1-s20-S2468227624002527-main_241123_065708

Scientific African 25 (2024) e02308
Contents lists available at ScienceDirect
Scientific African
journal homepage: www.elsevier.com/locate/sciaf
Protein biomarkers for diagnosis of breast cancer

Emeka Eze Joshua Iweala a, b , Doris Nnenna Amuji a, b, * , Faith Chinasaokwu Nnaji a, b
a
Department of Biochemistry, College of Science and Technology, Covenant University, Ota, Nigeria
b
Covenant Applied Informatics and Communication Africa Centre of Excellence (CApIC-ACE), Covenant University, Ota, Nigeria
A R T I C L E I N F O A B S T R A C T
Editor: DR B Gyampoh Breast cancer remains a major global health challenge, demanding better diagnostic tools.
Traditional methods like mammography have limitations, highlighting the need for specific, non-
Keywords: invasive approaches. Protein biomarkers offer a promising avenue for early and accurate detec
Estrogen receptor tion, potentially leading to improved patient outcomes and personalized treatment. This review
Point-of-care diagnosis
explores key protein biomarkers, including Estrogen Receptor (ER), Progesterone Receptor (PR),
Personalized treatment
Human Epidermal Growth Factor Receptor 2 (HER-2), and Cancer Antigen 27.29(CA27.29),
Protein biomarker
Proteomic methodologies focusing on the proteomic methodologies used in their discovery and validation. However,
challenges exist, such as variability in biomarker expression and limitations in abundance, sta
bility, and specificity, which hinder clinical use. The review discusses innovative strategies to
overcome these challenges, emphasizing the importance of translating biomarker research into
practical applications for personalized medicine in breast cancer diagnosis and therapy. This
exploration contributes to the evolving field of breast cancer diagnostics, paving the way for
future discoveries and improved patient care.
Introduction
Breast cancer has emerged as a significant global health concern, particularly affecting women, with occasional occurrences in
men. Despite advancements in healthcare, the disease remains a leading cause of mortality, with over 600,000 reported deaths
attributed to breast cancer [34,124]. Projections suggest that within the next decade, one in every eight women (12.9 %) may be
diagnosed with breast cancer [19]. The International Agency for Research on Cancer (IARC) reported more than 2.26 million cases of
breast cancer in 2020, marking the first time breast cancer has surpassed lung cancer in incidence rates [76]. While mammography
stands as a widely used method for breast cancer detection, its effectiveness is hindered by errors in both negative and positive ob
servations, as well as excessive detection rates ranging from 1 to 10 % [3,105]. Consequently, biomarkers are urgently needed to
Abbreviations: CTC, Circulating Tumor Cells; PTM, Posttranslational protein Modification; ER, Estrogen Receptor; PR, Progesterone Receptor;
HER-2, Human Epidermal Growth Factor Receptor; TNBC, Triple Negative Breast Cancer; HE4, Human Epididymis Protein 4; CA 15–3, Cancer
Antigen 15–3; AR, Androgen Receptor; ELISA, Enzyme-linked Immunosorbent Assay; IHC, Immunohistochemistry; DHT, Dihydrotestosterone;
RPPA, Reverse Phase Protein Array; CEA, Carcinoembryonic Antigen; LC-MS/MS, Liquid Chromatography-Mass Spectrometry; ddELISA, Droplet
digital ELISA; IARC, International Agency for Research on Cancer; FDA, Food and Drug Administration; DFS, Disease-free survival; NST, No Special
Type; VEGF, Vascular epithelial growth factor; EpCAM, Epithelial Cell Adhesion Molecule; PSMA, Prostate-Specific Membrane Antigen; EVs,
Extracellular Vesicles.
* Corresponding author: Department of Biochemistry, College of Science and Technology, Covenant University, KM. 10 Idiroko Road, Canaan
Land 112104, Ota, Ogun State, Nigeria.
E-mail address: doris.amujipgs@stu.cu.edu.ng (D.N. Amuji).
https://doi.org/10.1016/j.sciaf.2024.e02308
Received 23 March 2024; Received in revised form 11 June 2024; Accepted 1 July 2024
Available online 3 July 2024
2468-2276/© 2024 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license
(http://creativecommons.org/licenses/by/4.0/).
