Biomarker Consortium

GI Cancer

Biliary Tract Cancer

V-RAF murine sarcoma viral oncogene homolog B1 (BRAF)

 gene belongs to the RAS-RAF-MEK-ERK signal transduction pathway, also called the MAPK pathway, and is involved in cell growth, differentiation, and proliferation.

Proteins within the MAPK pathway are predominantly serine/threonine kinases that propagate the transduction pathway or activate downstream signal proteins for metabolism, mitosis, motility, and apoptosis.

The 3 RAF proteins in humans are ARAF, BRAF, and CRAF; ARAF and CRAF require additional phosphorylation for full activity; subsequently, BRAF is the most active in humans.

 mutation is V600E, which flips the protein into a constitutively active conformation.

The mutated V600E protein stimulates ERK signaling in vivo.

Other mutations cause reduced kinase activity, which activates wild-type CRAF and results in the same outcome of increased ERK signaling and tumorigenesis.

 V600E mutation status does not appear to correlate to survivability in biliary tract cancer (BTC).

 However, the power of studies to date has been low, so this conclusion may have limited applicability in light of future research.

BRAF V600E mutations are shown to be present in approximately 

Patients with unresectable or metastatic biliary tract cancers (BTCs) who are candidates for systemic therapy should receive comprehensive molecular profiling at diagnosis.

Common testing modalities are next-generation sequencing (NGS) or polymerase chain reaction (PCR) testing of tumor tissue.

 NGS can also be used on cell-free DNA to detect 

 Experts recommend BRAF testing for patients with unresectable or metastatic gallbladder cancer, intrahepatic CCA, and extrahepatic CCA.

 mutations appear exclusive to cholangiocarcinomas (CCAs), indicating that testing may provide nonspecific diagnostic support.

 Most intrahepatic CCAs are detected after the tumor is unresectable.

 mutations provides an additional treatment option prior to metastasis.

The first and currently only FDA-approved treatment for subsequent-line therapy for 

mutated BTC is dabrafenib with trametinib.

 Dabrafenib is an inhibitor of mutated BRAF proteins, which are constitutively active; they stimulate tumor cell proliferation via the MAPK pathway.

 Trametinib is a reversible inhibitor of mitogen-activated protein kinase kinase 1 (MEK1) and MEK2.

 MEK proteins are upstream regulators of the MAPK pathway.

 Tumor cells that initially respond to dabrafenib can develop resistance via the MEK protein, providing the rationale for combination therapy.

Learn more about Dabrafenib and Trametinib >

NCCN. Clinical Practice Guidelines in Oncology. Biliary tract cancer, version 1.2024. Accessed March 9, 2024. 

https://www.nccn.org/professionals/physician_gls/pdf/btc.pdf

TAFINLAR (dabrafenib). Prescribing information. Novartis; 2024. Accessed March 11, 2024. 

https://www.novartis.com/us-en/sites/novartis_us/files/tafinlar.pdf

MEKINIST (trametinib). Prescribing information. Novartis; 2024. Accessed March 11, 2024. 

https://www.novartis.com/us-en/sites/novartis_us/files/mekinist.pdf

Winstead E. FDA approves dabrafenib–trametinib for BRAF-positive cancers. National Cancer Institute. July 21, 2022. Accessed March 9, 2024. 

https://www.cancer.gov/news-events/cancer-currents-blog/2022/fda-dabrafenib-trametinib-braf-solid-tumors

Wan PTC, Garnett MJ, Roe SM, et al; Cancer Genome Project. Mechanism of activation of the RAF-ERK signaling pathway by oncogenic mutations of B-RAF. 

 2004;116(6):855-867. doi:10.1016/s0092-8674(04)00215-6

Cargnello M, Roux PP. Activation and function of the MAPKs and their substrates, the MAPK-activated protein kinases. 

 2011;75(1):50-83. doi:10.1128/MMBR.00031-10

Goeppert B, Frauenschuh L, Renner M, et al. BRAF V600E-specific immunohistochemistry reveals low mutation rates in biliary tract cancer and restriction to intrahepatic cholangiocarcinoma. 

