Phase II Study of Gemcitabine, Carboplatin, and Iniparib As Neoadjuvant Therapy for Triple-Negative and BRCA1/2 Mutation–Associated Breast Cancer With Assessment of a Tumor-Based Measure of Genomic Instability: PrECOG 0105

Abstract

Purpose

This study was designed to assess efficacy, safety, and predictors of response to iniparib in combination with gemcitabine and carboplatin in early-stage triple-negative and BRCA1/2 mutation–associated breast cancer.

Patients and Methods

This single-arm phase II study enrolled patients with stage I to IIIA (T ≥ 1 cm) estrogen receptor–negative (≤ 5%), progesterone receptor–negative (≤ 5%), and human epidermal growth factor receptor 2–negative or BRCA1/2 mutation–associated breast cancer. Neoadjuvant gemcitabine (1,000 mg/m² intravenously [IV] on days 1 and 8), carboplatin (area under curve of 2 IV on days 1 and 8), and iniparib (5.6 mg/kg IV on days 1, 4, 8, and 11) were administered every 21 days for four cycles, until the protocol was amended to six cycles. The primary end point was pathologic complete response (no invasive carcinoma in breast or axilla). All patients underwent comprehensive BRCA1/2 genotyping, and homologous recombination deficiency was assessed by loss of heterozygosity (HRD-LOH) in pretreatment core breast biopsies.

Results

Among 80 patients, median age was 48 years; 19 patients (24%) had germline BRCA1 or BRCA2 mutations; clinical stage was I (13%), IIA (36%), IIB (36%), and IIIA (15%). Overall pathologic complete response rate in the intent-to-treat population (n = 80) was 36% (90% CI, 27 to 46). Mean HRD-LOH scores were higher in responders compared with nonresponders (P = .02) and remained significant when BRCA1/2 germline mutations carriers were excluded (P = .021).

Conclusion

Preoperative combination of gemcitabine, carboplatin, and iniparib is active in the treatment of early-stage triple-negative and BRCA1/2 mutation–associated breast cancer. The HRD-LOH assay was able to identify patients with sporadic triple-negative breast cancer lacking a BRCA1/2 mutation, but with an elevated HRD-LOH score, who achieved a favorable pathologic response. Confirmatory controlled trials are warranted.

INTRODUCTION

Breast cancers that arise in BRCA1 and BRCA2 mutation carriers are characterized by homologous recombination DNA repair deficiency. The BRCA1 and BRCA2 gene products are critical for DNA double-strand break repair, and as such, DNA repair–targeted therapeutics have been investigated to exploit the inherent homologous recombination deficiency of these tumors to therapeutic advantage.¹ Preclinical studies have demonstrated that BRCA1/2-deficient breast tumors exhibit differential chemosensitivity compared with BRCA1/2-proficient cancers, with greater sensitivity to platinum and gemcitabine and less sensitivity to taxanes.^2–5 In clinical studies, data have shown high-level activity of cisplatin as neoadjuvant therapy in BRCA1 mutation carriers and poorer response rates and progression-free survival in BRCA1 mutation carriers (compared with noncarriers) with metastatic hormone receptor–negative breast cancer treated with taxane therapy.⁶

Sporadic triple-negative breast cancer (TNBC) shares many pathologic and molecular features with breast cancers caused by hereditary BRCA1 germline mutations.⁷ On the basis of this and the observations described, the hypothesis emerged that sporadic TNBC may possess similar DNA repair defects and demonstrate similar chemosensitivity profiles as BRCA1 mutation–associated breast tumors. Preclinically, basal-like breast cancer cell lines, like BRCA1-deficient cancer cell lines, demonstrate increased sensitivity to poly (ADP-ribose) polymerase (PARP) inhibition, cisplatin, and gemcitabine and are deficient in base excision repair, leading to enhanced sensitivity to oxidative DNA damage.^8,9

Therefore, we set out to investigate a neoadjuvant combination chemotherapy regimen targeting DNA repair defects in early-stage TNBC and BRCA1/2 mutation–associated breast cancers. Specifically, PrECOG 0105 was designed to assess the efficacy and safety of and predictors of response to iniparib (BSI-201; sanofi-aventis, Paris, France) in combination with gemcitabine and carboplatin. Over the course of this study, knowledge regarding the mechanism of action of iniparib evolved. Initially investigated as a PARP1 inhibitor, it was subsequently demonstrated that iniparib does not possess characteristics typical of the PARP inhibitor class.^10–12 Preclinically, the metabolites of iniparib are believed to be involved in the uncoupling of electron transport from oxidative phosphorylation, which in turn produces reactive oxygen species at cytotoxic levels.¹³

