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Breakdown of the different tumor types among double-PIK3CA mutant samples based on their clonality and cis/trans status.

Purpose:To comprehensively characterize tissue-specific and molecular subclasses of multiple PIK3CA (multi-PIK3CA) mutations and assess their impact on potential therapeutic outcomes.Experimental Design:We profiled a pan-cancer cohort comprised of 352,392 samples across 66 tumor types using a targeted hybrid capture-based next-generation sequencing panel covering at least 324 cancer-related genes. Molecularly defined subgroups, allelic configuration, clonality, and mutational signatures were identified and tested for association with PI3K inhibitor therapeutic response.Results:Multi-PIK3CA mutations are found in 11% of all PIK3CA-mutant tumors, including 9% of low tumor mutational burden (TMB) PIK3CA-mutant tumors, and are enriched in breast and gynecologic cancers. Multi-PIK3CA mutations are frequently clonal and in cis on the same allele and occur at characteristic positions across tumor types. These mutations tend to be mutually exclusive of mutations in other driver genes, and of genes in the PI3K pathway. Among PIK3CA-mutant tumors with a high TMB, 18% are multi-PIK3CA mutant and often harbor an apolipoprotein B mRNA-editing enzyme, catalytic polypeptide (APOBEC) mutational signature. Despite large differences in specific allele combinations comprising multi-PIK3CA mutant tumors, especially across cancer types, patients with different classes of multi-PIK3CA mutant estrogen receptor–positive, HER2-negative breast cancers respond similarly to PI3K inhibition.Conclusions:Our pan-tumor study provides biological insights into the genetic heterogeneity and tissue specificities of multi-PIK3CA mutations, with potential clinical utility to guide PI3K inhibition strategies.

Table S1: Clinico-demographic characteristics of single- and multi-PIK3CA mutant tumors; Table S2: Prevalence of single and multi-PIK3CA mutated tumors across all tumor types; Table S3: Prevalence of single and multi-PIK3CA mutated tumors across pan-tumor histological subtypes; Table S4: Prevalence of single and multi-PIK3CA mutated tumors in different receptor subgroups of breast cancer; Table S5: Clonality predictions for double PIK3CA mutated tumors in different tumor types; Table S6: Prediction of cis/trans orientation for double PIK3CA mutated tumors in different tumor types; Table S7: Patterns of co-occurrence and mutual exclusivity between multi-PIK3CA mutations and other gene alterations in the overall cohort and specific tumor types; Table S8: Comparison of co-occurring alterations in multi-PIK3CA and single-PIK3CA mutant tumors; Table S9: Prevalence of multi-PIK3CA mutations based on status of tumor mutational burden; Table S10: Breakdown of PIK3CA mutations in different classes of mutational signatures in patients with a high tumor mutational burden; Table S11: Prevalence of single- and multi-PIK3CA mutations based on tumor mutational burden and APOBEC signatures; Table S12: Prevalence of specific pairs of PIK3CA mutations across different tumor types; Table S13: Prevalence of specific pairs of PIK3CA mutations at the codon level, across different tumor types; Table S14: Codon enrichment between single and multi-PIK3CA mutant tumors across different tumor types.

Purpose:To comprehensively characterize tissue-specific and molecular subclasses of multiple PIK3CA (multi-PIK3CA) mutations and assess their impact on potential therapeutic outcomes.Experimental Design:We profiled a pan-cancer cohort comprised of 352,392 samples across 66 tumor types using a targeted hybrid capture-based next-generation sequencing panel covering at least 324 cancer-related genes. Molecularly defined subgroups, allelic configuration, clonality, and mutational signatures were identified and tested for association with PI3K inhibitor therapeutic response.Results:Multi-PIK3CA mutations are found in 11% of all PIK3CA-mutant tumors, including 9% of low tumor mutational burden (TMB) PIK3CA-mutant tumors, and are enriched in breast and gynecologic cancers. Multi-PIK3CA mutations are frequently clonal and in cis on the same allele and occur at characteristic positions across tumor types. These mutations tend to be mutually exclusive of mutations in other driver genes, and of genes in the PI3K pathway. Among PIK3CA-mutant tumors with a high TMB, 18% are multi-PIK3CA mutant and often harbor an apolipoprotein B mRNA-editing enzyme, catalytic polypeptide (APOBEC) mutational signature. Despite large differences in specific allele combinations comprising multi-PIK3CA mutant tumors, especially across cancer types, patients with different classes of multi-PIK3CA mutant estrogen receptor–positive, HER2-negative breast cancers respond similarly to PI3K inhibition.Conclusions:Our pan-tumor study provides biological insights into the genetic heterogeneity and tissue specificities of multi-PIK3CA mutations, with potential clinical utility to guide PI3K inhibition strategies.

