Comparison of Genetic and Epigenetic Alterations of Primary Tumors and Matched Plasma Samples in Patients with Colorectal Cancer

BackgroundAlthough recent advances in circulating DNA analysis allow the prediction of tumor genomes by noninvasive means, some challenges remain, which limit the widespread introduction of cfDNA in cancer diagnostics. We analyzed the status of the two best characterized colorectal cancer (CRC) genetic and epigenetic alterations in a cohort of CRC patients, and then compared the degree to which the two patterns move from tissue to plasma in order to improve our understanding of biology modulating the concordance between tissues and plasma methylation and mutation profiles.MethodsPlasma and tumor tissues were collected from 85 patients (69±14 years, 56 males). KRAS and SEPT9 status was assessed by allele refractory mutation system quantitative PCR and quantitative methylation-specific PCR, respectively. Six of the most common point mutations at codon 12 and 13 were investigated for KRAS analysis.ResultsKRAS mutations and SEPT9 promoter methylation were present in 34% (29/85) and in 82% (70/85) of primary tumor tissue samples. Both genetic and epigenetic analyses of cfDNA revealed a high overall concordance and specificity compared with tumor-tissue analyses. Patients presenting with both genetic and epigenetic alterations in tissue specimens (31.8%, 27/85) were considered for further analyses. The median methylation rates in tumour tissues and plasma samples were 64.5% (12.2–99.8%) and 14.5% (0–45.5%), respectively. The median KRAS mutation load (for matched mutations) was 33.6% (1.8–86.3%) in tissues and 2.9% (0–17.3) in plasma samples. The plasma/tissue (p/t) ratio of SEPT9 methylation rate was significantly higher than the p/t ratio of KRAS mutation load, especially in early stage cancers (p=0.0108).ConclusionThe results of this study show a discrepant rate of epigenetic vs. genetic alterations moving from tissue to plasma. Many factors could affect mutation cfDNA analysis, including both presence of tumor clonal heterogeneity and strict compartmentalization of KRAS mutation profile. The present study highlights the importance of considering the nature of the alteration when analyzing tumor-derived cfDNA.     

A Meta-Analysis of Retinoblastoma Copy Numbers Refines the List of Possible Driver Genes Involved in Tumor Progression

BackgroundWhile RB1 loss initiates retinoblastoma development, additional somatic copy number alterations (SCNAs) can drive tumor progression. Although SCNAs have been identified with good concordance between studies at a cytoband resolution, accurate identification of single genes for all recurrent SCNAs is still challenging. This study presents a comprehensive meta-analysis of genome-wide SCNAs integrated with gene expression profiling data, narrowing down the list of plausible retinoblastoma driver genes.MethodsWe performed SCNA profiling of 45 primary retinoblastoma samples and eight retinoblastoma cell lines by high-resolution microarrays. We combined our data with genomic, clinical and histopathological data of ten published genome-wide SCNA studies, which strongly enhanced the power of our analyses (N = 310).ResultsComprehensive recurrence analysis of SCNAs in all studies integrated with gene expression data allowed us to reduce candidate gene lists for 1q, 2p, 6p, 7q and 13q to a limited gene set. Besides the well-established driver genes RB1 (13q-loss) and MYCN (2p-gain) we identified CRB1 and NEK7 (1q-gain), SOX4 (6p-gain) and NUP205 (7q-gain) as novel retinoblastoma driver candidates. Depending on the sample subset and algorithms used, alternative candidates were identified including MIR181 (1q-gain) and DEK (6p gain). Remarkably, our study showed that copy number gains rarely exceeded change of one copy, even in pure tumor samples with 100% homozygosity at the RB1 locus (N = 34), which is indicative for intra-tumor heterogeneity. In addition, profound between-tumor variability was observed that was associated with age at diagnosis and differentiation grades.InterpretationSince focal alterations at commonly altered chromosome regions were rare except for 2p24.3 (MYCN), further functional validation of the oncogenic potential of the described candidate genes is now required. For further investigations, our study provides a refined and revised set of candidate retinoblastoma driver genes.     

