Exploiting Matrix Stiffness to Overcome Drug Resistance.

Drug resistance is arguably one of the biggest challenges facing cancer research today. Understanding the underlying mechanisms of drug resistance in tumor progression and metastasis are essential in developing better treatment modalities. Given the matrix stiffness affecting the mechanotransduction capabilities of cancer cells, characterization of the related signal transduction pathways can provide a better understanding for developing novel therapeutic strategies. In this review, we aimed to summarize the recent advancements in tumor matrix biology in parallel to therapeutic approaches targeting matrix stiffness and its consequences in cellular processes in tumor progression and metastasis. The cellular processes governed by signal transduction pathways and their aberrant activation may result in activating the epithelial-to-mesenchymal transition, cancer stemness, and autophagy, which can be attributed to drug resistance. Developing therapeutic strategies to target these cellular processes in cancer biology will offer novel therapeutic approaches to tailor better personalized treatment modalities for clinical studies.

Tumor evolution metrics predict recurrence beyond 10 years in locally advanced prostate cancer.

Cancer evolution lays the groundwork for predictive oncology. Testing evolutionary metrics requires quantitative measurements in controlled clinical trials. We mapped genomic intratumor heterogeneity in locally advanced prostate cancer using 642 samples from 114 individuals enrolled in clinical trials with a 12-year median follow-up. We concomitantly assessed morphological heterogeneity using deep learning in 1,923 histological sections from 250 individuals. Genetic and morphological (Gleason) diversity were independent predictors of recurrence (hazard ratio (HR) = 3.12 and 95% confidence interval (95% CI) = 1.34-7.3; HR = 2.24 and 95% CI = 1.28-3.92). Combined, they identified a group with half the median time to recurrence. Spatial segregation of clones was also an independent marker of recurrence (HR = 2.3 and 95% CI = 1.11-4.8). We identified copy number changes associated with Gleason grade and found that chromosome 6p loss correlated with reduced immune infiltration. Matched profiling of relapse, decades after diagnosis, confirmed that genomic instability is a driving force in prostate cancer progression. This study shows that combining genomics with artificial intelligence-aided histopathology leads to the identification of clinical biomarkers of evolution.

Optimizing cancer therapy: a review of the multifaceted effects of metronomic chemotherapy.

Metronomic chemotherapy (MCT), characterized by the continuous administration of chemotherapeutics at a lower dose without prolonged drug-free periods, has garnered significant attention over the last 2 decades. Extensive evidence from both pre-clinical and clinical settings indicates that MCT induces distinct biological effects than the standard Maximum Tolerated Dose (MTD) chemotherapy. The low toxicity profile, reduced likelihood of inducing acquired therapeutic resistance, and low cost of MCT render it an attractive chemotherapeutic regimen option. One of the most prominent aspects of MCT is its anti-angiogenesis effects. It has been shown to stimulate the expression of anti-angiogenic molecules, thereby inhibiting angiogenesis. In addition, MCT has been shown to decrease the regulatory T-cell population and promote anti-tumor immune response through inducing dendritic cell maturation and increasing the number of cytotoxic T-cells. Combination therapies utilizing MCT along with oncolytic virotherapy, radiotherapy or other chemotherapeutic regimens have been studied extensively. This review provides an overview of the current status of MCT research and the established mechanisms of action of MCT treatment and also offers insights into potential avenues of development for MCT in the future.

The Effect of QR Code-Supported Patient Training on Total Knee Arthroplasty-Related Problems and Emergency Department Admission Rate.

Knee arthroplasty surgery, which is increasingly performed due to increased life expectancy, has positive outcomes, although it can also cause serious health problems following surgery. This study was conducted to evaluate the impact of patient-related education via a QR code on total knee arthroplasty problems and emergency department referral rates. Participants were randomly assigned to intervention (n = 51) and control (n = 51) groups. The intervention group received QR code-supported training. The outcomes were assessed at baseline (preoperative), discharge, and postoperative sixth week. In the intervention group, significantly fewer problems related to total knee arthroplasty occurred at discharge and in week 6, and a higher level of functionality was noted ( P < .05). In week 6, the rate of emergency department admissions was lower, and mean scores for satisfaction with patient training were higher in the intervention group ( P < .05). In conclusion, QR code-supported patient training was effective in reducing the physiological and psychosocial problems related to total knee arthroplasty and the emergency department referral rates. In addition, it provided functional improvement and increased satisfaction with patient training.