Understanding the differences in biomarker prevalence that may exist among diverse populations is invaluable to accurately forecast biomarker-driven clinical trial enrollment metrics and to advance inclusive research and health equity. This study evaluated the frequency and types ofAnalyses were conducted using real-world genomic data from adult patients with BC treated in an academic or community setting in the United States and whose tumor tissue was submitted for comprehensive genomic profiling.Of 36,151 patients with BC (median age, 58 years; 99% female), the breakdown by predicted genetic ancestry was 75% European, 14% African, 6% Central/South American, 3% East Asian, and 1% South Asian. We demonstrated that patients of African ancestry are less likely to have tumors that harborThis analysis established that tumor

PURPOSE Understanding the differences in biomarker prevalence that may exist among diverse populations is invaluable to accurately forecast biomarker-driven clinical trial enrollment metrics and to advance inclusive research and health equity. This study evaluated the frequency and types of PIK3CA mutations ( PIK3CAmut) detected in predicted genetic ancestry subgroups across breast cancer (BC) subtypes. METHODS Analyses were conducted using real-world genomic data from adult patients with BC treated in an academic or community setting in the United States and whose tumor tissue was submitted for comprehensive genomic profiling. RESULTS Of 36,151 patients with BC (median age, 58 years; 99% female), the breakdown by predicted genetic ancestry was 75% European, 14% African, 6% Central/South American, 3% East Asian, and 1% South Asian. We demonstrated that patients of African ancestry are less likely to have tumors that harbor PIK3CAmut compared with patients of European ancestry with estrogen receptor–positive/human epidermal growth factor receptor 2–negative (ER+/HER2–) BC (37% [949/2,593] v 44% [7,706/17,637]; q = 4.39E-11) and triple-negative breast cancer (8% [179/2,199] v 14% [991/7,072]; q = 6.07E-13). Moreover, we found that PIK3CAmut were predominantly composed of hotspot mutations, of which mutations at H1047 were the most prevalent across BC subtypes (35%-41% ER+/HER2– BC; 43%-61% HER2+ BC; 40%-59% triple-negative breast cancer). CONCLUSION This analysis established that tumor PIK3CAmut prevalence can differ among predicted genetic ancestries across BC subtypes on the basis of the largest comprehensive genomic profiling data set of patients with cancer treated in the United States. This study highlights the need for equitable representation in research studies, which is imperative to ensuring better health outcomes for all.