Soluble AXL: A Possible Circulating Biomarker for Neurofibromatosis Type 1 Related Tumor Burden

Neurofibromatosis type 1 (NF1) is the most common tumor predisposition disorder affecting 1/3500 worldwide. Patients are at risk of developing benign (neurofibromas) and malignant peripheral nerve sheath tumors (MPNST). The AXL receptor tyrosine kinase has been implicated in several kinds of cancers, but so far no studies have investigated the role of AXL in NF1 related tumorigenesis. Recently, the soluble fraction from the extracellular domain of AXL (sAXL) has been found in human plasma, and its level was correlated to poor prognosis in patients with renal cancer. Compared to normal human Schwann cells, a significantly high expression level of AXL was found in three of the four MPNST cell lines and two of the three primary MPNST tissues. Similarly, the level of sAXL in conditioned media corresponded to the protein and mRNA levels of AXL in the MPNST cell lines. Furthermore, in two different human MPNST xenograft models, the human sAXL could be detected in the mouse plasma. Its level was proportionate to the size of the xenograft tumors, while no human sAXL was detect prior to the formation of the tumors. Treatment with a newly developed photodynamic therapy, prevented further tumor growth and resulted in drastically reduced the levels of sAXL compared to that of the control group. Finally, the level of sAXL was significantly increased in patients with plexiform tumors compared to patients with only dermal neurofibromas, further supporting the role of sAXL as a marker for NF1 related tumor burden.     

Clinical utility of tumor genomic profiling in patients with high plasma circulating tumor DNA burden or metabolically active tumors

Background

This retrospective study was undertaken to determine if the plasma circulating tumor DNA (ctDNA) level and tumor biological features in patients with advanced solid tumors affected the detection of genomic alterations (GAs) by a plasma ctDNA assay.

Method

Cell-free DNA (cfDNA) extracted from frozen plasma (N?=?35) or fresh whole blood (N?=?90) samples were subjected to a 62-gene hybrid capture-based next-generation sequencing assay FoundationACT. Concordance was analyzed for 51 matched FoundationACT and FoundationOne (tissue) cases. The maximum somatic allele frequency (MSAF) was used to estimate the amount of tumor fraction of cfDNA in each sample. The detection of GAs was correlated with the amount of cfDNA, MSAF, total tumor anatomic burden (dimensional sum), and total tumor metabolic burden (SUVmax sum) of the largest ten tumor lesions on PET/CT scans.

Results

FoundationACT detected GAs in 69 of 81 (85%) cases with MSAF >?0. Forty-two of 51 (82%) cases had ≥?1 concordance GAs matched with FoundationOne, and 22 (52%) matched to the National Comprehensive Cancer Network (NCCN)-recommended molecular targets. FoundationACT also detected 8 unique molecular targets, which changed the therapy in 7 (88%) patients who did not have tumor rebiopsy or sufficient tumor DNA for genomic profiling assay. In all samples (N?=?81), GAs were detected in plasma cfDNA from cancer patients with high MSAF quantity (P?=?0.0006) or high tumor metabolic burden (P?=?0.0006) regardless of cfDNA quantity (P?=?0.2362).

Conclusion

This study supports the utility of using plasma-based genomic assays in cancer patients with high plasma MSAF level or high tumor metabolic burden.

     

