Expert Perspectives on Molecular Testing in Non–Small Cell Lung Cancer

Comprehensive discussion on molecular testing strategies utilized in non–small cell lung cancer to help inform treatment decisions.


Stephen Liu, MD: In this particular case, bilateral lung nodules as large as 2 cm, enlarged lymph nodes, 2 brain metastases, they have a transbronchial biopsy that does confirm adenocarcinoma. The PD-L1 level is 20% by IHC [immunohistochemistry]. I think that’s listed here because that comes back pretty quickly. Immunohistochemistry can be back as soon as the next day. Tissue is sent for molecular testing, blood is sent for molecular testing. Let’s pause here and talk about testing. I think this is critical because our focus here is ALK [fusions]. In order to treat ALK properly, we have to find it when it’s there. Jyoti, are you testing for every genetic marker, or are you doing sort of a stepwise sequential approach?

Jyoti Patel, MD: Over the past few years, we’ve adopted comprehensive testing for everyone. With limited tissue and in the interest of time, and with the fact that we have so many markers that are actionable, our approach is to do multiplex testing, which includes RNA.

Stephen Liu, MD: So you’re doing DNA and RNA multiplex testing. Next-generation sequencing [NGS], is that done in house at Northwestern [University], or is that through a commercial laboratory?

Jyoti Patel, MD: It is done in house, and we have put in place the ability to have it reflexively ordered and done by the pathologist. That saves time for, at the time of histologic confirmation, slides are obtained and sent to the lab without that delay of another person needing to order.

Stephen Liu, MD: Jillian, at Georgetown [University], when you see a new patient here, what’s your genetic testing approach?

Jillian E. Thompson, NP: Typically, we’re going to make sure that we, as well, do DNA and RNA sequencing, next-generation sequencing, and we also do liquid biopsies. We’re doing that from the start to get those results to have comprehensive testing to see any possible targeted mutations.

Stephen Liu, MD: We’ve got 2 votes for DNA and RNA sequencing on tissue up front. Ross, a similar approach?

Ross Camidge, MD, PhD: Similar approach. The only thing we supplement with in addition to the PD-L1 testing is, I do still have concerns that next-generation sequencing doesn’t [catch] amplification very well. Now, does that matter here? There’s nothing licensed, but MET and HER2 amplification are 2 things to watch. So we send those off to FISH [fluorescence in situ hybridization] testing in addition to the NGS.

Stephen Liu, MD: Is your NGS in house or is it commercial?

Ross Camidge, MD, PhD: It’s in house.

Stephen Liu, MD: Both DNA and RNA?

Ross Camidge, MD, PhD: Yes.

Stephen Liu, MD: We’ve thrown these letters around a bit, but for our audience, what’s the importance of doing RNA in addition to DNA? Ross, do you want to explain that?

Ross Camidge, MD, PhD: What we’re really talking about is a next-generation sequencing assay, where we’re talking about the difference in the starting material. Either it’s the genomic DNA from the cancer, or it’s the RNA extracted from the cancer, converted into DNA, and then analyzed. The RNA, because you essentially get rid of the introns, allows you greater sensitivity for picking up fusions, which are essentially 2 genes stuck together. If you’re sequencing through the DNA, there can be a whole bunch of junk stuck in the middle. Also, for MET exon 14 skip mutations, where there’s a hundred different mutations you can get, but the end result is, there’s a bit missing from the RNA transcript. So, it’s really about sensitivity, particularly when you have fusions in MET exon 14. We had a paper a few years ago using MET exon 14 as an example, that if you used a DNA-based extraction, your pickup rate was about 1% or 2%. If you used RNA, your pickup rate was about 4%, which is supposed to be the real rate.

Stephen Liu, MD: Jyoti, I just want to be clear here, it’s RNA in addition to DNA, not RNA or DNA, correct?

Jyoti Patel, MD: Absolutely.

Stephen Liu, MD: We talked about NGS as our preferred modality, giving us the greatest sensitivity in identifying these alterations, but is there any role for some of the older tests like FISH, IHC, PCR [polymerase chain reaction]? Jillian, what do you think?

Jillian E. Thompson, NP: I think there is room for that, especially if you can get those results back quicker, as opposed to the week and a half or 2 weeks for the other results. And those can, in our case, can be done in house.

Stephen Liu, MD: There is a bit more speed, PCR and FISH could come back in a day or two. Ross, any role in your practice for these older assays?