A major goal of this study was to identify markers of response to this neoadjuvant therapy among patients with TNBC. Given the clinical potential of DNA repair–targeted therapeutics, many groups have focused on developing methods to characterize changes in the genomic landscape resulting from underlying homologous recombination defects in cancers.^14–17 In addition to BRCA1 and BRCA2, there are many DNA repair genes that may be altered by germline or somatic mutations, rearrangements, DNA methylation, or dysregulated mRNA expression that are hypothesized to result in impairment of the homologous recombination pathway. The homologous recombination deficiency loss of heterozygosity (HRD-LOH) assay allows for the detection of HRD regardless of etiology or mechanism as measured by levels of genomic LOH.¹⁴ During assay development, LOH regions of intermediate size were observed more frequently in ovarian tumors with defective BRCA1/2. On the basis of this finding, the HRD-LOH assay was developed and represents a count of the number of LOH regions of intermediate size (> 15 Mb and < whole chromosome) observed in the tumor genome. In this article, we report the first assessment to our knowledge of the HRD-LOH biomarker in TNBC and evaluate its ability to distinguish responders from nonresponders treated with neoadjuvant platinum-based therapy.

PATIENTS AND METHODS

Patients, Study Design, and Treatment Regimen

This single-arm phase II neoadjuvant study enrolled patients with newly diagnosed, treatment-naive stage I to IIIA (T size ≥ 1 cm by magnetic resonance imaging [MRI]) TNBC (estrogen receptor [ER] ≤ 5%, progesterone receptor [PR] ≤ 5%, and human epidermal growth factor receptor 2 [HER2] negative [0 or 1+ by immunohistochemistry or fluorescent in situ hybridization nonamplified]) or BRCA1/2 mutation–associated breast cancer. A core biopsy of the primary breast tumor was required for research purposes. All patients underwent comprehensive BRCA1 and BRCA2 genotyping.

Patients were treated with carboplatin intravenously (IV) at an area under the curve of 2 on days 1 and 8, gemcitabine 1,000 mg/m² IV on days 1 and 8, and iniparib 5.6 mg/kg IV on days 1, 4, 8, and 11 every 21 days before definitive surgery. In the original protocol, patients at Stanford University were treated with four cycles of therapy (total of 13 patients) before the protocol was amended to increase treatment duration to six cycles and expand the trial to multiple centers within PrECOG, with the goal of treating 80 patients with the six-cycle regimen. After completion of surgery, adjuvant systemic therapy and radiotherapy were recommended at the discretion of the treating physician.

This protocol was approved by the institutional review board at each participating center. Informed consent was obtained from all patients.

Study End Points and Statistical Considerations

The primary end point was pathologic complete response (pCR) by central assessment, defined as the absence of invasive carcinoma in the breast and axillary lymph nodes. The extent of residual disease was assessed using the residual cancer burden (RCB) index.¹⁸ This index has been validated as an independent prognostic marker of distant relapse-free survival in patients with breast cancer treated with neoadjuvant chemotherapy (RCB 0, complete pathologic response; RCB I, minimal residual disease; RCB II, moderate residual disease; and RCB III, extensive residual disease). Additional secondary end points included safety of the combination, radiographic response by MRI (central review), rate of conversion to breast conservation eligibility, and correlation of baseline tumor gene expression and gene copy number profiles with treatment response.

Per protocol, the primary analysis was to combine patients enrolled to receive four or six cycles of neoadjuvant therapy. At the time the study was expanded, the revised design was based solely on the patients to be treated with six cycles. Therefore, both the 6-cycle cohort alone and the combined cohorts are reported. The primary analysis included all patients registered with the intent to treat (ITT). Efficacy analyses were performed for all eligible patients and safety analyses for all patients who received at least one dose of trial therapy. Assuming that 76 of 80 patients were eligible and treated, the regimen would be deemed of interest if the lower bound of the 90% exact binomial CI for the pCR rate exceeded 25%. This design had an 87.5% power to detect a 15% absolute improvement in pCR over historical data with cisplatin alone,¹⁹ using a binomial test with a one-sided α level of 5%.

