Biomarker Consortium

GI Cancer

Colorectal Cancer

DNA mismatch repair/microsatellite instability.

DNA mismatch repair (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 one (or more) MMR genes can result from either somatic or germline mutations, as well as from epigenetic silencing.

 arises from inherited (germline) mutations in four MMR genes (MLH1, MSH2, MSH6, and PMS2) and stands as the predominant genetically linked predisposition to colorectal cancer (CRC), accounting for approximately

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

Microsatellite instability (MSI) refers to a hyper-mutable phenotype that results when MMR genes are inactive or faulty.MSI status is generally categorized into three types: high microsatellite instability (MSI-H), low microsatellite instability (MSI-L) and microsatellite stability (MSS). 

All newly diagnosed patients with CRC should undergo MMR or MSI testing.

Several techniques are used to test for the presence of MMR deficiency (dMMR) in CRC, including immunohistochemical (IHC) or polymerase chain reaction- (PCR-) based assays as well as NGS based testing.

Universal testing is recommended for all newly diagnosed patients with CRC. IHC assays test for the protein expression of 4 MMR genes known to be mutated in Lynch Syndrome. 

Under normal physiological conditions all 4 MMR proteins are expressed. 

However, the loss of expression of one or more of these suggests the presence of a MMR gene mutation and warrants further testing, including screening for the presence of BRAF V600E mutations.

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

Based findings from the CheckMate-142 trial, nivolumab received accelerated approval as a single agent in this setting; approval was later extended to nivolumab in combination with ipilimumab in 2018.

Pembrolizumab received approval as a first-line agent for the treatment of dMMR/MSI-H metastatic CRC in 2020 based on safety and efficacy demonstrated in the KEYNOTE-177 trial.

Subsequently, in 2021, dostarlimab received accelerated approval for dMMR recurrent or advanced solid tumors with no satisfactory alternative treatment options based on findings from GARNET.

Approved immunotherapy agents act by inhibiting the programmed death ligand 1 (PD-L1) receptor found on T cells, thereby restoring the anti-tumor immune response.

Learn more about Nivolumab and Ipilimumab >

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

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

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

Peters AC, Young LC, Maeda T, et al. Mammalian DNAmismatch repair protects cells from UVB-induced DNA damage by facilitating apoptosis and p53 activation.

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

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

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

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

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

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

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.

National Comprehensive Cancer Network. Clinical Practice Guidelines in Oncology. Colon Cancer, v1.2024. Accessed April 10, 2024. 

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

National Comprehensive Cancer Network. Clinical Practice Guidelines in Oncology. Rectal Cancer, v1.2024. Accessed April 10, 2024.

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

KEYTRUDA (pembrolizumab). Prescribing information. Merck & Co, Inc; 2018. Accessed June 12, 2024. 

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

OPDIVO (nivolumab). Prescribing information. Princeton, NJ, USA: Bristol-Myers Squibb Company; 2024. 

https://packageinserts.bms.com/pi/pi_opdivo.pdf

YERVOY (ipilimumab). Prescribing information. Princeton, NJ, USA: Bristol-Myers Squibb Company; 2023. 

https://packageinserts.bms.com/pi/pi_yervoy.pdf

JEMPERLI (dostarlimab). Prescribing information. GlaxoSmithKline LLC; 2024. Accessed September 9, 2024. 

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

FDA.gov. FDA grants nivolumab accelerated approval for MSI-H or dMMR colorectal cancer. Updated August 1, 2017. Accessed April 10, 2024. 

FDA.gov. FDA grants accelerated approval to ipilimumab for MSI-H or dMMR metastatic colorectal cancer. Updated July 11, 2018. Accessed April 10, 2024.

FDA.gov. FDA approves pembrolizumab for first-line treatment of MSI-H/dMMR colorectal cancer. Updated June 30, 2020. Accessed April 10, 2024.

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

dMMR/MSI-H

Neurotrophic-tropomyosin receptor tyrosine kinase (NTRK)

The neurotrophic-tropomyosin receptor tyrosine kinase 1 (

genes encode for proteins TRKA, TRKB, and TRKC, respectively.

NTRK proteins localize to cell membranes, and their endogenous ligands are neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3, neurotrophin-4, and neurotrophin-5.

 gene activation activates downstream signaling pathways RAS-RAF-MEK-ERK, phospholipase C, and phosphatidylinositol 3 kinase (PI3K).

