Philippe Bertheau

Effect of mutated TP53 on response of advanced breast cancers to high-dose chemotherapy

TP53 activation by genotoxic drugs can induce apoptosis or cell-cycle arrest. Thus, whether the gene is mutated or wild type could affect the response of a tumour to chemotherapy. Clinical data are unclear, possibly as a result of heterogeneity of tumours, drugs, methods of assessing response, or TP53 status. We studied 50 non-inflammatory, locally advanced breast cancers that had been treated with high doses of a combination of epirubicin and cyclophosphamide. We noted eight complete responses, which all occurred in the 14 patients with tumours containing mutated TP53 (p<0.0001). In high-grade, advanced breast cancers, inactivation of the TP53 pathway could greatly improve the response to this chemotherapy regimen.

TP53 status and response to chemotherapy in breast cancer.

Despite its central role in the control of apoptosis, senescence and cell cycle arrest, the tumor suppressor protein p53 remains an enigma for its possible role in predicting response to chemotherapy in cancer patients. Many studies remained inconclusive, others showed a better response for tumors with normal p53, and some recent studies showed adverse effects of normal p53 for response to treatment. p53 is not only a powerful pro-apoptotic factor in response to drug-induced DNA damages but also a potential inducer of cell cycle arrest, protecting tumor cells from further cytotoxic damages. Our review describes the classical as well as the more recent concepts. In order to draw definite conclusions, future works should use more reliable methods to assess the TP53 status and should address more homogeneous tumor subpopulations treated with homogeneous chemotherapy regimens.

p53 in breast cancer subtypes and new insights into response to chemotherapy.

Despite an obvious central role of p53 in the hallmarks of cancer, TP53 status is not yet used for the management of breast cancer. Recent findings may lead to reconsider the role of p53 in breast cancer. TP53 mutations are the most frequent genetic alterations in breast cancer, observed in 30% of breast carcinomas. Their distribution is highly linked to molecular tumor subtypes found in 26% of luminal tumors (17% of luminal A, 41% of luminal B), in 50% of HER2 amplified tumors, in 69% of molecular apocrine breast carcinomas and in 88% of basal-like carcinomas. The type of mutation is linked to the tumor subtype with higher frequency of base-pair substitutions in luminal tumors, whereas molecular apocrine and basal-like tumors present much higher frequency of complex mutations (deletions/insertions). The timing of TP53 mutation also depends on the tumor subtype, being the first important event in luminal tumors but occurring after PTEN loss in basal-like tumors. Regarding response to cytotoxic chemotherapy, the situation is far from the p53-dependent apoptosis paradigm with subsequent clinical response. We reported that TP53 mutated non inflammatory locally advanced breast carcinomas had a high rate of complete pathological response to dose-dense doxorubicin-cyclophosphamide chemotherapy, while TP53 wild-type (WT) tumors never achieved complete response. Using human breast cancer xenograft models, we suggested that this could be due to the induction of senescence in TP53 WT tumor cells. A recent work confirmed these findings in MMTV-Wnt1 mammary tumors, showing that growth arrest and senescent phenotype, not apoptosis, were induced in TP53 WT tumors following doxorubicin treatment, while lack of arrest in mutant tumors resulted in aberrant mitoses, cell death and a superior clinical response. Furthermore, in ER positive (ER(+)) breast tumors, it has been recently reported that ER represses the p53-mediated apoptotic response induced by DNA damage. Taken together, these data can help to better understand p53-mediated response to doxorubicin-based chemotherapy in breast cancer: in ER(+) TP53 WT breast cancers, ER-induced inhibition of p53 apoptotic response would lead preferentially to tumor cell senescence and subsequent resistance to treatment. Conversely, in ER negative (ER(-)) TP53 mutated breast cancers, accumulation of genetic abnormalities would lead to mitotic catastrophe and subsequent better response. In view of these recent results, p53 impact in breast cancer should be reconsidered.

Molecular apocrine breast cancers are aggressive estrogen receptor negative tumors overexpressing either HER2 or GCDFP15

IntroductionMolecular apocrine (MA) tumors are estrogen receptor (ER) negative breast cancers characterized by androgen receptor (AR) expression. We analyzed a group of 58 transcriptionally defined MA tumors and proposed a new tool to identify these tumors.MethodsWe performed quantitative reverse transcription PCR (qRT-PCR) for ESR1, AR, FOXA1 and AR-related genes, and immunohistochemistry (IHC) for ER, PR, Human Epidermal Growth Factor Receptor 2 (HER2), CK5/6, CK17, EGFR, Ki67, AR, FOXA1 and GCDFP15 and we analyzed clinical features.ResultsMA tumors were all characterized by ESR1(-) AR(+) FOXA1(+) and AR-related genes positive mRNA profile. IHC staining on these tumors showed 93% ER(-), only 58% AR(+) and 90% FOXA1(+). 67% and 57% MA tumors were HER2(3+) and GCDFP15(+), respectively. Almost all MA tumors (94%) had the IHC signature HER2(3+) or GCDFP15(+) but none of the 13 control basal-like (BL) tumors did. Clinically, MA tumors were rather aggressive, with poor prognostic factors.ConclusionMA tumors could be better defined by their qRT-PCR-AR profile than by AR IHC. In addition, we found that HER2 or GCDFP15 protein overexpression is a sensitive and specific tool to differentiate MA from BL in the context of ER negative tumors. A composite molecular and IHC signature could, therefore, help to identify MA tumors in daily practice.

p53 dependent cell-cycle arrest triggered by chemotherapy in xenografted breast tumors

