Issue 05 | May 2017

Welcome to Strand Genomics-A Monthly E-zine from Strand Life Sciences

Strand Life Sciences welcomes you to Strand Genomics, our monthly E-zine that includes articles of interest to physicians. This e-zine brings the latest news in the world of genetic diagnostics, to your doorstep. The E-zine features carefully crafted articles as well as curated news in the field of cancer therapy and genetic analyses to support the implementation of personalized medical care. We invite you to peruse as well as share these articles. Please also feel free to write back to us with comments and questions at strandlive@strandls.com

Liquid Biopsy Aided Genetic Analyses for Personalized Management of Cancer

Prof. Vijay Chandru
Chairman and Managing Director, Strand Life Sciences

Abstract

  • Liquid biopsy enabled analysis of circulating  tumor DNA (ctDNA) offers a chance to create personalized cancer management options.
  • Liquid biopsy based analyses offer a chance to assess patient responses to administered therapy at customised time intervals.
  • Liquid biopsy tests therefore offer an extremely convenient and minimally-invasive alternative technique to obtain a ‘biopsy’ of deep-seated, solid tumors, as and when required.
  • Liquid biopsy based DNA analyses therefore represent a huge leap forward in enabling delivery of cancer care.

Introduction

Personalized treatment is the next frontier in management of cancer patients. Oncologists have known for the past few decades that a ‘One Therapy Fits All’ approach does not work with all cancer patients and eventually cancers of all kinds develop resistance to known therapies. The ability to obtain a ‘genetic signature’ of cancers on a per patient basis has the potential for delivery of personalized treatment to each and every cancer patient.

Liquid biopsy is a game changer that can solve these issues. Liquid biopsy tests for cancers are essentially blood tests that can be used to identify such genetic signatures. Most cancer cells shed DNA into the bloodstream. This DNA can be harvested and then analyzed to build a catalog (list) of mutations that are present in the cancer cells of a patient. Effectively, this catalog is the genetic identifier of a patient’s cancer, akin to a label or a tag for the cancer.

Cancer Therapy Can Drain Resources

The fight against cancer is an emotional and financial rollercoaster ride. All of us have lost a loved one to this disease. Suppose a person is suffering from lung-related problems and his doctor suspects the presence of cancer. Now this person has to undergo a biopsy to obtain some lung tissue. It is known that in~30% of lung cancer cases, getting a sold tissue biopsy of good quality is the first hurdle. Even is a biopsy is obtained and the diagnosis of lung cancer is confirmed, the patient has to face the next hurdle of expensive chemotherapy and radiotherapy. Even after extensive therapy, there is always a nagging doubt that the cancer has not all gone away and there will be a relapse. Currently, a PET-CT scan, done once a year, is the only method of checking if the cancer is likely to come back or not. Going through the whole cycle of obtaining a solid tissue biopsy and therapy is definitely a hugely expensive and emotionally-draining experience.

Liquid Biopsy Can Provide Answers Within An Actionable Timeframe

In such a setting, liquid biopsy tests can offer significant relief. Liquid biopsy can provide enough cancer DNA for the initial diagnosis a well as for second, third and fourth rounds of biopsies. Not only is it easy to draw a small amount of blood, it is also convenient and can be done at a patient’s home as well. Moreover, liquid biopsy tests specifically developed for lung cancer can help an oncologist choose one amongst a set of new, targeted drugs that are highly specific for lung cancer.

Similarly, after the first round of therapy, all that a person has to do is provide another liquid biopsy sample. Scientists can estimate the remaining cancer load in the liquid biopsy and can understand whether this person is becoming resistant to prior therapy or not.

Liquid biopsy tests therefore offer an extremely convenient and minimally-invasive alternative technique to obtain a ‘biopsy’ of deep-seated, solid tumors, as and when required.

Liquid biopsy tests are inexpensive, no exposure to radiation involved, and a readout of the patient status can be obtained within 5 working days. Moreover, there is a limit to how many PET/CT scans a person can undergo, whereas there is no limit to the number of times a small amount of blood can be drawn from a person. Some early scientific studies have also shown that liquid biopsy tests provide an indication of cancer up to 9 months ahead of detection of cancer by scans and other diagnostic tools.

Given that liquid biopsy tests can provide a glimpse of tumor DNA, throughout the course of the disease, oncologists can now understand how and when a patient’s cancer responds to chemotherapy, why tumor cells stop responding to chemotherapy and how early should a patient be switched over to better and specific drugs. The ability to track changes in a patient’s cancer cells will now allow doctors and scientists to literally ‘walk in-step with a patient’s cancer’ and decide how and when to change chemotherapy drugs for that particular patient.