 2014;27(7):1028-1034. doi:10.1038/modpathol.2013.206

Lamarca A, Barriuso J, McNamara MG, Valle JW. Molecular targeted therapies: Ready for “prime time” in biliary tract cancer. 

 2020;73(1):170-185. doi:10.1016/j.jhep.2020.03.007

Subbiah V, Kreitman RJ, Wainberg ZA, et al.  Dabrafenib plus trametinib in BRAFV600E-mutated rare cancers: the phase 2 ROAR trial. 

2023;29(5):1103-1112. doi: 10.1038/s41591-023-02321-8.

BRAF V600E

 gene, located on chromosome 2 (2q34), encodes the IDH1 enzyme, a dimeric cytosolic NADP-dependent isocitrate dehydrogenase.

The IDH1 enzyme catalyzes the conversion of alpha-ketoglutarate to D-2-hydroxyglutarate (2-HG), a metabolite that plays a key role in chromatin regulation and cellular differentiation.

 gene result in elevated accumulation of 2-HG, disrupting normal differentiation, leading to the buildup of hepatic progenitor cells and contributing to malignant transformation into intrahepatic cholangiocarcinoma (CCA).

 mutations have been identified in approximately 

 of intrahepatic CCAs but are rare in other anatomical subsites.

mutations in intrahepatic CCA occur at codon 132 (R132X).

Patients with unresectable or metastatic biliary tract cancers (BTCs) who are eligible for systemic therapy should undergo comprehensive molecular profiling at the time of diagnosis.

 mutations can be conducted through tumor next-generation sequencing (NGS) using a multigene panel or via hotspot mutation analysis. Additionally, cell-free DNA (cfDNA) testing is capable of detecting IDH1 hotspot mutations.

 The National Comprehensive Cancer Network (NCCN) guidelines recommend testing for 

 mutations in patients with unresectable or metastatic intrahepatic or extrahepatic CCA and advise considering this testing for those with unresectable or metastatic gallbladder cancer.

 On August 25, 2021, the FDA approved ivosidenib for the treatment of locally advanced or metastatic CCA with an 

 mutation, the first and only IDH1-targeted therapy. Approval was based on the findings from study AG120-C-005 (NCT02989857), which evaluated ivosidenib in 185 patients with locally advanced or metastatic CCA with an 

 mutation who had progressed on 1 or 2 prior regimens, including at least 1 gemcitabine- or 5-flurouracil-containing regimen.

Ivosidenib is a small molecule inhibitor that specifically binds to and inhibits the mutant IDH1 enzyme that is responsible for converting alpha-ketoglutarate to 2-HG. By lowering 2-HG levels, ivosidenib helps restore normal cellular differentiation and can reverse the effects of the 

 mutation. This process may slow the growth of cancer cells and promote a more normal cellular phenotype.

Online Mendelian Inheritance in Man, OMIM®. Johns Hopkins University, Baltimore, MD. Isocitrate dehydrogenase, NADP(+), 1; IDH1. MIM Number: 147700. Updated April 17, 2024. Accessed October 4, 2024. 

Aguado-Fraile E, Tassinari A, Ishii Y, et al. Molecular and morphological changes induced by ivosidenib correlate with efficacy in mutant-

 2021;17(16):2057-2074. doi:10.2217/fon-2020-1274

Farshidfar F, Zheng S, Gingras MC, et al. Integrative genomic analysis of cholangiocarcinoma identifies distinct IDH-mutant molecular profiles. 

 2017;18(11):2780-2794. doi:10.1016/j.celrep.2017.02.033

Boscoe AN, Rolland C, Kelley RK. Frequency and prognostic significance of isocitrate dehydrogenase 1 mutations in cholangiocarcinoma: a systematic literature review. 

 2019;10(4):751-765. doi:10.21037/jgo.2019.03.10

Javle M, Bekaii-Saab T, Jain A, et al. Biliary cancer: utility of next-generation sequencing for clinical management.

