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High-Throughput Sequencing of T-Cell Receptors May Transform Diagnosis and Treatment of Cutaneous T-Cell Lymphomas


High-throughput sequencing of T-cell receptors may be a solution to some of the challenges confronting oncologists who treat cutaneous T-cell lymphomas. According to experts who presented talks on high-throughput sequencing of T-cell receptors at the 3rd World Congress of Cutaneous Lymphoma (sponsored by the International Society for Cutaneous Lymphomas), high-throughput sequencing can reliably aid in the diagnosis of cutaneous T-cell lymphomas, differentiating them from nonmalignant skin dyscrasias, and can monitor the effects of treatment. With other techniques, the ability to do both things has been limited.

Diagnosis of cutaneous T-cell lymphomas and monitoring of treatment have been hampered by the variability in presentation and histopathology of this cancer, lack of standardized definitions, their relative infrequency, and a clinical presentation that often mimics benign skin conditions such as psoriasis.

Identifying Malignant Clones

Ilain Kirsch, MD

Ilain Kirsch, MD

High-throughput sequencing of T-cell receptors is currently used only by a “handful of academic centers” for cutaneous T-cell lymphomas, noted Ilain Kirsch, MD, of Adaptive Biotechnologies, Seattle, which markets high-throughput immune receptor sequencing. A recent study by Dr. Kirsch and colleagues showed that high-throughput sequencing of T-cell receptors facilitated the diagnosis of cutaneous T-cell lymphomas and was superior to polymerase chain reaction in identifying malignant clones.1

High-throughput sequencing of T-cell receptors was able to identify malignant T-cell clones at sites of skin lesions in all 39 patients included in the study, whereas polymerase chain reaction correctly identified 27 of the 39 patients (70%). Furthermore, 10 of the 12 patients with detectable clonality on high-throughput sequencing of T-cell receptors (but negative on polymerase chain reaction) had early-stage disease (stage IA or IB), allowing for earlier treatment.

“It’s better to catch these patients early and treat them before they become more advanced. Earlier treatment may not necessarily result in a cure, but hopefully many patients will at least have a better quality of life and relief from some of the signs or symptoms of their disease,” Dr. Kirsch explained.

This same study found that high-throughput sequencing of T-cell receptors was able to accurately assess response to therapy and facilitated diagnosis of recurrences in patients with cutaneous T-cell lymphomas. Moreover, high-throughput sequencing of T-cell receptors provided insights into the cell origin (mature T cells) and location of malignant cutaneous T-cell lymphoma.

High-Throughput Sequencing vs Polymerase Chain Reaction


A goal at our center is to develop and implement high-throughput sequencing of T-cell receptors as a powerful tool to improve the diagnosis, follow-up, and prognosis of patients with cutaneous T-cell lymphomas.
— John T. O’Malley, MD, PhD

“High-throughput sequencing has advantages over [polymerase chain reaction] of the T-cell receptor,” said John T. O’Malley, MD, PhD, of Dana-Farber/Brigham and Women’s Cancer Center, Harvard Medical School, Boston, who works in the laboratory of ­Rachael A. Clark, MD, PhD. At the 3rd World Congress of Cutaneous Lymphoma, Dr. O’Malley presented results of a study comparing high-throughput sequencing of T-cell receptors with polymerase chain reaction T-cell receptors in 46 patients with cutaneous T-cell lymphoma.2

“A goal at our center is to develop and implement high-throughput sequencing of T-cell receptors as a powerful tool to improve the diagnosis, follow-up, and prognosis of patients with cutaneous T-cell lymphomas,” Dr. O’Malley told listeners.

He and his colleagues (among them Dr. Kirsch) sought to determine whether high-throughput sequencing of T-cell receptors could provide more rapid and accurate diagnoses and which cutaneous T-cell lymphoma therapies kill malignant clones vs only suppress visible inflammation. In addition, they worked to identify how to use high-throughput sequencing of T-cell receptors to discriminate patients with early-stage cutaneous T-cell lymphoma at high risk for eventual disease progression (which applies to about 20% of patients diagnosed with this type of cancer).