Impaired Pten Expression in Human Malignant Peripheral Nerve Sheath Tumours

Malignant peripheral nerve sheath tumours (MPNST) are aggressive sarcomas that develop in about 10% of patients with the genetic disease neurofibromatosis type 1 (NF1). Molecular alterations contributing to MPNST formation have only partially been resolved. Here we examined the role of Pten, a key regulator of the Pi3k/Akt/mTOR pathway, in human MPNST and benign neurofibromas. Immunohistochemistry showed that Pten expression was significantly lower in MPNST (n = 16) than in neurofibromas (n = 16) and normal nervous tissue. To elucidate potential mechanisms for Pten down-regulation or Akt/mTOR activation in MPNST we performed further experiments. Mutation analysis revealed absence of somatic mutations in PTEN (n = 31) and PIK3CA (n = 38). However, we found frequent PTEN promotor methylation in primary MPNST (11/26) and MPNST cell lines (7/8) but not in benign nerve sheath tumours. PTEN methylation was significantly associated with early metastasis. Moreover, we detected an inverse correlation of Pten-regulating miR-21 and Pten protein levels in MPNST cell lines. The examination of NF1−/− and NF1+/+Schwann cells and fibroblasts showed that Pten expression is not regulated by NF1. To determine the significance of Pten status for treatment with the mTOR inhibitor rapamycin we treated 5 MPNST cell lines with rapamycin. All cell lines were sensitive to rapamycin without a significant correlation to Pten levels. When rapamycin was combined with simvastatin a synergistic anti-proliferative effect was achieved. Taken together we show frequent loss/reduction of Pten expression in MPNST and provide evidence for the involvement of multiple Pten regulating mechanisms.     

Combined multimodal ctDNA analysis and radiological imaging for tumor surveillance in Non-small cell lung cancer

BackgroundRadiology is the current standard for monitoring treatment responses in lung cancer. Limited sensitivity, exposure to ionizing radiations and related sequelae constitute some of its major limitation. Non-invasive and highly sensitive methods for early detection of treatment failures and resistance-associated disease progression would have additional clinical utility.MethodsWe analyzed serially collected plasma and paired tumor samples from lung cancer patients (61 with stage IV, 48 with stages I-III disease) and 61 healthy samples by means of next-generation sequencing, radiological imaging and droplet digital polymerase chain reaction (ddPCR) mutation and methylation assays.ResultsA 62% variant concordance between tumor-reported and circulating-free DNA (cfDNA) sequencing was observed between baseline liquid and tissue biopsies in stage IV patients. Interestingly, ctDNA sequencing allowed for the identification of resistance-mediating p.T790M mutations in baseline plasma samples for which no such mutation was observed in the corresponding tissue. Serial circulating tumor DNA (ctDNA) mutation analysis by means of ddPCR revealed a general decrease in ctDNA loads between baseline and first reassessment. Additionally, serial ctDNA analyses only recapitulated computed tomography (CT) -monitored tumor dynamics of some, but not all lesions within the same patient. To complement ctDNA variant analysis we devised a ctDNA methylation assay (^methcfDNA) based on methylation-sensitive restriction enzymes. cfDNA methylation showed and area under the curve (AUC) of > 0.90 in early and late stage cases. A decrease in ^methcfDNA between baseline and first reassessment was reflected by a decrease in CT-derive tumor surface area, irrespective of tumor mutational status.ConclusionTaken together, our data support the use of cfDNA sequencing for unbiased characterization of the molecular tumor architecture, highlights the impact of tumor architectural heterogeneity on ctDNA-based tumor surveillance and the added value of complementary approaches such as cfDNA methylation for early detection and monitoring     

Comparison of ESR1 Mutations in Tumor Tissue and Matched Plasma Samples from Metastatic Breast Cancer Patients

BACKGROUND: ESR1 mutation in circulating cell-free DNA (cfDNA) is emerging as a noninvasive biomarker of acquired resistance to endocrine therapy, but there is a paucity of data comparing the status of ESR1 gene in cfDNA with that in its corresponding tumor tissue. The objective of this study is to validate the degree of concordance of ESR1 mutations between plasma and tumor tissue. METHODS: ESR1 ligand-binding domain mutations Y537S, Y537N, Y537C, and D538G were analyzed using droplet digital PCR in 35 patients with metastatic breast cancer (MBC) (35 tumor tissue samples and 67 plasma samples). RESULTS: Of the 35 paired samples, 26 (74.3%) were concordant: one patient had detectable ESR1 mutations both plasma (ESR1 Y537S/Y537N) and tumor tissue (ESR1 Y537S/Y537C), and 25 had WT ESR1 alleles in both. Nine (25.7%) had discordance between the plasma and tissue results: five had mutations detected only in their tumor tissue (two Y537S, one Y537C, one D538G, and one Y537S/Y537N/D538G), and four had mutations detected only in their plasma (one Y537S, one Y537N, and two Y537S/Y537N/D538G). Furthermore, longitudinal plasma samples from 19 patients were used to assess changes in the presence of ESR1 mutations during treatment. Eleven patients had cfDNA ESR1 mutations over the course of treatment. A total of eight of 11 patients with MBC with cfDNA ESR1 mutations (72.7%) had the polyclonal mutations. CONCLUSION: We have shown the independent distribution of ESR1 mutations between plasma and tumor tissue in 35 patients with MBC.     