Tumor-Microenvironment-on-Chip Platform for Assessing Drug Response in 3D Dynamic Culture.

Microphysiological systems involving microfluidic 3D culture of cancer cells have emerged as a versatile toolkit to study tumor biological problems and evaluate potential treatment strategies. Incorporation of microfluidic technologies in 3D tissue culture offers opportunities for realistic simulation of tumor microenvironment in vitro by facilitating a dynamic culture environment mimicking features of human physiology such as reconstituted ECM, interstitial flow, and gradients of drugs and biomacromolecules. This protocol describes development of 3D microfluidic cell culture based on Tumor-Microenvironment-on-Chip (T-MOC) platform modeling tumor blood and lymphatic capillary vessels and the interstitial space in between. Based on earlier applications of T-MOC for transport characteristics, drug response, and tumor-stroma interactions in mammary carcinoma and pancreatic adenocarcinoma, this protocol provides detailed description of device fabrication, on-chip 3D culture, and drug treatment assays. This protocol can easily be adapted for applications involving other cancer types.

Characterization of dabrafenib-induced drug insensitivity via cellular barcoding and collateral sensitivity to second-line therapeutics.

Drug insensitivity is arguably one of the biggest challenges in cancer therapeutics. Although effective therapeutic solutions in cancer are limited due to the emergence of drug insensitivity, exploiting evolutionary understanding in this context can provide potential second-line therapeutics sensitizing the drug insensitive populations. Targeted therapeutic agent dabrafenib is used to treat CRC patients with BRAF V600E genotype and insensitivity to dabrafenib is often observed. Understanding underlying clonal architecture of dabrafenib-induced drug insensitivity and identification of potential second-line therapeutics that could sensitize dabrafenib insensitive populations remain to be elucidated. For this purpose, we utilized cellular barcoding technology to decipher dabrafenib-induced clonal evolution in BRAF V600E mutant HT-29 cells. This approach revealed the detection of both pre-existing and de novo barcodes with increased frequencies as a result of dabrafenib insensitivity. Furthermore, our longitudinal monitoring of drug insensitivity based on barcode detection from floating DNA within used medium enabled to identify temporal dynamics of pre-existing and de novo barcodes in relation to dabrafenib insensitivity in HT-29 cells. Moreover, whole-exome sequencing analysis exhibited possible somatic CNVs and SNVs contributing to dabrafenib insensitivity in HT-29 cells. Last, collateral drug sensitivity testing demonstrated oxaliplatin and capecitabine, alone or in combination, as successful second-like therapeutics in inducing collateral sensitivity in dabrafenib-insensitive HT-29 cells. Overall, our findings demonstrate clonal dynamics of dabrafenib-insensitivity in HT-29 cells. In addition, oxaliplatin and capecitabine, alone or in combination, were successful second-line therapeutics in inducing collateral sensitivity in dabrafenib-insensitive HT-29 cells.

Does YouTube Provide High-Quality Information for Patients Regarding Night Guards, Especially for Over-the-Counter Ones?

The aim of this study was to evaluate the quality of the information on YouTube regarding night guards (NGs). YouTube was systematically searched using the keyword "night guards." Two independent reviewers examined the first 100 videos and exclusion criteria were applied. Descriptive characteristics of the remaining 60 videos were recorded. In addition, the purpose, target audience, and source of the included videos were collected. A 12-point content scale (CS) was used to evaluate video content, and the Global Quality Scale (GQS) was used to determine video quality. Statistical analyses were performed using the Kruskal-Wallis and Mann-Whitney tests, and the correlation between scores was evaluated using Spearman rho. Of the included videos, 50% were uploaded by dentists/health institutions, 26% by commercial sources and 24% by laypersons. The aim of 80% of the videos was to inform laypeople and 14% to inform professionals only. The content discussed the most (59.3%) was the production stages of NGs. The mean CS and GQS score of the videos were 2.06 ± 1.35 (poor) and 1.71 ± 0.88 (generally poor), respectively. A positive correlation was found between the CS and GQS scores ( r = 0.447). YouTube videos were found to be poor in terms of both content and quality. Since NGs for treating bruxism will always be a trending topic for patients on social media, the content of YouTube videos should be checked and enriched by professionals so that patients can access accurate information, especially about NGs obtained over the counter.