Summary

  • Genetic analyses are set to become companion diagnostic tests for therapies in many diseases, apart from cancer, in the forthcoming years.
  • Liquid biopsy based DNA analyses therefore represent a huge leap forward in enabling delivery of healthcare.
  • Liquid biopsy tests can provide efficient cancer progression profiles in a minimally-invasive manner and serve as reliable therapeutic decision-support aids.
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Monitoring Clonal Evolution Of Cancers Using Liquid Biopsies

Adapting targeting treatment to cancer progression

Treatment with targeted therapies places tumour cells under selective pressure, thereby triggering clonal evolution that can be captured using liquid biopsy approaches. The data obtained can provide important insights into mechanisms of resistance, and can guide therapeutic decision-making. This schematic depicts the dynamic changes in the abundance of tumour-cell subclones harbouring different mutations in a patient with metastatic colorectal cancer treated with anti-EGFR antibodies. Monitoring of a clonal mutation in APC that is present in essentially all tumour cells (clone 1, blue line) tracks tumour burden, while subclonal mutations (KRASG12D, KRASQ61H, and EGFRG465R in clones 2, 3 and 4, respectively) provide a measure of clonal evolution during therapy. Subclonal mutations can be lesion-specific, leading to variations in responses at different disease sites. In this hypothetical patient, primary treatment with the anti-EGFR antibody leads to a substantial decline in tumour burden through targeting of the majority of tumour cells, but outgrowth of the resistant KRASG12D-mutant subclone eventually causes tumour regrowth, necessitating a therapeutic switch. The change in therapy decreases the size of the KRASG12D-cell population, but resistant subclones (clones 3 and 4) with other mutations expand and drive tumour growth. A third-line of treatment restricts the growth of clone 3, but clone 4 continues to proliferate. (Siravegna, G. et al., 2017, doi:10.1038/nrclinonc.2017.14)

ctDNA: Prognostic Marker for Lung Cancer?

Dr. Shefali Sabharanjak
Strand Life Sciences

Abstract

  • In NSCLC patients, presence of ctDNA at baseline and follow-up timepoints, is associated with reduced PFS and OS.
  • Presence of ctDNA correlates with incidence of liver metastases and high proliferative capacity of tumor cells.
  • Occurrence of ctDNA is independent of the type of genetic mutations that cause NSCLC
  • Assessment of ctDNA at 6 week intervals, during the course of therapy, can provide insights into tumor burden and possible outcomes.

Introduction

Lung cancer has the dual distinction of high incidence and high fatality. Estimates from GLOBOCAN indicate that in 2008, 1.6 million new lung cancer cases were recorded around the globe and 1.37 million deaths worldwide, were attributed to lung cancer (Furrukh et al. 2013). The median survival time for lung cancer (non-small cell lung cancer, NSCLC) is 7.1 months without therapy (Wao et al. 2013) and 10 months with chemo- as well as targeted therapy (Furrukh et al. 2013). Given that survival statistics are not optimistic, development of prognostic tools that can provide rapid and quick insights about tumor load would be highly valuable.

Assessment of ctDNA, facilitated by liquid biopsy, can serve as one such prognostic marker. We have shared preliminary results from Strand’s internal study about ctDNA being a possible prognostic marker for several cancers, in the previous issue of Strand Genomics. Early results from Strand’s tumor concordance study show that patients with high ctDNA load were lost to cancer despite the histological characterization of tumors as early or non-invasive tumors.

Recently, another study with lung cancer patients has highlighted the prognostic value of monitoring ctDNA during the course of treatment. One hundred and nine patients with newly diagnosed non-small cell lung cancer (NSCLC) were included in this study. Tumor biopsies were analyzed using NGS to identify mutations (KRAS, EGFR, TP53, EGFR(T790M), BRAF, PIK3CA, SMAD4, STK11, to name a few) that could be identified using digital PCR (dPCR) on ctDNA. Plasma samples were collected prior to commencing chemotherapy or treatment with EGFR TKIs (T0- baseline), at 6±2 weeks (E1, First Evaluation) and at time of progression (ToP)(Pécuchet et al. 2016).

Results from the follow-up period (median 18.8 months) show that presence of ctDNA was a strong prognostic indicator for progression of the disease. Negative or low baseline ctDNA was associated with low tumor load (Odds ratio (OR) 0.24 [95% CI 0.08–0.70]) as well as lower chances of bone metastasis (OR 0.34 [95% CI 0.14–0.83]). Conversely, the presence of intermediate to high levels of ctDNA at baseline was correlated with reduced overall survival (OS) (median 13.6 versus 21.5 mo, p = 0.03, Figure 1B) as well as reduced progression-free survival (PFS) (median 4.9 versus 10.4 mo, p < 0.001, Figure 1A).

