Tumor-Infiltrating Lymphocytes Prognostic in the Metastatic and Neoadjuvant Breast Cancer Settings

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Studies presented at the 2016 San Antonio Breast Cancer Symposium added to growing evidence that tumor-infiltrating lymphocytes are important prognostic factors in breast cancer. One investigation evaluated their impact in advanced HER2-positive breast cancer, finding a linear relationship between tumor-infiltrating lymphocyte levels and overall survival.1 A second study, a meta-analysis of neoadjuvant trials, found them to be a strong predictor of pathologic complete response in all breast cancer subtypes and to be associated with a survival benefit in HER2-positive and triple-negative disease.2

Tumor-infiltrating lymphocytes represent the formation of an adaptive immune response to tumor development. They have been shown to predict for response to neoadjuvant therapy in HER2-positive and triple-negative breast cancer, but their role in luminal tumors and advanced breast cancer has been less clear. The impact of tumor-infiltrating lymphocytes is increasingly relevant in light of emerging immunotherapies in advanced breast cancer.

CLEOPATRA Trial Substudy

A higher level of tumor-infiltrating lymphocytes in patients with advanced HER2-positive breast cancer on the CLEOPATRA trial was associated with overall survival and with several clinicopathologic factors, according to the first study to investigate these relationships in the context of first-line dual HER2 blockade.

We saw a significant association with improved survival, with every 10% rise in stromal tumor-infiltrating lymphocyte levels equating to an 11% reduction in the risk of death.
— Stephen J. Luen, MD

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“We found a significant association with overall survival and a nonsignificant trend for improved progression-free survival,” said Stephen J. Luen, MD, of the Peter MacCallum Cancer Centre in Victoria, Australia. “Each 10% increase in stromal tumor-infiltrating lymphocytes was associated with an 11% reduction in the risk of death. The prognostic effect was not different according to treatment arm, however, so there is no predictive effect with regard to pertuzumab [Perjeta] treatment.”

The phase III CLEOPATRA trial randomized 808 patients with previously untreated HER2-positive advanced breast cancer to treatment with trastuzumab (Herceptin) and docetaxel, or to the same combination plus pertuzumab.3 Median progression-free survival was improved by 6.3 months and overall survival by 15.7 months with the addition of pertuzumab, establishing a new first-line standard.

Dr. Luen and colleagues investigated the prognostic association between stromal tumor-infiltrating lymphocytes and survival in the CLEOPATRA population, asking whether the benefit of pertuzumab differed by level of tumor-infiltrating lymphocytes (ie, would be predictive). In prospectively collected pretreatment tumor samples, they measured tumor-infiltrating lymphocytes in the stroma as a continuous variable, per 10% increments. They had 678 samples, of which 155 were freshly obtained and 519 were archival samples (4 were from unknown sources). They also had 20 paired primary and metastatic samples.

Multiple Associations Observed

Levels of tumor-infiltrating lymphocytes were significantly higher in tumors that were estrogen receptor–negative (P < .001). They were highest in tumors from Asian patients and lowest in tumors from African Americans. Dr. Luen noted that only a small percentage of tumors were from African Americans. Age, tumor grade, and presence of visceral disease at screening were not significantly associated with levels of tumor-infiltrating lymphocytes. Freshly obtained samples had significantly lower levels (P < .001), and there was a trend toward lower levels in metastatic samples (P = .07), with the exception of high levels (30%) in lung metastases.

Tumor-Infiltrating Lymphocytes in Breast Cancer

  • Two studies further validated the concept that high levels of tumor-infiltrating lymphocytes in breast tumors are prognostic of outcomes.
  • An analysis of samples from CLEOPATRA found high levels of these immunologic factors to be associated with overall survival; patients were treated with single or dual HER2 blockade.
  • A meta-analysis of six German studies in the neoadjuvant setting found high levels associated with the achievement of complete pathologic response in all molecular subtypes and with disease-free and overall survival in triple-negative and HER2-positive disease.

The survival analysis showed a nonsignificant trend for an association between increasing levels and better progression-free survival (hazard ratio [HR] = 0.95, P = .06) and a statistically significant association with overall survival (HR = 0.89, P = .001).

“We saw a significant association with improved survival, with every 10% rise in stromal tumor-infiltrating lymphocyte levels equating to an 11% reduction in the risk of death,” Dr. Luen said. “This linear association is consistent with previous reports in early breast cancer.”

The lowest overall survival (36.3 months) was seen in patients who received trastuzumab alone and had tumor-infiltrating lymphocyte levels ≤ 20%. The best survival (not reached) was in patients receiving pertuzumab who had the higher level, > 20%.