RESULTS

Patient and Tumor Characteristics

A total of 93 patients were treated in the study. Thirteen patients were enrolled onto the four-cycle protocol, and 80 patients were enrolled onto the six-cycle protocol. All patients in the four-cycle group completed treatment. Of the 80 patients in the six-cycle group, 11 (13.8%) discontinued treatment prematurely: five (6.3%) because of progressive disease, five because of unacceptable toxicity (four with wild-type BRCA, one with mutant BRCA), and one because of a protocol violation (patient lost to follow-up with mutant BRCA). There were no ineligible patients, so the ITT and safety populations included the same number of patients.

Patient characteristics are detailed in Table 1. All enrolled patients were women, and the majority were white (72%). Most patients had clinical stage II breast cancer (72%), and most had TNBC, except for three BRCA1/2 mutation carriers who had ER-positive and/or PR-positive/HER2-negative breast cancer. A total of 19 patients (24%) treated with six cycles had a deleterious germline mutation in BRCA1, BRCA2, or both genes. Sixty-five patients treated with six cycles of therapy had fresh frozen tumor tissue available for gene expression profiling (Affymetrix U133 plus 2.0; Santa Clara, CA), and of these, 78% were basal-like by PAM50 analysis²⁰ (Data Supplement). Fifty-one of the 65 samples passed the ER filter used by the Vanderbilt TNBC type calculator (http://cbc.mc.vanderbilt.edu/tnbc/), allowing for triple-negative molecular subtype assignment with the following distribution: basal-like 1 (BL1), n = 8 (16%); basal-like 2 (BL2), n = 2 (4%); immunomodulatory (IM), n = 14 (27%); luminal androgen receptor (LAR), n = 4 (8%); mesenchymal, n = 14 (27%); mesenchymal stem-like (MSL), n = 3 (6%); and unstable, n = 6 (12%; Data Supplement).²¹

Table 1.

Patient Demographic and Clinical Characteristics

Characteristic	Patients Receiving Six Cycles (n = 80)		All Patients (N = 93)
	No.	%	No.	%
Female sex	80	100.0	93	100.0
Race
Asian	10	12.5	11	11.8
Black or African American	7	8.8	9	9.7
Other	1	1.3	2	2.2
Unknown	4	5.0	4	4.3
White	58	72.5	67	72.0
Ethnic group
Hispanic or Latino	7	8.8	10	10.8
Not Hispanic or Latino	72	90.0	82	88.2
Unknown	1	1.3	1	1.1
Age at screening, years
Median	48.0		48.0
Range	26-74		26-74
Clinical stage
I	10	13	12	13
IIA	29	36	35	38
IIB	29	36	32	34
IIIA	12	15	14	15
Breast cancer subtype
Triple negative	77	96	90	97
ER and/or PR positive/HER2 negative	3	4	3	3
BRCA1/2 mutation status*
BRCA1 mutation	14	18	15	16
BRCA2 mutation	4	5	4	4
BRCA1 and BRCA2 mutations	1	1	1	1

Abbreviations: ER, estrogen receptor; HER2, human epidermal growth factor 2; PR, progesterone receptor.

^*Germline.

Response Data

The primary efficacy results for pathologic response in the ITT population are summarized in Table 2. Among all 80 enrolled patients treated with six cycles of therapy, 29 patients (36.3%) achieved a pCR. Among those wild type for BRCA1/2, the pCR rate was 33%. Among BRCA1/2 mutation carriers, the pCR rate was 47%, and in BRCA1/2 mutation carriers with TNBC, it was 56%. One BRCA1 mutation carrier had bilateral breast cancer and achieved a pCR in both breasts, although this was counted as one response. Among all 93 patients, 31 (33.3%) achieved a pCR (90% CI, 25.3% to 42.2%). Rates of combined RCB 0/1 are also detailed in these subgroups.

Table 2.

Pathologic Response in ITT Population

Response	All Patients		BRCA1/2 Wild Type		BRCA1/2 Mutant		TN and BRCA1/2 Mutant
	No.	%	No.	%	No.	%	No.	%
Patients receiving six cycles	n = 80		n = 61		n = 19		n = 16
pCR, RCB 0	29	36	20	33	9*	47	9*	56
90% CI, %	27 to 46		23 to 44		27 to 68		33 to 77
RCB 0 or 1	45	56	31	51	14	74	12	75
90% CI, %	46 to 66		40 to 62		52 to 89		52 to 91
Patients receiving four and six cycles	n = 93		n = 73		n = 20		n = 17
pCR, RCB 0	31	33	22	30	9	45	9	53
90% CI, %	25 to 42		21 to 40		26 to 65		31 to 74
RCB 0 or 1	51	55	36	49	15	75	13	76
90% CI, %	46 to 64		39 to 60		54 to 90		54 to 92

Abbreviations: ITT, intent to treat; pCR, pathologic complete response; RCB, residual cancer burden; TN, triple negative.

