Resistance Mutations Pose Complex Challenges for Pathologists


The emergence of resistance mutations in patients with cancer who receive targeted therapies is an expected development that will require new diagnostic methods of identifying the mechanisms through which these alterations occur, according to Fei Dong, MD, during the 2019 Association for Molecular Pathology Annual Meeting.<br /> &nbsp;

Fei Dong, MD

The emergence of resistance mutations in patients with cancer who receive targeted therapies is an expected development that will require new diagnostic methods of identifying the mechanisms through which these alterations occur, according to Fei Dong, MD.

Large-scale next-generation sequencing (NGS) studies, such as The Cancer Genome Atlas, have resulted in the characterization of most primary targetable oncogenic mutations in common cancer types, Dong said during a presentation at the 2019 Association for Molecular Pathology Annual Meeting.1He finds novel resistance mutations to be an interesting and developing area.

&ldquo;In the last few years within our field we&rsquo;ve seen tremendous change,&rdquo; said Dong, an assistant professor of pathology at Brigham and Women's Hospital in Boston, Massachusetts. &ldquo;Specifically, we&rsquo;ve seen the transition from single-gene tests to NGS-based genotyping and cancer diagnostics. This has led to increased complexity in variant analysis and clinical interpretation.&rdquo;

In an interview withTargeted Oncology, Dong said that resistance mutations present new challenges for molecular pathologists. &ldquo;The emergence of resistance mutations means that we have to optimize our assays to detect the new areas within the genes that we were previously testing to identify mutations that are known to be mechanistically important and we have to adjust our assays appropriately as new mechanisms of resistance pop up,&rdquo; he said.

During his presentation, Dong stressed that cancers are characterized by clonal evolution. &ldquo;Cancer is a clonal process, but within a cancer clone there can be genetically defined subclones, and during the process of cancer evolution there can be branch events that represent key events in either cancer progression or metastasis, or when the cancer is exposed to selection pressure, such as exposure to certain types of therapies,&rdquo; he said.

To illustrate that point, Dong discussed 3 areas: resistance to tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML), lung cancer, and colorectal cancer (CRC); mechanisms that cause resistance to hormone therapy in breast cancer and PARP inhibitor therapy inBRCA-deficient tumors; and emerging evidence of immune evasion in response to immune checkpoint inhibitors (ICIs).

In CML, Dong detailed the development of TKIs that inhibit the BCR-ABL tyrosine kinase, starting with the discovery in 1960 of the Philadelphia chromosome abnormality that causes the alteration. &ldquo;The story of BCR-ABL is really tremendous and it&rsquo;s classic,&rdquo; he said. &ldquo;It&rsquo;s a paradigm within our field about targeted therapies against genetic alterations and it&rsquo;s also a story that spans over 50 years.&rdquo;

Resistance mutations that develop in response to imatinib (Gleevec) therapy, the first drug targeting BCR-ABL to gain FDA approval, were first observed inABL1in 2002, Dong said. Molecular analyses identified kinase domain mutations, particularly at T315I, as the cause of resistance, and ponatinib (Iclusig) was shown to specifically target the aberration.2

In non—small cell lung cancer (NSCLC), the correlation betweenEGFRmutation and gefitinib (Iressa) therapy was first established in 2004, according to Dong. The T790M mutation was subsequently identified as conferring resistance to EGFR inhibitors. Osimertinib (Tagrisso) was approved in 2015 specifically for patients with the T790M mutation who had progressed on prior EGFR TKI therapy; it has since been approved for first-line therapy in those with metastatic NSCLC that harborsEGFRexon 19 deletions or exon 21 L858R mutations.3Before osimertinib gained approval though, investigators had already found acquired resistance mutations to the agent inEGFRC797S by conducting NGS testing on cell-free plasma DNA samples.4

InALKfusion—positive NSCLC, multiple resistance mutations have been characterized in patients who receive ALK-targeting TKIs, most frequently in L1196M and G1269A. Investigators found that each ALK inhibitor has a &ldquo;distinct spectrum&rdquo; of resistance mutations; 1 abberation, a mutation inALKG1202R, confers high-level resistance to second-generation ALK inhibitors.5