E.E.J. Iweala et al. Scientific African 25 (2024) e02308
facilitate early detection, prognosis, and recurrence prediction. Biomarkers, such as those derived from affected tumor cells or other
tissues, play a crucial role in molecular diagnostics and enable liquid biopsies, a valuable clinical approach leveraging circulating
tumor cells (CTCs), Deoxyribonucleic acid (DNA), and ribonucleic acid (RNA) detectable in blood samples [10]. Despite advancements
in understanding the molecular pathways involved in breast cancer, a universal biomarker applicable to all breast cancer subtypes
remains elusive. Molecular biological markers encompass diverse entities, including DNA, RNA, peptides, proteins, lipids, and me
tabolites. Notably, proteins offer heightened sensitivity, enabling the detection of minute samples for early diagnosis and disease
detection. As such, recent efforts have focused on identifying novel protein biomarkers through proteomic techniques [8]. The pursuit
of precision medicine has gained momentum globally, with initiatives like the United States National Institutes of Health’s All of Us
program aiming to personalize treatment based on individuals’ biological and pathological characteristics. In this context, proteomics,
which integrates protein and genome analysis, has emerged as a prominent approach to elucidating protein biomarkers.
Post-translational protein modifications (PTMs) associated with cancer development further underscore the significance of proteomics
in identifying potential biomarkers [86]. Recent discoveries, such as TOR1B’s potential as a biomarker for predicting bone metastasis
in breast cancer patients, underscore the evolving landscape of personalized healthcare [89]. Protein biomarkers offer distinct ad
vantages in disease identification due to their ease of measurement compared to other biological markers. Genomic approaches enable
the identification of gene modulation, while proteomic approaches highlight variations in protein expression levels, aiding in the
identification of disease hallmarks [20,66,104]. The present study synthesized and consolidated current research findings on protein
biomarkers for breast cancer diagnosis and elucidated the proteomic methodologies used to identify and validate them. By providing a
comprehensive review of the existing literature, we seek to enhance our understanding of the role of protein biomarkers in breast
cancer detection and prognosis. Our analysis focused on key protein biomarkers involved in critical pathways such as cell proliferation,
survival, and invasion.
The current landscape of protein biomarkers
Biomarkers are crucial in disease diagnosis, offering precise information on prognosis and treatment response [17]. In breast
cancer, protein biomarkers play a pivotal role, reliably revealing phenotypical variations and contributing to the heterogeneity of
breast cancer [11]. Despite their importance, only a few have gained approval from the Federal and Drug Administration (FDA) for
breast cancer diagnosis [135]. Protein biomarker expressions aid in diagnosis and serve as therapeutic targets. Their ease of analysis
and wide applicability across healthcare sectors make them invaluable in personalized medicine [14]. However, the complexity of
cancer development and the significant variations in protein levels among patients necessitate a personalized approach to treatment.
This has led to the emergence of precision medicine as a guiding principle in cancer research and therapy [41]. Specific protein
biomarkers, as listed in (Table 1), play crucial roles in breast cancer diagnosis and treatment. These biomarkers guide treatment
decisions, predict patient prognosis, and inform personalized treatment strategies, improving patient outcomes.
Traditional biomarkers
Traditional biomarkers like CA27.29, CA 15–3, and CEA have been pivotal in guiding treatment decisions and predicting prognosis.
For example, CA27.29, or MUC1, is a serum biomarker in breast cancer prognosis and treatment evaluation [52]. This glycoprotein,
detected using BR 27.29 monoclonal antibodies, is crucial for monitoring disease progression and chemotherapy response, especially
in advanced stages where traditional diagnostic methods may be inconclusive [58]. Elevated levels of CA27.29 are associated with
disease progression, making it valuable in detecting treatment failure and recurrence during follow-up care. However, its clinical
utility is tempered by limitations in specificity, leading to false positives in benign conditions and other cancers [7]. Therefore, while
CA27.29 is unsuitable for initial breast cancer screening, it excels in monitoring metastatic breast cancer treatment efficacy when used
alongside clinical assessments and imaging [52]. Comparative studies with biomarkers like CA 15–3 and CEA have shown CA27.29 to
have comparable sensitivity, particularly in node-positive breast cancer patients, highlighting its role in risk stratification and disease
Table 1
Key breast cancer protein biomarkers and their clinical significance.
Biomarker Type of biomarker Clinical utility FDA approval Ref.