Current Technologies and Future Perspectives in Immunotherapy towards a Clinical Oncology Approach.

Immunotherapy is now established as a potent therapeutic paradigm engendering antitumor immune response against a wide range of malignancies and other diseases by modulating the immune system either through the stimulation or suppression of immune components such as CD4+ T cells, CD8+ T cells, B cells, monocytes, macrophages, dendritic cells, and natural killer cells. By targeting several immune checkpoint inhibitors or blockers (e.g., PD-1, PD-L1, PD-L2, CTLA-4, LAG3, and TIM-3) expressed on the surface of immune cells, several monoclonal antibodies and polyclonal antibodies have been developed and already translated clinically. In addition, natural killer cell-based, dendritic cell-based, and CAR T cell therapies have been also shown to be promising and effective immunotherapeutic approaches. In particular, CAR T cell therapy has benefited from advancements in CRISPR-Cas9 genome editing technology, allowing the generation of several modified CAR T cells with enhanced antitumor immunity. However, the emerging SARS-CoV-2 infection could hijack a patient's immune system by releasing pro-inflammatory interleukins and cytokines such as IL-1ß, IL-2, IL-6, and IL-10, and IFN-γ and TNF-α, respectively, which can further promote neutrophil extravasation and the vasodilation of blood vessels. Despite the significant development of advanced immunotherapeutic technologies, after a certain period of treatment, cancer relapses due to the development of resistance to immunotherapy. Resistance may be primary (where tumor cells do not respond to the treatment), or secondary or acquired immune resistance (where tumor cells develop resistance gradually to ICIs therapy). In this context, this review aims to address the existing immunotherapeutic technologies against cancer and the resistance mechanisms against immunotherapeutic drugs, and explain the impact of COVID-19 on cancer treatment. In addition, we will discuss what will be the future implementation of these strategies against cancer drug resistance. Finally, we will emphasize the practical steps to lay the groundwork for enlightened policy for intervention and resource allocation to care for cancer patients.

Can natural language processing serve as a consultant in oral surgery?

OBJECTIVE: In this comprehensive evaluation, ten experienced oral surgeon experts posed a total of twenty oral surgery-related questions, including dental implant and tooth extractions, to three distinct Natural Language Processing (NLP)-based chatbot platforms: ChatGPT, Microsoft Bing, and Google Bard. The study aimed to assess the effectiveness of these chatbots in responding to specialized medical questions. MATERIALS AND METHODS: Two primary evaluation metrics were employed: a Likert Scale (LS) for measuring the accuracy and completeness of responses and a Global Quality Scale (GQS) for evaluating the clarity of responses. Statistical analyses, including one-way analysis of variance (ANOVA) and Post Hoc Tukey, were conducted to assess and compare the performance of the chatbots as rated by the experts. RESULTS: The results of the study revealed significant differences in the performance of the chatbots. ChatGPT statistically achieved a better mean LS score of 1.4000±0.15986 than Microsoft Bing (1.8750±0.18143) and Google Bards (2.0500±0.12472) (P < 0.001). Additionally, ChatGPT statistically achieved a higher GQS score of 4.4200±0.30111 than Microsoft Bing (3.7550±0.28621) and Google Bards (3.5250±0.22392) (P < 0.001). CONCLUSIONS: These findings showed the substantial advantage of ChatGPT in effectively addressing oral surgery-related questions with superior accuracy, completeness, and clarity. The study highlights the potential of advanced NLP platforms to enhance information retrieval and communication within the field of oral surgery, reinforcing the utility of such technologies in medical and surgical domains.

Investigation of evolutionary dynamics for drug resistance in 3D spheroid model system using cellular barcoding technology.

