First-In-Human Phase I Study of Tinengotinib (TT-00420), a Multiple Kinase Inhibitor, as a Single Agent in Patients With Advanced Solid Tumors.

PURPOSE: This first-in-human phase I dose-escalation study evaluated the safety, pharmacokinetics, and efficacy of tinengotinib (TT-00420), a multi-kinase inhibitor targeting fibroblast growth factor receptors 1-3 (FGFRs 1-3), Janus kinase 1/2, vascular endothelial growth factor receptors, and Aurora A/B, in patients with advanced solid tumors. PATIENTS AND METHODS: Patients received tinengotinib orally daily in 28-day cycles. Dose escalation was guided by Bayesian modeling using escalation with overdose control. The primary objective was to assess dose-limiting toxicities (DLTs), maximum tolerated dose (MTD), and dose recommended for dose expansion (DRDE). Secondary objectives included pharmacokinetics and efficacy. RESULTS: Forty-eight patients were enrolled (dose escalation, n = 40; dose expansion, n = 8). MTD was not reached; DRDE was 12 mg daily. DLTs were palmar-plantar erythrodysesthesia syndrome (8 mg, n = 1) and hypertension (15 mg, n = 2). The most common treatment-related adverse event was hypertension (50.0%). In 43 response-evaluable patients, 13 (30.2%) achieved partial response (PR; n = 7) or stable disease (SD) ≥ 24 weeks (n = 6), including 4/11 (36.4%) with FGFR2 mutations/fusions and cholangiocarcinoma (PR n = 3; SD ≥ 24 weeks n = 1), 3/3 (100.0%) with hormone receptor (HR)-positive/HER2-negative breast cancer (PR n = 2; SD ≥ 24 weeks n = 1), 2/5 (40.0%) with triple-negative breast cancer (TNBC; PR n = 1; SD ≥ 24 weeks n = 1), and 1/1 (100.0%) with castrate-resistant prostate cancer (CRPC; PR). Four of 12 patients (33.3%; HR-positive/HER2-negative breast cancer, TNBC, prostate cancer, and cholangiocarcinoma) treated at DRDE had PRs. Tinengotinib's half-life was 28-34 hours. CONCLUSIONS: Tinengotinib was well tolerated with favorable pharmacokinetic characteristics. Preliminary findings indicated potential clinical benefit in FGFR inhibitor-refractory cholangiocarcinoma, HER2-negative breast cancer (including TNBC), and CRPC. Continued evaluation of tinengotinib is warranted in phase II trials.

Single cell biology-a Keystone Symposia report.

Single cell biology has the potential to elucidate many critical biological processes and diseases, from development and regeneration to cancer. Single cell analyses are uncovering the molecular diversity of cells, revealing a clearer picture of the variation among and between different cell types. New techniques are beginning to unravel how differences in cell state-transcriptional, epigenetic, and other characteristics-can lead to different cell fates among genetically identical cells, which underlies complex processes such as embryonic development, drug resistance, response to injury, and cellular reprogramming. Single cell technologies also pose significant challenges relating to processing and analyzing vast amounts of data collected. To realize the potential of single cell technologies, new computational approaches are needed. On March 17-19, 2021, experts in single cell biology met virtually for the Keystone eSymposium "Single Cell Biology" to discuss advances both in single cell applications and technologies.

Interactions between cancer cells and immune cells drive transitions to mesenchymal-like states in glioblastoma.

The mesenchymal subtype of glioblastoma is thought to be determined by both cancer cell-intrinsic alterations and extrinsic cellular interactions, but remains poorly understood. Here, we dissect glioblastoma-to-microenvironment interactions by single-cell RNA sequencing analysis of human tumors and model systems, combined with functional experiments. We demonstrate that macrophages induce a transition of glioblastoma cells into mesenchymal-like (MES-like) states. This effect is mediated, both in vitro and in vivo, by macrophage-derived oncostatin M (OSM) that interacts with its receptors (OSMR or LIFR) in complex with GP130 on glioblastoma cells and activates STAT3. We show that MES-like glioblastoma states are also associated with increased expression of a mesenchymal program in macrophages and with increased cytotoxicity of T cells, highlighting extensive alterations of the immune microenvironment with potential therapeutic implications.