CA 27.29 Serum-based Prognosis, Treatment response Approved [52]

CA 15–3 Serum-based Diagnosis, Prognosis Approved [21,72]
CEA Serum-based Prognosis, Treatment response Approved [48]
Estrogen receptor Hormone-based Diagnosis, Prognosis Approved [135,99]
Progesterone receptor Hormone-based Diagnosis, Prognosis Approved [15,75]
HER-2 Tissue-based Diagnosis, Prognosis, Treatment response Approved [5,73]
Ki67 Tissue-based Diagnosis, Prognosis, Treatment choice Investigational [6,65]
Androgen receptor Hormone-based Diagnosis, Prognosis, Treatment response Investigational [53,70]
Fatty acid synthase Tissue-based and Serum-based Prognosis Investigational [59]
BRCA 1 and BRCA 2 Tissue-based Diagnosis, Prognosis Investigational [55]
EpCAM Serum-based Diagnosis, Prognosis Investigational [80]
Cyclin D1 Tissue-based Diagnosis, Prognosis Investigational [2,109]
Mammaglobin A Serum-based Diagnosis, Prognosis Investigational [45,78]
HE4 Serum-based Diagnosis, Prognosis Investigational [1,79,126]
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management [7,52]. Genetic variations in CA27.29 underscore its impact on clinical interpretation and stability, influencing its role in
personalized medicine [67]. In contrast, CA 15–3, another widely used biomarker, is FDA-approved for monitoring treatment response
and recurrence in advanced breast cancer patients but lacks sensitivity for early-stage detection [72,110]. Combining CA 15–3 with
other biomarkers like tumor-associated proteins (TAP) has shown promise in enhancing diagnostic accuracy for early breast cancer
detection [21]. Meanwhile, carcinoembryonic antigen (CEA) serves a similar role in treatment response monitoring but suffers from
non-specific elevation in other cancers and benign conditions, limiting its utility in breast cancer screening [77].
Established biomarkers
Biomarkers like ER, PR, and HER-2 are well-established biomarkers recommended by Expert panels for use in clinical laboratories
for breast cancer diagnosis and treatment decisions. They are actively employed in clinical settings.
Estrogen receptor
The estrogen receptor is a well-established breast cancer biomarker. it is a useful biomarker for predicting a breast cancer patient’s
responsiveness to hormone treatment in an adjuvant setting and disease progression. A large body of research using a merged dataset
concluded that approximately 80 % of all breast cancers are estrogen-positive (ER+). It is clearly defined, nonaggressive, and a better
prognostic sign than estrogen-negative (ER) breast cancer [135]. In patients with estrogen-positive breast cancer, abridged BH3
interacting domain death agonists and 14–3-3 proteins such as gamma, epsilon, tau, beta/alpha, and seta/deta isoforms serve as
neoadjuvant markers for treatment response [41,64]. Research studies in the late nineteenth century were the first to depict the
regression of metastatic tumors of patients after ovariectomy showing hormonal control of breast cancers [36]. ER is one of the major
ovarian hormones accumulating in the reproductive organs. Clinical trials conducted on 380 patients indicated that 55–60 % of ER+
and 8 % of ER- breast cancer regressed in response to endocrine therapies regardless of the treatment type [51]. ER detection in breast
cancer patients is vital for evaluating prognosis and treatment choice in clinical practice [99]. In clinical settings, ER is employed as
one of the main predictive biomarkers of endocrine therapy and its testing is compulsory in all breast cancer [127]. Many hormone
receptor-positive early-stage breast cancer is treated mainly with adjuvant endocrine therapy, thus making ER a valuable biomarker
for prognostic stratification of breast cancer. Others include PR, HER-2, and proliferative index (Ki67). Testing of ER in invasive breast
cancers using validated immunohistochemistry (IHC) has been a gold standard for the prediction of patients who are likely to benefit
from endocrine therapy, and this assay is continuously recommended by the Expert panel. No other assay has taken place for this
purpose yet. also, ER-positive breast cancer is those with 1–100 % of tumor nuclei positive [9].
