Search

Human breast cancer (BRCA) shows tremendous genomic, gene expression, clinical, and phenotypic heterogeneity. Known driver gene alterations can only explain a portion of this heterogeneity, some of which likely arise from variation in the target cell for transformation, in addition to incompletely understood gene copy number and epigenetic alterations. These factors are difficult to identify with certainty using human patient samples due to widely varying germline genetic backgrounds, thousands of gene copy and epigenetic changes per sample, and, unknown target cell transformation. Activating mutations in the p110α catalytic subunit of PI3K are one of the most common genetic alterations in human BRCA. Here, we report results from two Sleeping Beauty (SB) transposon-accelerated mouse models of Pik3ca-mutant mammary cancer showing how genotype-phenotype correlations can be drawn providing strong candidates for mediating tumor phenotypes, including estrogen-receptor (ER)-dependent gene expression, high cell cycle activity, and immune cell exclusion. We used SB transposon mutagenesis in mice on a Pik3caH1047R activated mutant background to model mammary cancer development in two different mammary epithelial compartments. Both the target cell for mutagenesis and the specific transposon-induced mutations correlated with specific tumor phenotypes, including whether the tumors were ER positive or negative. RNA sequencing of tumors revealed novel genotype-phenotype correlations implicating specific transposon-altered gene drivers of high cell cycle activity, ER-dependent gene expression, and white blood cell exclusion from the tumor. Many transposon-implicated genes are altered at the gene copy number or epigenetic/methylation level in human BRCA, and several were functionally validated. These models provide a source of genetically heterogenous mouse mammary tumors with a uniform initiating mutation, Pik3caH1047R, useful for identifying cooperating pathways and drivers of specific tumor phenotypes. Citation Format: Morito Kurata, Emiily Pope, Jingmin Shu, Wenlin Yuan, Wendy Hudson, Mark Sokolowski, Setareh Bagherzadeh, Zora Modrusan, Eric Stawiski, Steffen Durinck, Sekar Seshigiri, Aaron Sarver, Nuri Temiz, David Largaespada. Discovery of cancer genes and pathways operative in PI3K-activated mammary cancer reveals clinically relevant genotype-phenotype correlations. [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-08-04.

The kidney is a complex organ that governs many physiological parameters. It is roughly divided into three parts, the renal pelvis, medulla, and cortex. Covering the cortex is the renal capsule, a serosal tissue that provides protection and forms a barrier for the kidney. Serosal tissues of many organs have been recently shown to play a vital role in homeostasis and disease. Analyses of the cells that reside in these tissues have identified distinct cell types with unique phenotypes. Surprisingly, despite the importance of serosal tissues, little is known about cells of the renal capsule. Here, we characterized this niche and found that it is mainly comprised of fibroblasts and macrophages, but also includes many other diverse cell types. Characterizing renal capsule-associated macrophages, we found that they consist of a distinct subset (i.e., TLF+macrophages) that is nearly absent in the kidney parenchyma. Injury, disease, and other changes within the kidney, affected the cell composition and phenotype of the renal capsule, indicating its dynamic and vibrant response to changes within the organ parenchyma. Lastly, we studied age-related changes in the renal capsule and found that aging affected the cell composition and pro-inflammatory phenotype of macrophages, increased CD8 T cells and other lymphocyte counts, and promoted a senescence-associated phenotype in fibroblasts. Taken together, our data illustrate the complexity and heterogeneity of the renal capsule and its underlying changes during aging and disease, improving our understanding of the kidney serosa that may be valuable for novel renal therapies.

By leveraging tumorgraft (patient-derived xenograft) RNA-sequencing data, we developed an empirical approach, DisHet, to dissect the tumor microenvironment (eTME). We found that 65% of previously defined immune signature genes are not abundantly expressed in renal cell carcinoma (RCC) and identified 610 novel immune/stromal transcripts. Using eTME, genomics, pathology, and medical record data involving >1,000 patients, we established an inflamed pan-RCC subtype (IS) enriched for regulatory T cells, natural killer cells, TH1 cells, neutrophils, macrophages, B cells, and CD8+ T cells. IS is enriched for aggressive RCCs, including BAP1-deficient clear-cell and type 2 papillary tumors. The IS subtype correlated with systemic manifestations of inflammation such as thrombocytosis and anemia, which are enigmatic predictors of poor prognosis. Furthermore, IS was a strong predictor of poor survival. Our analyses suggest that tumor cells drive the stromal immune response. These data provide a missing link between tumor cells, the TME, and systemic factors.Significance: We undertook a novel empirical approach to dissect the renal cell carcinoma TME by leveraging tumorgrafts. The dissection and downstream analyses uncovered missing links between tumor cells, the TME, systemic manifestations of inflammation, and poor prognosis. Cancer Discov; 8(9); 1142–55. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 1047

Supplementary Figures S1-S2 and Supplementary Tables S2-S4.