While the prognostic effect was stronger in the pertuzumab arm, there was no significant interaction between the effect of tumor-infiltrating lymphocytes and pertuzumab treatment. “Importantly, patients derived a benefit from pertuzumab irrespective of the level of tumor-infiltrating lymphocytes, so they should not be used to determine which patients can be spared pertuzumab,” he said.

The findings show that the positive influence of preexisting antitumor immunity persists in the advanced setting, and strategies to augment immunity may further improve survival, he suggested.

Neoadjuvant Therapy and Lymphocyte Levels

Our results suggest tumor-infiltrating lymphocytes are a strong predictive marker for response to neoadjuvant chemotherapy in all molecular subtypes, and this predictive effect is translated into a survival benefit in HER2-positive and triple-negative breast cancers.
— Carsten Denkert, MD

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The neoadjuvant study, led by Carsten Denkert, MD, of Charité University Hospital, Berlin, evaluated stromal tumor-infiltrating lymphocytes in 3,771 patients from 6 prospective neoadjuvant clinical trials of the German Breast Group, documenting their relevance for pathologic complete response, disease-free survival, and overall survival, by molecular subtype.

“Our results suggest that tumor-infiltrating lymphocytes are a strong predictive marker for response to neoadjuvant chemotherapy in all molecular subtypes, and this predictive effect is translated into a survival benefit in HER2-positive and triple-negative breast cancers. In contrast, a survival benefit was not observed in luminal breast cancers, suggesting a different biology of the immunologic infiltrate in this subtype,” Dr. Denkert reported.

In the complete cohort, increased tumor-infiltrating lymphocytes (≥ 60%) were observed in 19% of tumors, and 44% of these patients achieved pathologic complete responses (P < .0005). Increased levels (≥ 60%) were observed in 30% of triple-negative tumors, 19% of HER2-positive tumors, and 13% of hormone receptor–positive/HER2-negative tumors.

In all three subtypes, increased levels were significantly associated with increased rates of pathologic complete response (P < .0005). In the multivariate analysis of tumor-infiltrating lymphocytes as a continuous variable, every 10% increase increased the probability of achieving a pathologic complete response by 17% (P < .0001 for all subtypes).

Interesting associations were observed between levels of tumor-infiltrating lymphocytes and disease-free and overall survival. In univariate and multivariate analyses, high levels of tumor-infiltrating lymphocytes were associated with improved disease-free and overall survival rates in the triple-negative and HER2-positive subsets.

“This was most clearly seen for disease-free survival, early in the therapeutic course,” Dr. Denkert said. “Improved survival can be explained by increased pathologic complete responses in this group. In these subtypes, tumor-infiltrating lymphocytes were a positive predictive factor as well as a positive prognostic factor,” he said.

In contrast, in luminal breast cancer, low levels were associated with improved survival, even though increased levels were linked to increased rates of pathologic complete response. “This was most clearly seen for overall survival, late in the therapeutic course,” he continued. This could be a chance finding or a sign of different biology of the immunologic infiltrate in this subtype. One hypothesis is that tumor-infiltrating lymphocytes may be linked to a reduced endocrine response.

“The role of tumor-infiltrating lymphocytes in resistance to different types of endocrine therapy should be investigated,” Dr. Denkert concluded. ■

Disclosure: Dr. Luen reported no potential conflicts of interest. Dr. Denkert is cofounder of Sividon Diagnostics, Cologne, and on the advisory board of AstraZeneca.


1. Luen SJ, Salgado R, Fox S, et al: Prognostic associations of tumor-infiltrating lymphocytes in advanced HER2-positive breast cancer treated with pertuzumab and trastuzumab. 2016 San Antonio Breast Cancer Symposium. Abstract S1-08. Presented December 7, 2016.

2. Denkert C, von Minckwitz G, Darb-Esfahani S, et al: Evaluation of tumor-infiltrating lymphocytes as predictive and prognostic biomarker in different subtypes of breast cancer treated with neoadjuvant therapy. 2016 San Antonio Breast Cancer Symposium. Abstract S1-09. Presented December 7, 2016.

3. Swain SM, Baselga J, Kim SB, et al: Pertuzumab, trastuzumab, and docetaxel in HER2-positive metastatic breast cancer. N Engl J Med 372:724-734, 2015.

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Expert Point of View: Melinda L. Telli, MD

A high level of evidence supports that [tumor-infiltrating lymphocytes] are prognostic. Some people are suggesting that they can now be incorporated into treatment decision-making, as we do with other routine prognostic factors such as estrogen receptor status and Ki67.




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