Background Mutations in p110α, encoded by PIK3CA, are present in ~40% of HR+/HER2- BCs. GDC-0077, a PI3Kα-selective inhibitor and mutant PI3Kα degrader, elicits antitumor activity in PIK3CAmut preclinical models as a single agent and when combined with endocrine therapy (ET). New evidence suggests BCs harboring multiple PIK3CAmut exhibit increased signaling through the PI3K/AKT pathway and are more sensitive to PI3Kα inhibitors compared with BCs with a single PIK3CAmut. We report a preliminary analysis of PIK3CAmut status with clinical outcomes from an ongoing study of GDC-0077 alone or with ET (letrozole/fulvestrant) ± palbociclib (palbo) in pts with PIK3CAmut HR+/HER2- mBC (NCT03006172). Methods Detectable PIK3CAmut from local tumor tissue/blood-based assay or tumor tissue by cobas PIK3CA assay were required to enroll. Plasma-derived circulating tumor (ct) DNA was collected at baseline (BL), cycle 1 day 15 (C1D15), and C2D1 (in the cohort where GDC-0077 starts at C1D15) to detect PIK3CAmut. Paired tumor samples were analyzed for Ki67 and pAKT/pS6 expression by immunohistochemistry. Single vs multiple PIK3CAmut was correlated with the percentage of pharmacodynamic (PD) inhibition of Ki67/pAKT/pS6 expression; with the PIK3CAmut allele frequency ratio between BL and C1D15 or C2D1 (MAFr15); with best overall response (BOR, RECIST v1.1); and with time on treatment (TOT) in days. Statistical analyses: Kruskal-Wallis and Mann-Whitney-Wilcoxon for group and pairwise comparisons, respectively, and two-sample proportion testing for categorical comparisons. Results Data cutoff was 03/20/2020. PIK3CAmut were detected in 87/103 (84.5%) pts with BL ctDNA available for sequencing. Multiple PIK3CAmut were detected in 21/87 (24.1%) BL ctDNA samples: 9 from pts treated with single-agent GDC-0077; 8 from pts treated with GDC-0077 + letrozole/fulvestrant; and 4 from pts treated with GDC-0077 + letrozole/fulvestrant + palbo. The median number of lines of prior therapy for metastatic disease was not different between pts with multiple (3.0 lines) vs single (2.5 lines) PIK3CAmut detected at BL (p = 0.205). Median percentage inhibition of Ki67/pAKT/pS6 expression was greater in pts with multiple (-65.8, -70.3, -66.8%, respectively) vs single (-42.1, -34.1, -29.5%) PIK3CAmut detected at BL (p = 0.095, 0.002, 0.056). Median MAFr15 was lower in pts with multiple (MAFr15 0.01) vs single PIK3CAmut (MAFr15 0.15) detected at BL (p = 0.004). Of 73 pts with both BL ctDNA-detected PIK3CAmut and BOR data, 16/16 (100%) with multiple PIK3CAmut experienced BOR of partial response (PR) or stable disease (SD) while 42/57 (73.7%) with single PIK3CAmut experienced BOR of PR or SD (p = 0.051). No pts with multiple PIK3CAmut detected experienced a BOR of progressive disease. Median TOT was greater in pts with multiple PIK3CAmut (196 days) vs single PIK3CAmut (140.5 days) detected at BL, but this was not significant (p = 0.1804). Conclusions The fraction of pts in which multiple PIK3CAmut were identified from BL ctDNA in this HR+/HER2- mBC dataset (24.1%) was slightly higher than reported elsewhere. This may be due to the method of detection (blood vs tissue) and/or the definition of multiple PIK3CAmut used. Pts in which multiple PIK3CAmut were detected by ctDNA exhibited greater depth of PD biomarker inhibition in tumors and experienced PR/SD more often compared with pts in which only one PIK3CAmut was detected. However, no significant associations were observed with the number of prior lines of therapy for metastatic disease or TOT. The dataset is currently too small to assess the impact of different treatment regimens in this study but will be re-evaluated as the data mature. Citation Format: Komal Jhaveri, Dejan Juric, Andrea Varga, Nicolas Turner, Peter Schmid, Cristina Saura, Mafalda Oliveira, Ian E Krop, Kevin Kalinsky, Antoine Italiano, Erika Hamilton, Valentina Gambardella, Andrés Cervantes, Philippe L Bedard, Bonnie P Liu, Jessica W Chen, Junko Aimi, Stephanie Royer-Joo, Jennifer L Schutzman, Katherine E Hutchinson. Preliminary correlative analysis of clinical outcomes with PIK3CA mutation (mut) status from a phase I/Ib study of GDC-0077 in patients (pts) with hormone receptor-positive/HER2-negative metastatic breast cancer (HR+/HER2- mBC) [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-12.

Preeclampsia is a multi-organ complication of pregnancy characterized by sudden hypertension and proteinuria that is among the leading causes of preterm delivery and maternal morbidity and mortality worldwide. The heterogeneity of preeclampsia poses a challenge for understanding its etiology and molecular basis. Intriguingly, risk for the condition increases in high-altitude regions such as the Peruvian Andes. To investigate the genetic basis of preeclampsia in a population living at high altitude, we characterized genome-wide variation in a cohort of preeclamptic and healthy Andean families (n = 883) from Puno, Peru, a city located above 3,800 meters of altitude. Our study collected genomic DNA and medical records from case-control trios and duos in local hospital settings. We generated genotype data for 439,314 SNPs, determined global ancestry patterns, and mapped associations between genetic variants and preeclampsia phenotypes. A transmission disequilibrium test (TDT) revealed variants near genes of biological importance for placental and blood vessel function. The top candidate region was found on chromosome 13 of the fetal genome and contains clotting factor genes PROZ, F7, and F10. These findings provide supporting evidence that common genetic variants within coagulation genes play an important role in preeclampsia. A selection scan revealed a potential adaptive signal around the ADAM12 locus on chromosome 10, implicated in pregnancy disorders. Our discovery of an association in a functional pathway relevant to pregnancy physiology in an understudied population of Native American origin demonstrates the increased power of family-based study design and underscores the importance of conducting genetic research in diverse populations.