^*One BRCA1 mutation carrier had bilateral TN breast cancer and achieved pCR in both breasts.

In the six-cycle and combined groups, respectively, 58 (72.5%; 90% CI, 63.1% to 80.6%) and 68 patients (73.1%; 90% CI, 64.5% to 80.6%) achieved an objective response by MRI. Among the 23 and 27 patients not eligible for breast-conservation surgery at baseline in the six-cycle and combined groups, 14 (60.9%; 90% CI, 41.7% to 77.8%) and 15 (55.6%; 90% CI, 38.2% to 72.0%), respectively, became eligible for breast conservation.

Adverse Events

All patients had at least one treatment-emergent adverse event (TEAE). The most common treatment-related TEAEs among all 93 patients and 80 patients treated with six cycles, respectively, were fatigue (84.9% and 85.0%), nausea (81.7% and 81.3%), neutropenia or neutrophil count decreased (49.5% and 53.8), alopecia (46.2% and 51.3%; [grade 2, 4.3% and 5.0%]), anemia (33.3% and 35%), dysgeusia (25.8% and 28.8%), diarrhea (24.7% and 26.3%), and rash (20.4% and 23.8%). All grade 4 TEAEs occurred in patients receiving six cycles of treatment. There were no deaths during the study. Table 3 summarizes all grade 3 to 4 adverse events possibly, probably, or definitely related to the combination treatment regimen. Notably, among 80 patients treated with six cycles of therapy, the rate of grade 2 (complete) alopecia was only 5%, and the rate of grade ≥ 2 neuropathy was 1%.

Table 3.

Grade 3 to 4 Adverse Events by CTCAE (version 3.0)

Adverse Event	Six Cycles (n = 80)				Four and Six Cycles (N = 93)
	Grade 3		Grade 4		Grade 3		Grade 4
	No.	%	No.	%	No.	%	No.	%
Neutropenia	33	41	6	8	36	39	6	6
Febrile neutropenia	0	0	0	0
ALT elevation	12	15	0	0	14	15	0	0
Anemia	8	10	0	0	8	9	0	0
AST elevation	7	9	0	0	7	8	0	0
Thrombocytopenia	4	5	2	3	4	4	2	2
Fatigue	2	3	0	0	3	3	0	0
Subdural hematoma	0	0	1	1	0	0	1	1
Cerebrovascular accident	0	0	1	1	0	0	1	1
Pulmonary embolism	0	0	1	1	0	0	1	1
Headache	1	1	0	0	1	1	0	0
Nausea	1	1	0	0	1	1	0	0
Vomiting	1	1	0	0	1	1	0	0
Flu-like illness	1	1	0	0	1	1	0	0
Urinary tract infection	1	1	0	0	1	1	0	0

NOTE. Adverse events possibly, probably, or definitely related to treatment with gemcitabine, carboplatin, and iniparib.

Abbreviation: CTCAE, Common Terminology Criteria for Adverse Events.

Correlative Studies

HRD-LOH.

Given the hypothesis of underlying DNA repair defects in sporadic TNBC, we set out to evaluate a novel measure of genomic instability to detect the accumulation of changes in the genomic landscape of a tumor attributable to defective homologous recombination DNA repair. Methods for determination of the HRD-LOH score are detailed in the Appendix (online only).

Among 80 patients treated in the six-cycle protocol, 77 (18 with mutant BRCA1/2) had sufficient DNA extracted from their tumor core biopsies to proceed with the HRD assay (Data Supplement). Of these, 66 samples (17 with mutant BRCA1/2) passed the quality filter based on the level of discrimination between balanced and unbalanced regions of the tumor genome, and 11 did not (including one with mutant BRCA1/2). One BRCA1 mutation carrier was excluded because she had no pathologic outcome data available, leaving 65 patients with HRD-LOH and response data. In total, four somatic mutations were identified in BRCA1 or BRCA2 on tumor sequencing; two of these patients achieved a pCR.