Aberrant cAMP Metabolism in NF1 Malignant Peripheral Nerve Sheath Tumor Cells

Malignant peripheral nerve sheath (MPNST) cell lines derived from patients with neurofibromatosis type 1 (NF!) were found to have basal cAMP levels which are two-fold higher than cAMP levels in normal human adult Schwann cells (nHSC). PCR analysis also revealed that normal adult human Schwann cells express mRNA for types Ill, IV, and IX adenylyl cyclase (AC) while NF1 MPNST cells express AC mRNA of types II, V, and VIII in addition to expressing all the isoforms of normal adult human Schwann cells. Further PCR analysis revealed that NF1 MPNST lines express mRNA for EP2 and EP4 prostaglandin receptors whereas nHSC only express mRNA for the EP2 receptor. Exogenous prostaglandins alone or in combination with PDGF BB induced greater increases in cAMP levels and proliferation of NF1 MPNST cells compared to nHSC. We conclude that aberrant cAMP signaling in NF1 MPNST cells contributes to tumor formation in NF1 patients.     

Somatic genome variations in vascular tissues and peripheral blood leukocytes in patients with atherosclerosis

The first data on the existence of multiple genomic rearrangements, such as copy number variation (CNV) and copy neutral loss of heterozygosity, in vascular tissues and peripheral blood leukocytes from patients with atherosclerosis, are presented. Compared to internal mammary arteries and peripheral blood leukocytes, right coronary arteries in the atherosclerotic plaque area presented with a higher CNV length and number of genes located in their vicinity. In each of the patients, 6–16% of CNVs were common to the three types of tissues examined. Therefore, most of the copy number variations in the tissues affected by atherosclerosis (from 68 to 91% in each of the patients) were of somatic origin. The gains in 3p21.31 (CACNA2D2), 7q32.1 (FLNC), 19p13.3 (C19orf29, PIP5K1C), and 21q22.3 (COL6A1) were detected in vascular tissues but not in peripheral blood leukocytes. Moreover, the gain in 7p15.2 (SKAP2), detected in the patients with atherosclerosis, did not overlap with any CNV regions currently reported in The Database of Genomic Variants. The loss of heterozygosity in 12 out of 13 chromosomal regions was copy neutral and covered tumor suppressor genes (SFRP1, CEBPD, RB1CC1, DIRAS3, TUSC3, and ZDHHC2).     

Comparative Oncogenomics Implicates the Neurofibromin 1 Gene (NF1) as a Breast Cancer Driver

Identifying genomic alterations driving breast cancer is complicated by tumor diversity and genetic heterogeneity. Relevant mouse models are powerful for untangling this problem because such heterogeneity can be controlled. Inbred Chaos3 mice exhibit high levels of genomic instability leading to mammary tumors that have tumor gene expression profiles closely resembling mature human mammary luminal cell signatures. We genomically characterized mammary adenocarcinomas from these mice to identify cancer-causing genomic events that overlap common alterations in human breast cancer. Chaos3 tumors underwent recurrent copy number alterations (CNAs), particularly deletion of the RAS inhibitor Neurofibromin 1 (Nf1) in nearly all cases. These overlap with human CNAs including NF1, which is deleted or mutated in 27.7% of all breast carcinomas. Chaos3 mammary tumor cells exhibit RAS hyperactivation and increased sensitivity to RAS pathway inhibitors. These results indicate that spontaneous NF1 loss can drive breast cancer. This should be informative for treatment of the significant fraction of patients whose tumors bear NF1 mutations.     