Ross Camidge, MD, PhD: Not really. We do have an immunohistochemistry assay for a rapid turnaround time. The reason I’ve I see it done is…there are many countries around the world that only have ALK FISH or only have ALK IHC. While most of the time you can believe a positive result, there is a small but significant false positive and false negative rate. The reason that’s important is, let’s say you have a patient who is called ALK+ by one of these other techniques, and they don’t respond to the initial ALK inhibitor. The first thing you should doubt is the diagnostic, not switch them to another ALK inhibitor.

Stephen Liu, MD: Absolutely. The response rates, as we see, are so high that when things don’t get better very quickly, we question that. I absolutely agree. Jyoti, what about in the neoadjuvant setting? I know that’s not our focus here today, but in the neoadjuvant setting, could these tests maybe play a role?

Jyoti Patel, MD: Sure, absolutely. Now that there has been a significant embrace of neoadjuvant immunotherapy in patients with resectable disease, I think it’s essential to exclude patients with EGFR and ALK translocations from that approach. So again, in these early biopsies, you want that information very quickly, and it may be that PCR and IHC are appropriate to exclude those patients and get them on to potential curative therapy.

We recently had a patient, a 90-plus-year-old, who had a very aggressive appearing cancer that was very poorly differentiated. We did NGS, and we actually detected an ALK fusion that was quite atypical, a PURA-ALK fusion, which has been described but not really quantified. So I went the other way to convince myself that this was the right thing to do. I did IHC, and it was strongly positive, and he subsequently had a great response to a TKI [tyrosine kinase inhibitor]. So it helps on both ends.

Stephen Liu, MD: It’s nice to have these. This case, interestingly, had tissue and blood sent off, so a liquid biopsy and ctDNA [circulating tumor DNA] testing. Jyoti, does that play a role in your initial work-up?

Jyoti Patel, MD: We have now adopted concurrent testing. Certainly there’s some debate about that. But in my mind, with the likelihood of having either insufficient tissue or a problem in getting results within 7 days, [which is what we] consider an appropriate time frame in someone who has rapidly accelerating disease, we’ve found that doing concurrent blood biopsy is quite helpful. It’s sort of that piece of interpreting a Venn diagram. Some patients will be positive on tissue, some patients will be positive on blood, most are positive on both. But with either positive assay, you can initiate the appropriate therapy.

Stephen Liu, MD: Jillian, what’s your approach on blood-based testing in the initial diagnosis setting?

Jillian E. Thompson, NP: We’ve made that a practice now, we are doing that with all of our patients at baseline. From once not having that and not doing it, now it has become a practice.

Stephen Liu, MD: Ross, is your approach similar?

Ross Camidge, MD, PhD: Yes. I’d like to point out one thing, obviously you’ve got the cream of the crop on this call. My biggest worry is that there are [people] in the community who are just sending the blood-based testing, it comes back with nothing, and they say “I’ve done the NGS,” but they didn’t have a driver [oncogene]. That is something we need to fix, because a negative doesn’t mean a negative on blood.

Stephen Liu, MD: Let’s state that again. We send off a liquid biopsy. That comes back pretty quickly. That’s one of the advantages, [results in] about a week. We get that result, with nothing detected. What should we do then, Ross?

Ross Camidge, MD, PhD: Jyoti and Jillian have already got the tissue up and running in parallel, so they’re sitting pretty. But other people, if they were putting all their eggs in the basket of the blood [testing], then they would probably have to send the tissue. Or they have to shoot from the hip, but bearing in mind that if the patient is a never or light smoker, they haven’t really done the full job of proving that they don’t have a driver oncogene.

Stephen Liu, MD: We believe a positive, and we have very few false positives. If we see an actionable target on a liquid biopsy, we go with it. If we don’t, we keep investigating and we look a bit further. I see very little harm in doing the liquid biopsy. It’s a relatively noninvasive test. I guess the only barrier would be one of cost, which is a real barrier. I think that when I look at liquid and tissue, they’re complementary. It wouldn’t be a problem if they were complimentary, but they’re not. There’s a fee associated with it.

Ross Camidge, MD, PhD: I saw what you did there. That was very clever.

Stephen Liu, MD: But when you think of overall, from a big picture standpoint, the cost of missing one of these alterations, I think is much greater. Because if we don’t see an ALK fusion and it’s there, we are going to treat this person with the wrong treatment, agreed?

Ross Camidge, MD, PhD: Yes.

Transcript edited for clarity.

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