High-throughput sequencing of T-cell receptors and polymerase chain reaction T-cell receptors were used to examine isolated DNA from blood and skin lesions from 46 patients with cutaneous T-cell lymphomas of all stages, patients with other inflammatory diseases, and healthy controls.

“We found that high-throughput sequencing could reliably distinguish between cutaneous T-cell lymphomas and other conditions,” said Dr. O’Malley. High-throughput sequencing was more sensitive and specific than polymerase chain reaction, detecting T-cell clones in all 46 patients and discriminating between cutaneous T-cell lymphomas and psoriasis, eczema, and healthy skin.

Assessing Therapeutic Responses

“Next, we wanted to use high-throughput sequencing to assess responses to therapy,” continued Dr. O’Malley. “Patients can respond wonderfully to topical therapy, but then skin lesions recur. This raises the question of whether malignant cells are still present in the treated skin lesions of the responding patients.”

The investigators assessed patients diagnosed with cutaneous T-cell lymphomas at the site of a skin lesion using high-throughput sequencing of T-cell receptors before and after three types of commonly used therapies: low-dose brachytherapy, psoralen plus ultraviolet light (PUVA) [in collaboration with Peter Wolf, MD, and Pablo
Vieyra Garcia, PhD
, at the Medical University of Graz, Austria], and topical resiquimod (a potent Toll-like receptor 7/8 agonist) [in collaboration with Alain Rook, MD, at the University of Pennsylvania].

Novel Sequencing Method in Cutaneous T-Cell Lymphoma

  • High-throughput sequencing of T-cell receptors can distinguish between cutaneous T-cell lymphoma and benign skin disease, allowing for accurate and timely diagnosis. It may prove to be superior to polymerase chain reaction high-throughput sequencing for diagnosis.
  • High-throughput sequencing of T-cell receptors may be a reliable way to monitor response to therapy.
  • Use of high-throughput sequencing of T-cell receptors is currently found in only a handful of academic centers, but it may become the wave of the future for cutaneous T-cell lymphomas.

For both low-dose brachytherapy and topical resiquimod, clinical response was correlated with eradication of the malignant clone, whereas this was not true for PUVA.

In 10 patients treated with low-dose brachytherapy, 4 had complete eradication of the malignant clone, sparing benign cells. In a small study of 12 patients treated with resiquimod (stages IB, IA, and IIA),3 9 had greater than 50% improvement in the treated skin lesions, with 2 of the complete responders having eradication of the malignant clone. The patients who had a partial response in the skin also showed a reduction in the malignant clone, with the exception of one patient who had an increase.

By contrast, eight patients with early-stage cutaneous T-cell lymphomas who were treated with twice-weekly PUVA had excellent clinical responses, but none of the patients had eradication of the malignant clone. Half of the patients had a similar malignant clone burden at the end of therapy.

“Clinical response to PUVA does correlate with reduction of benign T cells. These data suggest that PUVA does not kill malignant T cells, but it does suppress inflammation. These insights suggest that high-throughput sequencing of T-cell receptors has the potential to categorize our therapies,” Dr. O’Malley continued.

“We may be able to differentiate between ‘cidal’ therapies (ie, low-dose brachytherapy and topical resiquimod gel) and static therapies (like PUVA). This whets the appetite that some therapies do a better job of eradicating the malignant clone, while PUVA makes the inflammation go away, but the clone remains in the skin lesion. The clinical relevance of this remains to be determined,” he added. ■

Disclosure: Dr. Kirsch is an employee of Adaptive Biotechnologies, which markets high-throughput sequencing. Dr. O’Malley reported no potential conflicts of interest.

References

1. Kirsch IR, Watanabe R, O’Malley JT, et al: TCR sequencing facilitates diagnosis and identifies mature T cells as the cell of origin in CTCL. Sci Transl Med 7:308ra158, 2015.

2. O’Malley JT, Kirsch F, Lowry E, et al: High-throughput T cell receptor sequencing transforms care of cutaneous T cell lymphoma patients. 3rd World Congress of Cutaneous Lymphomas. Abstract G-05. Presented October 27, 2016.

3. Rook A, Gelfland J, Wysocka M, et al: Topical resiquimod can induce disease regression, eradicate malignant T cells and enhance T cell effector functions in CTCL. Blood 126:1452-1461, 2015.


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