The major long‐term prognostic factor for breast cancer patients treated by first‐line chemotherapy is response to treatment. We have previously shown that complete responses to high doses epirubicin‐cyclophosphamide were observed only in human tumors bearing a TP53 mutation. Three xenografted human breast tumors, 2 of them with a TP53 mutation and one of them without, were studied for their immediate response to this drug association. Cell cycle, cellular senescence and cell death were characterized and quantified on tissue section before and after treatment. The TP53 wild‐type tumor showed a strong early induction of senescence‐like phenotype with overexpression of SA‐β‐gal and p21CIP1. In contrast both TP53 mutated tumors showed no sign of cell cycle arrest or senescence. Conversely, abnormal mitoses strongly increased, only in TP53 mutated tumors. Thus, in these in vivo models, epirubicin‐cyclophosphamide treatment induces senescence‐like features in TP53 wild‐type tumor, likely accounting for cell cycle arrest and subsequent resistance to treatment. Conversely in TP53 mutated tumors, chemotherapy induces mitotic catastrophe and tumor death, accounting for complete response to this association exclusively in patients with TP53 mutated tumors.

Differential regulation of sunitinib targets predicts its tumor-type-specific effect on endothelial and/or tumor cell apoptosis

AbstractPurposenSunitinib is an inhibitor of tyrosine-kinase receptors, and no biomarker predictive of sunitinib response is available. The purpose of this preclinical study was to show whether sunitinib molecular targets could be used as biomarkers to assess tumor response to sunitinib in human cancer cell line xenografts of three different tumor types.MethodsUsing mice xenografted with liver, breast and renal carcinoma cell lines, we sequentially analyzed the effect of 7-day sunitinib treatment on tumor and vascular compartments.ResultsIn all xenografts, microvessel damage occurred from Day 1. Tumor damage also occurred in liver, breast, but not in renal xenografts. Using specific human and mouse probes for genes encoding sunitinib targets, we showed a significant relation between apoptotic tumor cell numbers and human PDGFRΒ and RET mRNA expression in liver cancer and to human VEGFR2 expression in breast cancer xenografts. In contrast, in renal cancer xenografts, vascular effect evaluated by measuring endothelial cell apoptosis was related to mouse Vegfr1, Vegfr2 and Vegfa-164 expression.ConclusionThis study identifies sunitinib vascular and tumor effects according to different tumor types and shows that sunitinib molecular targets used as biomarkers enable assessment of therapeutic response.

Functional TP53 mutations have no impact on response to cytotoxic agents in metastatic colon cancer

BACKGROUNDnSurvival of metastatic colon cancer (mCC) patients has considerably improved with optimization of new drugs regimen. Inactivation of TP53 pathway by TP53 mutations is observed in nearly half of colorectal tumors. The impact of such mutations has been poorly studied in the metastatic setting.nnnMETHODSnThe files of 254 mCC treated in a single institution at Saint-Louis hospital between January 1999 and April 2011 were retrospectively reviewed. Tissue samples for analysis of TP53 mutations were available for 68 patients, performed using FASAY. The prognostic value of TP53 status was evaluated by comparing progression free survival (PFS) and overall survival (OS) in the group of TP53-mutated and wild type patients.nnnRESULTSnPFS was 6.9 months and OS 21.7 months in the whole population. There was no statistical difference in TP53-mutated and wild type groups in term of PFS (HR=1.04; IC 95%=0.6-1.79) and OS (HR=0.99; IC 95%=0.53-1.55) whatever the chemotherapy regimen (oxaliplatin- or irinotecan-based). Only BRAF V600 mutation was demonstrated to be a poor prognostic factor for PFS and OS, and CEA level for OS.nnnCONCLUSIONSnRoutine determination of TP53 mutations, even with a highly sensitive method, cannot be recommended to predict chemotherapy response in mCC.

p53 in the Clinic: A Pathologist’s View

The p53 protein has many biological effects and a high prevalence of mutations in human cancer. Pathologists use this biomarker in several different situations, to help for the differential diagnosis of reactive/dysplastic/benign/malignant lesions, to better characterize malignant tumours, and to help in predicting response to treatment. p53 status of dysplastic and borderline lesions can most often be assessed only by immunohistochemistry (IHC) since these microscopical lesions are too small for advanced molecular assays. p53 protein status determined by IHC has however a smaller sensitivity compared to TP53 status assessed by gene sequencing. This explains why p53 as a biomarker remains not so widely used by pathologists except for several well-defined indications detailed in this chapter. P63, a member of the p53 family, is also frequently used in diagnostic pathology as a differentiation marker and will be discussed in this chapter.

Principe et applications de la microdissection laser en histopathologie

Resume Les tissus biologiques normaux sont constitues de differentes populations de cellules adoptant une architecture souvent tres intriquee. Ceci entraine une grande heterogeneite dans la composition de chaque tissu. La microdissection laser permet, sous controle de la vue et de l’expertise du pathologiste, de selectionner une population ou une region cellulaire d’interet sur tous types de prelevements tissulaires. Elle repond ainsi au besoin de s’affranchir de l’heterogeneite cellulaire caracterisant aussi les tissus cancereux ou sont disseminees cellules stromales, cellules precancereuses, cellules cancereuses et cellules normales residuelles. Grâce aux developpements technologiques qui n’ont fait que croitre durant la decennie qui vient de s’ecouler dans le domaine de la biologie moleculaire, cette technique a permis de preciser le role important des acides nucleiques (ADN et ARN) et de certaines proteines qualifiees de marqueurs biologiques dans des mecanismes physiopathologiques. C’est en pathologie tumorale que la microdissection laser demontre a l’heure actuelle ses performances dans de nombreux champs d’application dont l’interet sera ici precise.