sTIL Levels Research Articles

Stromal tumor infiltrating lymphocytes in hormone receptor positive/HER2 negative metastatic breast cancer

The immune landscape of hormone receptor positive, HER2-negative metastatic breast cancer (HR+/HER2- mBC), the most common subtype of BC, remains understudied. This is mainly due to reduced sample acquisition opportunities from metastases as compared to primary tumors. In this study, we explored stromal tumor-infiltrating lymphocytes (sTIL) in metastatic samples collected through our post-mortem tissue donation program UPTIDER (NCT04531696). sTIL were scored as a continuous parameter according to international guidelines on 427 metastases and 38 primary untreated tumors acquired from 20 patients with HR+/HER2- mBC. ER-status was evaluated on 362 metastases with a cut-off for positivity set at 1% according to ASCO/CAP guidelines. Our analyses show that 54% and 15% of metastases had sTIL levels >1% and >5% respectively. sTIL levels tended to be lower in metastases as compared to their respective primary tumor (Estimate: -2.83, 95%CI: -5.77-0.11, p:0.07). sTIL levels were lower in metastases from invasive lobular carcinoma than in metastases from invasive breast carcinoma of no special type (Estimate: -1.67, 95%CI: -2.35--0.98, p:<0.001). A loss of ER expression was observed in 14% of all metastases, yet a negative ER-status was not significantly associated with increased sTIL levels. Finally, sTIL levels were significantly higher in lung and axillary lymph node metastases compared to all metastases. While these analyses were conducted on multiple metastases obtained at the end of life after several lines of treatment, the data provides novel and valuable insights into the state of immune infiltration in patients with metastatic HR+/HER2- BC.

Tumor-Infiltrating Lymphocytes in Patients With Stage I Triple-Negative Breast Cancer Untreated With Chemotherapy

The absolute benefit of chemotherapy for all patients with stage I triple-negative breast cancer (TNBC) is unclear, and biomarkers are not currently available for selecting patients with an excellent outcome for whom neoadjuvant or adjuvant chemotherapy may have negligible benefit. High levels of stromal tumor-infiltrating lymphocytes (sTILs) are associated with favorable survival in TNBC, but data solely in stage I TNBC are lacking. To examine the outcomes of patients of all ages with stage I TNBC solely and who received neither neoadjuvant nor adjuvant chemotherapy, according to centrally reviewed sTIL levels at prespecified cutoffs. This cohort study used the Netherlands Cancer Registry to identify patients diagnosed with stage I TNBC between January 1, 2005, and December 31, 2015, who were not treated with chemotherapy. Only patients who did not receive neoadjuvant and/or adjuvant chemotherapy were selected. The clinical data were matched with their corresponding pathology data provided by the Dutch Pathology Registry. Data analysis was performed between February and October 2023. The primary end point was breast cancer-specific survival (BCSS) at 5, 10, and 15 years for the prespecified sTIL level cutoffs of 30%, 50%, and 75%. Hematoxylin and eosin-stained slides were used for central review of histologic subtype, grade, and lymphovascular invasion. The International Immuno-Oncology Biomarker Working Group guidelines were used to score the sTIL levels; these levels were determined for 1041 patients. Of a total of 4511 females with stage I TNBC, patients who were not treated with chemotherapy were selected and tissue blocks requested; sTILs were scored in 1041 patients (mean [SD] age at diagnosis, 64.4 [11.1] years, median follow-up 11.4 [95% CI, 10.9-11.9] years) who were included in the analyses.. Most tumors (952 [91.5%]) were invasive carcinomas of nonspecial histologic subtype. Most patients (548 [52.6%]) had pT1cN0 tumors. Median (range) sTIL level was 5% (1%-99%). A total of 775 patients (74.4%) had sTIL levels below 30%, 266 (25.6%) had 30% or greater, 203 (19.5%) had 50% or greater, and 141 (13.5%) had 75% or greater. Patients with pT1abN0 tumors had a more favorable outcome vs patients with pT1cN0 tumors, with a 10-year BCSS of 92% (95% CI, 89%-94%) vs 86% (95% CI, 82%-89%). In the overall cohort, sTIL levels of at least 30% were associated with better BCSS compared with sTIL levels less than 30% (96% and 87%, respectively; hazard ratio [HR], 0.45; 95% CI, 0.26-0.77). High sTIL levels of 50% or greater were associated with a better outcome than low sTIL levels of less than 50% (HR, 0.27; 95% CI, 0.10-0.74) in patients with pT1C tumors, with a 10-year BCSS of 95% increasing to 98% with sTIL levels of 75% or greater. Results of this study showed that patients with stage I TNBC and high level of sTILs who did not receive neoadjuvant or adjuvant chemotherapy had excellent 10-year BCSS. The findings further support the role of sTILs as integral biomarkers in prospective clinical trials of therapy optimization for this patient population.

A real-world clinicopathological model for predicting pathological complete response to neoadjuvant chemotherapy in breast cancer.

This study aimed to develop and validate a clinicopathological model to predict pathological complete response (pCR) to neoadjuvant chemotherapy (NAC) in breast cancer patients and identify key prognostic factors. This retrospective study analyzed data from 279 breast cancer patients who received NAC at Zhejiang Provincial People's Hospital from 2011 to 2021. Additionally, an external validation dataset, comprising 50 patients from Lanxi People's Hospital and Second Affiliated Hospital, Zhejiang University School of Medicine from 2022 to 2023 was utilized for model verification. A multivariate logistic regression model was established incorporating clinical, ultrasound features, circulating tumor cells (CTCs), and pathology variables at baseline and post-NAC. Model performance for predicting pCR was evaluated. Prognostic factors were identified using survival analysis. In the 279 patients enrolled, a pathologic complete response (pCR) rate of 27.96% (78 out of 279) was achieved. The predictive model incorporated independent predictors such as stromal tumor-infiltrating lymphocyte (sTIL) levels, Ki-67 expression, molecular subtype, and ultrasound echo features. The model demonstrated strong predictive accuracy for pCR (C-statistics/AUC 0.874), especially in human epidermal growth factor receptor 2 (HER2)-enriched (C-statistics/AUC 0.878) and triple-negative (C-statistics/AUC 0.870) subtypes, and the model performed well in external validation data set (C-statistics/AUC 0.836). Incorporating circulating tumor cell (CTC) changes post-NAC and tumor size changes further improved predictive performance (C-statistics/AUC 0.945) in the CTC detection subgroup. Key prognostic factors included tumor size >5cm, lymph node metastasis, sTIL levels, estrogen receptor (ER) status and pCR. Despite varied pCR rates, overall prognosis after standard systemic therapy was consistent across molecular subtypes. The developed predictive model showcases robust performance in forecasting pCR in NAC-treated breast cancer patients, marking a step toward more personalized therapeutic strategies in breast cancer.