The NTRK proteins are involved in the growth and proliferation of neural cells in particular, and gene fusions can induce tumorigenesis in neural and non-neural cell cancers.

 gene fusions are drivers for tumorigenesis and lead to constitutively active downstream signaling of various growth pathways.

 gene fusions are present in approximately 

1% of solid tumors but are much less common in CRCs, with an estimated prevalence ranging from 0.35% to 0.7%.

 gene fusions in CRC makes it challenging to elucidate clinicopathological characteristics. 

In a 2022 study, researchers analyzed a cohort of 16 patients with NTRK+ colorectal cancer (CRC) and found that of the three genes, 

 rearrangements were the most common, and TPM3 was the most common fusion partner in introns 7-11. NTRK+ CRC variants were more likely to have microsatellite instability.

Given the availability of targeted TRK inhibitors, patients with metastatic colorectal cancer should undergo molecular testing for 

Feasible testing methods include immunohistochemistry (ICH), fluorescence in situ hybridization (FISH), DNA-based next-generation sequencing, and RNA-based NGS.

The ideal approach appears to be RNA-based NGS because of the higher rates of false positives and lack of evidence of functional transcription present in other methods.

Guidelines endorse using multiple testing methodologies, depending on the accessibility of assays and available testing material.

There are currently two treatment options for unresectable or metastatic solid tumors with 

 gene fusions: larotrectinib and entrectinib. 

Notably, NTRK inhibitors have demonstrated effectiveness only in cases with 

While these targeted therapies are only appropriate for a small subset of CRC patients, they do offer an additional option with an attractive safety profile and an overall response rate of 57-79%.

The FDA has approved entrectinib for the treatment of adult and pediatric patients 12 years of age and older with solid tumors that have a 

 gene fusion without a known acquired resistance mutation, are metastatic, or where surgical resection is likely to result in severe morbidity; it is also approved for situations where there are no satisfactory alternative treatments or the patient has progressed following treatment. 

This indication has received accelerated approval based on the overall response rate and duration of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in confirmatory trials.

Entrectinib is an inhibitor of the tropomyosin receptor tyrosine kinases (TRK) TRKA, TRKB, and TRKC (encoded by the NTRK genes 

 respectively), proto-oncogene tyrosine-protein kinase ROS1 (ROS1), and anaplastic lymphoma kinase (ALK). Entrectinib also inhibits JAK2 and TNK2.

Fusion proteins that include TRK, ROS1, or ALK kinase domains can drive tumorigenic potential through hyperactivation of downstream signaling pathways leading to unconstrained cell proliferation. Entrectinib demonstrated in vitro and in vivo inhibition of cancer cell lines derived from multiple tumor types harboring 

https://www.gene.com/patients/medicines/rozlytrek

The FDA has approved larotrectinib for the treatment of adult and pediatric patients with solid tumors that have a 

 gene fusion without a known acquired resistance mutation, are metastatic, or where surgical resection is likely to result in severe morbidity; it is also approved for situations where there are no satisfactory alternative treatments or the patient has progressed following treatment.

Larotrectinib is an inhibitor of the tropomyosin receptor kinases (TRK), TRKA, TRKB, and TRKC. TRKA, B, and C are encoded by the genes 

Chromosomal rearrangements involving in-frame fusions of these genes with various partners can result in constitutively-activated chimeric TRK fusion proteins that can act as an oncogenic driver, promoting cell proliferation and survival in tumor cell lines. In in vitro and in vivo tumor models, larotrectinib demonstrated anti-tumor activity in cells with constitutive activation of TRK proteins resulting from gene fusions, deletion of a protein regulatory domain, or in cells with TRK protein overexpression.

https://hcp.vitrakvi-us.com/support-and-resources/resources

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 Therapeutics.

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

Solomon JP, Hechtman JF. Detection of NTRK Fusions: Merits and Limitations of Current Diagnostic Platforms. 

2019;79(13):3163-3168. doi:10.1158/0008-5472.CAN-19-0372

National Comprehensive Cancer Network. NCCN guidelines version 1.2024: Colon Cancer. 

Wang H, Li Z, Ou Q, et al. NTRK fusion positive colorectal cancer is a unique subset of CRC with high TMB and microsatellite instability. 

 2022;11(13):2541-2549. doi:10.1002/cam4.4561

Cocco E, Benhamida J, Middha S, et al. Colorectal Carcinomas Containing Hypermethylated MLH1 Promoter and Wild-Type BRAF/KRAS Are Enriched for Targetable Kinase Fusions. 