Currently available mRNA vaccines are extremely safe and effective to prevent severe SARS-CoV-2 infections. However, the emergence of novel variants of concerns has highlighted the importance of high population-based vaccine rates to effectively suppress viral transmission and breakthrough infections. While initially left out from vaccine efforts, children have become one of the most affected age groups and are key targets to stop community and household spread. Antibodies are central for vaccine induced protection and emerging data points to the importance of additional Fc effector functions like opsononophagocytosis or cytotoxicity, particularly in the context of variants of concern that escape neutralizing antibodies. Here, we observed delayed induction and reduced magnitude of vaccine induced antibody titers in children 5-11 years receiving two doses of the age recommended 10 μg dose of the Pfizer SARS-CoV-2 BNT162b2 vaccine compared to adolescents (12-15 years) or adults receiving the 30 μg dose. Conversely, children mounted equivalent or more robust neutralization and opsonophagocytic functions at peak immunogenicity, pointing to a qualitatively more robust humoral functional response in children. Moreover, broad cross-variants of concern responses were observed across children, with enhanced IgM and parallel IgG cross-reactivity to variants of concern (VOCs) in children compared to adults. Collectively, these data argue that despite the lower magnitude of the BNT162b2 induced antibody response in children, vaccine induced immunity in children target VOCs broadly and exhibit enhanced functionality that may contribute to attenuation of disease.

Recoding viral genomes by introducing numerous synonymous nucleotide substitutions that create suboptimal codon pairs provides new live-attenuated vaccine candidates. Because recoding typically involves a large number of nucleotide substitutions, the risk of de-attenuation is presumed to be low. However, this has not been thoroughly studied. We previously generated human respiratory syncytial virus (RSV) in which the NS1, NS2, N, P, M and SH ORFs were codon-pair deoptimized (CPD) by 695 synonymous nucleotide changes (Min A virus). Min A exhibited a global reduction in transcription and protein synthesis, was restricted for replication in vitro and in vivo, and exhibited moderate temperature sensitivity. Here, we show that under selective pressure by serial passage at progressively increasing temperatures, Min A regained replication fitness and lost its temperature sensitivity. Whole-genome deep sequencing identified numerous missense mutations in several genes, in particular ones accumulating between codons 25 and 34 of the phosphoprotein (P), a polymerase cofactor and chaperone. When re-introduced into Min A, these P mutations restored viral transcription to wt level, resulting in increased protein expression and RNA replication. Molecular dynamic simulations suggested that these P mutations increased the flexibility of the N-terminal domain of P, which might facilitate its interaction with the nucleoprotein N, and increase the functional efficiency of the RSV transcription/replication complex. Finally, we evaluated the effect of the P mutations on Min A replication and immunogenicity in hamsters. Mutation P[F28V] paradoxically reduced Min A replication but not its immunogenicity. The further addition of one missense mutation each in M and L generated a version of Min A with increased genetic stability. Thus, this study provides further insight into the adaptability of large-scale recoded RNA viruses under selective pressure and identified an improved CPD RSV vaccine candidate., Author summary Synonymous recoding of viral genomes by codon-pair deoptimization (CPD) generates live-attenuated vaccines presumed to be genetically stable due to the high number of nucleotide substitutions. However, their actual genetic stability under selective pressure was largely unknown. In a recoded human respiratory syncytial virus (RSV) mutant called Min A, six of 11 ORFs were CPD, reducing protein expression and inducing moderate temperature sensitivity and attenuation. When passaged in vitro under selective pressure, Min A lost its temperature-sensitive phenotype and regained fitness by the acquisition of numerous mutations, in particular missense mutations in the viral phosphoprotein (P), a polymerase cofactor and a chaperone for soluble nucleoprotein. These P mutations increased RSV gene transcription globally, thereby increasing RSV protein expression, RNA replication, and virus particle production. Thus, the P mutations increased the efficiency of the RSV transcription/replication complex, compensating for the reduced protein expression due to CPD. In addition, introduction of the P mutations into Min A generated a live-attenuated vaccine candidate with increased genetic stability. Surprisingly, this vaccine candidate exhibited increased attenuation and, paradoxically, exhibited increased immunogenicity per plaque-forming unit in hamsters. This study provides insights into the adaptability of large-scale recoded RNA viruses and identified an improved CPD RSV vaccine candidate.