In Figure 1, the distribution of HRD-LOH scores among responders in blue and nonresponders in gold is depicted. Mean HRD-LOH scores were higher in responders compared with nonresponders (15.7 v 12.5; P = .020; Table 4). Importantly, mean HRD-LOH scores were similar in BRCA1/2-mutant versus intact responders. When those with BRCA1/2 germline mutations were excluded, mean HRD-LOH scores in BRCA1/2 wild-type responders were higher than in wild-type nonresponders (16.1 v 12.3; P = .021). When the data were analyzed using a cutoff of ≥ 10 as indicative of homologous recombination deficiency, responders were more likely to exhibit HRD-LOH scores ≥ 10 compared with nonresponders in all patients and BRCA1/2 wild-type patients (P = .0026 and .0024, respectively). As shown in Figure 2, rates of favorable pathologic response, defined as RCB of 0 or 1, were 66% versus 20% for patients with an HRD-LOH score ≥ 10 compared with < 10; 81% versus 47% for patients harboring an underlying germline BRCA1/2 mutation compared with wild type; and 66% versus 8% for patients with a high HRD-LOH score or BRCA1/2 mutation compared with patients with a low HRD-LOH score who were also BRCA1/2 wild type, respectively (Fig 2). When data were analyzed using RCB 0 (pCR) versus no pCR, mean HRD-LOH scores were 16.0 among those with a pCR and 13.4 among those with no pCR (P = .058). When looking at HRD-LOH as a dichotomous variable (< 10 v ≥ 10), patients with a pCR were more likely to exhibit HRD-LOH scores ≥ 10 compared with patients who did not achieve a pCR (P = .0012).

Fig 1.

Homologous recombination deficiency–loss of heterozygosity (HRD-LOH) score distribution among responders (residual cancer burden, 0 or 1) and nonresponders (residual cancer burden, 2 or 3). Plot shows distribution of HRD-LOH scores among responders in blue (BRCA1/2 mutant and wild type) and nonresponders in gold. Size of each dot corresponds to number of patients with that score; larger dot indicates more patients with that HRD-LOH score.

Table 4.

Association of Response (RCB 0 or 1) and HRD-LOH Score (n = 65 of 80 patients receiving six cycles)

Response	HRD-LOH Score	P*
	Mean
All patients (n = 65)		.020
Responders (n = 36)	15.7
Nonresponders (n = 29)	12.5
BRCA1/2 wild type (n = 49)		.021
Responders (n = 23)	16.1
Nonresponders (n = 26)	12.3

Abbreviations: HRD-LOH, homologous recombination deficiency–loss of heterozygosity; RCB, residual cancer burden.

^*Wilcoxon rank sum test.

^†Fisher's exact test.

Fig 2.

Rate of favorable response (residual cancer burden [RCB], 0 or 1) by homologous recombination deficiency–loss of heterozygosity (HRD-LOH) score, germline BRCA1/2 status, and their combination (n = 65). Proportion of patients achieving favorable response, defined as RCB index score of 0 or 1 based on (A) HRD-LOH score low (< 10) versus high (≥ 10), (B) germline BRCA1/2 status wild type (WT) versus mutant, and (C) combination of HRD-LOH low (< 10) and germline BRCA1/2 WT versus HRD-LOH high (≥ 10) or germline BRCA1/2 mutant. Total of 65 of 80 patients in six-cycle protocol had HRD-LOH data available.

In an attempt to better understand the underlying mechanism for a high HRD score and/or clinical response in patients with TNBC without germline or tumor BRCA1/2 mutations, a preliminary analysis of tumor BRCA1 promoter methylation and subsequent gene expression was performed for 25 of 45 BRCA1/2 wild-type patients with HRD-LOH data. In total, 15 samples had BRCA1 promoter methylation, and all had an HRD-LOH score ≥ 10. Nine (60%) of 15 were responders (RCB 0 or 1; Data Supplement).

Gene expression.