2016;122(24):3838-3847. doi:10.1002/cncr.30254

NCCN. Clinical Practice Guidelines in Oncology. Biliary tract cancers, version 4.2024. Accessed October 4, 2024. 

FDA approves ivosidenib for advanced or metastatic cholangiocarcinoma. Updated February 1, 2022. Accessed October 4, 2025. 

https://www.fda.gov/drugs/resources-information-approved-drugs/fda-approves-ivosidenib-advanced-or-metastatic-cholangiocarcinoma

TIBSOVO (ivosidenib). Prescribing information. Servier Pharmaceuticals LLC; 2023. 

https://www.tibsovopro.com/pdf/prescribinginformation.pdf

IDH1

Neurotrophic tropomyosin receptor kinase (NTRK)

 genes encode for tropomyosin receptor kinases (TRKs) that attach to cell membranes and are involved in normal nervous system development.

Under normal circumstances, neurotrophins activate TRK receptors, which lead to downstream ERK signaling, cellular proliferation, and growth.

This receptor family has 3 known members: TRKA, TRKB, and TRKC.

Their respective endogenous ligands are nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophins 3, 4, and 5.

Other pathways implicated in broad NTRK signaling include mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K)/AKT.

 gene fusions are a driver gene for tumorigenesis.

Oncogenesis occurs via hybrid gene fusions that result in the rearrangement of NTRK sequences to attach to constitutively activated genes.

 gene fuses with an N-terminal target, which results in an overexpression of the kinase domain of kinase function by the chimeric protein.

 gene fusions in solid tumors is approximately 1% but is more prevalent in certain rare tumors.

 of BTC, and the rarity of these oncogenes limits our ability to form precise incidence estimates.

 Patients with unresectable or metastatic biliary tract cancers (BTCs) who are candidates for systemic therapy should receive comprehensive molecular profiling at diagnosis.

 fusions, National Comprehensive Cancer Network guidelines recommend multigene next-generation sequencing (NGS), preferably using a transcriptome-based approach.

While immunohistochemistry, fluorescence in situ hybridization (FISH), and reverse transcriptase–chain polymerase reaction have been used to detect 

 fusions, these methods are not mentioned in the guidelines for BTC.

 gene fusions are found in multiple unresectable or metastatic biliary tract cancers including gallbladder malignancies, intrahepatic cholangiocarcinomas (CCAs), and extrahepatic CCAs.

 Testing for these gene fusions is recommended for these 3 types of biliary tract cancer (BTC).

Two drugs have been approved by the FDA for the treatment of 

 gene fusion–positive, unresectable or metastatic solid tumors including BTC.

 Larotrectinib was approved in 2018, and entrectinib was approved in 2019.

 Entrectinib is an inhibitor of TRKA, TRKB, and TRK3; larotrectinib is a selective pan-TRK inhibitor.

 Both have shown specificity to patients with 

 gene fusions with little to no activity when fusions are absent.

 Both TRK inhibitors have a favorable safety profile with a 57% to 75% response rate in pretreated, NTRK-positive tumors.

 Larotrectinib and entrectinib are tyrosine kinase inhibitors specifically targeting TRKA, TRKB, and TRKC, anaplastic lymphoma kinase (ALK), and the proto-oncogene tyrosine-protein kinase 1 (ROS1).

Farha N, Dima D, Ullah F, Kamath S. Precision oncology targets in biliary tract cancer. 

2023;15(7):2105. doi:10.3390/cancers15072105

Manea CA, Badiu DC, Ploscaru IC, et al. A review of NTRK fusions in cancer. 

 2022;79:103893. doi:10.1016/j.amsu.2022.103893

Okamura R, Boichard A, Kato S, Sicklick JK, Bazhenova L, Kurzrock R. Analysis of NTRK alterations in pan-cancer adult and pediatric malignancies: Implications for NTRK-targeted therapeutic

Lange A, Lo HW. Inhibiting TRK proteins in clinical cancer therapy. 

. 2018;10(4):105. doi:10.3390/cancers10040105

Demols A, Rocq L, Perez-Casanova L, et al. A two-step diagnostic approach for NTRK gene fusion detection in biliary tract and pancreatic adenocarcinomas. 