Protein isoforms produced by alternative splicing (AS) of many genes have been implicated in several aspects of cancer genesis and progression. These observations motivated a genome-wide assessment of AS in breast cancer. We accomplished this by measuring exon level expression in 31 breast cancer and nonmalignant immortalized cell lines representing luminal, basal, and claudin-low breast cancer subtypes using Affymetrix Human Junction Arrays. We analyzed these data using a computational pipeline specifically designed to detect AS with a low false-positive rate. This identified 181 splice events representing 156 genes as candidates for AS. Reverse transcription-PCR validation of a subset of predicted AS events confirmed 90%. Approximately half of the AS events were associated with basal, luminal, or claudin-low breast cancer subtypes. Exons involved in claudin-low subtype–specific AS were significantly associated with the presence of evolutionarily conserved binding motifs for the tissue-specific Fox2 splicing factor. Small interfering RNA knockdown of Fox2 confirmed the involvement of this splicing factor in subtype-specific AS. The subtype-specific AS detected in this study likely reflects the splicing pattern in the breast cancer progenitor cells in which the tumor arose and suggests the utility of assays for Fox-mediated AS in cancer subtype definition and early detection. These data also suggest the possibility of reducing the toxicity of protein-targeted breast cancer treatments by targeting protein isoforms that are not present in limiting normal tissues. Mol Cancer Res; 8(7); 961–74. ©2010 AACR.

Supplementary Table S1.

Supplementary Figure 1 from Temporal Dissection of Tumorigenesis in Primary Cancers

Supplementary Figure 2 from Temporal Dissection of Tumorigenesis in Primary Cancers

Supplementary Figure Legends 1-4, Methods from Temporal Dissection of Tumorigenesis in Primary Cancers

List of UTSW KCP patients with RNA-Seq and exome-seq. Sheet 1 contains information of 35 patients whose matched trio RNA-Seq data were used in the primary DisHet analyses. Sheet 2 contains information of patients whose bulk tumors were analyzed by RNA-Seq and whose follow up data are available. There are overlap between patients from these two sheets. Sheet 3 contains the information of tumorgraft data used in Fig. 5bc.

The full technical details of DisHet

Annotation of eTME genes. eTME genes (3x) are shown in the table. The "eTME (20x)" column shows eTME genes that satisfy the 20x cutoff. The other columns show the interpretation of the eTME genes using BLUEPRINT data, the Winslow signatures, Immunome signatures, and the ESTIMATE signature (immune and stromal signatures combined).

Supplementary Table 2 from Temporal Dissection of Tumorigenesis in Primary Cancers

Supplementary Figure 3 from Temporal Dissection of Tumorigenesis in Primary Cancers

Supplementary Figure 4 from Temporal Dissection of Tumorigenesis in Primary Cancers

Gene ontology analysis shows enriched GO terms of the 904 eTME genes (20x cutoff) (first sheet), the 778 and the 610 eTME genes (20x cutoff) that are not captured by the Immunome alone and Immunome+ESTIMATE+Winslow gene signatures (second and third sheets).

Supplementary Table 1 from Temporal Dissection of Tumorigenesis in Primary Cancers

Sup. Fig. 1 Simulation analyses show accurate dissection of bulk tumor RNA-Seq data by DisHet. Sup. Fig. 2 Validating immune/stroma component expression dissected by DisHet. Sup. Fig. 3 Comparison of dissection performance of DeMix and DisHet. Sup. Fig. 4 Principal Component Analysis (PCA) shows the clustering of TCGA pan-RCC patients tumors by the tumor-specific genes and eTME-specific genes, respectively. Sup. Fig. 5 CD4 and CD8 immunohistochemistry staining in the Six2-Cre;VhlF/F;Pbrm1F/F and Six2-Cre;VhlF/F;Bap1F/+. Supl. Fig. 6 Clinical manifestations of eTME-NIS patients.