Complex evolutionary dynamics governing the drug resistance is one of the major challenges in cancer treatment. Understanding these mechanisms requires a sequencing technology with higher resolution to delineate whether pre-existing or de novo drug mechanisms are behind the drug resistance. Combining this technology with clinically very relevant model system, namely 3D spheroids, better mimicking tumorigenesis and drug resistance have so far been lacking. Thus, we sought to establish dabrafenib and irinotecan resistant derivatives of barcoded 3D spheroids with the ultimate aim to quantify the selection-induced clonal dynamics and identify the genomic determinants in this model system. We found that dabrafenib and irinotecan induced drug resistance in 3D-HT-29 and 3D-HCT-116 spheroids are mediated by pre-existing and de novo resistant barcodes, indicating the presence of polyclonal drug resistance in this system. Moreover, whole-exome sequencing analysis found chromosomal gains and mutations associated with dabrafenib and irinotecan resistance in 3D-HT-29 and 3D-HCT-116 spheroids. Last, we show that dabrafenib and irinotecan resistance are also mediated by multiple drug resistance by detection of upregulation of the drug efflux pumps, ABCB1 and ABCG2, in our spheroid model system. Overall, we present the quantification of drug resistance and evolutionary dynamics in spheroids for the first time using cellular barcoding technology and the underlying genomic determinants of the drug resistance in our model system.

Identification of collateral sensitivity and evolutionary landscape of chemotherapy-induced drug resistance using cellular barcoding technology.

Background: One of the most significant challenges impeding cancer treatment effectiveness is drug resistance. Combining evolutionary understanding with drug resistance can pave the way for the identification of second-line drug options that can overcome drug resistance. Although capecitabine and irinotecan are commonly used therapeutic agents in the treatment of CRC patients, resistance to these agents is common. The underlying clonal dynamics of resistance to these agents using high-resolution barcode technology and identification of effective second-line drugs in this context remain unclear. Methods and materials: Caco-2 and HT-29 cell lines were barcoded, and then capecitabine and irinotecan resistant derivatives of these cell lines were established. The frequencies of barcodes from resistant cell lines and harvested medium, longitudinally, were determined. Collateral drug sensitivity testing was carried out on resistant Caco-2 and HT-29 cell lines using single agents or drug combinations. The SyngeryFinder tool was used to analyse drug combination testing. Results: In Caco-2 and HT-29 cell lines, barcode frequency measurements revealed clonal dynamics of capecitabine and irinotecan formed by both pre-existing and de novo barcodes, indicating the presence of polyclonal drug resistance. The temporal dynamics of clonal evolution in Caco-2 and HT-29 cell lines were demonstrated by longitudinal analysis of pre-existing and de novo barcodes from harvested medium. In Caco-2 and HT-29 cell lines, collateral drug sensitivity revealed a number of drugs that were effective alone and in combination. Conclusion: The use of barcoding technology reveals the clonal dynamics of chemotherapy-induced drug resistance not only from harvested cell populations, but also from longitudinal sampling throughout the course of clonal evolution. Second-line drugs that sensitize drug-resistant CRC cell lines are identified through collateral drug testing.

Pan-Cancer Analysis of the COVID-19 Causal Gene SLC6A20.

Genome-wide association studies demonstrated that the chromosome 3p31.21 locus was linked to the severity of COVID-19 disease. The SLC6A20 gene was reported to be one of the critical causal genes regulated by this locus. Various studies focused on demonstrating the severity of COVID-19 in cancer patients and reported that elevated SARS-CoV-2-associated gene expression might contribute to increased susceptibility for COVID-19 in cancer patients. Given that pan-cancer association for the COVID-19 causal gene SLC6A20 is lacking, we aimed to perform systematic profiling of SLC6A20 in different malignancies. Human Protein Atlas, UALCAN, and Hepatocellular Carcinoma (HCCDB) databases were used to assess SLC6A20 gene expression changes in The Cancer Genome Atlas samples with respect to their normal counterparts. GEPIA and TIMER2.0 databases were used to determine the correlation between SLC6A20 and COVID-19-associated genes. Different databases were used for identification of the correlation of SCL6A20 with infiltrating immune cells. The canSAR database was utilized to determine the association of SCL6A20 with immune profiling in different malignancies. The STRING database was utilized to determine the protein network interacting with SLC6A20. Here, we showed SLC6A20 mRNA expression in pan-cancer samples and their normal counterparts. Increased SCL6A20 expression was associated with tumor grade, and there was a positive correlation with SARS-CoV-2-associated genes. Furthermore, SLC6A20 expression was positively correlated with infiltrating neutrophils and immune-related signatures. Lastly, SLC6A20 expression was found to be associated with the angiotensin converting enzyme 2 homologue, TMEM27, suggesting a potential link between SLC6A20 and COVID-19. Taken together, these results suggest that elevated SLC6A20 levels might be partly responsible for increased susceptibility of cancer patients to COVID-19 disease. Therapeutic intervention strategies against SLC6A20 in cancer patients, alongside other treatment modalities, might offer a benefit in delaying COVID-19 disease.

