from healthcare/public health accounts had the lowest rate of unverifiable information (12.3%, p: 0.04). The number of likes and retweets per tweet was not associated with a difference in either false or unverifiable content. The keyword "COVID-19" had the lowest rate of misinformation and unverifiable information, while the keywords "#2019_ncov" and "Corona" were associated with the highest amount of misinformation and unverifiable content respectively. Conclusions Medical misinformation and unverifiable content pertaining to the global COVID-19 epidemic are being propagated at an alarming rate on social media. We provide an early quantification of the magnitude of misinformation spread and highlight the importance of early interventions in order to curb this phenomenon that endangers public safety at a time when awareness and appropriate preventive actions are paramount.