Single-cell transcriptomics in cancer: computational challenges and opportunities.

Intratumor heterogeneity is a common characteristic across diverse cancer types and presents challenges to current standards of treatment. Advancements in high-throughput sequencing and imaging technologies provide opportunities to identify and characterize these aspects of heterogeneity. Notably, transcriptomic profiling at a single-cell resolution enables quantitative measurements of the molecular activity that underlies the phenotypic diversity of cells within a tumor. Such high-dimensional data require computational analysis to extract relevant biological insights about the cell types and states that drive cancer development, pathogenesis, and clinical outcomes. In this review, we highlight emerging themes in the computational analysis of single-cell transcriptomics data and their applications to cancer research. We focus on downstream analytical challenges relevant to cancer research, including how to computationally perform unified analysis across many patients and disease states, distinguish neoplastic from nonneoplastic cells, infer communication with the tumor microenvironment, and delineate tumoral and microenvironmental evolution with trajectory and RNA velocity analysis. We include discussions of challenges and opportunities for future computational methodological advancements necessary to realize the translational potential of single-cell transcriptomic profiling in cancer.

Spatial transcriptome profiling by MERFISH reveals subcellular RNA compartmentalization and cell cycle-dependent gene expression.

The expression profiles and spatial distributions of RNAs regulate many cellular functions. Image-based transcriptomic approaches provide powerful means to measure both expression and spatial information of RNAs in individual cells within their native environment. Among these approaches, multiplexed error-robust fluorescence in situ hybridization (MERFISH) has achieved spatially resolved RNA quantification at transcriptome scale by massively multiplexing single-molecule FISH measurements. Here, we increased the gene throughput of MERFISH and demonstrated simultaneous measurements of RNA transcripts from ∼10,000 genes in individual cells with ∼80% detection efficiency and ∼4% misidentification rate. We combined MERFISH with cellular structure imaging to determine subcellular compartmentalization of RNAs. We validated this approach by showing enrichment of secretome transcripts at the endoplasmic reticulum, and further revealed enrichment of long noncoding RNAs, RNAs with retained introns, and a subgroup of protein-coding mRNAs in the cell nucleus. Leveraging spatially resolved RNA profiling, we developed an approach to determine RNA velocity in situ using the balance of nuclear versus cytoplasmic RNA counts. We applied this approach to infer pseudotime ordering of cells and identified cells at different cell-cycle states, revealing ∼1,600 genes with putative cell cycle-dependent expression and a gradual transcription profile change as cells progress through cell-cycle stages. Our analysis further revealed cell cycle-dependent and cell cycle-independent spatial heterogeneity of transcriptionally distinct cells. We envision that the ability to perform spatially resolved, genome-wide RNA profiling with high detection efficiency and accuracy by MERFISH could help address a wide array of questions ranging from the regulation of gene expression in cells to the development of cell fate and organization in tissues.

A Murine Model of Chronic Lymphocytic Leukemia Based on B Cell-Restricted Expression of Sf3b1 Mutation and Atm Deletion.

SF3B1 is recurrently mutated in chronic lymphocytic leukemia (CLL), but its role in the pathogenesis of CLL remains elusive. Here, we show that conditional expression of Sf3b1-K700E mutation in mouse B cells disrupts pre-mRNA splicing, alters cell development, and induces a state of cellular senescence. Combination with Atm deletion leads to the overcoming of cellular senescence and the development of CLL-like disease in elderly mice. These CLL-like cells show genome instability and dysregulation of multiple CLL-associated cellular processes, including deregulated B cell receptor signaling, which we also identified in human CLL cases. Notably, human CLLs harboring SF3B1 mutations exhibit altered response to BTK inhibition. Our murine model of CLL thus provides insights into human CLL disease mechanisms and treatment.

Durvalumab for recurrent or metastatic head and neck squamous cell carcinoma: Results from a single-arm, phase II study in patients with ≥25% tumour cell PD-L1 expression who have progressed on platinum-based chemotherapy.