Immune selection determines tumor antigenicity and influences response to checkpoint inhibitors.

In cancer, evolutionary forces select for clones that evade the immune system. Here we analyzed >10,000 primary tumors and 356 immune-checkpoint-treated metastases using immune dN/dS, the ratio of nonsynonymous to synonymous mutations in the immunopeptidome, to measure immune selection in cohorts and individuals. We classified tumors as immune edited when antigenic mutations were removed by negative selection and immune escaped when antigenicity was covered up by aberrant immune modulation. Only in immune-edited tumors was immune predation linked to CD8 T cell infiltration. Immune-escaped metastases experienced the best response to immunotherapy, whereas immune-edited patients did not benefit, suggesting a preexisting resistance mechanism. Similarly, in a longitudinal cohort, nivolumab treatment removes neoantigens exclusively in the immunopeptidome of nonimmune-edited patients, the group with the best overall survival response. Our work uses dN/dS to differentiate between immune-edited and immune-escaped tumors, measuring potential antigenicity and ultimately helping predict response to treatment.

Penetrance of Gastric Adenocarcinoma Susceptibility Genes: A Systematic Review.

BACKGROUND: Gastric adenocarcinoma (GAC) is the fifth most common cancer in the world, and the presence of germline pathogenic variants has been linked with approximately 5% of gastric cancer diagnoses. Multiple GAC susceptibility genes have been identified, but information regarding the risk associated with pathogenic variants in these genes remains obscure. We conducted a systematic review of existing studies reporting the penetrance of GAC susceptibility genes. METHODS: A structured search query was devised to identify GAC-related papers indexed in MEDLINE/PubMed. A semi-automated natural language processing algorithm was applied to identify penetrance papers for inclusion. Original studies reporting the penetrance of GAC were included and the full-text articles were independently reviewed. Summary statistics, effect estimates, and precision parameters from these studies were compiled into a table using a predetermined format to ensure consistency. RESULTS: Forty-five studies were identified reporting the penetrance of GAC among patients harboring mutations in 13 different genes: APC, ATM, BRCA1, BRCA2, CDH1, CHEK2, MLH1, MSH2, MSH6, PMS2, MUTYH-Monoallelic, NBN, and STK11. CONCLUSION: Our systematic review highlights the importance of testing for germline pathogenic variants in patients before the development of GAC. Management of patients who harbor a pathogenic mutation is multifactorial, and clinicians should consider cancer risk for each applicable gene-cancer association throughout the screening and management process. The scarcity of studies we found investigating the risk of GAC among patients with pathogenic variants in GAC susceptibility genes highlights the need for more investigations that focus on producing robust risk estimates for gene-cancer associations.

Increased TRIM31 gene expression is positively correlated with SARS-CoV-2 associated genes TMPRSS2 and TMPRSS4 in gastrointestinal cancers.

Besides typical respiratory symptoms, COVID-19 patients also have gastrointestinal symptoms. Studies focusing on the gastrointestinal tumors derived from gastrointestinal tissues have raised a question whether these tumors might express higher levels of SARS-CoV-2 associated genes and therefore patients diagnosed with GI cancers may be more susceptible to the infection. In this study, we have analyzed the expression of SARS-CoV-2 associated genes and their co-expressions in gastrointestinal solid tumors, cancer cell lines and patient-derived organoids relative to their normal counterparts. Moreover, we have found increased co-expression of TMPRSS2-TMPRSS4 in gastrointestinal cancers suggesting that SARS-CoV-2 viral infection known to be mediated by this protease pair might facilitate the effects of viral infection in GI cancer patients. Further, our findings also demonstrate that TRIM31 expression is upregulated in gastrointestinal tumors, while the inhibition of TRIM31 significantly altered viral replication and viral processes associated with cellular pathways in gastrointestinal cancer samples. Taken together, these findings indicate that in addition to the co-expression of TMPRSS2-TMPRSS4 protease pair in GI cancers, TRIM31 expression is positively correlated with this pair and TRIM31 may play a role in providing an increased susceptibility in GI cancer patients to be infected with SARS-CoV-2 virus.

YouTube™ as a Source of Information for Patients Regarding Dental Implant Failure: A Content Analysis.