Gene expression signatures, stromal tumor infiltrating lymphocytes (sTILs), and change in tumor cellularity to predict pathological complete response (pCR) after 12 week de-escalated neoadjuvant endocrine therapy (ET) vs paclitaxel + dual HER2 blockade in the WSG-TP-II trial.

519 Background: There are limited data on predictive biomarkers for de-escalated ET or chemotherapy with dual anti-HER2 blockade in HR+/HER2+ early breast cancer (BC). In this translational pre-planned project of the WSG-TP-II phase II-trial (NCT03272477), we aimed to identify associations of gene expression signatures and sTILs with pCR. Methods: Patients with cT1c-cT4c, cN0-3 centrally confirmed HR+/HER2+ BC were randomized to 12 weeks of standard ET (n = 100) or paclitaxel (Pac; n = 107). All patients received trastuzumab + pertuzumab (T+P) q3w as neoadjuvant and adjuvant treatment. Gene expression signatures were analyzed using NanoString BC360 panel in baseline biopsies (T+P+ET: n = 72; T+P+Pac: n = 78). sTILs were analyzed in 93 (T+P+ET) and 97 patients (T+P+Pac) at baseline and in 65 (T+P+ET) and 57 patients (T+P+Pac) at week 3. Impacts of standardized BC360 gene expression signatures and sTILs on pCR (ypT0/is ypN0; primary endpoint) expressed in odds ratios (OR) were estimated by logistic regression analysis. Results: pCR rate in patients with BC360 analysis was 39.3% (T+P+ET: 23.6%; T+P+Pac: 53.9%). Overall, ERBB2 (OR 2.37; 95%CI 1.55, 3.62) and cytotoxic cells signature (OR 1.42; 95%CI 1.02, 1.99) were favorable for pCR, while apoptosis (OR 0.64; 95%CI 0.44, 0.92), estrogen receptor 1 (OR 0.60; 95%CI 0.42, 0.85), estrogen receptor signaling (OR 0.64; 95%CI 0.45, 0.91), and progesterone receptor (OR 0.67; 95%CI 0.47, 0.94) signatures were unfavorable. Analyzing by treatment arm, a similar pattern was observed in the T+P+Pac arm (ERBB2: OR 2.01; apoptosis: OR 0.59; estrogen- and progesterone-related signatures: OR 0.41-0.58), but not in the T+P+ET arm where only ERBB2 was prognostic for pCR (OR 7.24; 95%CI 2.12, 24.05). Baseline ≥30% sTIL levels (vs &lt; 30% sTILs; n = 16 vs n = 174) were associated with pCR (OR 5.16; 95%CI 1.60, 16.66); significance was not achieved in either trial arm. Compared to &lt; 30% sTILs (n = 90), low tumor cellularity at week 3 precluding sTILs analysis ( &lt; 500 invasive tumor cells, n = 22), but not ≥30% sTILs (n = 10), was prognostic for pCR in all patients (OR 9.47; 95%CI 2.94, 30.50) and in individual trial arms (OR 6.00-11.79). Conclusions: This hypothesis-generating translational results suggest that gene expression signatures (particularly ERBB2), baseline sTILs, and low tumor cellularity at week 3 predict pCR after ET + double HER2 blockade. Future neoadjuvant trials are needed to prospectively test the use of baseline gene expression and sTILs analysis, and early on-treatment cellularity measurement to select patients with HR+/HER2+ tumors for de-escalated endocrine-therapy-based approaches. Clinical trial information: NCT03272477 .

Abstract P5-02-03: Combined biomarker analysis for prediction of pathological complete response (pCR) after 12 weeks of pembrolizumab + trastuzumab + pertuzumab in HER2-enriched early breast cancer: Keyriched-1 trial

Abstract Background In unselected HER2+ early breast cancer (EBC), de-escalated chemotherapy-free neoadjuvant therapy (NAT) with dual HER2-blockade induces pCR rates of only 20%-40%. In order to achieve pCR rates by de-escalated therapy comparable to those achieved by chemotherapy-based regimens, patient selection and more effective chemotherapy-free regimens are thus key. KEYRICHED-1 (NCT03988036), a single-arm phase 2 study, is the first trial to investigate chemotherapy-free NAT with dual HER2 blockade and pembrolizumab in HER2-enriched HER2+ EBC. In a translational subproject, we analyzed gene signatures together with tumor cell proliferation and spatiotemporal immune cell profiling to identify predictive factors for pCR. Methods 48 pre- and postmenopausal patients with newly diagnosed HER2 2+ (ISH positive) or 3+ EBC (stage I-III) and HER2-enriched (HER2-E) subtype by PAM50 were included in the study. All patients received 4 cycles of pembrolizumab (200 mg), trastuzumab biosimilar ABP 980 (loading dose (LD) 8 mg/kg bodyweight (BW), maintenance dose (MD) 6 mg/kg BW), and pertuzumab (LD 840 mg/kg BW, MD 420 mg/kg BW) q21d. Primary objective was pCR (centrally confirmed absence of invasive tumor in breast and lymph nodes: ypT0/is, ypN0). NanoString Breast Cancer 360 panel was performed in baseline biopsies (n=42). ≥30% Ki67 decrease, &amp;lt; 500 invasive tumor cells or no evidence of tumor in week 3 biopsies (on treatment) were classified as early response. sTILs were analyzed at baseline (n=42) and week 3 (n=28). Ongoing analyses include whole exome sequencing and multiplexed immunohistochemistry for expression of PD1, PDL1, CD4, CD8, CD68, and CD20 levels in tumor and stroma at baseline and at week 3. Impact of standardized expression of single genes, signatures, and sTILs on pCR was evaluated with univariable and multivariate logistic regression analyses and summarized with odds ratios (OR) and 95% confidence intervals (95%CI). Results 42 patients with BC360 and sTILs data at baseline were included in the analysis. Median age was 55 years (range: 22-83), 11 patients (31%) had node-positive EBC. At baseline, 28 patients had sTIL levels ≥30% and 14 had sTILs &amp;lt; 30%; the corresponding pCR rates were 57.1% (n=16) and 28.6% (n=4, p=0.108). At week 3 (on treatment), 16 patients had sTIL levels ≥30%, 50% (n=8) had a pCR vs 8.3% in those with &amp;lt; 30% sTILs (one patient out of 12, p=0.039). 37 patients had early response, 54.1% of them (n=20) had a pCR vs 0% in early non-responders (n=5, p=0.049). In univariate analysis, IDO1, ERBB2, IFNγ, cytotoxic cells, cytotoxicity, CD8 T-cells, TIGIT, and tumor inflammation signatures were statistically significantly associated with pCR (OR 2.3-3.6); ERBB2, IDO1, IFNγ and CD8 T-cells remained significant after adjusting for hormone receptor (HR) and central HER2 status (OR 2.2-4.3). 70 single genes were predictive for pCR; none of them remained significant after false discovery rate adjustment (25%). In multivariable analysis for baseline markers including signatures, sTILs, HR and central HER2 status, only ERBB2 (OR 8.7, 95%CI 1.9-39.0, p=0.0046) and cytotoxic cells signatures (OR 4.6, 95%CI 1.6-13.5, p=0.0059) were predictive for pCR. Results of whole exome sequencing, and multiplexed immunohistochemistry analysis of immune cell markers will be presented at the Symposium. Conclusions Biomarker analysis in the unique KEYRICHED-1 cohort revealed that early response at week 3, ERBB2 and immune related signatures as well as on-therapy sTIL levels predict pCR after a chemotherapy-free combination of immunotherapy and dual HER2 blockade in HER2-enriched EBC. These results pave the way for validation in larger de-escalation trials investigating short, chemotherapy-free regimens in selected patients with HER2+ EBC. Funding for this research was provided by MSD Sharp &amp; Dohme GmbH. Citation Format: Monika Graeser, Sherko Kuemmel, Oleg Gluz, Friedrich Feuerhake, Valery Volk, Daniel Ulbrich-Gebauer, Claudia Biehl, Mattea Reinisch, Athina Kostara9, Iris Scheffen, Kerstin Luedtke-Heckenkamp, Andreas Hartkopf, Felix Hilpert, Angela Kentsch, Carsten Ziske, Reinhard Depenbusch, Michael Braun, Jens-Uwe Blohmer, Christine zu Eulenburg, Matthias Christgen, Ronald Kates, Stephan Bartels, Hans-Heinrich Kreipe, Enrico Pelz, Peter Schmid, Nadia Harbeck. Combined biomarker analysis for prediction of pathological complete response (pCR) after 12 weeks of pembrolizumab + trastuzumab + pertuzumab in HER2-enriched early breast cancer: Keyriched-1 trial [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-02-03.