 2023;28(7):e520-e525. doi:10.1093/oncolo/oyad075

NCCN. Clinical Guidelines in Oncology. Pancreatic adenocarcinoma, version 1.2024. Accessed June 11, 2024. 

https://www.nccn.org/professionals/physician_gls/pdf/pancreatic.pdf

ROZLYTREK (entrectinib). Prescribing information. Genentech; 2019. Accessed May 5, 2024. 

https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/212725s000lbl.pdf

VITRAKVI (larotrectinib). Prescribing information. Loxo Oncology; 2018. Accessed May 5, 2024. 

https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/211710s000lbl.pdf

Hong DS, DuBois SG, Kummar S, et al. Larotrectinib in patients with TRK fusion-positive solid tumours: a pooled analysis of three phase 1/2 clinical trials. 

. 2020;21(4):531-540. doi:10.1016/S1470-2045(19)30856-3

NTRK fusions

DNA mismatch repair/microsatellite instability

DNA MMR is involved in preserving genomic fidelity by repairing DNA after mismatching errors after replication or recombination.

MMR is also involved in cell cycle regulation and apoptosis in response to DNA damage.

Four genes regulate MMR mechanisms, and the dimer proteins they form are responsible for mismatch recognition, insertion-deletion loops, base excision, and other functions. Biallelic inactivation of 1 or more MMR genes can result from either somatic or germline mutations as well as from epigenetic silencing.

Germline mutations in MMR genes are frequently associated with Lynch syndrome, which is known to increase the risk of several types of cancers.

Microsatellites are short tandem nucleotide repeats (1-6 nucleotides) scattered throughout the genome and prone to high rates of mutation.

MSI refers to a hypermutable phenotype that results when MMR genes are inactive or faulty. 

MSI status is generally categorized into 3 types: high MSI (MSI-H), low MSI (MSI-L), and microsatellite stability (MSS). 

The incidence of advanced biliary tract cancers (BTCs) with MMR-deficiency (dMMR)/MSI-H is approximately 

All patients with unresectable or metastatic gallbladder cancer, intrahepatic CCA, or extrahepatic CCA should receive MSI or MMR testing at diagnosis.

 Several techniques are used to test for the presence of dMMR in colorectal cancer, including immunohistochemistry (IHC) or polymerase chain reaction (PCR)–based assays as well as next-generation sequencing (NGS) testing. IHC assays assess expression of MMR proteins, PCR evaluates tumor DNA for microsatellite repeats, and NGS identifies inactivating mutations in MMR genes. MSI-H status correlates with the loss expression in 2 of 4 MMR proteins and abnormal microsatellites in 2 or more regions.

 The National Comprehensive Cancer Network guidelines recommended MMR and MSI testing in patients with unresectable or metastatic BTCs including intrahepatic cholangiocarcinoma, extrahepatic cholangiocarcinoma, and gallbladder cancer.

The FDA has approved several immunotherapy agents for the treatment of patients with dMMR/MSI-H solid tumors.

 In 2023, pembrolizumab received approval for the treatment of adult and pediatric patients who have unresectable or metastatic MSI-H or dMMR solid tumors as determined by an FDA-approved test, who have progressed following prior treatment, and who have no satisfactory alternative treatment options.

 Subsequently, based on KEYNOTE-966 study (NCT04003636) findings, pembrolizumab was approved as a first-line therapy option to be used in combination with gemcitabine and cisplatin for locally advanced unresectable or metastatic BTC.

 Dostarlimab has also received accelerated approval for dMMR recurrent or advanced solid tumors with no satisfactory alternative treatment options based on findings from the GARNET trial (NCT02715284).

 Dostarlimab and pembrolizumab act by inhibiting the programmed death ligand 1 (PD-L1) receptor found on T cells, thereby restoring the antitumor immune response.

Modrich P, Lahue R. Mismatch repair in replication fidelity, genetic recombination, and cancer biology. 