Pheochromocytoma in Neurofibromatosis Type 1: When Should it Be Suspected

ObjectivesNeurofibromatosis type 1 (NF1) carries an increased risk of pheochromocytoma. Most experts recommend that NF1 patients be screened for pheochromocytoma if hypertension develops. We sought to compare NF1 and non-NF1 patients with pheochromocytoma.MethodsRetrospective analysis of a prospectively collected database of all patients undergoing pheochromocytoma resection by a single surgeon from 2003-2012. Statistical significance was evaluated using Fisher’s exact test for categorical variables and the Wilcoxon rank sum test for continuous variables.ResultsOf 56 patients undergoing pheochromocytoma resection, 6 (11%) had NF1. All 6 (100%) NF1 patients had pheochromocytoma diagnosed incidentally during work-up for another condition, whereas 28 of 50 (56%) non-NF1–associated pheochromocytomas were diagnosed incidentally (P = .071). Hypertension was present in 1 (17%) NF1 patient and in 37 (74%) of the non-NF1 patients (P = .011). Tumors were significantly smaller in NF1 patients compared with non-NF1 patients (median tumor dimension, 2.75 cm vs. 5.9 cm, respectively;P = .014).ConclusionAlthough NF1 patients have a well-known increased risk of developing pheochromocytoma, in the current series, all NF1 patients referred to the surgeon for adrenalectomy had pheochromocytoma diagnosed incidentally. Nevertheless, NF1 patients had significantly smaller tumors and less hypertension than other patients treated for pheochromocytoma, perhaps due to a higher frequency of imaging occasioned by their other neoplasms. The common recommendation to screen for pheochromocytoma when hypertension develops would have failed to spur screening in 83% of these NF1 patients. Routine screening for pheochromocytoma in all NF1 patients may be warranted after evaluating whether this is cost-effective in reducing morbidity and mortality. (Endocr Pract. 2014;20:792-796)     

High-resolution aCGH and expression profiling identifies a novel genomic subtype of ER negative breast cancer

Background

The characterization of copy number alteration patterns in breast cancer requires high-resolution genome-wide profiling of a large panel of tumor specimens. To date, most genome-wide array comparative genomic hybridization studies have used tumor panels of relatively large tumor size and high Nottingham Prognostic Index (NPI) that are not as representative of breast cancer demographics.

Results

We performed an oligo-array-based high-resolution analysis of copy number alterations in 171 primary breast tumors of relatively small size and low NPI, which was therefore more representative of breast cancer demographics. Hierarchical clustering over the common regions of alteration identified a novel subtype of high-grade estrogen receptor (ER)-negative breast cancer, characterized by a low genomic instability index. We were able to validate the existence of this genomic subtype in one external breast cancer cohort. Using matched array expression data we also identified the genomic regions showing the strongest coordinate expression changes ('hotspots'). We show that several of these hotspots are located in the phosphatome, kinome and chromatinome, and harbor members of the 122-breast cancer CAN-list. Furthermore, we identify frequently amplified hotspots on 8q22.3 (EDD1, WDSOF1), 8q24.11-13 (THRAP6, DCC1, SQLE, SPG8) and 11q14.1 (NDUFC2, ALG8, USP35) associated with significantly worse prognosis. Amplification of any of these regions identified 37 samples with significantly worse overall survival (hazard ratio (HR) = 2.3 (1.3-1.4) p = 0.003) and time to distant metastasis (HR = 2.6 (1.4-5.1) p = 0.004) independently of NPI.

Conclusion

We present strong evidence for the existence of a novel subtype of high-grade ER-negative tumors that is characterized by a low genomic instability index. We also provide a genome-wide list of common copy number alteration regions in breast cancer that show strong coordinate aberrant expression, and further identify novel frequently amplified regions that correlate with poor prognosis. Many of the genes associated with these regions represent likely novel oncogenes or tumor suppressors.     