BACKGROUND: Patients with recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC) progressing on platinum-based chemotherapy have poor prognoses and limited therapeutic options. Programmed cell death-1 (PD-1) and its ligand 1 (PD-L1) are frequently upregulated in HNSCC. The international, multi-institutional, single-arm, phase II HAWK study (NCT02207530) evaluated durvalumab monotherapy, an anti-PD-L1 monoclonal antibody, in PD-L1-high patients with platinum-refractory R/M HNSCC. PATIENTS AND METHODS: Immunotherapy-naïve patients with confirmed PD-L1-high tumour cell expression (defined as patients with ≥25% of tumour cells expressing PD-L1 [TC ≥ 25%] using the VENTANA PD-L1 [SP263] Assay) received durvalumab 10 mg/kg intravenously every 2 weeks for up to 12 months. The primary end-point was objective response rate; secondary end-points included progression-free survival (PFS) and overall survival (OS). RESULTS: Among evaluable patients (n = 111), objective response rate was 16.2% (95% confidence interval [CI], 9.9-24.4); 29.4% (95% CI, 15.1-47.5) for human papillomavirus (HPV)-positive patients and 10.9% (95% CI, 4.5-21.3) for HPV-negative patients. Median PFS and OS for treated patients (n = 112) was 2.1 months (95% CI, 1.9-3.7) and 7.1 months (95% CI, 4.9-9.9); PFS and OS at 12 months were 14.6% (95% CI, 8.5-22.1) and 33.6% (95% CI, 24.8-42.7). Treatment-related adverse events were 57.1% (any grade) and 8.0% (grade ≥3); none led to death. At data cut-off, 24.1% of patients remained on treatment or in follow-up. CONCLUSION: Durvalumab demonstrated antitumour activity with acceptable safety in PD-L1-high patients with R/M HNSCC, supporting its ongoing evaluation in phase III trials in first- and second-line settings. In an ad hoc analysis, HPV-positive patients had a numerically higher response rate and survival than HPV-negative patients.

Afatinib versus gemcitabine/cisplatin for first-line treatment of Chinese patients with advanced non-small-cell lung cancer harboring EGFR mutations: subgroup analysis of the LUX-Lung 6 trial.

INTRODUCTION: Non-small-cell lung cancer (NSCLC) is the leading cause of cancer death in China. Four epidermal growth factor receptor (EGFR)-targeted tyrosine kinase inhibitors - afatinib, erlotinib, icotinib, and gefitinib - are available for first-line treatment of NSCLC in China; however, there are few data to guide treatment choice. The Phase III LUX-Lung 6 trial compared afatinib with platinum-based chemotherapy for first-line treatment of patients from Southeast Asia with EGFR mutation-positive advanced NSCLC. This post hoc analysis assessed the findings from LUX-Lung 6 in Chinese patients. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: NCT01121393. MATERIALS AND METHODS: Previously untreated patients with EGFR mutation-positive stage IIIB/IV lung adenocarcinoma were randomized 2:1 to receive afatinib or ≤6 cycles of gemcitabine/ cisplatin. The key outcomes were progression-free survival (PFS; primary), objective response rate, disease control rate, overall survival (OS), duration of response and disease control, patient-reported outcomes, and safety. Three hundred and twenty-seven patients from mainland China were treated (89.8% of overall LUX-Lung 6 population; afatinib 217, gemcitabine/cisplatin 110). RESULTS: PFS was significantly longer with afatinib than gemcitabine/cisplatin (median 11.0 versus 5.6 months; hazard ratio [HR], 0.30 [95% CI, 0.21, 0.43]; P,0.0001). Overall, there was no significant difference in OS between treatment arms; however, in a subgroup analysis, afatinib significantly improved OS versus gemcitabine/cisplatin in patients with an EGFR Del19 mutation (median 31.6 versus 16.3 months; HR, 0.61 [95% CI, 0.41, 0.91]; P=0.0146). Afatinib was well tolerated, with most treatment-related adverse events (TRAEs) being of grade 1 or 2 severity. The most common grade 3/4 TRAEs with afatinib were rash/acne (15.9%/0.5%), stomatitis (6.1%/0%), and diarrhea (5.6%/0%). TRAEs leading to permanent discontinuation were reported in 12 patients (5.6%) receiving afatinib and 43 (41.7%) receiving gemcitabine/cisplatin. Afatinib significantly improved PFS compared with standard first-line chemotherapy in Chinese patients with EGFR mutation-positive NSCLC and demonstrated a manageable safety profile. CONCLUSION: The findings support the rationale for using afatinib as a first-line treatment option for this patient population.