 2019;79(6):1047-1053. doi:10.1158/0008-5472.CAN-18-3126

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

https://www.gene.com/download/pdf/rozlytrek_prescribing.pdf

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

https://labeling.bayerhealthcare.com/html/products/pi/vitrakvi_PI.pdf

FDA approves larotrectinib for solid tumors with NTRK gene fusions | FDA. Accessed March 18, 2024. 

https://www.fda.gov/drugs/fda-approves-larotrectinib-solid-tumors-ntrk-gene-fusions

Drilon A, Laetsch TW, Kummar S, et al. Efficacy of Larotrectinib in TRK Fusion–Positive Cancers in Adults and Children. 

 2018;378(8):731-739. doi:10.1056/NEJMoa1714448

FDA approves entrectinib for NTRK solid tumors and ROS-1 NSCLC | FDA. Accessed March 19, 2024. 

https://www.fda.gov/drugs/resources-information-approved-drugs/fda-approves-entrectinib-ntrk-solid-tumors-and-ros-1-nsclc

Demetri GD, Paz-Ares L, Farago AF, et al. Efficacy and safety of entrectinib in patients with NTRK fusion-positive (NTRK-fp) tumors: pooled analysis of STARTRK-2, STARTRK-1 and ALKA-372-001. 

 2018;29:viii713. doi:10.1093/annonc/mdy424.017

NTRK fusions

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

The RAS-RAF-MEK-ERK pathway is an important protein kinase cascade that regulates cell differentiation, proliferation, and survival in response to hormones, cytokines, and growth factors.

There are 3 functional RAF proteins in humans, including V-Raf murine sarcoma viral oncogene homolog B (BRAF).

BRAF is a serine/threonine-specific protein kinase with 3 conserved domains: CR1, CR2, and CR3.

The wild-type BRAF protein forms a strong hydrophobic interaction in residues 596-600, which keeps it in its inactive state. 

BRAF becomes activated once the activation loop is phosphorylated, destabilizing the hydrophobic site and enabling downstream kinase activity.

 mutation is an acquired gene mutation identified most commonly in older patients (>60 years), proximal tumor location, locally advanced characteristics, and poor cell differentiation.

 5% to 9% of patients with colorectal cancer.

 V600E mutations are associated with worse survival after recurrence in patients with stage III colon cancer and are associated with a shorter median progression-free and median overall survival than patients with wild-type 

 can result in constitutive activation of the 

 mutations are more common in cancer types that also harbor 

 mutations, including colorectal cancer, borderline ovarian cancers, and malignant melanoma.

This association occurs because activation of the RAS-RAF-MEK-ERK pathway can be achieved by cancer cells through mutation at various points along the pathway.

 gain synergistic effects when combined with other agents that target kinases in the same pathway.

All patients with metastatic CRC should receive molecular testing for 

The preferred method for molecular testing is broad profiling using an NGS panel in a CLIA-certified laboratory, though immunohistochemistry (IHC) is also an option.

Testing can be performed on either the primary tumor or metastatic lesions.

All patients with mCRC should undergo testing for 

 mutations, HER2 amplifications, and MSI/MMR status; testing may be performed individually or via NGS (preferred).

A single drug regimen is FDA-approved for treating adult patients with mCRC with a 

 V600E mutation detected by an FDA-approved test after prior therapy: encorafenib in combination with cetuximab.

NCCN guidelines version 1.2024 also recommend encorafenib in combination with panitumumab.

In combination with cetuximab and mFOLFOX6, for the treatment of patients with metastatic colorectal cancer (mCRC) with a BRAF V600E mutation, as detected by an FDA‑approved test. *

This indication is approved under accelerated approval based on response rate and durability of response. Continued approval for this indication may be contingent upon verification and description of clinical benefit in a confirmatory trial(s).

In combination, for the treatment of adult patients with metastatic colorectal cancer (CRC) with a

 V600E mutation, as detected by an FDA-approved test, after prior therapy.

Encorafenib is a kinase inhibitor of BRAF V600E, wild-type BRAF, and CRAF.

Cetuximab is an epidermal growth factor receptor (EGFR) inhibitor.

Learn more about Encorafenib and Cetuximab >

https://braftovi.pfizerpro.com/mektovi-m/material

Clancy C, Burke JP, Kalady MF, Coffey JC. BRAF mutation is associated with distinct clinicopathological characteristics in colorectal cancer: a systematic review and meta-analysis. 

