Background. Breast cancer is the most common solid tumor diagnosed in women, both in Italy and worldwide. Most breast cases are classified as hormone receptor-positive (HR+) HER2-negative (HER2-). The cornerstone of metastatic HR+/HER2- breast cancer treatment is endocrine therapy, often used in association with targeted therapy. Despite the effectiveness of current therapies, all patients will sooner or later experience disease progression due to the development of endocrine resistance. Consequently, it is crucial to dynamically recharacterize the biology of metastatic breast cancer in order to potentially guide treatment sequencing; in this context liquid biopsy might represent a useful approach. Within this context, the multicenter prospective observational study CHAMBER is being conducted to dynamically investigate, using liquid biopsy, mutations affecting genes of interest for endocrine resistance and tumorigenesis, particularly ESR1 and PIK3CA. The present study evaluates the feasibility of serial liquid biopsies and analyses preliminary data from the CHAMBER study, allowing for a first comparison with data available in literature. Materials and methods. Plasma samples were collected during routine venipuncture for each patient; the timepoint for each sample collection was established at every disease progression detected clinically or through imaging techniques. To identify circulating tumor DNA (ctDNA), the samples were hybridized with 45 target genes from the Kapa Custom HyperChoice panel and subsequently sequenced using NGS with the HighOutput NextSeq kit. The company GenomeUp handled the alignment and variant calling processes. The alignment and variant calling were managed by GenomeUp. Only pathogenic and likely pathogenic variants were considered in the analysis. Results. To date, 93 patients have been enrolled at the IOV IRCSS in Padova. This paper presents preliminary data from samples collected from 8 patients. Serial sampling was performed a minimum of 1 time (in 2 patients) and a maximum of 4 times. Analysis of the samples from the 6 patients undergoing serial liquid biopsies revealed an apparent association between increased ctDNA levels and disease progression. Furthermore, considering the mutations under investigation, the mutation in ESR1 was detected in 50% of the patients, all of whom had been previously treated with aromatase inhibitors. This percentage is high, exceeding the ones reported in literature, and it is consistent with the fact that this population has undergone significant pretreatment with endocrine therapy. The mutation in PIK3CA was identified in 75% of the patients, with 3 cases occurring in the known hotspots H1047L, H1047R, E545K. Five out of the six patients had previously been treated with fulvestrant (as monotherapy or in combination), supporting the hypothesis in the literature that activation of the PI3K pathway seems to be involved in mechanisms of resistance to fulvestrant therapy. Conclusions. The preliminary results of our study require further investigation with future data that will emerge from the remaining enrolled population. However, the information obtained so far supports the use of liquid biopsy as a valuable tool for monitoring the dynamic progression of the disease, with the future goal of using this method to promptly adjust therapy and provide significant clinical benefits to patients.
Introduzione. Il carcinoma della mammella è la malattia oncologica più frequente nelle pazienti di sesso femminile, sia in Italia che nel mondo. La maggior parte dei casi di carcinoma mammario rientra nel sottotipo a recettori ormonali positivi (HR+) HER2-negativo (HER2-). Il cardine del trattamento del carcinoma mammario HR+/HER2- in stadio metastatico è la terapia endocrina, coadiuvata dall’uso di farmaci a bersaglio molecolare. Tuttavia, nonostante l’efficacia delle terapie attuali, i pazienti andranno incontro ad una progressione di malattia a causa della comparsa di resistenza ai trattamenti (tra cui la resistenza alla terapia ormonale, anche detta endocrino resistenza). Conseguentemente, si rende rilevante ricaratterizzare in modo dinamico la biologia della malattia metastatica al fine di determinare più specificatamente la corretta sequenza di farmaci da adottare; una metodica potenzialmente utile è la biopsia liquida. In questo contesto nasce lo studio multicentrico osservazionale prospettico CHAMBER che indaga, grazie all’utilizzo della biopsia liquida, le mutazioni a carico di geni di interesse per l’endocrino resistenza e la tumorigenesi, in particolare ESR1 e PIK3CA. Il presente elaborato valuta la fattibilità di biopsie liquide seriate e analizza i primi dati derivati dallo studio CHAMBER, permettendo così un preliminare confronto con i dati di letteratura. Materiali e metodi. Pazienti con carcinoma mammario metastatico HR+/HER2-, arruolate nello studio CHAMBER, sono state sottoposte a prelievo di plasma durante i prelievi ematici di routine. Il timepoint per ogni prelievo è stato stabilito ad ogni progressione di malattia rilevata clinicamente o mediante tecniche di imaging. Il DNA tumorale circolante (ctDNA) estratto dai campioni di plasma è stato analizzato in NGS, mediante ibridazione con un pannello Kapa Custom HyperChoice (45 geni target) e successivo sequenziamento usando il kit HighOutput NextSeq; l’azienda GenomeUp ha gestito l’allineamento e la chiamata delle varianti. Nell’analisi sono state considerate solo le varianti