. 1996; 65:101-33. doi:10.1146/annurev.bi.65.070196.000533

Marti TM, Kunz C, Fleck O. DNA mismatch repair and mutation avoidance pathways. 

Bellacosa A. Functional interactions and signaling properties of mammalian DNA mismatch repair proteins. 

. 2001; 8:1076-1092. doi:10.1038/sj.cdd.4400948

Peters AC, Young LC, Maeda T, Tron VA, Andrew SE. Mammalian DNA mismatch repair protects cells from UVB-induced DNA damage by facilitating apoptosis and p53 activation. 

 2003; 2:427-435. doi:10.1016/s1568-7864(03)00003-x

 1996; 65:101-133. doi:10.1146/annurev.bi.65.070196.000533

Herman JG, Umar A, Polyak K, et al. Incidence and functional consequences of hMLH1 promoter hypermethylation in colorectal carcinoma. 

 1998;95:6870-6875. doi:10.1073/pnas.95.12.6870

Hampel H, Frankel WL, Martin E, et al. Feasibility of screening for Lynch syndrome among patients with colorectal cancer. 

 2008;26:5783-5788. doi:10.1200/JCO.2008.17.5950

Lynch HT, de la Chapelle A. Hereditary colorectal cancer. 

 2003;348:919-932. doi:10.1056/NEJMra012242

Aaltonen LA, Salovaara R, Kristo P, et al. Incidence of hereditary nonpolyposis colorectal cancer and the feasibility of molecular screening for the disease. 

 1998;338:1481-1487. doi:10.1056/NEJM199805213382101

Hampel H, Frankel WL, Martin E, et al. Screening for the Lynch syndrome (hereditary nonpolyposis colorectal cancer). 

 2005;352:1851-1860. doi:10.1056/NEJMoa043146

Bonneville R, Krook MA, Chen HZ, et al. Detection of microsatellite instability biomarkers via next-generation sequencing. 

 2020;2055:11-32. doi:10.1007/978-1-4939-9773-2_5

Li K, Luo H, Huang L, Luo H, Zhu X. Microsatellite instability: a review of what the oncologist should know. 

 2020;20:16. doi:10.1186/s12935-019-1091-8

NCCN. Clinical Practice Guidelines in Oncology. Biliary tract cancers, version 1.2024. Accessed April 10, 2024. 

KEYTRUDA (pembrolizumab). Prescribing information. Merck & Co; 12018. Accessed April 10, 2024. 

https://www.merck.com/product/usa/pi_circulars/k/keytruda/keytruda_pi.pdf

JEMPERLI (dostarlimab). Prescribing information. GlaxoSmithKline; 2024. Accessed April 10, 2024. 

https://gskpro.com/content/dam/global/hcpportal/en_US/Prescribing_Information/Jemperli/pdf/JEMPERLI-PI-MG.PDF

FDA converts to full approval indication for KETRUDA (pembrolizumab) for certain adult and pediatric patients with advanced microsatellite instability-high (MSI-H) or mismatch repair deficient (dMMR) solid tumors. Press release. Merck. March 29, 2023. Accessed April 10, 2024. 

https://www.merck.com/news/fda-converts-to-full-approval-indication-for-keytruda-pembrolizumab-for-certain-adult-and-pediatric-patients-with-advanced-microsatellite-instability-high-msi-h-or-mismatch-repair-deficient/

FDA approves pembrolizumab with chemotherapy for biliary tract cancer. FDA. November 1, 2023. Accessed April 10, 2024. 

https://www.fda.gov/drugs/resources-information-approved-drugs/fda-approves-pembrolizumab-chemotherapy-biliary-tract-cancer

FDA grants accelerated approval to dostarlimab-gxly for dMMR advanced solid tumors. FDA. Updated July 18, 2023. Accessed April 10, 2024. 

https://www.fda.gov/drugs/resources-information-approved-drugs/fda-grants-accelerated-approval-dostarlimab-gxly-dmmr-advanced-solid-tumors

MSI-H/dMMR

 gene was first discovered and characterized in 1985 on chromosome 10.

 encodes a transmembrane tyrosine kinase with multiple active extracellular sites including a cysteine-rich domain and cadherin-like domain 1 (CLD1), CLD2, CLD3, and CLD4.