Impact of stromal tumor-infiltrating lymphocytes (sTILs) on response to neoadjuvant chemotherapy in triple-negative early breast cancer in the WSG-ADAPT TN trial

BackgroundHigher density of stromal tumor-infiltrating lymphocytes (sTILs) at baseline has been associated with increased rates of pathological complete response (pCR) after neoadjuvant chemotherapy (NACT) in triple-negative breast cancer (TNBC). While evidence supports favorable association of pCR with survival in TNBC, an independent impact of sTILs (after adjustment for pCR) on survival is not yet established. Moreover, the impact of sTIL dynamics during NACT on pCR and survival in TNBC is unknown.MethodsThe randomized WSG-ADAPT TN phase II trial compared efficacy of 12-week nab-paclitaxel with gemcitabine versus carboplatin. This preplanned translational analysis assessed impacts of sTIL measurements at baseline (sTIL-0) and after 3 weeks of chemotherapy (sTIL-3) on pCR and invasive disease-free survival (iDFS). Predictive performance of sTIL-0 and sTIL-3 for pCR was quantified by ROC analysis and logistic regression; Kaplan–Meier estimation and Cox regression (with mediation analysis) were used to determine their impact on iDFS.ResultsFor prediction of pCR, the AUC statistics for sTIL-0 and sTIL-3 were 0.60 and 0.63, respectively, in all patients; AUC for sTIL-3 was higher in NP/G. The positive predictive value (PPV) of “lymphocyte-predominant” status (sTIL-0 ≥ 60%) at baseline was 59.3%, though only 13.0% of patients had this status. To predict non-pCR, the cut point sTIL-0 ≤ 10% yielded PPV = 69.5% while addressing 33.8% of patients.Higher sTIL levels (particularly at 3 weeks) were independently and favorably associated with better iDFS, even after adjusting for pCR. For example, the adjusted hazard ratio for 3-week sTILs ≥ 60% (vs. < 60%) was 0.48 [0.23–0.99]. Low cellularity in 3-week biopsies was the strongest individual predictor for pCR (in both therapy arms), but not for iDFS.ConclusionThe independent impact of sTILs on iDFS suggests that favorable immune response can influence key tumor biological processes for long-term survival. The results suggest that the reliability of pCR following neoadjuvant therapy as a surrogate for survival could vary among subgroups in TNBC defined by immune response or other factors. Dynamic measurements of sTILs under NACT could support immune response-guided patient selection for individualized therapy approaches for both very low levels (more effective therapies) and very high levels (de-escalation concepts).Trial registration: Clinical trials No: NCT01815242, retrospectively registered January 25, 2013.

BI-RADS Ultrasound Lexicon Descriptors and Stromal Tumor-Infiltrating Lymphocytes in Triple-Negative Breast Cancer

Increased levels of stromal tumor-infiltrating lymphocytes (sTILs) have recently been considered a favorable independent prognostic and predictive biomarker in triple-negative breast cancer (TNBC). The purpose of this study was to determine the relationship between BI-RADS (Breast Imaging Reporting and Data System) ultrasound lexicon descriptors and sTILs in TNBC. Patients with stage I-III TNBC were evaluated within a single-institution neoadjuvant clinical trial. Two fellowship-trained breast radiologists used the BI-RADS ultrasound lexicon to assess pretreatment tumor shape, margin, echo pattern, orientation, posterior features, and vascularity. sTILs were defined as low <20 or high ≥20 on the pretreatment biopsy. Fisher's exact tests were used to assess the association between lexicon descriptors and sTIL levels. The 284 patients (mean age 52 years, range 24-79 years) were comprised of 68% (193/284) with low-sTIL tumors and 32% (91/284) with high-sTIL tumors. TNBC tumors with high sTILs were more likely to have the following features: (1) oval/round shape than irregular shape (p = 0.003), (2) circumscribed or microlobulated margins than spiculated, indistinct, or angular margins (p = 0.0005); (3) complex cystic and solid pattern than heterogeneous pattern (p = 0.006); and (4) posterior enhancement than shadowing (p = 0.002). There was no significant association between sTILs and descriptors for orientation and vascularity (p = 0.06 and p = 0.49, respectively). BI-RADS ultrasound descriptors of the pretreatment appearance of a TNBC tumor can be useful in discriminating between tumors with low and high sTIL levels. Therefore, there is a potential use of ultrasound tumor characteristics to complement sTILs when used as stratification factors in treatment algorithms for TNBC.

Long-term outcomes of patients with node-negative (N0), triple-negative breast cancer (TNBC) who did not receive adjuvant chemotherapy according to stromal TILs (sTILs).