In CRC, an analysis of circulating tumor DNA from patients who initially responded to EGFR TKIs showed mutations inKRASand theEGFRectodomain as well asKRASandMETamplification. Investigators also showed thatKRASmutations changed dynamically with treatment, Dong said.6

Turning his attention to other types of therapy, Dong pointed out that mutations in theESR1ligand-binding domain, most commonly in codons 536 to 538 and E380Q, are associated with treatment with aromatase inhibitors (AIs). Nevertheless, patients withESR1mutations still respond to estrogen receptor therapy, although study findings show that progression-free survival (PFS) is higher for those treated with fulvestrant (Faslodex), a selective estrogen receptor degrader, compared with the AI exemestane (HR, 0.52; 95% CI, 0.30-0.92;P= .02).7

For agents that target DNA repair pathways, Dong noted that secondaryBRCAmutations have been implicated in resistance to PARP inhibitors. AlthoughBRCA1/2-mutated tumors are responsive to PARP inhibition in several malignancies, preclinical studies in pancreatic cancer cell lines show that acquired deletion of a frameshift mutation inBRCA2confers resistance to PARP therapy.8

In terms of ICI immunotherapies, understanding of mechanisms of resistance is in the early stages, Dong said. In one study, investigators examined paired tumor specimens before and after therapy from patients with NSCLC who developed resistance after receiving a single-agent PD-1 inhibitor or a combination of PD-1 and CTLA-4 ICIs. They found that resistant clones had lost 7 to 18 mutation-associated neoantigens, damaging the T cells&rsquo; ability to recognize the tumor cells.8

Overall, Dong said, genomic changes that occur as a result of acquired resistance are more complex types of mutations that are important to identify for selecting therapies for later lines of treatment. &ldquo;The identification of these resistance mutations is not just for academic purposes,&rdquo; he said. &ldquo;The appropriate clinical variant annotations by us can guide the appropriate treatment selections for the patient in terms of secondary and tertiary therapy. As a field, we should anticipate the need for new methods to identify genetic resistance mechanisms.&rdquo;


  1. Dong F. Novel mechanisms of acquired resistance to targeted therapies in cancer. Presented at: 2019 Association for Molecular Pathology Annual Meeting; November 7-9, 2019; Baltimore, MD.
  2. O'Hare T, Shakespeare WC, Zhu X, et al. AP24534, a pan-BCR-ABL inhibitor for chronic myeloid leukemia, potently inhibits the T315I mutant and overcomes mutation-based resistance.Cancer Cell.2009;16(5):401-412. doi: 10.1016/j.ccr.2009.09.028.
  3. Tagrisso [prescribing information]. Wilmington, DE: AstraZeneca Pharmaceuticals LP; 2018. Accessed November 8, 2019.
  4. Thress KS, Paweletz CP, Felip E, et al. Acquired EGFR C797S mutation mediates resistance to AZD9291 in non-small cell lung cancer harboring EGFR T790M.Nat Med. 2015;21(6):560-562. doi: 10.1038/nm.3854.
  5. Gainor JF, Dardaei L, Yoda S, et al. Molecular mechanisms of resistance to first- and second-generation ALK inhibitors in ALK-rearranged lung cancer.Cancer Discov.2016;6(10):1118-1133. doi: 10.1158/2159-8290.CD-16-0596.
  6. Siravegna G, Mussolin B, Buscarino M, et al. Clonal evolution and resistance to EGFR blockade in the blood of colorectal cancer patients [erratum inNat Med. 2015;21(7):827].Nat Med.2015;21(7):795-801. doi: 10.1038/nm.3870.
  7. Fribbens C, O&rsquo;Leary B, Kilburn L, et al. Plasma ESR1 mutations and the treatment of estrogen-receptor positive advanced breast cancer.J Clin Oncol.2016;34(25):2961-2968. doi: 10.1200/JCO.2016.67.3061.
  8. Edwards SL, Brough R, Lord CJ, et al. Resistance to therapy caused by intragenic deletion in BRCA2.Nature. 2008;451(7182):1111-1115. doi: 10.1038/nature06548.
  9. Anagnostou V, Smith KN, Forde PM, et al. Evolution of neoantigen landscape during immune checkpoint blockade in non-small cell lung cancer.Cancer Discov.2017;7(3):264-276. doi: 10.1158/2159-8290.CD-16-0828.
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