Figure 1. Presence of ctDNA is associated with reduced PFS and OS in NSCLC patients (Pécuchet et al. 2016).

Moreover, multivariate analyses showed that Hazard Ratios for OS as well as PFS were 1.82 and (95% CI 1.01–3.55, p = 0.045) and 2.14 (95% CI, 1.30–3.67 p = 0.002) (Pécuchet et al. 2016), when ctDNA was detected in patients at baseline as well as throughout the course of treatment and follow-up.

High ctDNA Levels Indicate Secondary Metastases and High Proliferative Potential

Patients in this study were segregated into high, intermediate and low tertiles based on the amount of ctDNA detected at baseline. Patients with high levels of ctDNA had higher tumor burden as assessed by size of tumor (RECIST criteria, see Figure 2A). Additionally, the incidence of liver metastases was higher in patients with high ctDNA (Figure 2B). The proliferative capacity of lung cancer was assessed by Ki67 immunohistochemistry of tumor biopsy samples (Figure 2C). Interestingly, the proliferative capacity of the tumors was correlated positively with high levels of ctDNA in circulation.

Figure 2. Correlation of levels of ctDNA and incidence of tumor burden (A), liver metastases (B) and proliferative potential (C)

Further analyses of data from the study also show that lower ctDNA at baseline as well as at E1 (6±2 weeks) was also associated with lower RECIST scores (Figure 3).

Figure 3. Change in levels of ctDNA correlates with RECIST scores of lung cancer patients.

Levels of ctDNA in this study were not correlated with the genetic mutation(s) identified from tissue biopsies (Supplementary data in the paper, available online). The presence or absence of ctDNA was a better prognostic marker, independent of the mutant genes that were identified using NGS as well as dPCR tests, from tumor tissue as well as from plasma samples (Pécuchet et al. 2016).

Results from this study suggest that periodic monitoring of patients using liquid biopsy based assessment of ctDNA can be a useful prognostic tool. Liquid-biopsy based genetic analyses have been useful in assessing the development of resistance to TKI therapy in lung cancer patients (Zheng et al. 2016; Remon et al. 2017). Additionally, assessment of ctDNA using other genes like KRAS and BRAF as markers can serve as a prognostic tool for patients receiving non-TKI therapy as well.

Summary

  • In NSCLC patients, presence of ctDNA at baseline and follow-up timepoints, is associated with reduced PFS and OS
  • Presence of ctDNA correlates with incidence of liver metastases and high proliferative capacity of tumor cells
  • Occurrence of ctDNA is independent of the type of genetic mutations that cause NSCLC
  • Assessment of ctDNA at 6 week intervals, during the course of therapy, can provide insights into tumor burden and possible outcomes

References

Furrukh, M. et al., 2013. Improving Outcomes in Advanced Lung Cancer: Maintenance therapy in non-small-cell lung carcinoma. Sultan Qaboos University medical journal, 13(1), pp.3–18. Available at: http://www.ncbi.nlm.nih.gov/pubmed/23573377 [Accessed April 28, 2017].

Pécuchet, N. et al., 2016. Base-Position Error Rate Analysis of Next-Generation Sequencing Applied to Circulating Tumor DNA in Non-Small Cell Lung Cancer: A Prospective Study. M. Ladanyi, ed. PLoS medicine, 13(12), p.e1002199. Available at: http://www.ncbi.nlm.nih.gov/pubmed/28027313 [Accessed April 24, 2017].

Remon, J. et al., 2017. Osimertinib benefit in EGFR -mutant NSCLC patients with T790M -mutation detected by circulating tumour DNA. Annals of Oncology, p.mdx017. Available at: http://www.ncbi.nlm.nih.gov/pubmed/28104619 [Accessed January 30, 2017].

Wao, H. et al., 2013. Survival of patients with non-small cell lung cancer without treatment: a systematic review and meta-analysis. Systematic reviews, 2, p.10. Available at: http://www.ncbi.nlm.nih.gov/pubmed/23379753 [Accessed April 28, 2017].

Zheng, D. et al., 2016. Plasma EGFR T790M ctDNA status is associated with clinical outcome in advanced NSCLC patients with acquired EGFR-TKI resistance. Scientific reports, 6, p.20913. Available at: http://www.ncbi.nlm.nih.gov/pubmed/26867973 [Accessed February 16, 2017].

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