548 Background: sTILs are a well-established prognostic and predictive biomarker in patients with operable TNBC receiving pre or postoperative systemic therapy. We1 and others2,3 have also shown that sTILs are prognostic in patients who did not receive adjuvant chemotherapy. Here, we detail the outcomes of systemically untreated patients with N0 TNBC according to sTIL score. We focused on the N0 subset as a group of patients who may be candidates for future prospective therapy de-escalation trials. Methods: From a clinically annotated cohort of 605 patients with centrally confirmed TNBC (ER/PR &lt; 1% and HER2 negative) with long-term outcomes data, we identified 182 patients treated with locoregional therapy only (breast surgery +/- radiation therapy and no chemotherapy). The clinicopathological characteristics of this cohort have previously been published1. In this analysis, we report the 5- and 10-year invasive disease-free survival (iDFS) and overall survival (OS) rates of patients with N0 TNBC according to sTIL levels. IDFS and OS were defined as per the STEEP classification and estimated using the Kaplan–Meier method. Comparisons of the survival distributions between groups were assessed by the log-rank test. sTILs were assessed as a continuous parameter according to the International TIL Working Group guidelines. For comparisons of outcomes between groups, tumors were classified as lymphocyte-predominant TNBC (defined as containing ≥50% sTILs) vs non-lymphocyte-predominant ( &lt; 50% sTILs). Results: Of 182 systemically untreated patients, 149 (82%) were N0 and most (78%) were post-menopausal. T stage distribution was T1: 68%, T2: 28%, T3/4: 4%. Among N0 patients, 31 (21%) had lymphocyte-predominant TNBC, and in this group the 5-year iDFS and OS were 89% (95% CI 76-100) and 96% (95% CI 89-100), while the 10-year iDFS and OS were 89% (95% CI 76-100) and 87% (95% CI 73-100), respectively. In contrast, outcomes for patients with non-lymphocyte predominant TNBC were significantly worse. For this group, 5-year iDFS and OS were 62% (95% CI 53-73) and 78% (95% CI 71-86) while the 10-year iDFS and OS were 45% (95% CI 36-58) and 66% (95% CI 68-76), respectively ( log-rank p = 0.02 for iDFS and log-rank p = 0.03 for OS). Conclusions: sTIL quantification identifies a subset of patients with early-stage N0 TNBC with an exceedingly good prognosis, even in the absence of adjuvant chemotherapy. These data provide support for the evaluation of sTILs as part of prospective investigation of systemic therapy de-escalation strategies in N0 TNBC.

Immune microenvironment characterisation and dynamics during anti-HER2-based neoadjuvant treatment in HER2-positive breast cancer

Despite their recognised role in HER2-positive (HER2+) breast cancer (BC), the composition, localisation and functional orientation of immune cells within tumour microenvironment, as well as its dynamics during anti-HER2 treatment, is largely unknown. We here investigate changes in tumour-immune contexture, as assessed by stromal tumour-infiltrating lymphocytes (sTILs) and by multiplexed spatial cellular phenotyping, during treatment with lapatinib-trastuzumab in HER2+ BC patients (PAMELA trial). Moreover, we evaluate the relationship of tumour-immune contexture with hormone receptor status, intrinsic subtype and immune-related gene expression. sTIL levels increase after 2 weeks of HER2 blockade in HR-negative disease and HER2-enriched subtype. This is linked to a concomitant increase in cell density of all four immune subpopulations (CD3+, CD4+, CD8+, Foxp3+). Moreover, immune contexture analysis showed that immune cells spatially interacting with tumour cells have the strongest association with response to anti-HER2 treatment. Subsequently, sTILs consistently decrease at the surgery in patients achieving pathologic complete response, whereas most residual tumours at surgery remain inflamed, possibly reflecting a progressive loss of function of T cells. Understanding the features of the resulting tumour immunosuppressive microenvironment has crucial implications for the design of new strategies to de-escalate or escalate systemic therapy in early-stage HER2+ BC.

Characterization of Stromal Tumor-infiltrating Lymphocytes and Genomic Alterations in Metastatic Lobular Breast Cancer.

Invasive lobular carcinoma (ILC) represents the second most common histologic breast cancer subtype after invasive ductal carcinoma (IDC). While primary ILC has been extensively studied, metastatic ILC has been poorly characterized at the genomic and immune level. We retrospectively assembled the multicentric EuroILC series of matched primary and metastatic samples from 94 patients with estrogen receptor (ER)-positive ILC. Stromal tumor-infiltrating lymphocytes (sTILs) were assessed by experienced pathologists. Targeted sequencing and low pass whole-genome sequencing were conducted to detect mutations and copy-number aberrations (CNAs). We compared the frequencies of the alterations in EuroILC with those from patients with ER-positive metastatic ILC (n = 135) and IDC (n = 563) from MSK-IMPACT. Low sTIL levels were observed in ILC metastases, with higher levels in the mixed nonclassic histology. Considering ILC metastases from EuroILC and MSK-IMPACT, we observed that >50% of tumors harbor genomic alterations that have previously been associated with endocrine resistance. A matched primary/metastasis comparison in EuroILC revealed mutations (AKT1, ARID1A, ESR1, ERBB2, or NF1) and CNAs (PTEN or NF1 deletion, CYP19A1 amplification) associated with endocrine resistance that were private to the metastasis in 22% (7/32) and 19% (4/21) of patients, respectively. An increase in CDH1, ERBB2, FOXA1, and TBX3 mutations, in CDH1 deletions and a decrease in TP53 mutations was observed in ILC as compared with IDC metastases. ILC metastases harbor genomic alterations that may potentially explain endocrine resistance in a large proportion of patients, and present genomic differences as compared with IDC metastases.

Analysis of the \u201ccentrosome-ome\u201d identifies MCPH1 deletion as a cause of centrosome amplification in human cancer

The centrosome is the microtubule organizing center of human cells and facilitates a myriad of cellular functions including organization of the mitotic spindle to ensure faithful chromosome segregation during mitosis, cell polarization and migration, and primary cilia formation. A numerical increase in centrosomes, or centrosome amplification (CA), is common in cancer and correlates with more aggressive clinical features and worse patient outcomes. However, the causes of CA in human cancer are unclear. Many previous studies have identified mechanisms of CA in cellulo, such as overexpression of PLK4, but it is unclear how often these are the primary mechanism in human disease. To identify a primary cause of CA, we analyzed The Cancer Genome Atlas (TCGA) genomic and transcriptomic data for genes encoding the 367 proteins that localize to the centrosome (the “centrosome-ome”). We identified the following candidates for primary causes of CA: gain-of-function alterations of CEP19, CEP72, CTNNB1, PTK2, NDRG1, SPATC1, TBCCD1; and loss-of-function alterations of CEP76, MCPH1, NEURL4, and NPM1. In cellulo analysis of these candidates revealed that loss of MCPH1/microcephalin caused the most robust increase in centriole number. MCPH1 deep gene deletions are seen in 5–15% of human cancers, depending on the anatomic site of the tumor. Mechanistic experiments demonstrated that loss of MCPH1 caused a CDK2-dependent increase in STIL levels at the centrosome to drive CA. We conclude that loss of MCPH1 is common in human cancer and is likely to be a cause of CA.