Progesterone receptor
Progesterone receptor, a protein modulated by estrogen, was the first biomarker for prognosis and predicting response to endocrine
therapies. Assessing PR protein is a gold standard for informed treatment decisions as its expression serves as a predictive biomarker as
regards tumor response to anti-cancer drugs and it also predicts functional and targetable ER in breast cancer [15]. However, there is
controversy in the clinical assessment of PR status, and as such, some laboratories do not assess it routinely, while in some, PR testing is
considered optional in breast cancer. This is major because of its predictive value uncertainty in adjuvant settings as almost all
PR-positive (PR+) tumors are ER+, and ER-, PR+ tumors are rare. studies have indicated that prolonged exposure to PR predisposes
one to breast cancer [51]. PR plays a crucial role in breast cancer development and progression with potential implications for hor
mone treatment and breast cancer prognosis [44]. PR overexpression is mainly observed in breast cancer subtypes with a good baseline
prognosis compared to those with a poor baseline prognosis, serving as a crucial prognostic biomarker [75]. In breast cancer, PR serves
as an important biomarker for DFS or overall survival rate. The two isoforms of PR (PRA and PRB) are co-expressed at the same level in
normal breast cancer. In many cases of ductal carcinomas in situ (DCIS) and invasive breast cancer, studies have found that there is a
higher proportion of PR-A isoform compared to PR-B. This means that PR-A is more predominant than PR-B in these lesions. Thus,
breast cancer treatment is likely to be affected by the ratio of PR-A to PR-B. However, there is a controversy on the viewpoint of PR
isoform’s impacts on breast cancer prognosis. Some studies demonstrated that elevated ratios of the two isoforms are associated with
poor prognosis and relapse after tamoxifen [51]. However, other research has indicated that a higher PR-A to PR-B ratio is associated
with favorable prognostic biomarkers and the presence of a luminal A subtype of breast cancer. A study of immunostaining of PR
showed that the absence or low expression of PR, which may be due to cell dedifferentiation in breast cancer, is associated with poorer
patient prognosis [120]. In approximately 65–75 % of breast cancer, PR positivity has been observed likewise in 80–90 % of ER-related
breast cancer cases [69]. However, there is a variation in the percentage based on the positivity cutoff used, and there has been a record
of equivocal on the predictive role of expression of PR in early-stage breast cancer. IHC is used to assess the expression of PR and this
technique ensures validity, reliability, and result consistency. In recent research of about 16,445 tumor cases that use IHC, 11.3 % PR
immunostaining was observed, and 57.4 % of breast cancer [120]. Low PR immunostaining correlates with advanced tumor stage,
metastasis, positive HER-2 status, and shorter overall survival in breast cancer of no special type (NST) [37,120]. The assessment of ER
and PR biomarkers in tumor samples has greatly helped in the reduction of mastectomies in many women in favor of lumpectomies and
hormone treatments, thus informing treatment decisions and reducing the risk of breast cancer relapse [9].
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Human epidermal growth factor receptor 2
HER-2, also called HER2/neu, belongs to the family of type 1 transmembrane tyrosine kinase receptors [126]. HER-2 is a crucial
modulator of the growth of a cell and embryogenic differentiation, and it is essential for the maturation of humans at puberty.
However, HER-2 signaling deregulation in mammary cells enhances breast carcinogenesis. About 25 % to 30 % of human metastatic
breast tumors overexpresses HER-2 [107]. HER-2 overexpression is a good predictor of decreased survival and reduced recurrence [82,
122]. This is so because cancers grow rapidly and are likely to metastasize compared to the ones that under-expressed HER-2. Hence,
HER-2 has become a vital treatment target for this breast cancer subset [83]. HER-2 level is usually evaluated either by IHC analysis for
expression of HER-2 or fluorescent in situ hybridization analysis (FISH) for HER-2 copy gene during breast cancer diagnosis. There is
the creation of an anti-cancerous drug for breast cancer patients as a result of the discovery of HER-2 or neu oncogene. United States
FDA approved some HER-2 targeted therapies for overexpressed HER-2 in metastasizing breast carcinoma, which are Trastuzumab
(Herceptin), Lapatinib (Tykerb), and pertuzumab [73,92,134]. However, overexpression of HER-2 is a predictive marker that can show
a good outcome when trastuzumab is used [5,135]. Trastuzumab, or Herceptin, is a humanized recombinant monoclonal antibody
targeting the extracellular HER-2 receptor [116].
Proliferation index (Ki67)
Ki67 is a proliferative protein marker associated with cell proliferation, and its expression level, measured as a percentage of
positively stained cells in a tumor sample, is used in clinical settings to assess tumor aggressiveness, predict patient outcomes and
therapeutic response in breast cancer [6]. Much attention has been directed to Ki67 as a prognostic biomarker in luminal breast cancer.