Phase Ib Study of High-dose Intermittent Afatinib in Patients With Advanced Solid Tumors.

BACKGROUND: The present phase Ib study assessed the maximum tolerated dose (MTD), safety, pharmacokinetics, and antitumor activity of high-dose intermittent (HDI) afatinib monotherapy for patients with advanced solid tumors. The planned focus was patients with epidermal growth factor receptor (EGFR) T790M+ non-small-cell lung cancer (NSCLC). MATERIALS AND METHODS: Eligible patients had histologically confirmed advanced solid tumors that were unsuitable for, or unresponsive to, standard therapy. The study used a 3+3 design with a starting dose of 90 mg/d for 3 days every 14 days (28-day cycles) and incremental dose escalations to 200 mg/d. RESULTS: Thirty-five patients (18 with NSCLC) were treated (6 at 90 mg; 3 at 120 mg; 9 at 150 mg; 11 at 160 mg; and 6 at 200 mg). One patient in the 90-mg cohort (grade 3 rash) and 2 patients in the 200-mg cohort (grade 3 diarrhea; grade 3 mucositis) experienced a dose-limiting toxicity. The MTD was 160 mg. The most common treatment-related adverse events were diarrhea (total, 88.6%; grade 3, 14.3%), rash/acne (total, 62.9%; grade 3, 2.9%), and fatigue (total, 40.0; grade 3, 0%). The maximum afatinib plasma concentration at the MTD was 313 ng/mL, exceeding the in vitro IC50 (inhibitor concentration decreased by one half) range for T790M inhibition. The trough levels suggested no systematic change in afatinib plasma concentrations during long-term treatment at this dosing schedule. Of the 13 T790M+ NSCLC patients, 1 achieved an objective response (7.7%). CONCLUSION: HDI afatinib was feasible and tolerable and could potentially be further explored for NSCLC indications, including patients with central nervous system disease, rare EGFR mutations, or T790M+ NSCLC intolerant of third-generation EGFR tyrosine kinase inhibitors.

Linking transcriptional and genetic tumor heterogeneity through allele analysis of single-cell RNA-seq data.

Characterization of intratumoral heterogeneity is critical to cancer therapy, as the presence of phenotypically diverse cell populations commonly fuels relapse and resistance to treatment. Although genetic variation is a well-studied source of intratumoral heterogeneity, the functional impact of most genetic alterations remains unclear. Even less understood is the relative importance of other factors influencing heterogeneity, such as epigenetic state or tumor microenvironment. To investigate the relationship between genetic and transcriptional heterogeneity in a context of cancer progression, we devised a computational approach called HoneyBADGER to identify copy number variation and loss of heterozygosity in individual cells from single-cell RNA-sequencing data. By integrating allele and normalized expression information, HoneyBADGER is able to identify and infer the presence of subclone-specific alterations in individual cells and reconstruct the underlying subclonal architecture. By examining several tumor types, we show that HoneyBADGER is effective at identifying deletions, amplifications, and copy-neutral loss-of-heterozygosity events and is capable of robustly identifying subclonal focal alterations as small as 10 megabases. We further apply HoneyBADGER to analyze single cells from a progressive multiple myeloma patient to identify major genetic subclones that exhibit distinct transcriptional signatures relevant to cancer progression. Other prominent transcriptional subpopulations within these tumors did not line up with the genetic subclonal structure and were likely driven by alternative, nonclonal mechanisms. These results highlight the need for integrative analysis to understand the molecular and phenotypic heterogeneity in cancer.

Phase I dose-escalation trial of afatinib, an irreversible ErbB family blocker, in combination with gemcitabine or docetaxel in patients with relapsed or refractory solid tumors.