Pembrolizumab plus chemotherapy as neoadjuvant treatment of high-risk, early-stage triple-negative breast cancer: results from the phase 1b open-label, multicohort KEYNOTE-173 study

The phase Ib KEYNOTE-173 study was conducted to assess the safety and preliminary antitumor activity of neoadjuvant chemotherapy plus pembrolizumab in high-risk, early-stage, non-metastatic triple-negative breast cancer (TNBC). Six pembrolizumab plus chemotherapy regimens were evaluated (cohorts A-F). All cohorts received a pembrolizumab 200-mg run-in dose (cycle 1), then eight cycles of pembrolizumab in combination with a taxane with or without carboplatin for 12 weeks, and then doxorubicin and cyclophosphamide for an additional 12 weeks before surgery. Primary end points were safety and recommended phase II dose (RP2D); secondary end points were pathological complete response (pCR) rate, objective response rate, and event-free and overall survival. Exploratory end points were the relationship between outcome and potential biomarkers, such as tumor programmed death ligand 1 (PD-L1) expression (combined positive score) and stromal tumor-infiltrating lymphocyte levels (sTILs). Sixty patients were enrolled between 18 February 2016, and 28 February 2017. Dose-limiting toxicities occurred in 22 patients, most commonly febrile neutropenia (n= 10 across cohorts). Four cohorts (B, C, D, F) did not meet the RP2D threshold; two cohorts did (A, E). The most common grade ≥3 treatment-related adverse event was neutropenia (73%). Immune-mediated adverse events and infusion reactions occurred in 18 patients (30%) and were grade ≥3 in six patients (10%). The pCR rate (ypT0/Tis ypN0) across all cohorts was 60% (range 49%-71%). Twelve-month event-free and overall survival rates ranged from 80% to 100% across cohorts (100% for four cohorts). Higher pre-treatment PD-L1 combined positive score, and pre- and on-treatment sTILs were significantly associated with higher pCR rates (P= 0.0127, 0.0059, and 0.0085, respectively). Combination neoadjuvant chemotherapy and pembrolizumab for high-risk, early-stage TNBC showed manageable toxicity and promising antitumor activity. In an exploratory analysis, the pCR rate showed a positive correlation with tumor PD-L1 expression and sTIL levels. ClinicalTrials.gov identifier: NCT02622074.

Abstract P4-10-18: Changes in tumour infiltrating lymphocytes during neoadjuvant endocrine therapy and possible clinical implications for guiding therapy

Abstract Background: Stromal Tumour Infiltrating Lymphocytes (sTILs) has been established as a predictive biomarker for response to neoadjuvant chemotherapy irrespective of subtype. While increased sTIL levels is associated with prolonged survival among patients with triple-negative and HER2 positive breast cancer increased sTILs has in Estrogen Receptor positive (ER+) breast cancer been suggested to be an adverse prognostic factor. It has been hypothesised that differences in the cellular composition of immune cells may explain the dissimilarity in prognostic impact according to breast cancer subtype. Here, we report data from a neoadjuvant phase II study, treating postmenopausal patients with primary ER+, HER2 negative, operable breast cancer with letrozole for four months by the Danish Breast Cancer Group (DBCG). We analysed the association of sTILs, and changes in sTILs during neoadjuvant endocrine treatment (NET), with pathological response and correlation with other clinicopathological variables such as Ki67, as potential biomarkers for risk-stratification of patients following NET. Method: 113 postmenopausal patients with ER+, HER2 negative breast cancer were treated with NET for four months. Endpoint was pathological response assessed by a modified Miller Payne scale and the Residual Cancer Burden (RCB) score. Pretherapeutic core biopsies and post therapeutic surgical specimens were assessed centrally for the percentage of sTILs on HE sections according to the International TILs working group guidelines. sTILs association to pathological response were assessed with univariate logistic regression. Odds Ratio (OR) was estimated with a 95 % confidence interval. Correlation of sTILs and Ki67 were tested with Pearson´s correlation coefficient. Changes in sTILs with a paired t-test. Level of significance was set to 5 %. Results: sTILs concentration increased with mean 6.8 % (p &amp;lt;.0001) during treatment in the assessable patients (range -39 % - 60 %). Forty-four percent had no pathological response as assessed by the modified Miller Payne scale and 11 % had RCB-Class III after NET. Increase in sTILs was significantly associated with poorer pathological response with OR=1.38 (95 % CI: 1.05 - 1.82; p = 0.03) per 10 % increase for no pathological response, and correspondingly OR=1.78 (95 % CI: 1.27 - 2.50; p &amp;lt;.0001) for higher level RCB-class per 10 % increase. The correlation between pre-therapeutic sTILs and Ki67 was moderate (Pearson 0.4; p = 0.0004), however the association grew stronger post treatment (Pearson 0.5; p &amp;lt;.0001). Conclusion: Increased sTILs activity during NET was associated with poor treatment response. An increase in sTILs during NET could be considered indictive for potential immunogenic tumours, suggesting that patients with increasing sTIL in residual disease after NET might derive benefit from the addition of immunotherapy. We found a significant correlation between sTILs and Ki67 levels. Ki67 has been established as a window of opportunity biomarker for antiproliferative response. Combining biomarkers of antiproliferative response such as Ki67 with biomarkers of immunogenetic significance might be important to determine optimal combination of adjuvant therapy. A window of opportunity study with an aromatase inhibitor could show if it is feasible to detect a rise in sTILs early enough to guide adjuvant treatment in the perioperative setting. Citation Format: Signe Korsgaard Skriver, Maj-Britt Jensen, Ann Soegaard Knoop, Bent Ejlertsen, Anne-Vibeke Laenkholm. Changes in tumour infiltrating lymphocytes during neoadjuvant endocrine therapy and possible clinical implications for guiding therapy [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-10-18.

Abstract P1-18-19: Phase II, open label, randomized, biomarker study of immune-mediated mechanism of action of neoadjuvant subcutaneous trastuzumab in patients with operable, locally advanced, or inflammatory HER2-positive breast cancer. ImmunHER trial on behalf of the Gruppo Oncologico Italiano di Ricerca Clinica (GOIRC)