Purpose: microRNAs have been shown to be involved in different human cancers. We therefore have performed expression profiles on a panel of pediatric tumors to identify cancer-specific microRNAs. We also investigated if microRNAs are coregulated with their host gene.Experimental Design: We performed parallel microRNAs and mRNA expression profiling on 57 tumor xenografts and cell lines representing 10 different pediatric solid tumors using microarrays. For those microRNAs that map to their host mRNA, we calculated correlations between them.Results: We found that the majority of cancer types clustered together based on their global microRNA expression profiles by unsupervised hierarchical clustering. Fourteen microRNAs were significantly differentially expressed between rhabdomyosarcoma and neuroblastoma, and 8 of them were validated in independent patient tumor samples. Exploration of the expression of microRNAs in relationship with their host genes showed that the expression for 43 of 68 (63%) microRNAs located inside known coding genes was significantly correlated with that of their host genes. Among these 43 microRNAs, 5 of 7 microRNAs in the OncomiR-1 cluster correlated significantly with their host gene MIRHG1 (P < 0.01). In addition, high expression of MIRHG1 was significantly associated with high stage and MYCN amplification in neuroblastoma tumors, and the expression level of MIRHG1 could predict the outcome of neuroblastoma patients independently from the current neuroblastoma risk-stratification in two independent patient cohorts.Conclusion: Pediatric cancers express cancer-specific microRNAs. The high expression of the OncomiR-1 host gene MIRHG1 correlates with poor outcome for patients with neuroblastoma, indicating important oncogenic functions of this microRNA cluster in neuroblastoma biology. (Clin Cancer Res 2009;15(17):5560–8)

Supplementary Table S1: X-ray crystallography data processing and structure refinement statistics Supplementary Table S2: BRAF and RAS mutational status of cell lines used in the study Supplementary Table: S3: Cellular IC50 (nM) of parental and ERKi-resistant lines to ERK inhibitors Supplementary Table S4: Mutations found in ERKi- resistant lines. Supplementary Table-S5- Copy number (log2 ratio) in ERKi-R lines compare to parental cell lines. Supplementary Table S6: Gene expression (RPKM) in parental and ERKi-R cells

Purpose: MAPK pathway inhibitors targeting BRAF and MEK have shown clinical efficacy in patients with RAF- and/or RAS-mutated tumors. However, acquired resistance to these agents has been an impediment to improved long-term survival in the clinic. In such cases, targeting ERK downstream of BRAF/MEK has been proposed as a potential strategy for overcoming acquired resistance. Preclinical studies suggest that ERK inhibitors are effective at inhibiting BRAF/RAS-mutated tumor growth and overcome BRAF or/and MEK inhibitor resistance. However, as observed with other MAPK pathway inhibitors, treatment with ERK inhibitors is likely to cause resistance in the clinic. Here, we aimed to model the mechanism of resistance to ERK inhibitors.Experimental Design: We tested five structurally different ATP-competitive ERK inhibitors representing three different scaffolds on BRAF/RAS-mutant cancer cell lines of different tissue types to generate resistant lines. We have used in vitro modeling, structural biology, and genomic analysis to understand the development of resistance to ERK inhibitors and the mechanisms leading to it.Results: We have identified mutations in ERK1/2, amplification and overexpression of ERK2, and overexpression of EGFR/ERBB2 as mechanisms of acquired resistance. Structural analysis of ERK showed that specific compounds that induced on-target ERK mutations were impaired in their ability to bind mutant ERK. We show that in addition to MEK inhibitors, ERBB receptor and PI3K/mTOR pathway inhibitors are effective in overcoming ERK-inhibitor resistance.Conclusions: These findings suggest that combination therapy with MEK or ERBB receptor or PI3K/mTOR and ERK inhibitors may be an effective strategy for managing the emergence of resistance in the clinic. Clin Cancer Res; 24(16); 4044–55. ©2018 AACR.