Background Afatinib, an irreversible ErbB family blocker, has shown synergistic antitumor activity and manageable tolerability in combination with chemotherapy. This phase I study assessed oral afatinib plus intravenous gemcitabine or docetaxel in patients with relapsed/refractory solid tumors. Methods Patients received afatinib (30, 40, or 50 mg) plus gemcitabine (1000 or 1250 mg/m2) or docetaxel (60 or 75 mg/m2). Dose escalation proceeded via a 3 + 3 design until the maximum tolerated dose (MTD) was reached. Adverse events (AEs), pharmacokinetics and antitumor activity were also assessed. Results Dose-limiting toxicities during Cycle 1 were reported in 6/39 patients receiving afatinib/gemcitabine (most commonly diarrhea, thrombocytopenia and vomiting) and 16/54 patients receiving afatinib/docetaxel (most commonly febrile neutropenia and stomatitis). The MTDs were established as afatinib 40 mg/gemcitabine 1000 mg/m2 and afatinib 30 mg/docetaxel 60 mg/m2. The most common drug-related AEs were diarrhea, asthenia and rash with afatinib/gemcitabine, and diarrhea, asthenia and stomatitis with afatinib/docetaxel. No relevant pharmacokinetic interactions were observed for either combination. Both combinations demonstrated clinical activity and durable disease control at the MTDs. Compared with the MTD, higher response rates were achieved with afatinib 30 mg/docetaxel 75 mg/m2 (28% vs 6%); however, this regimen was associated with problematic febrile neutropenia, an expected AE with docetaxel, that is often managed with growth factor support. Conclusions Afatinib/gemcitabine and afatinib/docetaxel demonstrated manageable safety profiles, with evidence of clinical efficacy at the MTDs. For afatinib/docetaxel, a dose level of afatinib 30 mg/docetaxel 75 mg/m2 produced higher response rates. Trial registration: NCT01251653 ( ClinicalTrials.gov ).

Continued use of afatinib with the addition of cetuximab after progression on afatinib in patients with EGFR mutation-positive non-small-cell lung cancer and acquired resistance to gefitinib or erlotinib.

OBJECTIVES: In a phase Ib trial, afatinib plus cetuximab demonstrated promising clinical activity (objective response rate [ORR]: 29%; median progression-free survival [PFS]: 4.7 months) in patients with epidermal growth factor receptor (EGFR) mutation-positive non-small-cell lung cancer (NSCLC) with acquired resistance to erlotinib or gefitinib. Here, a separate cohort exploring afatinib plus cetuximab after progression on afatinib is reported. MATERIALS AND METHODS: Patients with EGFR mutation-positive NSCLC who progressed on erlotinib or gefitinib received afatinib 40mg daily until progression, followed by afatinib daily plus cetuximab 500mg/m2 every 2 weeks until progression or intolerable adverse events (AEs). Endpoints included safety, ORR, and PFS. RESULTS: Thirty-seven patients received afatinib monotherapy. Two (5%) patients responded; median PFS was 2.7 months. Thirty-six patients transitioned to afatinib plus cetuximab. Four (11%) patients responded; median PFS was 2.9 months. Median PFS with afatinib plus cetuximab for patients who received afatinib monotherapy for ≥12 versus <12 weeks was 4.9 versus 1.8 months (p=0.0354), and for patients with T790M-positive versus T790M-negative tumors was 4.8 versus 1.8 months (p=0.1306). Fifty percent of patients receiving afatinib plus cetuximab experienced drug-related grade 3/4 AEs. The most frequent drug-related AEs (any grade) were diarrhea (70%), rash (49%), and fatigue (35%) with afatinib monotherapy and rash (69%), paronychia (39%), and dry skin (36%) with afatinib plus cetuximab. CONCLUSION: Sequential EGFR blockade with afatinib followed by afatinib plus cetuximab had a predictable safety profile and demonstrated modest activity in patients with EGFR mutation-positive NSCLC with resistance to erlotinib or gefitinib. CLINICALTRIALS. GOV IDENTIFIER: NCT01090011.

Integrated single-cell genetic and transcriptional analysis suggests novel drivers of chronic lymphocytic leukemia.