Diagnostic Accuracy of PET/CT-Guided Percutaneous Biopsies for Malignant Peripheral Nerve Sheath Tumors in Neurofibromatosis Type 1 Patients

Background

Malignant peripheral nerve sheath tumors (MPNST) are one of the most frequent causes of death in patients with neurofibromatosis type 1 (NF1). Early detection is crucial because complete surgical resection is the only curative treatment. It has been previously reported that an ¹⁸F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) image with a T/L (Tumor/Liver) SUV_max ratio > 1.5 provides a high negative predictive value; however, it is not specific enough to make a NF1-related MPNST diagnosis. A formal proof of malignant transformation from a histological analysis is necessary before surgical excision because the procedure can cause mutilation. The objective of the present work was to investigate the effectiveness of and complications associated with PET/CT-guided percutaneous biopsies for an NF1-related MPNST diagnosis.

Methods

PET/CT-guided percutaneous biopsy procedures performed on 26 NF1 patients with a clinical suspicion of MPNST and a suspect lesion from a PET/CT scan (T/L SUV_max ratio > 1.5) were retrospectively evaluated. The localization of the suspected malignant site was determined using PET/CT. A stereotactic (ultrasonic and CT control) core biopsy technique was used with a local anesthesia.

Results

The first PET/CT-guided percutaneous biopsies enabled a pathological diagnosis for all of the patients (no "inconclusive " results were obtained), and no secondary procedures were needed. Among the 26 patients, the histopathological results from the biopsy were malignant in 17 cases and benign (BPNST with atypical cells) in nine cases. No complications from the diagnostic procedure were observed. A surgical resection was performed in 18 patients (seven benign and 11 malignant biopsies), removing the fine needle biopsy scar. In addition, six locally advanced/metastatic MPNST were treated with chemo/radiotherapy, and two BPNST had no progression after a follow-up of 14 and 39 months, respectively. The PET/CT-guided percutaneous biopsy gave 25 accurate diagnoses and one false negative (BPNST with atypical cells on the biopsy and MPNST on the operated tumor), resulting in a diagnostic accuracy rate of 96%. This false negative case may be explained by the high heterogeneity of the tumor: benign areas were contiguous with the malignant ones and associated with inflammation.

Conclusions

PET/CT-guided percutaneous biopsies are an effective and relatively non-traumatic procedure for diagnosis of NF1-related MPNST. It is the most reliable approach for early detection of MPNST.     

Cancer Special Issue: Early detection and minimal residual disease

Beryne Odeny discusses PLOS Medicine’s Special Issue on early cancer detection and minimal residual disease.