Supplementary Data from microRNA Profiling Identifies Cancer-Specific and Prognostic Signatures in Pediatric Malignancies

Supplementary Material from In vitro differential sensitivity of melanomas to phenothiazines is based on the presence of codon 600 BRAF mutation

Supplemental legend

Fig. S1. ERK inhibitors and their effect on MAPK mutant cell lines. A. Structure of ERK inhibitors used in this study. B. Dose response curve of ERK inhibitors on the viability of A375, IPC298, SKMEL30, HCT116, MIA PaCa2 and Panc1 cells. Fig. S2. Western blot analysis of parental, VI-3-R and G994-R HCT116 (A) or MIA PaCa2 (B) cells treated with indicated ERK inhibitors. Fig. S3. Western blot analysis of HCT116 cells (A) and SKMEL30 (B) stably expressing indicated ERK1 or ERK2 mutants. Fig. S4. Copy number and expression analysis of MITF in SKMEL30-ERKi-R cells. A. MITF copy gain in SKMEL30-V11e-R, -VI-3-R and -G994-R. B. MITF copy number vs. expression in SKMEL30-V11e-R, VI-3-R and G994-R cells. Fig. S5. Dose response curve of ERK2 overexpressing IPC298 cells treated with MEK inhibitors GDC-0973 or AZD6244 (A) and ERK2 amplified IPC298-G994-R (B) or MIA PaCa2-S984-R (C) cells treated with indicated ERK-inhibitors from alternate scaffold class. Fig. S6. Dose response curve of parental or ERKi-resistant HCT116 (A) and MIA PaCa2 (B) cells treated with indicated inhibitors. The data shown are same as in Fig. 6B for HCT116 (A) and Fig. 6C for MIA PaCa2 (B) grouped by drug treatment.

Snakebite envenoming is a serious and neglected tropical disease that kills ~100,000 people annually. High-quality, genome-enabled comprehensive characterization of toxin genes will facilitate development of effective humanized recombinant antivenom. We report a de novo near-chromosomal genome assembly of Naja naja, the Indian cobra, a highly venomous, medically important snake. Our assembly has a scaffold N50 of 223.35 Mb, with 19 scaffolds containing 95% of the genome. Of the 23,248 predicted protein-coding genes, 12,346 venom-gland-expressed genes constitute the ‘venom-ome’ and this included 139 genes from 33 toxin families. Among the 139 toxin genes were 19 ‘venom-ome-specific toxins’ (VSTs) that showed venom-gland-specific expression, and these probably encode the minimal core venom effector proteins. Synthetic venom reconstituted through recombinant VST expression will aid in the rapid development of safe and effective synthetic antivenom. Additionally, our genome could serve as a reference for snake genomes, support evolutionary studies and enable venom-driven drug discovery., Analysis of a near-chromosomal genome assembly and transcriptome profiling of the Indian cobra identifies genes expressed in the venom glands. These data should help develop a new antivenom.

The underrepresentation of non-Europeans in human genetic studies so far has limited the diversity of individuals in genomic datasets and led to reduced medical relevance for a large proportion of the world’s population. Population-specific reference genome datasets as well as genome-wide association studies in diverse populations are needed to address this issue. Here we describe the pilot phase of the GenomeAsia 100K Project. This includes a whole-genome sequencing reference dataset from 1,739 individuals of 219 population groups and 64 countries across Asia. We catalogue genetic variation, population structure, disease associations and founder effects. We also explore the use of this dataset in imputation, to facilitate genetic studies in populations across Asia and worldwide., Using whole-genome sequencing data from 1,739 individuals, the GenomeAsia 100K Project catalogues genetic variation, population structure and disease associations to facilitate genetic studies in Asian populations and increase representation in genetics studies worldwide.

Metastasis is the principal cause of cancer related deaths. Tumor invasion is essential for metastatic spread. However, determinants of invasion are poorly understood. We addressed this knowledge gap by leveraging a unique attribute of kidney cancer. Renal tumors invade into large vessels forming tumor thrombi (TT) that migrate extending sometimes into the heart. Over a decade, we prospectively enrolled 83 ethnically-diverse patients undergoing surgical resection for grossly invasive tumors at UT Southwestern Kidney Cancer Program. In this study, we perform comprehensive histological analyses, integrate multi-region genomic studies, generate in vivo models, and execute functional studies to define tumor invasion and metastatic competence. We find that invasion is not always associated with the most aggressive clone. Driven by immediate early genes, invasion appears to be an opportunistic trait attained by subclones with diverse oncogenomic status in geospatial proximity to vasculature. We show that not all invasive tumors metastasize and identify determinants of metastatic competency. TT associated with metastases are characterized by higher grade, mTOR activation and a particular immune contexture. Moreover, TT grade is a better predictor of metastasis than overall tumor grade, which may have implications for clinical practice., Tumour thrombi (TT) are intravascular extensions of renal cell carcinomas (RCC) which often lead to distant metastases. Here the authors examine the determinants of vascular invasion and metastasis in a unique cohort of RCC patients with TT using multi-region genomics and in vivo models.