Background: Mutations in PIK3CA, encoding the catalytic subunit p110α of PI3K, are present in ~40% of ER+/HER- BC. Taselisib (TAS) is a potent and selective β-sparing PI3K inhibitor that improves outcomes in combination with endocrine therapy in pts with PIK3CAmut advanced breast cancer (aBC). To better understand the molecular alterations associated with response to PI3K inhibition, we profiled the genomic landscape of ctDNA collected from PIK3CAmut, ER+, HER2- aBC pts immediately prior to treatment (tx) with TAS or placebo (PBO) plus fulvestrant (FUL). Methods: Pts were enrolled as part of a phase III randomized study of TAS or PBO plus FUL in ER+, HER2- PIK3CAmut aBC (SANDPIPER, NCT02340221). Pre-tx ctDNA samples from 508 pts underwent comprehensive genomic profiling using the FoundationOne Liquid NGS assay at Foundation Medicine, Inc.; the 339 pts with PIK3CAmut ctDNA were further analyzed herein. The top mutated genes were analyzed for prognostic value against investigator-assessed progression-free survival (PFS) for both tx regimens using Cox proportional hazards regression modeling. As analysis was exploratory, no adjustments were made for multiple testing. Results: The top altered genes were TP53 (44%), ESR1 (37%), CDH1 (17%), FGFR1 (12%), NF1 (11%), CHEK2 (10%), and PTEN (9%). In pts treated with PBO+FUL, alterations in PTEN (HR 2.8; 95% CI 1.4-5.7; p=0.0107) and TP53 (HR 2.0; 95% CI 1.3-3.1; p=0.0025) were associated with a worse prognosis compared to pts with no mutation detected (NMD) in these genes. In pts treated with TAS+FUL, alterations in FGFR1 (HR 2.4; 95% CI 1.5-3.7; p=0.0006), TP53 (HR 1.9; 95% CI 1.4-2.6; p=0.0001) and PTEN (HR 1.8; 95% CI 1.1-2.8; p=0.0265) were associated with a worse prognosis compared to pts with NMD in these genes. Alterations in ESR1, CDH1, or CHEK2 were not associated with prognosis (p≥0.05) in either tx arm. A trend towards worse prognosis was observed in pts with NF1 altered ctDNA treated with PBO+FUL (HR 2.1; 95% CI 1.1-4.1; p=0.0527), which was not observed in pts treated with TAS+FUL (HR 0.97; 95% CI 0.57-1.65; p=0.901). Within the NF1-altered subgroup, a significant PFS difference was observed between TAS- vs PBO-treated pts (HR 0.28; 95% CI 0.11-0.67; p=0.0058; median 5.65 vs 1.94 months, respectively). Conclusions: We report that the most frequently mutated genes identified are consistent with previous studies in pts with ER+, HER2- aBC. This analysis shows that alterations in TP53 and PTEN were associated with poor prognosis in both tx arms, and FGFR1 alterations were associated with a poor prognosis in TAS+FUL treated pts. Further, NF1 alterations were associated with a poor prognosis in PBO+FUL treated pts, an association that was not observed with TAS+FUL. These findings may inform future rational combination strategies for the clinical development of PI3K inhibitors. Citation Format: Jessica W. Chen, Susan Dent, William Jacot, Javier Cortés, Ian E. Krop, Thomas J. Stout, Frauke Schimmoller, Heidi M. Savage, Katherine E. Hutchinson, Timothy R. Wilson. The genomic landscape and prognostic implications of somatic alterations in patients (pts) with ER+, HER2-, PIK3CA mutated (mut) advanced breast cancer treated with taselisib and fulvestrant [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5165.