Intra-tumoral genetic heterogeneity has been characterized across cancers by genome sequencing of bulk tumors, including chronic lymphocytic leukemia (CLL). In order to more accurately identify subclones, define phylogenetic relationships, and probe genotype-phenotype relationships, we developed methods for targeted mutation detection in DNA and RNA isolated from thousands of single cells from five CLL samples. By clearly resolving phylogenic relationships, we uncovered mutated LCP1 and WNK1 as novel CLL drivers, supported by functional evidence demonstrating their impact on CLL pathways. Integrative analysis of somatic mutations with transcriptional states prompts the idea that convergent evolution generates phenotypically similar cells in distinct genetic branches, thus creating a cohesive expression profile in each CLL sample despite the presence of genetic heterogeneity. Our study highlights the potential for single-cell RNA-based targeted analysis to sensitively determine transcriptional and mutational profiles of individual cancer cells, leading to increased understanding of driving events in malignancy.

Transcriptomic Characterization of SF3B1 Mutation Reveals Its Pleiotropic Effects in Chronic Lymphocytic Leukemia.

Mutations in SF3B1, which encodes a spliceosome component, are associated with poor outcome in chronic lymphocytic leukemia (CLL), but how these contribute to CLL progression remains poorly understood. We undertook a transcriptomic characterization of primary human CLL cells to identify transcripts and pathways affected by SF3B1 mutation. Splicing alterations, identified in the analysis of bulk cells, were confirmed in single SF3B1-mutated CLL cells and also found in cell lines ectopically expressing mutant SF3B1. SF3B1 mutation was found to dysregulate multiple cellular functions including DNA damage response, telomere maintenance, and Notch signaling (mediated through KLF8 upregulation, increased TERC and TERT expression, or altered splicing of DVL2 transcript, respectively). SF3B1 mutation leads to diverse changes in CLL-related pathways.

Cell-Type-Specific Alternative Splicing Governs Cell Fate in the Developing Cerebral Cortex.

Alternative splicing is prevalent in the mammalian brain. To interrogate the functional role of alternative splicing in neural development, we analyzed purified neural progenitor cells (NPCs) and neurons from developing cerebral cortices, revealing hundreds of differentially spliced exons that preferentially alter key protein domains-especially in cytoskeletal proteins-and can harbor disease-causing mutations. We show that Ptbp1 and Rbfox proteins antagonistically govern the NPC-to-neuron transition by regulating neuron-specific exons. Whereas Ptbp1 maintains apical progenitors partly through suppressing a poison exon of Flna in NPCs, Rbfox proteins promote neuronal differentiation by switching Ninein from a centrosomal splice form in NPCs to a non-centrosomal isoform in neurons. We further uncover an intronic human mutation within a PTBP1-binding site that disrupts normal skipping of the FLNA poison exon in NPCs and causes a brain-specific malformation. Our study indicates that dynamic control of alternative splicing governs cell fate in cerebral cortical development.

Clonal evolution in patients with chronic lymphocytic leukaemia developing resistance to BTK inhibition.

Resistance to the Bruton's tyrosine kinase (BTK) inhibitor ibrutinib has been attributed solely to mutations in BTK and related pathway molecules. Using whole-exome and deep-targeted sequencing, we dissect evolution of ibrutinib resistance in serial samples from five chronic lymphocytic leukaemia patients. In two patients, we detect BTK-C481S mutation or multiple PLCG2 mutations. The other three patients exhibit an expansion of clones harbouring del(8p) with additional driver mutations (EP300, MLL2 and EIF2A), with one patient developing trans-differentiation into CD19-negative histiocytic sarcoma. Using droplet-microfluidic technology and growth kinetic analyses, we demonstrate the presence of ibrutinib-resistant subclones and estimate subclone size before treatment initiation. Haploinsufficiency of TRAIL-R, a consequence of del(8p), results in TRAIL insensitivity, which may contribute to ibrutinib resistance. These findings demonstrate that the ibrutinib therapy favours selection and expansion of rare subclones already present before ibrutinib treatment, and provide insight into the heterogeneity of genetic changes associated with ibrutinib resistance.

Effects of Ketoconazole on the Pharmacokinetics of Lenvatinib (E7080) in Healthy Participants.