Significance Genetic heterogeneity is a significant driver of antibiotic resistance in bacteria. Understanding copy number (CN) heterogeneity is important because minority subclones with increased CN can drive resistance during antibiotic exposure, but revert and escape detection during clinical susceptibility testing. Despite its clinical relevance, CN variation has eluded quantification at single-molecule resolution. Here, we report nanopore sequencing of arylomycin-resistant mutants carrying tandem repeats ranging in size from 4.8 to 50.0 kb and encompassing the arylomycin target gene lepB. Reads spanning individual repeat arrays show vast differences in CN, underscoring the importance of amplifications in driving the emergence of genetic heterogeneity. This is a direct observation of cell-to-cell CN differences in an antibiotic-resistant bacterial population., Tandem gene amplification is a frequent and dynamic source of antibiotic resistance in bacteria. Ongoing expansions and contractions of repeat arrays during population growth are expected to manifest as cell-to-cell differences in copy number (CN). As a result, a clonal bacterial culture could comprise subpopulations of cells with different levels of antibiotic sensitivity that result from variable gene dosage. Despite the high potential for misclassification of heterogenous cell populations as either antibiotic-susceptible or fully resistant in clinical settings, and the concomitant risk of inappropriate treatment, CN distribution among cells has defied analysis. Here, we use the MinION single-molecule nanopore sequencer to uncover CN heterogeneity in clonal populations of Escherichia coli and Acinetobacter baumannii grown from single cells isolated while selecting for resistance to an optimized arylomycin, a member of a recently discovered class of Gram-negative antibiotic. We found that gene amplification of the arylomycin target, bacterial type I signal peptidase LepB, is a mechanism of unstable arylomycin resistance and demonstrate in E. coli that amplification instability is independent of RecA. This instability drives the emergence of a nonuniform distribution of lepB CN among cells with a range of 1 to at least 50 copies of lepB identified in a single clonal population. In sum, this remarkable heterogeneity, and the evolutionary plasticity it fuels, illustrates how gene amplification can enable bacterial populations to respond rapidly to novel antibiotics. This study establishes a rationale for further nanopore-sequencing studies of heterogeneous cell populations to uncover CN variability at single-molecule resolution.

By leveraging tumorgraft (patient-derived xenograft) RNA-sequencing data, we developed an empirical approach, DisHet, to dissect the tumor microenvironment (eTME). We found that 65% of previously defined immune signature genes are not abundantly expressed in renal cell carcinoma (RCC) and identified 610 novel immune/stromal transcripts. Using eTME, genomics, pathology, and medical record data involving >1,000 patients, we established an inflamed pan-RCC subtype (IS) enriched for regulatory T cells, natural killer cells, TH1 cells, neutrophils, macrophages, B cells, and CD8+ T cells. IS is enriched for aggressive RCCs, including BAP1-deficient clear-cell and type 2 papillary tumors. The IS subtype correlated with systemic manifestations of inflammation such as thrombocytosis and anemia, which are enigmatic predictors of poor prognosis. Furthermore, IS was a strong predictor of poor survival. Our analyses suggest that tumor cells drive the stromal immune response. These data provide a missing link between tumor cells, the TME, and systemic factors. Significance: We undertook a novel empirical approach to dissect the renal cell carcinoma TME by leveraging tumorgrafts. The dissection and downstream analyses uncovered missing links between tumor cells, the TME, systemic manifestations of inflammation, and poor prognosis. Cancer Discov; 8(9); 1142–55. ©2018 AACR. This article is highlighted in the In This Issue feature, p. 1047