PLOS Medicine’s editorial team, together with guest editors, Chris Abbosh, Sarah-Jane Dawson and Charles Swanton, are delighted to disseminate several high-quality translational research and clinical studies on advances in early cancer detection. In 2020 alone, there were upward of 19 million new cancer cases and 10 million cancer deaths, worldwide [1]. Cancer kills more people than HIV/AIDS, tuberculosis and malaria combined and should be a top health priority, regardless of region or country [2]. Early detection of cancer and identification of minimal residual disease (MRD), post-treatment, are key to timely treatment and cure. This issue features robust studies that bring cutting edge, and potentially scalable, innovations that have the potential to inform research, policy, and clinical cancer management.Three studies in this issue center on innovations for detection of MRD. Yaqi Wang and colleagues found that combining circulating tumor DNA (ctDNA) and Magnetic Resonance Imaging (MRI) improved prediction of response to neoadjuvant chemoradiotherapy in patients with locally advanced rectal cancer (LARC) before surgery [3]. This combined model also improved stratification of patients at high risk of recurrence, and clearly has important clinical implications for management of LARC as it could potentially inform guidelines on patient selection for non-operative management and targeted treatment strategies for those with highly recurrent diseases. Jeanne Tie and colleagues confirmed the prognostic utility of post-surgery and post-chemotherapy ctDNA in determining the risk of relapse among patients with colorectal cancer with liver metastases (CRCLM) [4]. They demonstrated the function of serial ctDNA measurement as an early marker of treatment of efficacy. This is a noteworthy advance that requires further study around optimized integration of ctDNA analyses in adjuvant chemotherapy for resectable CRCLM. Pradeep S. Chauhan and colleagues applied next-generation sequencing (NGS) for urine tumor DNA (utDNA) detection to assess MRD in patients with muscle-invasive bladder cancer who received neoadjuvant chemotherapy [5]. They found that MRD detection prior to radical cystectomy correlated with pathologic response and may be used to identify candidates for bladder sparing treatment. Urine tumor DNA also offers the ability to determine tumor mutational burden and can therefore facilitate personalized immunotherapy.Two studies in this issue focused on early cancer detection. Jeffrey J. Szymanski and colleagues investigated the use of plasma cell-free DNA (cfDNA) ultra-low-pass whole genome sequencing (ULP-WSG) to distinguish the malignant peripheral nerve sheath tumor (MPNST) from its benign precursor lesion–plexiform neurofibroma–in patients with Neurofibromatosis type1(NF1) [6]. This provides a strong evidence base for use of plasma cfDNA in liquid biopsy to distinguish early between benign and malignant tumors of this hereditary cancer. This is proof of concept that cfDNA can be leveraged as a biomarker for monitoring treatment response in patients with MPNST. Brian D Nicholson and colleagues demonstrated that risk scores based on combinations of risk factors and routine blood tests can be used to stratify patients with unexpected weight loss based on their risk of cancer [7]. They found that these combined risk scores showed superior clinical utility–compared to the symptoms-only model–to discriminate between patients with and without cancer. In this, they clearly demonstrate innovation in the use of routine clinical tools at scale. This type of model could potentially be scaled-up in under-resourced settings.With growing global interest in cancer diagnostics and treatment, these robust assays and tools are a welcome addition to the early cancer detection armamentarium, prior to and post-treatment. Further innovation around low-cost technologies and tools for early detection that can be rapidly tested and scaled up will further galvanize, the universal commitment to defeat cancer in both high and low resource settings.     

DUF1220-Domain Copy Number Implicated in Human Brain-Size Pathology and Evolution

DUF1220 domains show the largest human-lineage-specific increase in copy number of any protein-coding region in the human genome and map primarily to 1q21, where deletions and reciprocal duplications have been associated with microcephaly and macrocephaly, respectively. Given these findings and the high correlation between DUF1220 copy number and brain size across primate lineages (R² = 0.98; p = 1.8 × 10⁻⁶), DUF1220 sequences represent plausible candidates for underlying 1q21-associated brain-size pathologies. To investigate this possibility, we used specialized bioinformatics tools developed for scoring highly duplicated DUF1220 sequences to implement targeted 1q21 array comparative genomic hybridization on individuals (n = 42) with 1q21-associated microcephaly and macrocephaly. We show that of all the 1q21 genes examined (n = 53), DUF1220 copy number shows the strongest association with brain size among individuals with 1q21-associated microcephaly, particularly with respect to the three evolutionarily conserved DUF1220 clades CON1(p = 0.0079), CON2 (p = 0.0134), and CON3 (p = 0.0116). Interestingly, all 1q21 DUF1220-encoding genes belonging to the NBPF family show significant correlations with frontal-occipital-circumference Z scores in the deletion group. In a similar survey of a nondisease population, we show that DUF1220 copy number exhibits the strongest correlation with brain gray-matter volume (CON1, p = 0.0246; and CON2, p = 0.0334). Notably, only DUF1220 sequences are consistently significant in both disease and nondisease populations. Taken together, these data strongly implicate the loss of DUF1220 copy number in the etiology of 1q21-associated microcephaly and support the view that DUF1220 domains function as general effectors of evolutionary, pathological, and normal variation in brain size.