In an exploratory analysis in the six-cycle group, we assessed response by RCB group (0 or 1 v 2 or 3) across the TNBC molecular subtypes. As previously reported,²² we also observed considerable variation in rates of favorable response across these subtypes, although numbers in some of the groups were small, and results must be interpreted cautiously (Data Supplement). When germline BRCA1/2 mutation status was overlaid on this, we observed that those with mutant BRCA1/2 were distributed across the various subtypes, with only one BRCA-mutant patient in the BL1 and BL2 groups combined. Mean HRD-LOH scores across the TNBC subtypes were as follows: BL1, 17 (range, 2 to 24; n = 8); BL2, 20 (n = 1); IM, 15.9 (range, 6 to 24; n = 10); LAR, 6.7 (range, 5 to 8; n = 3); mesenchymal, 13.1 (range, 6 to 25; n = 14); MSL, 13 (range, 10 to 16; n = 2); and unstable, 10.6 (range, 6 to 17; n = 6). Appendix Table A1 (online only) lists the molecular characteristics of the five patients with progressive disease.

DISCUSSION

The study met its primary end point, with a pCR rate of 36% (90% CI, 27% to 46%) among patients treated with six cycles of therapy and a pCR rate of 33.3% (90% CI, 25.3% to 42.2%) in all patients. Given the nonrandomized nature of this study, the relative contribution of iniparib therapy cannot be assessed. Nevertheless, this platinum-based, non–anthracycline- and non–taxane-based regimen was well tolerated and produced pathologic responses that were on par with those recently reported with third-generation anthracycline/taxane-based regimens (pCR, 26% to 39%).^23,24 In molecularly defined subgroups of our TNBC cohort selected for DNA repair deficiencies using BRCA1/2 mutation status and/or the HRD-LOH assay, we observed favorable pathologic response rates (RCB, 0 or 1) ≥ 65%, supporting the targeted nature of the therapy. Our results do not have direct impact on clinical practice today, but they strongly suggest that patient selection based on underlying DNA repair deficiency in future randomized trials of standard versus DNA repair defect–targeted therapy in TNBC should be pursued. A strength of this study is the comprehensive molecular phenotyping that was performed on all patient tumors. Microarray gene expression analysis confirmed that the majority of patients selected for triple-negative status based on immunohistochemical characterization were basal-like (78% by PAM50). Further subtyping using the Vanderbilt gene expression criteria suggest that some patients with TNBC may exhibit particular chemosensitivity (IM and MSL groups), whereas others are more resistant (LAR) and that these differences are not directly related to BRCA1/2 germline mutation status.

This study also comprehensively evaluated BRCA1/2 germline mutation status of all enrolled patients as well as pretreatment tumor biopsy samples for BRCA1/2 mutations and a measure of genomic instability using a novel diagnostic approach. The pCR rate with this platinum-based regimen was highest in patients with germline BRCA1/2 mutations, although comparisons in our study were underpowered. Among patients lacking a germline BRCA1/2 mutation, the HRD-LOH assay was able to identify additional patients with sporadic TNBC and an elevated HRD-LOH score associated with underlying defects in homologous recombination who achieved a favorable pathologic response. Therefore, the HRD-LOH assay seems to be a powerful diagnostic tool for assessing DNA repair capacity of tumors, as reflected by a so-called genomic scar, without knowledge of an underlying genetic cause and without the need for assessing a DNA damage–inducible response to therapy (eg, gamma-H2AX focus formation). Prospective evaluation of the HRD-LOH assay is necessary to confirm whether this biomarker is prognostic or truly predictive of therapeutic benefit to DNA-damaging therapy such as platinum as well as newer-generation PARP inhibitors. The molecular mechanism for HRD in our patients with TNBC without a BRCA1/2 mutation is not known, but epigenetic downregulation of BRCA1 expression secondary to promoter methylation may explain this in part.

Whether the carboplatin and gemcitabine backbone of our regimen represents an improved therapy for the HRD-selected patients with TNBC in our trial, or whether these patients simply respond better to any cytotoxic regimen, cannot yet be assessed. To date, the results of three randomized neoadjuvant TNBC platinum studies have been reported, all of which examined an add-on approach of carboplatin to anthracycline- and taxane-based therapy. The GEICAM (Grupo Español de Investigación en Cáncer de Mama) 2006-03 study of epirubicin and cyclophosphamide followed by docetaxel with or without carboplatin showed no improvement in pCR with the addition of carboplatin.²⁵ In the phase II GeparSixto trial, which assessed a regimen of dose-intense anthracycline- and taxane-based chemotherapy with bevacizumab with or without carboplatin, patients receiving carboplatin achieved a pCR rate of 53.2% compared with 36.9% in patients who did not.²⁶ In the phase II CALGB 40603 (Cancer and Leukemia Group B) study of standard anthracycline/taxane-based chemotherapy with or without carboplatin and with or without bevacizumab, the addition of carboplatin significantly increased the pCR rate (breast and axilla) in the per-protocol population by 13% (pCR with carboplatin, 54% v without, 41%).²³ Toxicity was increased and early discontinuation was more common in carboplatin-treated patients.