A propensity for rewiring genetic and epigenetic regulatory networks, thus enabling sustained cell proliferation, suppression of apoptosis, and the ability to evade the immune system, is vital to cancer cell propagation. An increased understanding of how this is achieved is critical for identifying or improving therapeutic interventions. In this study, using acute myeloid leukemia (AML) human cell lines and a custom CRISPR/Cas9 screening platform, we identify the H3K9 methyltransferase SETDB1 as a novel, negative regulator of innate immunity. SETDB1 is overexpressed in many cancers, and loss of this gene in AML cells triggers desilencing of retrotransposable elements that leads to the production of double-stranded RNAs (dsRNAs). This is coincident with induction of a type I interferon response and apoptosis through the dsRNA-sensing pathway. Collectively, our findings establish a unique gene regulatory axis that cancer cells can exploit to circumvent the immune system.

Background Baboons are a widely used nonhuman primate model for biomedical, evolutionary, and basic genetics research. Despite this importance, the genomic resources for baboons are limited. In particular, the current baboon reference genome Panu_3.0 is a highly fragmented, reference-guided (i.e., not fully de novo) assembly, and its poor quality inhibits our ability to conduct downstream genomic analyses. Findings Here we present a de novo genome assembly of the olive baboon (Papio anubis) that uses data from several recently developed single-molecule technologies. Our assembly, Panubis1.0, has an N50 contig size of ∼1.46 Mb (as opposed to 139 kb for Panu_3.0) and has single scaffolds that span each of the 20 autosomes and the X chromosome. Conclusions We highlight multiple lines of evidence (including Bionano Genomics data, pedigree linkage information, and linkage disequilibrium data) suggesting that there are several large assembly errors in Panu_3.0, which have been corrected in Panubis1.0.

The cellular origin of sporadic pancreatic neuroendocrine tumors (PNETs) is obscure. Dysregulated hormone production and the absence of obvious driver mutations mask tumor origins although they presumably arise from endocrine cell lineages such as glucagon-producing alpha cells or insulin-expressing beta cells. A reciprocal feedback circuit involving pancreatic glucagon and hepatic amino acids regulates alpha cell mass such that loss of hepatic glucagon receptor (GCGR) signaling drives alpha cell hyperproliferation and tumor formation. This genetically-defined system is an ideal model for parsing the mechanisms of PNET origin and progression. Utilizing a combination of genetically engineered Gcgr knockout mice and GCGR-inhibiting antibodies, we found that elevated plasma amino acids drive reversion to a proliferative SLC38A5+ embryonic progenitor-like status. Chronic activation of this mTOR-dependent state drives tumor initiation and single cell profiling reveals tumors originate from a subpopulation of alpha cells. Profiling RNA, tumor signaling, and immune content of biopsies from rare human PNETs found in individuals with germ-line GCGR loss-of-function variants showed active mTOR and prominent tumor cell-associated expression of amino acid transporter SLC7A8. This work, for the first time, identifies metabolic control of an adult-to-progenitor alpha cell identity switch and shows that chronic maintenance of progenitor identity in the adult organ drives proliferation and tumor initiation.

Despite widespread use of CRISPR, comprehensive data on the frequency and impact of Cas9-mediated off-targets in modified rodents are limited. Here we present deep-sequencing data from 81 genome-editing projects on mouse and rat genomes at 1,423 predicted off-target sites, 32 of which were confirmed, and show that high-fidelity Cas9 versions reduced off-target mutation rates in vivo. Using whole-genome sequencing data from ten mouse embryos, treated with a single guide RNA (sgRNA), and from their genetic parents, we found 43 off-targets, 30 of which were predicted by an adapted version of GUIDE-seq.

Recent work has demonstrated that two archaic human groups (Neanderthals and Denisovans) interbred with modern humans and contributed to the contemporary human gene pool. These findings relied on the availability of high-coverage genomes from both Neanderthals and Denisovans. Here we search for evidence of archaic admixture from a worldwide panel of 1,667 individuals using an approach that does not require the presence of an archaic human reference genome. We find no evidence for archaic admixture in the Andaman Islands, as previously claimed, or on the island of Flores, where Homo floresiensis fossils have been found. However, we do find evidence for at least one archaic admixture event in sub-Saharan Africa, with the strongest signal in Khoesan and Pygmy individuals from Southern and Central Africa. The locations of these putative archaic admixture tracts are weighted against functional regions of the genome, consistent with the long-term effects of purifying selection against introgressed genetic material.