Abstract Background: Tumor-infiltrating lymphocytes (TILs) have been reported to be associated with increased therapeutic efficacy of trastuzumab/pertuzumab-based neoadjuvant therapy (NT) in patients (pts) with HER2-positive breast cancer (BC). Subcutaneous (SC) trastuzumab has non-inferior efficacy to intravenous (IV) administration, with a similar safety profile. Interestingly, SC trastuzumab has been observed to be more immunogenic than IV trastuzumab and act at different immunologic levels. Therefore, by modifying the modality of administration of trastuzumab, it could be possible to interfere with different immune pathways and exert a favorable immunomodulation in HER2-positive BC. Methods: In this non-comparative, phase II, neoadjuvant, randomized study, pts were eligible if they had previously untreated, histologically confirmed, locally advanced, inflammatory, or early-stage HER2-positive BC. Pts were treated with FEC (fluorouracil 500 mg/m2; epirubicin 75 mg/m2; cyclophosphamide 500 mg/m2) q21 × 3 cycles. Then, they were randomly assigned (1:1) to receive: docetaxel (75 mg/m2) plus pertuzumab (840 mg loading dose (LD), then 420 mg) plus IV trastuzumab (8 mg/kg LD, then 6 mg/kg) q21 × 4 cycles (arm A) or, docetaxel plus pertuzumab plus SC trastuzumab (fixed dose of 600 mg) q21 × 4 cycles (arm B). After surgery, pts received trastuzumab q21 × 14 cycles using the same formulation (SC or IV) of the preoperative phase. The primary endpoint was the rate of stromal TILs (sTILs) on residual disease after surgery. Tumor biopsy and posttreatment surgical samples were centrally analyzed for TILs. Blood samples were also collected during NT for tumor-specific lymphocyte cell activity analysis. Feasibility, efficacy and safety were also evaluated. ClinicalTrials.gov: NCT03144947. Results: Between November 2016 and September 2017, according to an adaptive Simon's two-stage optimal design, we enrolled 65 pts, of whom two were deemed ineligible for the study. Thus, 63 pts (31 in arm A and 32 in arm B) were assessed for the primary and secondary endpoints. The pathologic complete response (pCR; no invasive tumor in breast and axilla) rates were 64.5% (95% CI, 47-81) in arm A, and 59.4% (95% CI, 42-76) in arm B. The most common adverse events of grade 3 or higher were neutropenia (15 [48.4%] pts in arm A, and 11 [34.4%] in arm B), neurotoxicity (1 [3.2%], and 2 [6.2%], respectively), and diarrhea (1 [3.2%], and 1 [3.1%], respectively). There were no events of congestive heart failure. At surgery, 11 pts in arm A and 13 pts in arm B were evaluable for TIL analysis. The median value of sTILs (7.5%) on pre-treatment tumor biopsies was used as the cut-off value, and high sTIL levels were observed in 27.3% and in 46.1% of residual tumors after treatment arm A and B, respectively. There was a positive correlation between pretreatment sTILs and PD-L1 expression on stromal immune cells (Kendall’s τ =0.80). Interestingly, a significant inverse correlation was observed between PD-L1 expression on pretreatment sTILs and the T cell co-receptor CD3 expressed on posttreatment sTILs (Pearson’s ρ = -0.70). This finding was particularly evident in the arm B group (ρ = -0.85). Conclusions: NT with either SC or IV trastuzumab in combination with pertuzumab and chemotherapy had a significant effect on sTIL expression at surgery. In particular, the SC trastuzumab-based arm exerted the most relevant enrichment of sTILS in posttreatment residual tumors. These findings suggest a role for the SC administration of trastuzumab in determining favorable variations of host immune response parameters among pts with HER2-positive early BC who had residual disease after NT. Citation Format: Antonino Musolino, Stefania Gori, Elisabetta Cretella, Alessandra Marabese, Luigi Cavanna, Antonio Frassoldati, Giancarlo Bisagni, Chiara Casarini, Emilio Bria, Luisa Carbognin, Elena Fiorio, Alba A Brandes, Claudio Zamagni, Lorenzo Gianni, Alberto Zambelli, Filippo Montemurro, Michele Tognetto, Renata Todeschini, Giuseppe Maglietta, Gabriele Missale, Enrico M Silini. Phase II, open label, randomized, biomarker study of immune-mediated mechanism of action of neoadjuvant subcutaneous trastuzumab in patients with operable, locally advanced, or inflammatory HER2-positive breast cancer. ImmunHER trial on behalf of the Gruppo Oncologico Italiano di Ricerca Clinica (GOIRC) [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P1-18-19.

Usefulness of imaging findings in predicting tumor-infiltrating lymphocytes in patients with breast cancer.

Tumor-infiltrating lymphocytes (TILs) have been determined as a new prognostic indicator of immunotherapy response in breast cancer (BC). The aim of this study is to investigate the effectiveness of imaging features in predicting the TIL levels in invasive BC patients. A total of 158 patients with invasive BC were included in our study. All lesions were evaluated based on the BIRADS lexicon. US was performed for all the patients and 89 of them underwent MRI. The histologic stromal TIL (sTIL) levels were assessed and associations between the sTIL levels and imaging features were evaluated. Tumors with high sTIL levels had more circumscribed margins, round shape, heterogeneous echogenicity, and larger size on ultrasonography (p < 0.005). There was a statistically significant positive correlation between the sTIL levels and ADC value (p < 0.001). Tumors with high sTIL levels had significantly more homogeneous enhancement than the tumors with low sTIL levels (p = 0.001). Logistic regression analysis showed that the ADC was the most statistically significant parameter in predicting the sTIL levels (the odds ratio was 90.952; p = 0.002). The optimal cutoff value for ADC in predicting low and high sTIL levels was found to be 0.87 × 10-3 mm2 s-1 (AUC = 0.726, 73% specificity, and 60% sensitivity). Imaging findings, especially the ADC, may play an important role as an adjunct tool in cases of uncertain situations and may improve the accuracy of biopsy results. The prediction of sTIL levels using imaging findings may give an opportunity to predict prognosis. • Preoperative assessment of TILs is an important biomarker of prognosis and treatment efficacy. • ADC value can be a useful tool in distinguishing high and low sTIL levels as a non-invasive method. • The prediction of sTIL levels using imaging findings may give an opportunity to predict prognosis and an optimal treatment for the BC patients.

252P - Clinical features of early-stage (I-III) triple-negative breast cancer (TNBC) patients with tumours exhibiting low-overall change in molecular profile after neoadjuvant therapy