Placing these results, along with the results of the GeparSixto and CALGB 40603 studies, in the broader context deserves careful consideration. In the conduct of PrECOG 0105, we found that certain toxicities commonly encountered with anthracycline/taxane-containing adjuvant therapy, specifically alopecia (grade 2, 5%), chemotherapy-induced amenorrhea, and peripheral neuropathy (grade 2, 1%; grade 3 to 4, 0%), were infrequent. In a randomized setting, we would expect that important differences in toxicity would be observed among patients treated with gemcitabine and carboplatin compared with a regimen such as paclitaxel and carboplatin followed by doxorubicin and cyclophosphamide. These toxicity differences are important and may affect endocrine symptom burden, reproductive concerns, and overall quality of life in this group of predominantly young, premenopausal breast cancer survivors.

In summary, our data from PrECOG 0105 support the notion that understanding the biology of TNBC and BRCA1- and BRCA2-mutant breast cancer allows for improved therapeutic strategies that target the DNA repair defects of these tumors. Future trials with treatment selection based on tumor DNA repair capacity in TNBC are currently in development and may lead to improved long-term outcomes.

Glossary Terms

BRCA1:

a tumor suppressor gene known to play a role in repairing DNA breaks. Mutations in this gene are associated with increased risks of developing breast or ovarian cancer.

BRCA2:

a tumor suppressor gene whose protein product is involved in repairing chromosomal damage. Although structurally different from BRCA1, BRCA2 has cellular functions similar to BRCA1. BRCA2 binds to RAD51 to fix DNA breaks caused by irradiation and other environmental agents. Also known as the breast cancer 2 early onset gene.

homologous recombination:

genetic recombination whereby nucleotide sequences are exchanged between two similar or identical strands of DNA to facilitate accurate repair of DNA double-strand breaks.

neoadjuvant therapy:

the administration of chemotherapy prior to surgery. Induction chemotherapy is generally designed to decrease the size of the tumor prior to resection and to increase the rate of complete (R0) resections.

triple-negative breast cancer (TNBC):

Breast tumors that are negative for estrogen and progesterone receptor expression, and that also underexpress HER-neu.

Appendix

Homologous Recombination Deficiency–Loss of Heterozygosity Assay

To generate a homologous recombination deficiency–loss of heterozygosity (HRD-LOH) assay score, DNA copy number was determined using genome-wide single-nucleotide polymorphism (SNP) data generated from Affymetrix (Santa Clara, CA) MIP arrays (n = 2) or a custom Agilent (Santa Clara, CA) SureSelect XT capture followed by sequencing on an Illumina (San Diego, CA) HiSeq2500 (n = 33) or both (n = 42). When data from both assays were available, the highest quality score was used for analysis. SNP data were analyzed using an algorithm that determines the most likely allele-specific copy number at each SNP location after accounting for contamination of the tumor sample with nontumor DNA.

The HRD-LOH score was calculated by counting the number of LOH regions that were > 15 Mb in length but shorter than the length of a complete chromosome. The correlation coefficient for the 30 samples with passing assays on both platforms was 0.93. Tumor sequence data for BRCA1 and BRCA2 were analyzed for the presence of variants from wild-type sequence. Variants were classified as deleterious or suspected deleterious based on previously described criteria. Read coverage across each exon was used to detect large rearrangements. BRCA1 promoter methylation was assessed by bisulfite conversion and polymerase chain reaction amplification of the proximal promoter region followed by next-generation sequencing.

In this analysis, we defined an HRD-LOH score < 10 as homologous recombination proficient and an HRD-LOH score ≥ 10 as homologous recombination deficient. This cutoff of 10 corresponds to the 10th percentile of HRD score distribution in a set of 260 ovarian and breast cancer tumors with deleterious mutations in the BRCA1 or BRCA2 gene or promoter methylation of the BRCA1 or RAD51C gene.

Table A1.