patogenetiche e verosimilmente patogenetiche. Risultati. Ad oggi presso lo IOV IRCSS di Padova sono state arruolate 93 pazienti con carcinoma mammario metastatico HR+/HER2-. In questo elaborato vengono presentati i dati preliminari emersi da campioni prelevati da 8 pazienti. I prelievi seriati sono stati effettuati da minimo di 1 (in 2 pazienti) ad un massimo di 4 volte; dall’analisi dei campioni delle 6 pazienti sottoposte a biopsie liquide seriate è emersa una associazione numerica tra l’aumento dei livelli di ctDNA e il progredire della malattia. Inoltre, considerando le mutazioni oggetto di indagine, la mutazione in ESR1 è stata rilevata nel 50% delle pazienti, tutte precedentemente trattate con inibitori delle aromatasi. Questa percentuale risulta elevata, superiore a quelle riportate in letteratura, ed è consistente con il fatto che si tratta di una popolazione significativamente pretrattata con terapia endocrina. La mutazione in PIK3CA è emersa nel 75% delle pazienti considerate, in 3 casi in uno degli hotspot noti (H1047L, H1047R e E545K). Cinque delle sei pazienti per le quali è stata rilevata una mutazione di PIK3CA erano state trattate in precedenza con fulvestrant, sia in monoterapia che in combinazione, avvalorando l’ipotesi presente in letteratura secondo cui l’attivazione della pathway di PI3K sembra essere implicata nei meccanismi di resistenza alla terapia con fulvestrant. Conclusioni. I risultati preliminari del nostro studio necessitano di ulteriori approfondimenti. Tuttavia, le informazioni fino ad ora emerse appaiono supportare il potenziale utilizzo della biopsia liquida come strumento utile per monitorare l’andamento dinamico della malattia, con l’obiettivo futuro di utilizzare questa metodica per modulare tempestivamente la terapia.
Evoluzione dinamica delle mutazioni di ESR1 e PIK3CA su DNA tumorale circolante nelle pazienti con carcinoma mammario HR+/HER2- metastatico: analisi dello studio osservazionale prospettico CHAMBER
MENEGALDO, VALENTINA
2022/2023
Abstract
Background. Breast cancer is the most common solid tumor diagnosed in women, both in Italy and worldwide. Most breast cases are classified as hormone receptor-positive (HR+) HER2-negative (HER2-). The cornerstone of metastatic HR+/HER2- breast cancer treatment is endocrine therapy, often used in association with targeted therapy. Despite the effectiveness of current therapies, all patients will sooner or later experience disease progression due to the development of endocrine resistance. Consequently, it is crucial to dynamically recharacterize the biology of metastatic breast cancer in order to potentially guide treatment sequencing; in this context liquid biopsy might represent a useful approach. Within this context, the multicenter prospective observational study CHAMBER is being conducted to dynamically investigate, using liquid biopsy, mutations affecting genes of interest for endocrine resistance and tumorigenesis, particularly ESR1 and PIK3CA. The present study evaluates the feasibility of serial liquid biopsies and analyses preliminary data from the CHAMBER study, allowing for a first comparison with data available in literature. Materials and methods. Plasma samples were collected during routine venipuncture for each patient; the timepoint for each sample collection was established at every disease progression detected clinically or through imaging techniques. To identify circulating tumor DNA (ctDNA), the samples were hybridized with 45 target genes from the Kapa Custom HyperChoice panel and subsequently sequenced using NGS with the HighOutput NextSeq kit. The company GenomeUp handled the alignment and variant calling processes. The alignment and variant calling were managed by GenomeUp. Only pathogenic and likely pathogenic variants were considered in the analysis. Results. To date, 93 patients have been enrolled at the IOV IRCSS in Padova. This paper presents preliminary data from samples collected from 8 patients. Serial sampling was performed a minimum of 1 time (in 2 patients) and a maximum of 4 times. Analysis of the samples from the 6 patients undergoing serial liquid biopsies revealed an apparent association between increased ctDNA levels and disease progression. Furthermore, considering the mutations under investigation, the mutation in ESR1 was detected in 50% of the patients, all of whom had been previously treated with aromatase inhibitors. This percentage is high, exceeding the ones reported in literature, and it is consistent with the fact that this population has undergone significant pretreatment with endocrine therapy. The mutation in PIK3CA was identified in 75% of the patients, with 3 cases occurring in the known hotspots H1047L, H1047R, E545K. Five out of the six patients had previously been treated with fulvestrant (as monotherapy or in combination), supporting the hypothesis in the literature that activation of the PI3K pathway seems to be involved in mechanisms of resistance to fulvestrant therapy. Conclusions. The preliminary results of our study require further investigation with future data that will emerge from the remaining enrolled population. However, the information obtained so far supports the use of liquid biopsy as a valuable tool for monitoring the dynamic progression of the disease, with the future goal of using this method to promptly adjust therapy and provide significant clinical benefits to patients.| File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.12608/47436