BackgroundSeth et al. (ASCO 19) identified four biologic classes based on longitudinal molecular profiling of TNBC tumors pre- and post- neoadjuvant adriamycin/cyclophosphamide (AC). The four groups were defined based on the enriched and depleted hallmark pathways that were observed. The low-overall change (LOC) group was defined as having no significant change in the pathways in pre- and post-AC biopsies. Of note, this group of patients had lower pathological complete response (pCR) rates. The aim of this study is to describe the clinical features of early-stage (I-III) TNBC patients with tumors exhibiting LOC in their biopsies with neoadjuvant therapy. MethodsWe analyzed the clinical characteristics of 48 patients with early stage (I-III) TNBC enrolled in the ARTEMIS trial (NCT02276443). All patients received neoadjuvant AC; 48% of patients were subsequently treated on a clinical trial and 52% received standard taxane-based chemotherapy. We compared the clinical characteristics of 17 patients in the LOC group with 31 patients in the other three biologic classes. Two-group comparison was performed using Chi-squared test. ResultsAge at diagnosis, histology, Vanderbilt subtype, Ki-67, sTIL levels and vimentin expression were similar between the groups. Median age at diagnosis in the LOC group was 58 (range, 27-74). All patients in this group were stage II-III, with no stage I identified. 59% of patients in the LOC group had stage III disease compared with 26% in the other groups (p=0.02), Also, 47% of patients in this group had AR≥10% compared to 19% in the other biological groups (p=0.04).Table252PTableVariableGroupLow-Overall Change Group (n=17)%Other Biologic Classes (n=31)%pAge at diagnosis≤60105925810.10>60741619StageI00619N/AII74117550.36III10598260.02HistologyInvasive ductal158826840.23Metaplastic00413Apocrine1600Other1613Vanderbilt subtypeBL3188260.24LAR42426M/MSL3181135IM212412Ki-67 (%)≥ 5084721680.16< 509531032Androgen Receptor Expression (%)≥ 108476190.04< 109532581Vimentin Expression (%)≥502124130.91<5015882787sTILLow (<5)74115480.37Moderate (≥5-30)9521135High (>30)0013 ConclusionsPatients with low-overall change in their tumors had later-stage disease and higher AR expression. Previous analysis by Seth et al. has demonstrated a lower pCR rate in this group with standard neoadjuvant chemotherapy. This analysis highlights the potential role of androgen deprivation in this class of tumors. Legal entity responsible for the studyThe authors. FundingThe University of Texas MD Anderson Cancer Center Moonshots Program and a CPRIT Multi-Investigator Research Award (MIRA). DisclosureS. Moulder: Research grant / Funding (self): Pfizer; Research grant / Funding (self): Genentech; Research grant / Funding (self): Novartis; Research grant / Funding (self): Seattle Genetics; Research grant / Funding (self): EMD Serono; Research grant / Funding (self): Bayer; Research grant / Funding (self): AstraZeneca. All other authors have declared no conflicts of interest.

SFI1 promotes centriole duplication by recruiting USP9X to stabilize the microcephaly protein STIL.

In mammals, centrioles participate in brain development, and human mutations affecting centriole duplication cause microcephaly. Here, we identify a role for the mammalian homologue of yeast SFI1, involved in the duplication of the yeast spindle pole body, as a critical regulator of centriole duplication in mammalian cells. Mammalian SFI1 interacts with USP9X, a deubiquitylase associated with human syndromic mental retardation. SFI1 localizes USP9X to the centrosome during S phase to deubiquitylate STIL, a critical regulator of centriole duplication. USP9X-mediated deubiquitylation protects STIL from degradation. Consistent with a role for USP9X in stabilizing STIL, cells from patients with USP9X loss-of-function mutations have reduced STIL levels. Together, these results demonstrate that SFI1 is a centrosomal protein that localizes USP9X to the centrosome to stabilize STIL and promote centriole duplication. We propose that the USP9X protection of STIL to facilitate centriole duplication underlies roles of both proteins in human neurodevelopment.

Abstract P1-08-02: Associations of epidemiologic and clinical features with intensity of immune infiltrates in postmenopausal breast cancer cases from the cancer prevention study-II cohort

Abstract Stromal tumor-infiltrating lymphocytes (sTILs) are often prominent in HER2-enriched and triple negative breast cancers and predict a favorable prognosis for these subtypes. Clinical and epidemiological determinants of sTILs have not been defined in large cohorts. Accordingly, we analyzed these associations for 702 eligible, postmenopausal invasive breast cancer cases from the Cancer Prevention Study (CPS)-II Nutrition cohort. CPS-II included 97,786 women who completed baseline and follow-up surveys since 1992. Women with self-reported breast cancer diagnoses were consented for medical record and tumor tissue retrieval. We analyzed pre-diagnostic data from the survey prior to diagnosis on personal history of benign breast disease (BBD), menopausal hormone use, alcohol intake, cigarette smoking status, waist circumference, body mass index, adult weight gain, nonsteroidal anti-inflammatory drugs, and physical activity. One pathologist (TG) evaluated sTILs using whole slide images of H&amp;E stained sections according to recommendations by the International TILs Working Group 2014. sTIL levels were dichotomized: none/minimal (0-10%) and moderate/high (&amp;gt;10%). We compared the sTIL levels by clinical and epidemiologic risk factors using chi-square statistics. Odds ratio (ORs) and 95% confidence intervals (CIs) for the associations of clinical and epidemiologic risk factors with sTILs were estimated with multivariable logistic regression. Mean age at diagnosis was 71.9 years; 44% of cancers were moderate grade, 74% were localized staged and 88% were luminal-like. Cancers with high/moderate sTIL levels (n=614), compared to none/minimal (n=88), were more likely to be high grade (54% vs. 20%; p-value &amp;lt;0.001), node-positive (41% vs. 22%; p-value &amp;lt;0.001), and non-luminal (36% vs. 8%; p-value &amp;lt;0.001). In a model mutually-adjusted for age at diagnosis and clinical factors, all clinical factors were associated with moderate/high sTIL levels: node-positive (OR=2.15, 95% CI 1.28 – 3.58), high grade (OR=3.98, 95% CI 1.88 – 9.06), and non-luminal subtype (OR=3.65, 95% CI 2.00 – 6.59). Age at diagnosis was not associated with sTIL levels (per year: OR=1.01, 95% CI 0.97 – 1.05). Women with BBD were less likely to have moderate/high sTIL levels than minimal/none sTIL levels (38% vs. 54%; age-adjusted OR=0.50, 95% CI 0.31 – 0.79). Ever smokers also were less likely to have moderate/high sTIL levels (38% vs. 49%) with an OR=0.61 (95% 0.38 – 0.97). Other risk factors were not significantly associated with sTIL levels. In this large epidemiologic cohort, non-luminal cancers had more sTILs than luminal cancers. Women with a personal history of BBD and ever cigarette smokers had significant lower levels of sTILs. Our study is among the first to examine pre-diagnostic risk factors in relation to sTILs and provides the impetus for larger, population-based studies with phenotypic characterization of immune cells. Citation Format: Gaudet MM, Mellow MM, Puvanesarajah S, Gapstur SM, Sherman ME, Gansler T. Associations of epidemiologic and clinical features with intensity of immune infiltrates in postmenopausal breast cancer cases from the cancer prevention study-II cohort [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-08-02.

