Elena Gallego

Resistance to Immune Checkpoint Blockade (ICB) constitutes the current limiting factor for the optimal implementation of this novel therapy, which otherwise demonstrates durable responses with acceptable toxicity scores. This limitation is exacerbated by a lack of robust biomarkers. In this study, we have dissected the basal TME composition at the gene expression and cellular levels that predict response to Nivolumab and prognosis. BCR, TCR and HLA profiling were employed for further characterization of the molecular variables associated with response. The findings were validated using a single-cell RNA-seq data of metastatic melanoma patients treated with ICB, and by multispectral immunofluorescence. Finally, machine learning was employed to construct a prediction algorithm that was validated across eight metastatic melanoma cohorts treated with ICB. Using this strategy, we have unmasked a major role played by basal intratumoral Plasma cells expressing high levels of IGKC in efficacy. IGKC, differentially expressed in good responders, was also identified within the Top response-related BCR clonotypes, together with IGK variants. These results were validated at gene, cellular and protein levels; CD138+ Plasma-like and Plasma cells were more abundant in good responders and correlated with the same RNA-seq-defined fraction. Finally, we generated a 15-gene prediction model that outperformed the current reference score in eight ICB-treated metastatic melanoma cohorts. The evidenced major contribution of basal intratumoral IGKC and Plasma cells in good response and outcome in ICB in metastatic melanoma is a groundbreaking finding in the field beyond the role of T lymphocytes.

Resistance to Immune Checkpoint Blockade (ICB) constitutes the current limiting factor for the optimal implementation of this novel therapy that otherwise demonstrates durable responses with acceptable toxicity scores. This limitation is exacerbated by a lack of robust biomarkers. In this study, we have dissected the basal TME composition at the gene expression and cellular levels that predict response to Nivolumab and prognosis. BCR, TCR and HLA profiling were employed for further characterization of the molecular variables associated with response. The findings were validated using a single-cell RNA-seq data of metastatic melanoma patients treated with ICB and by multispectral immunofluorescence. Finally, machine learning was employed to construct a prediction algorithm that was validated across eight metastatic melanoma cohorts treated with ICB. Using this strategy, we have unmasked a major role played by basal intratumoral plasma cells expressing high levels of IGKC in efficacy. IGKC, differentially expressed in good responders, was also identified within the Top response-related BCR clonotypes, together with IGK variants. These results were validated at gene, cellular and protein levels; CD138+ Plasma-like and Plasma cells were more abundant in good responders and correlated with the same RNA-seq defined fraction. Finally, we generated a 15-genes prediction model that outperformed the current reference score in eight ICB-treated metastatic melanoma cohorts. The evidenced major contribution of basal intratumoral IGKC and plasma cells in good response and outcome in ICB in metastatic melanoma is a groundbreaking finding in the field beyond the role of T lymphocytes.