. 2013;15(12):e711-e718. doi:10.1111/codi.12427

Wan PTC, Garnett MJ, Roe SM, et al. 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

Cutler RE, Stephens RM, Saracino MR, Morrison DK. Autoregulation of the Raf-1 serine/threonine kinase. 

. 1998;95(16):9214-9219. doi:10.1073/pnas.95.16.9214

NCCN. Clinical Practice Guidelines in Oncology. Colon cancer, version 1.2024. 

Sinicrope FA, Shi Q, Allegra CJ, et al. Association of DNA mismatch repair and mutations in BRAF and KRAS with survival after recurrence in stage III colon cancers. 

. 2017;3(4):472-480. doi:10.1001/jamaoncol.2016.5469

Tol J, Nagtegaal ID, Punt CJ. BRAF mutation in metastatic colorectal cancer. 

. 2009;361(1):98-99. doi:10.1056/nejmc0904160

BRAFTOVI (Encorafenib). Prescribing information. Array BioPharma, Inc; 2023. 

https://labeling.pfizer.com/ShowLabeling.aspx?id=12990

Davies H, Bignell GR, Cox C, et al. Mutations of the 

. 2002;417(6892):949-954. doi:10.1038/nature00766

Tabernero J, Grothey A, Van Cutsem E, et al. Encorafenib plus cetuximab as a new standard of care for previously treated BRAF V600E-mutant metastatic colorectal cancer: Updated survival results and subgroup analyses from the BEACON study. 

2021;39(4):273-284. doi:10.1200/JCO.20.02088

FDA approves encorafenib in combination with cetuximab for metastatic colorectal cancer with a BRAF V600E mutation. US FDA. Updated April 9, 2020. Accessed May 21, 2024. 

https://www.fda.gov/drugs/resources-information-approved-drugs/fda-approves-encorafenib-combination-cetuximab-metastatic-colorectal-cancer-braf-v600e-mutation

ERBITUX (cetuximab). Prescribing information. Eli Lilly and Company; 2021. 

https://uspl.lilly.com/erbitux/erbitux.html#pi

BRAF V600E

Human epidermal growth factor receptor2 (HER2); erb-b2 avian erythroblastic leukemia viral oncogene homolog 2 (ERB-B2) 

 oncogene encodes a transmembrane tyrosine kinase receptor that is part of the epidermal growth factor receptor family.

HER2 initiates downstream signaling pathways related to cell proliferation by forming homodimers or heterodimers with other EGFR family members.

Amplification or overexpression of HER2 results in the increased activation of mitogenic signaling, leading to uncontrolled cell proliferation and tumorigenesis. 

 amplification is more frequently observed in distal (rectal) and left-sided tumors, while being less common in proximal and right-sided CRC.

-amplified CRC is also associated with a greater number of metastatic sites, particularly lung and brain metastases. 

While the prognostic significance of HER2 amplification in CRC remains unclear, it may serve as a predictive marker for resistance to anti-EGFR therapies.

 amplification or overexpression occurs in approximately

 3-5% of metastatic CRC, with a higher reported prevalence in RAS/BRAF wild type tumors

Testing for HER2 overexpression is generally performed via immunohistochemistry (IHC) assays, with positivity defined as 3+ staining in greater than 50% of tumor cells.

Reflex testing with fluorescence in situ hybridization (FISH) is recommended in patients with IHC 2+ to identify 

 amplification, with positivity defined as a HER2:CEP17 ratio greater than or equal to 2 in more than 50% of tumor cell.

Next generation sequencing (NGS) may also be used to identify 

 amplifications, and MSI/MMR status; testing may be performed individually or via NGS (preferred).

The FDA has approved two targeted therapies for HER2+ metastatic CRC, tucatinib in combination with trastuzumab and trastuzumab deruxtecan.

On January 19, 2023, the FDA granted accelerated approval to tucatinib in combination with the anti-HER2 monoclonal antibody, trastuzumab, for RAS wild-type HER2+ unresectable or metastatic CRC that has progressed on chemotherapy with fluoropyrimidine, oxaliplatin, and irinotecan.

 This approval was based on findings from the phase 2 MOUNTAINEER study (NCT03043313).

On April 5, 2024, the Food and Drug Administration granted accelerated approval to fam-trastuzumab deruxtecan-nxki (Enhertu, Daiichi Sankyo, Inc.) for adult patients with unresectable or metastatic HER2-positive (IHC3+) solid tumors who have received prior systemic treatment and have no satisfactory alternative treatment options.