BACKGROUND: Lenvatinib is an oral, multitargeted, tyrosine kinase inhibitor under clinical investigation in solid tumors. In vitro evidence indicates that lenvatinib metabolism may be modulated by ketoconazole, an inhibitor of CYP3A4 and p-glycoprotein. METHODS: In this Phase I, single-center, randomized, open-label, two-period, crossover study, healthy adults (18-55 years; N = 18) were randomized to one of two sequences (ketoconazole → placebo or vice versa). Ketoconazole (400 mg) or placebo was administered orally once daily for 18 days; a 5 mg dose of lenvatinib was orally administered on Day 5 of each treatment period. Blood samples were collected over 14 days and lenvatinib plasma concentrations measured by high-performance liquid chromatography/tandem mass spectrometry. RESULTS: Systemic exposure to lenvatinib increased slightly (15-19%) with coadministration of ketoconazole. Although the 90% confidence interval (CI) for area under the plasma concentration-time curve (AUC) was within the prespecified bioequivalence interval of 80-125%, Cmax slightly exceeded the 125% CI bound (134%). No changes in tmax, tlag, or t½ were observed. Thirteen subjects (72%) experienced treatment-emergent adverse events (11 mild, 2 moderate), most commonly headache (22%) and diarrhea (17%). CONCLUSIONS: Lenvatinib exposure was slightly increased by ketoconazole; however, the magnitude of the change was relatively small, and likely not clinically meaningful.

Locally disordered methylation forms the basis of intratumor methylome variation in chronic lymphocytic leukemia.

Intratumoral heterogeneity plays a critical role in tumor evolution. To define the contribution of DNA methylation to heterogeneity within tumors, we performed genome-scale bisulfite sequencing of 104 primary chronic lymphocytic leukemias (CLLs). Compared with 26 normal B cell samples, CLLs consistently displayed higher intrasample variability of DNA methylation patterns across the genome, which appears to arise from stochastically disordered methylation in malignant cells. Transcriptome analysis of bulk and single CLL cells revealed that methylation disorder was linked to low-level expression. Disordered methylation was further associated with adverse clinical outcome. We therefore propose that disordered methylation plays a similar role to that of genetic instability, enhancing the ability of cancer cells to search for superior evolutionary trajectories.

Effect of rifampicin on the pharmacokinetics of lenvatinib in healthy adults.

BACKGROUND AND OBJECTIVES: Lenvatinib is an oral, multitargeted tyrosine kinase inhibitor under clinical investigation in solid tumours. This study evaluated the influence of P-glycoprotein (P-gp) inhibition (single-dose rifampicin) and simultaneous cytochrome P450 3A4 (CYP3A4)/P-gp induction (multiple-dose rifampicin) on lenvatinib pharmacokinetics. METHODS: This Phase I, single-centre, single-dose (lenvatinib mesylate 24 mg), open-label, sequential study enrolled 15 healthy volunteers. Three regimens were administered over three periods: Period (P) 1 (Days 1-8), P2 (Days 15-22) and P3 (Days 29-50), with a 14-day (first dose) and 28-day (second dose) washout period after lenvatinib mesylate administration (Day 1, Day 15 and Day 43). In P2, a single oral dose of rifampicin (600 mg) was coadministered with lenvatinib. In P3, rifampicin was administered daily (600 mg) for 21 days (Days 29-49). Serial blood samples were collected, and plasma concentrations of total (protein bound + unbound) and free (unbound) lenvatinib and total metabolites (M1, M2, M3 and M5) were measured by validated high-performance liquid chromatography/tandem mass spectrometry. RESULTS: Single-dose rifampicin (P-gp inhibition) increased area under the plasma concentration-time curve from time zero to infinity (AUC0-∞) of free and total lenvatinib by 32 and 31 %, respectively. Multiple-dose rifampicin (simultaneous P-gp and CYP3A4 induction) decreased lenvatinib AUC0-∞ (total: 18 %; free: 9 %). Treatment-emergent adverse events were mild or moderate and occurred in 7 subjects (47 %). CONCLUSION: Lenvatinib exposure was increased by P-gp inhibition; however, based on free concentrations, simultaneous P-gp and CYP3A4 induction results met the prespecified bioequivalence 90 % confidence interval. Overall, the magnitude of these changes was relatively small, and likely not clinically meaningful.