Under normal circumstances, RET protein is involved in the development of the kidneys and enteric nervous system as well as the homeostasis of neural, neuroendocrine, and hematopoietic cells.

Four glial cell line–derived neurotrophic factor (GDNF) family ligands (GFL) are endogenous ligands for RET; these include GDNF, persephin, artemin, and neurturin.

Rather than bind directly to RET proteins, GFLs first bind to GDNF family receptor-α (GFRα) coreceptors, and this complex mediates RET homodimerization.

The activated complex autophosphorylates and activates signal transduction pathways MAPK, RAS, ERK, PI3K, AKT, phospholipase C-γ, and JAK-STAT.

 gene fusions are found most frequently in thyroid cancers (5% to 10%) and non–small cell lung cancer (1% to 2%) while the incidence in biliary tract cancer (BTC) is relatively low (

Oncogenic RET proteins are involved in tumor cell proliferation, invasion, differentiation, and migration, and oncogenesis occurs via 2 primary mechanisms.

The first mechanism is via constitutive activation of kinase activity, and the second is chromosomal rearrangements that produce a chimeric protein fusion of the 

unresectable or metastatic BTC who are candidates for systemic therapy should receive comprehensive molecular profiling at the time of diagnosis.

 The preferred test is next-generation sequencing (NGS), because traditional testing methods have limited utility for detecting 

The National Comprehensive Cancer Network (NCCN) guidelines recommend testing for 

gene fusions for all patients with unresectable or metastatic gallbladder cancer, intrahepatic cholangiocarcinoma (CCA), and extrahepatic CCA.

 Currently, the only FDA-approved treatment for adult patients with locally advanced or metastatic BTC tumors with a 

 Pralsetinib is a selective RET tyrosine kinase inhibitor mentioned within NCCN guidelines for BTC but does not currently have approval for treating tumor-agnostic solid tumors including BTCs.

 Pralsetinib is recommended under certain circumstances as first-line or subsequent-line systemic therapy.

 Selpercatinib is a selective RET kinase inhibitor that inactivates wild-type RET and various mutated RET isoforms.9,10 Selpercatinib is well tolerated with a permanent discontinuation rate of 8% and a dose interruption rate of 64%.

Zhao L, Wang N, Zhang D, Jia Y, Kong F. A comprehensive overview of the relationship between RET gene and tumor occurrence. 

 2023;13:1090757. doi:10.3389/fonc.2023.1090757

Belli C, Penault-Llorca F, Ladanyi M, et al. ESMO recommendations on the standard methods to detect RET fusions and mutations in daily practice and clinical research. 

 2021;32(3):337-350. doi:10.1016/j.annonc.2020.11.021

Ibanez CF. Structure and physiology of the RET receptor tyrosine kinase. 

. 2013;5(2):a009134-a009134. doi:10.1101/cshperspect.a009134

Drilon A, Hu ZI, Lai GGY, Tan DSW. Targeting RET-driven cancers: Lessons from evolving preclinical and clinical landscapes. 

 2018;15(3):151-167. doi:10.1038/nrclinonc.2017.175

Carlomagno F. Thyroid cancer: Role of RET and beyond. 

Kato S, Subbiah V, Marchlik E, Elkin SK, Carter JL, Kurzrock R. RET aberrations in diverse cancers: next-generation sequencing of 4,871 patients. 

 2017;23(8):1988-1997. doi:10.1158/1078-0432.CCR-16-1679

NCCN. Clinical Practice Guidelines in Oncology. Biliary Tract Cancers, version 4.2024. Accessed Oct 2, 2024. 

Shi M, Wang W, Zhang J, et al. Identification of RET fusions in a Chinese multicancer retrospective analysis by next-generation sequencing. 

 2022;113(1):308-318. doi:10.1111/cas.15181

Biliary Tract Cancer

Futibatinib

Initial US Approval

Indications

Recommended Dose/Route

Pivotal Studies

Safety

References