Molecular profiling of tumor specimens is a key contributor in the application of precision medicine toward patient care in oncology. The presence of genetic mutational data may provide insight into the etiology of the cancer and inform about the available therapeutic options for patients diagnosed with various cancer types. However, the ability to centrally organize, structure, and visualize this genetic data can be hindered by the lack of necessary computational infrastructure. Here we present a somatic tumor data visualization portal titled STAMPede that utilizes tumor sequencing data from an in-house solid tumor oncology sequencing panel at Stanford Health Care. STAMPede is intended to provide Stanford healthcare providers and clinical researchers an easy to navigate web-based portal to query and display gene-, variant-, and cancer-level summary statistics.

Successful implementation of precision oncology requires both the deployment of nucleic acid sequencing panels to identify clinically actionable biomarkers, and the efficient screening of patient biomarker eligibility to on-going clinical trials and therapies. This process is typically performed manually by biocurators, geneticists, pathologists, and oncologists; however, this is a time-intensive, and inconsistent process amongst healthcare providers. We present the development of a feature matching algorithmic pipeline that identifies patients who meet eligibility criteria of precision medicine clinical trials via genetic biomarkers and apply it to patients undergoing treatment at the Stanford Cancer Center. This study demonstrates, through our patient eligibility screening algorithm that leverages clinical sequencing derived biomarkers with precision medicine clinical trials, the successful use of an automated algorithmic pipeline as a feasible, accurate and effective alternative to the traditional manual clinical trial curation.

Successful implementation of precision oncology requires both the deployment of nucleic acid sequencing panels to identify clinically actionable biomarkers, and the efficient screening of patient biomarker eligibility to on-going clinical trials and therapies. This process is typically performed manually by biocurators, geneticists, pathologists, and oncologists; however, this is a time-intensive, and inconsistent process amongst healthcare providers. We present the development of a feature matching algorithmic pipeline that identifies patients who meet eligibility criteria of precision medicine clinical trials via genetic biomarkers and apply it to patients undergoing treatment at the Stanford Cancer Center. This study demonstrates, through our patient eligibility screening algorithm that leverages clinical sequencing derived biomarkers with precision medicine clinical trials, the successful use of an automated algorithmic pipeline as a feasible, accurate and effective alternative to the traditional manual clinical trial curation.

Tendons and ligaments are critical components of the musculoskeletal system, yet the pathways specifying this lineage remain poorly defined. Through a screen of known bioactive chemicals in zebrafish, we identified a new pathway regulating tendon cell induction. We established that statin, through inhibition of the mevalonate pathway, causes an expansion of the tendon progenitor population. Co-expression and live imaging studies indicate that the expansion does not involve an increase in cell proliferation, but rather results from re-specification of cells from the neural crest-derived sox10+/sox9a+ skeletal lineage. The effect on tendon cell expansion is specific to the geranylgeranylation branch of the mevalonate pathway and mediated by inhibition of Rac activity. This work establishes a novel role for the mevalonate pathway and Rac activity in regulating specification of the tendon lineage.

Despite the importance of tendons and ligaments for transmitting movement and providing stability to the musculoskeletal system, their development is considerably less well understood than that of the tissues they serve to connect. Zebrafish have been widely used to address questions in muscle and skeletal development, yet few studies describe their tendon and ligament tissues. We have analyzed in zebrafish the expression of several genes known to be enriched in mammalian tendons and ligaments, including scleraxis (scx), collagen 1a2 (col1a2) and tenomodulin (tnmd), or in the tendon-like myosepta of the zebrafish (xirp2a). Co-expression studies with muscle and cartilage markers demonstrate the presence of scxa, col1a2 and tnmd at sites between the developing muscle and cartilage, and xirp2a at the myotendinous junctions. We determined that the zebrafish craniofacial tendon and ligament progenitors are neural crest derived, as in mammals. Cranial and fin tendon progenitors can be induced in the absence of differentiated muscle or cartilage, although neighboring muscle and cartilage are required for tendon cell maintenance and organization, respectively. By contrast, myoseptal scxa expression requires muscle for its initiation. Together, these data suggest a conserved role for muscle in tendon development. Based on the similarities in gene expression, morphology, collagen ultrastructural arrangement and developmental regulation with that of mammalian tendons, we conclude that the zebrafish tendon populations are homologous to their force-transmitting counterparts in higher vertebrates. Within this context, the zebrafish model can be used to provide new avenues for studying tendon biology in a vertebrate genetic system.