Characteristics of Patients With Progressive Disease

Patient	ER (%)	PR (%)	BRCA1/2 Germline Mutation Status	HRD-LOH Score	TNBC Subtype
1	0	0	Negative	9	Mesenchymal
2	0	0	Negative	24	Basal-like 1
3	0	0	Negative	9	Sample did not pass ER filter (high degree of contamination)
4	0	0	Negative	8	LAR
5	1	1	Negative	11	No gene expression data available

Abbreviations: ER, estrogen receptor; HRD-LOH, homologous recombination deficiency–loss of heterozygosity; LAR, luminal androgen receptor; PR, progesterone receptor; TNBC, triple-negative breast cancer.

AUTHORS' DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

Disclosures provided by the authors are available with this article at www.jco.org.

AUTHOR CONTRIBUTIONS

Conception and design: Melinda L. Telli, Allison W. Kurian, Robert W. Carlson, Anne-Renee Hartman, Judith Manola, James M. Ford

Financial support: Melinda L. Telli, James M. Ford

Administrative support: Melinda L. Telli, Pei-Jen Chang, Joseph A. Sparano, James M. Ford

Provision of study materials or patients: Melinda L. Telli, Allison W. Kurian, Irene L. Wapnir, Robert W. Carlson, Joseph A. Sparano, Bobbie Head, Lori J. Goldstein, Barbara Haley, Shaker R. Dakhil, James M. Ford

Collection and assembly of data: Melinda L. Telli, Kristin C. Jensen, Allison W. Kurian, Jafi A. Lipson, Kirsten Timms, Elizabeth A. Schackmann, Irene L. Wapnir, Pei-Jen Chang, Bobbie Head, Barbara Haley, Shaker R. Dakhil, Judith Manola, James M. Ford

Data analysis and interpretation: Melinda L. Telli, Kristin C. Jensen, Shaveta Vinayak, Patrick J. Flaherty, Kirsten Timms, Victor Abkevich, Joseph A. Sparano, Lori J. Goldstein, Julia E. Reid, Anne-Renee Hartman, Judith Manola, James M. Ford

Manuscript writing: All authors

Final approval of manuscript: All authors

AUTHORS' DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

Phase II Study of Gemcitabine, Carboplatin, and Iniparib As Neoadjuvant Therapy for Triple-Negative and BRCA1/2 Mutation–Associated Breast Cancer With Assessment of a Tumor-Based Measure of Genomic Instability: PrECOG 0105

The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or jco.ascopubs.org/site/ifc.

Melinda L. Telli

Consulting or Advisory Role: Oncoplex DX, Vertex

Research Funding: sanofi-aventis (Inst), Novartis (Inst), Abbott Laboratories (Inst), Calithera Biosciences (Inst), PharmaMar (Inst), Myriad Genetics (Inst), Biomarin (Inst)

Kristin C. Jensen

No relationship to disclose

Shaveta Vinayak

Travel, Accommodations, Expenses: Incyte

Allison W. Kurian

Research Funding: Myriad Genetics (Inst), Invitae (Inst)

Jafi A. Lipson

Employment: Genentech (I)

Research Funding: Hologic (Inst)

Patrick J. Flaherty

No relationship to disclose

Kirsten Timms

Employment: Myriad Genetics

Stock or Other Ownership: Myriad Genetics

Research Funding: Myriad Genetics

Patents, Royalties, Other Intellectual Property: Myriad Genetics (Inst)

Victor Abkevich

Employment: Myriad Genetics

Stock or Other Ownership: Myriad Genetics

Elizabeth A. Schackmann

No relationship to disclose

Irene L. Wapnir

No relationship to disclose

Robert W. Carlson

Research Funding: sanofi-aventis (Inst)

Pei-Jen Chang

No relationship to disclose

Joseph A. Sparano

No relationship to disclose

Bobbie Head

No relationship to disclose

Lori J. Goldstein

No relationship to disclose

Barbara Haley

Research Funding: Novartis (Inst), Pfizer (Inst), Roche (Inst)

Shaker R. Dakhil

No relationship to disclose

Julia E. Reid

Employment: Myriad Genetics

Stock or Other Ownership: Myriad Genetics

Anne-Renee Hartman

Employment: Myriad Genetics

Stock or Other Ownership: Myriad Genetics

Judith Manola

Research Funding: sanofi-aventis (Inst)

James M. Ford

Research Funding: sanofi-aventis (Inst), Myriad Genetics (Inst), Invitae (Inst), Varian (Inst)