ABSTRACT: The aim of this study is to investigate the quality of the information YouTube TM offers to patients concerning dental implant failure. YouTube TM was searched systematically using the keyword 'dental implant failure'. The first 100 videos were viewed by two independent researchers. For each video, its purpose, target audience and source were also recorded. A 10 point content scale (CS) was used to evaluate the video content. The Global Quality Scale (GQS) was also used to determine the quality of videos. Statistical analyses were performed using the Kruskal Wallis and Mann Whitney test and correlation coefficient analyses were performed using Spearman's Rho. While 92.2% of the videos included in the study were uploaded by dentists/health institutions, only 3.1% were uploaded by laypersons. Of the videos, 40.6% were aimed at informing laypersons and 56.3% targetted professionals. The content which was discussed most (71.9%) was 'definition of a dental implant' followed by 'reasons for failure' (65.6%). The mean CS and GQS score of the videos were 3.75 ± 2.35 (moderate) and 2.07 ± 1.05 (generally poor), respectively. There was a positive correlation between the CS and GQS score (r = 0.620). The quality of information on YouTube TM regarding dental implant failure was found inadequate. Thus, the information currently available online needs to be constantly checked and improved by professionals. In addition, clinicians should improve the ways in which they use YouTube TM to better inform patients about the causes, risks and treatment choices involved with failed dental implants.

Inhibition of Wnt signalling by Notch via two distinct mechanisms.

Notch and Wnt are two essential signalling pathways that help to shape animals during development and to sustain adult tissue homeostasis. Although they are often active at the same time within a tissue, they typically have opposing effects on cell fate decisions. In fact, crosstalk between the two pathways is important in generating the great diversity of cell types that we find in metazoans. Several different mechanisms have been proposed that allow Notch to limit Wnt signalling, driving a Notch-ON/Wnt-OFF state. Here we explore these different mechanisms in human cells and demonstrate two distinct mechanisms by which Notch itself, can limit the transcriptional activity of ß-catenin. At the membrane, independently of DSL ligands, Notch1 can antagonise ß-catenin activity through an endocytic mechanism that requires its interaction with Deltex and sequesters ß-catenin into the membrane fraction. Within the nucleus, the intracellular domain of Notch1 can also limit ß-catenin induced transcription through the formation of a complex that requires its interaction with RBPjκ. We believe these mechanisms contribute to the robustness of cell-fate decisions by sharpening the distinction between opposing Notch/Wnt responses.

Systems Biology and Experimental Model Systems of Cancer.

Over the past decade, we have witnessed an increasing number of large-scale studies that have provided multi-omics data by high-throughput sequencing approaches. This has particularly helped with identifying key (epi)genetic alterations in cancers. Importantly, aberrations that lead to the activation of signaling networks through the disruption of normal cellular homeostasis is seen both in cancer cells and also in the neighboring tumor microenvironment. Cancer systems biology approaches have enabled the efficient integration of experimental data with computational algorithms and the implementation of actionable targeted therapies, as the exceptions, for the treatment of cancer. Comprehensive multi-omics data obtained through the sequencing of tumor samples and experimental model systems will be important in implementing novel cancer systems biology approaches and increasing their efficacy for tailoring novel personalized treatment modalities in cancer. In this review, we discuss emerging cancer systems biology approaches based on multi-omics data derived from bulk and single-cell genomics studies in addition to existing experimental model systems that play a critical role in understanding (epi)genetic heterogeneity and therapy resistance in cancer.

Subclonal reconstruction of tumors by using machine learning and population genetics.

Most cancer genomic data are generated from bulk samples composed of mixtures of cancer subpopulations, as well as normal cells. Subclonal reconstruction methods based on machine learning aim to separate those subpopulations in a sample and infer their evolutionary history. However, current approaches are entirely data driven and agnostic to evolutionary theory. We demonstrate that systematic errors occur in the analysis if evolution is not accounted for, and this is exacerbated with multi-sampling of the same tumor. We present a novel approach for model-based tumor subclonal reconstruction, called MOBSTER, which combines machine learning with theoretical population genetics. Using public whole-genome sequencing data from 2,606 samples from different cohorts, new data and synthetic validation, we show that this method is more robust and accurate than current techniques in single-sample, multiregion and longitudinal data. This approach minimizes the confounding factors of nonevolutionary methods, thus leading to more accurate recovery of the evolutionary history of human cancers.