Abstract P2-08-47: Evaluation of stromal tumor-infiltrating lymphocytes (sTIL) and tertiary lymphoid structures (TLS) in early breast cancer patients with triple negative breast cancer(TNBC) included in a prospective study of neoadjuvant chemotherapy (NAC) with Epirubicin and cyclophosphamide (EC) and carboplatin-paclitaxel (PC) (BSMO 2014-01)

Abstract Background: BSMO 2014-01 is a completed prospective phase 2 study evaluating the efficacy of neoadjuvant EC and PC. One of the secondary endpoints was the correlation of sTIL with response, pCR and survival. We also assessed the relationship between sTIL and TLS in the diagnostic biopsies. Methods: Stromal TIL (sTIL) were evaluated on H&amp;E stained tumor biopsies before the start of the NAC according to the criteria described by Salgado et al(1). Scores were defined as "low" or "high" if lymphocytic infiltration in the stroma around the tumor was ≤ 10% or &amp;gt; 10%. TLS are ectopic lymph node-like structures recently identified in breast cancer. TLS were counted using a dual IHC stain for CD3 (T cells) and CD20 (B cells) and categorized as "little" if the TLS occupied &amp;lt; 10% or "moderate to abundant" if they occupied ≥ 10% of the adjacent tissue. The correlation between sTIL and pathologic parameters was analyzed using the chi-square test; DFS and OS between the groups was estimated by using the log-rank test. Results: So far we could quantify the number of sTIL in 38 out of 63 TNBC pts treated with neoadjuvant EC-PC. Twenty eight pts had a high sTIL score and 10 pts had a low sTIL score. The high-sTIL group (19/28) achieved a numerical higher pathologic complete remission (pCR) rate than the low-sTIL group (5/10) (p=0.3); both groups had a comparable disease free survival of 28.6 mths and 26.7 mths respectively (p=0.7). The overall survival was similar:29 mths and 27.8 mths respectively (p=0.8). Stromal TLS were identified in 10 out 23 samples available for this analysis and we could demonstrate a positive correlation between high levels of sTIL and high levels of moderate to abundant TLS(CD3) in the adjacent tissue in six out of the ten samples in which TLS were present (p=0.1). Conclusion: These preliminary results could not confirm the results published by Denkert et al earlier this year(2). A trend for correlation of the presence of high sTIL with moderate to abundant levels of TLS was found. Analysis on the remaining samples of all patients included in the study and correlation with outcome is ongoing and these completed results will be presented. (1)Assessing Tumor-infiltrating lymphocytes in solid tumors. Hendry S, Salgado R et al. Adv Anat Pathol 2017; 235-251. (2)Tumour-infiltrating lymphocytes and prognosis in different subtypes of breast cancer. Denkert C et al. Lancet Oncol 2018; 40-50. Citation Format: Fontaine C, Van den Eynden G, de Wind R, Boisson A, Renard V, Van den Bulck H, Vuylsteke P, Glorieux P, Dopchie C, Decoster L, De Grève J, Awada A, Wildiers H, Willard-Gallo K. Evaluation of stromal tumor-infiltrating lymphocytes (sTIL) and tertiary lymphoid structures (TLS) in early breast cancer patients with triple negative breast cancer(TNBC) included in a prospective study of neoadjuvant chemotherapy (NAC) with Epirubicin and cyclophosphamide (EC) and carboplatin-paclitaxel (PC) (BSMO 2014-01) [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-08-47.

Abstract P3-10-09: Relationship between tumor infiltrating lymphocytes (TILs) and response to pembrolizumab (Pembro)+chemotherapy (Chemo) as neoadjuvant treatment (NAT) for triple-negative breast cancer (TNBC): phase Ib KEYNOTE-173 trial

Abstract Background:Increasing quantities of stromal TILs (sTILs) are associated with higher pathologic complete response (pCR) rates with conventional chemo in early-stage TNBC. We evaluated the association between sTILs and PD-L1 expression with response to pembro+chemo as NAT for TNBC in the KEYNOTE-173 trial (NCT02622074). Methods: sTILs were quantified using light microscopy of H&amp;E-stained slides from pretreatment and on-treatment (during first 3 weeks of pembro monotherapy) tumor biopsies by a pathologist blind to response data. Pretreatment PD-L1 expression was assessed using the PD-L1 IHC 22C3 pharmDx assay and reported as combined positive score (CPS). Endpoints were pCR rate by ypT0 ypN0 and ypT0/Tis ypN0 and objective response rate (ORR; RECIST v1.1) after the first 4 cycles of NAT (taxane±carboplatin+pembro) by MRI. sTILs and PD-L1 CPS were evaluated as continuous variables. Association between sTILs and PD-L1 CPS with response was assessed using logistic regression and area under the reciever operating curve (AUROC) analyses, with a 1-sided alpha level of 0.10. Correlation between PD-L1 and sTILs was assessed by Spearman's rank correlation coefficient. Multivariate analysis included sTILs (pretreatment and on-treatment) and PD-L1 CPS. Likelihood ratio tests were used to evaluate the added value of factors in predicting pCR rate. Results: Of 60 total pts, 34 had tumors evaluated for pretreatment sTILs, 52 for PD-L1 CPS, and 33 for both sTILs and CPS. On-treatment sTILs were evaluated in 31 pts. Overall pCR rates were 56.7% and 60% by ypT0 ypN0 and ypT0/Tis ypN0, respectively; ORR was 78.3%. In pts evaluated for sTILs and CPS (individually), pCR rates and ORR were comparable with overall pCR rates and ORR. There was a significant correlation between pretreatment sTILs and PD-L1 CPS (ρ=0.65, P&amp;lt;0.001).Higher pretreatment sTILs were significantly associated with response: ypT0 ypN0 P= 0.011; ypT0/Tis ypN0 P=0.006; ORR P=0.061. On-treatment sTILs were also significantly associated with response: ypT0 ypN0 P=0.061; ypT0/Tis ypN0 P=0.041; ORR P=0.031. Pretreatment PD-L1 CPS was significantly associated with response: ypT0 ypN0 P=0.073; ypT0/is ypN0 P=0.030; and ORR P=0.021. AUROC of pretreatment sTIL association with pCR was numerically higher than with on-treatment sTILs and PD-L1 CPS (0.69 vs 0.61 vs 0.56 for ypT0ypN0 and 0.72 vs 0.67 vs 0.62 for ypT0/Tis ypN0). Responders had higher median pretreatment sTIL levels vs nonresponders: 45% [10, 75] vs 10% [5, 20] for pCR rate by ypT0 ypN0 and 52.5% [10, 73.8] vs 10% [5, 20] for pCR rate by ypT0/Tis ypN0; 25% [5, 70] vs 10% [6.3, 27.5] for ORR. In multivariate analysis, only pretreatment sTILs were significant for both pCR endpoints (ypT0 ypN0 P=0.031; ypT0/Tis ypN0 P=0.034). Likelihood ratio tests demonstrated that for both pCR endpoints, PD-L1 CPS (P=0.683/P=0.422) and on-treatment sTILs (P=0.984/P=0.568) did not add significantly more value to pretreatment sTILs when predicting pCR. Conclusions:Higher quantities of pretreatment sTILs and PD-L1 CPS and on-treatment sTILs were significantly associated with higher pCR rates and ORR in primary TNBC treated with pembro and NAT. Citation Format: Loi S, Schmid P, Cortés J, Park YH, Muñoz-Couselo E, Kim S-B, Sohn J, Im S-A, Holgado E, Foukakis T, Kuemmel S, Dent R, Wang A, Aktan G, Karantza V, Salgado R. Relationship between tumor infiltrating lymphocytes (TILs) and response to pembrolizumab (Pembro)+chemotherapy (Chemo) as neoadjuvant treatment (NAT) for triple-negative breast cancer (TNBC): phase Ib KEYNOTE-173 trial [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-09.