Efficacy was evaluated in 192 adult patients with previously treated unresectable or metastatic HER2-positive (IHC 3+) solid tumors who were enrolled in one of three multicenter trials: DESTINY-PanTumor02 (NCT04482309), DESTINY-Lung01 (NCT03505710), and DESTINY-CRC02 (NCT04744831).

Adult patients with unresectable or metastatic HER2-positive (IHC 3+) solid tumors who have received prior systemic treatment and have no satisfactory alternative treatment options.

Trastuzumab deruxtecan (T-DXd) is a HER2-directed ADC consisting of a humanized anti-HER2 IgG1 antibody linked to the topoisomerase I inhibitor payload, deruxtecan, via a cleavable tetrapeptide linker.

Upon binding HER2 on tumor cells, T-DXd undergoes internalization and intracellular linker cleavage, thereby releasing the DXd payload and subsequently causing DNA damage and apoptotic cell death.

Learn more about Fam-Trastuzumab Deruxtecan nxki (T-DXd) >

https://www.enhertuhcp.com/en/support-and-resources/for-patients-and-caregivers

In combination with trastuzumab for the treatment of adult patients with RAS wild-type HER2-positive unresectable or metastatic colorectal cancer that has progressed following treatment with fluoropyrimidine-, oxaliplatin-, and irinotecan-based chemotherapy.

Tucatinib is a tyrosine kinase inhibitor of HER2. 

In vitro, tucatinib inhibits phosphorylation of HER2 and HER3, resulting in inhibition of downstream MAPK and AKT signaling and cell proliferation, and showed anti-tumor activity in HER2 expressing tumor cells.

In vivo, tucatinib inhibited the growth of HER2 expressing tumors. 

The combination of tucatinib and trastuzumab showed increased anti-tumor activity in vitro and in vivo compared to either drug alone.

https://www.tukysa.com/mbc/resources-and-support 

Rubin I and Yarden, Y. The basic biology of HER2. 

Schechter AL, Stern DF, Vaidyanathan L, et al. The neu oncogene: an erb-B-related gene encoding a 185,000-Mr tumour antigen. 

Ahcene Djaballah S, Daniel F, Milani A, Ricagno G, Lonardi S. HER2 in colorectal cancer: the long and winding road from negative predictive factor to positive actionable target. 

Sartore-Bianchi A, Lonardi S, Martino C, et al. Pertuzumab and trastuzumab emtansine in patients with HER2-amplified metastatic colorectal cancer: the phase II HERACLES-B trial. 

Siena S, Di Bartolomeo M, Raghav K, et al; DESTINY-CRC01 investigators. Trastuzumab deruxtecan (DS-8201) in patients with HER2-expressing metastatic colorectal cancer (DESTINY-CRC01): a multicentre, open-label, phase 2 trial. 

Strickler JH, Ng K, Cercek A, et al. MOUNTAINEER: open-label, phase II study of tucatinib combined with trastuzumab for HER2-positive metastatic colorectal cancer (SGNTUC-017, trial in progress). 

Sartore-Bianchi A, Amatu A, Porcu L, et al. HER2 positivity predicts unresponsiveness to EGFR-targeted treatment in metastatic colorectal cancer. 

Raghav K, Loree JM, Morris JS, et al. Validation of HER2 amplification as a predictive biomarker for anti-epidermal growth factor receptor antibody therapy in metastatic colorectal cancer. 

National Comprehensive Cancer Network. Clinical Practice Guidelines in colon cancer, version 1.2024. Accessed April 22, 2024. 

National Comprehensive Cancer Network. Clinical Practice Guidelines in rectal cancer, version 1.2024. Accessed April 22, 2024. 

TUKYSA (tucatinib). Prescribing information. Bothell, WA, USA: Seagen Inc., 2023. 

https://docs.seagen.com/TUKYSA_Full_Ltr_Master.pdf

ENHERTU (fam-trastuzumab deruxtecan-nxki). Prescribing information. Basking Ridge, NJ, USA: Daiichi Sankyo, Inc., 2024. 

https://daiichisankyo.us/prescribing-information-portlet/getPIContent?productName=Enhertu&inline=true

Colorectal Cancer

Nivolumab and Ipilimumab

Initial US Approval

Indications

Dose Reductions for Adverse Reactions

Pivotal Studies

Safety

References