Introduction: Recurrence risk in breast cancer varies throughout the follow-up time. We examined if these changes are related to the level of expression of the proliferation pathway and intrinsic subtypes. Methods: Expression of estrogen and progesterone receptor, Ki-67, human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor (EGFR) and cytokeratin 5/6 (CK 5/6) was performed on tissue-microarrays constructed from a large and uniformly managed series of early breast cancer patients (N = 1,249). Subtype definitions by four biomarkers were as follows: luminal A (ER + and/or PR+, HER2-, Ki-67 <14), luminal B (ER + and/or PR+, HER2-, Ki-67 ≥14), HER2-enriched (any ER, any PR, HER2+, any Ki-67), triple-negative (ER-, PR-, HER2-, any Ki-67). Subtype definitions by six biomarkers were as follows: luminal A (ER + and/or PR+, HER2-, Ki-67 <14, any CK 5/6, any EGFR), luminal B (ER + and/or PR+, HER2-, Ki-67 ≥14, any CK 5/6, any EGFR), HER2-enriched (ER-, PR-, HER2+, any Ki-67, any CK 5/6, any EGFR), Luminal-HER2 (ER + and/or PR+, HER2+, any Ki-67, any CK 5/6, any EGFR), Basal-like (ER-, PR-, HER2-, any Ki-67, CK5/6+ and/or EGFR+), triple-negative nonbasal (ER-, PR-, HER2-, any Ki-67, CK 5/6-, EGFR-). Each four- or six-marker defined intrinsic subtype was divided in two groups, with Ki-67 <14% or with Ki-67 ≥14%. Recurrence hazard rate function was determined for each intrinsic subtype as a whole and according to Ki-67 value. Results: Luminal A displayed a slow risk increase, reaching its maximum after three years and then remained steady. Luminal B presented most of its relapses during the first five years. HER2-enriched tumors show a peak of recurrence nearly twenty months post-surgery, with a greater risk in Ki-67 ≥14%. However a second peak occurred at 72 months but the risk magnitude was greater in Ki-67 <14%. Triple negative tumors with low proliferation rate display a smooth risk curve, but with Ki-67 ≥14% show sharp peak at nearly 18 months. Conclusions: Each intrinsic subtype has a particular pattern of relapses over time which change depending on the level of activation of the proliferation pathway assessed by Ki-67. These findings could have clinical implications both on adjuvant treatment trial design and on the recommendations concerning the surveillance of patients.

Cyclooxygenase-2 (COX2) is an enzyme that plays a role in different stages of the carcinogenic process and has prognostic and predictive values that have not yet been established. The objective of this study was to evaluate the role of COX2 overexpression in advanced squamous cell carcinoma of the larynx that has been treated with a phonation conservation protocol.

The prognosis and need or not for adjuvant therapy in patients with small breast tumors (≤1cm N0) is the subject of controversy as regards the clinical benefit obtained, toxicity, and the economical costs generated. A retrospective analysis was made of 238 patients with early-stage breast cancer (pT1≤1 cm N0M0) diagnosed between January 1993 and May 2008. As regards the systemic adjuvant treatments provided, (a) 122 (51%) received no treatment, (b) 102 (43%) received hormone therapy, (c) 9 (4%) chemotherapy, and (d) 5 (2%) received both hormone therapy and chemotherapy. An analysis was made of disease-free survival (DFS) and breast cancer-specific survival in our series of patients, and of their correlation to clinicopathological factors (age, tumor size, histological grade, estrogen receptor (ER) expression, HER-2 overexpression, and systemic adjuvant therapy). The median follow-up of this cohort was 63months (range 5-145). Some type of relapse was recorded in 4.2% of the patients (six patients presented local recurrence in all cases subjected to rescue treatment with surgery and/or radiotherapy, three patients developed distant metastases, and one patient presented a resected local recurrence followed by systemic relapse). The 5year DFS was 96%, and the 5year breast cancer-specific survival was 99.6%. A univariate analysis was made of the clinicopathological variables and their association to DFS. None of the variables was seen to be significantly correlated to shorter DSF except for an association between HER-2 overexpression and poor outcome borderline significance (p=0.07). The prognosis of our pT1≤1cm N0M0 tumors was excellent, although the absence of systemic adjuvant therapy in one-half of the patients.

Mutational analysis of the KRAS gene has recently been established as a complementary in vitro diagnostic tool for the identification of patients with colorectal cancer who will not benefit from anti-epidermal growth factor receptor (EGFR) therapies. Assessment of the mutation status of KRAS might also be of potential relevance in other EGFR-overexpressing tumors, such as those occurring in breast cancer. Although KRAS is mutated in only a minor fraction of breast tumors (5%), about 60% of the basal-like subtype express EGFR and, therefore could be targeted by EGFR inhibitors. We aimed to study the mutation frequency of KRAS in that subtype of breast tumors to provide a molecular basis for the evaluation of anti-EGFR therapies.