Ki67 IHC serves as a prognostic biomarker in early breast cancer (stage I and II), guiding decisions on adjuvant chemotherapy [32]. It
aids in predicting or monitoring chemotherapy response. Particularly in ER-positive, HER2-negative breast cancers, Ki67 helps
distinguish luminal A from luminal B subtypes, impacting treatment decisions [69,90]. High Ki67 is associated with a favorable
response to chemotherapy, notably in TNBC [12]. However, it is not yet a well-established biomarker and also has limited assessment
in routine practice. The utilization of Ki67 is contentious because its assessment relies on subjective interpretation, potentially
stemming from its questionable analytical validity. This issue hampers the broad acceptance of this biomarker for guiding treatment
decisions in breast cancer [69]. A recent study indicated that high Ki67 is linked to worse prognosis in breast cancer patients, and it also
serves as a marker for treatment outcomes in luminal breast cancer [69]. Of recent, clinical trials have been conducted for Ki67 as a
predictive biomarker for chemotherapy and recurrence-free survival in neoadjuvant endocrine therapy for hormone receptor-positive
breast cancer. European Institute of Oncology clinical trial showed that luminal B breast cancer with positive lymph node metastasis
expressing high Ki67 of above 32 % can benefit from adjuvant chemotherapy. Other previous clinical trials include PACS01,
BCIRG001, and IBCSG. The PACS01 trial showed that an elevated Ki67 index (≥20 %) was associated with greater effectiveness of
docetaxel in adjuvant treatment for ER-positive cancer of the breast. The BCIRG001 trial discovered that a combination of docetaxel,
doxorubicin, and cyclophosphamide (TAC) had a substantial complementing effect on hormonal therapy for patients with an elevated
Ki67 index (≥13 %), ER positivity, and lymph node positivity. IBCSG trials VIII and IX found an association between a high Ki67 index
(≥19 %) and poor DFS in individuals with endocrine-reactive cancer of the breast [132]. Although the Ki67 index was informative for
prognosis and risk stratification in all three trials, its predictive value for response to adjuvant chemotherapy was not observed
uniformly. In the monarchE trial for node-positive, luminal HER2-negative early breast cancer, a Ki67 score of ≥20 % identified
patients at high risk of recurrence. Abemaciclib plus endocrine therapy was found beneficial compared to endocrine therapy alone for
these patients. This study marked the first instance of Ki67 becoming a biomarker influencing therapy choice, leading to FDA approval
inclusion [65,111].
Emerging biomarkers
While established markers like CA 15–3, ER, PR, and HER-2 have undoubtedly played a significant role in breast cancer diagnosis
and treatment, the evolving landscape of the disease demands even more accurate and informative tools. This ongoing search has
identified promising novel protein biomarkers with exciting potential for improved early detection and personalized treatment
strategies. Recent research has revealed some novel markers, such as Fatty acid synthase (FASN), BRCA 1 and BRCA 2, Epithelial cell
adhesion molecule (EpCAM), androgen receptor (AR), mammaglobin-A, cyclin D1, and Human epididymis protein 4 (HE4), offering a
glimpse into the future of breast cancer diagnosis.
Fatty acid synthase
FASN is a novel protein biomarker of breast cancer. It is an enzyme crucial for synthesizing fatty acids and controlling gene
expression in cell death and DNA repair. FASN has been identified as a potential biomarker and therapeutic target in breast cancer.
Elevated levels of FASN are linked to the proliferation, metastasis, resistance to chemotherapy, and recurrence of cancer cells in
various cancers, including breast cancer [94]. A recent study observed a direct relationship between FASN expression and the Ki-67
proliferation index in TNBC. This implies that FASN can be a valuable biomarker for this breast cancer subtype [94]. Furthermore, a
study also suggests that simultaneous assessment of mesenchymal-epithelial transition factor (MET) and FASN expression levels can
serve as predictive indicators for TNBC prognosis, and utilizing both biomarkers together may enhance prognosis assessment accuracy
[59]. Moreover, research is exploring the potential use of FASN in both tissue samples and blood.
4
Androgen receptor
AR, a transcription factor stimulated by androgens such as dihydrotestosterone (DHT) in females and testosterone in males, has
gained increased attention as a promising biomarker of breast cancer [46]. Studies have shown that AR is frequently expressed in
breast cancer and has been implicated in all stages of breast cancer development. The AR expression level varies across various breast
cancer subtypes, with 60 %− 80 % positivity [53]. The prognostic role of AR was studied in 395 breast cancer patients using micro
arrays, and it revealed that high expression of AR is linked to favorable overall survival and good DFS. Thus, AR expression can be a
standalone prognostic biomarker for overall survival and DFS [53]. A study also suggested that AR expression level can be a predictive
biomarker guiding decisions regarding neoadjuvant chemotherapy in the luminal A subtype [70]. Research indicates that the AR could
act as a tumor suppressor in breast cancer cases where estrogen receptor alpha is present (ERα+). At the same time, it may function as a
tumor promoter in breast cancer cases lacking estrogen receptor alpha (ERα-), including HER-2 positive and TNBC, serving as a poor
prognostic factor [130,131]. Furthermore, the androgen receptor has been identified as a favorable positive prognostic factor in breast
cancer cases that express estrogen receptors (ER+) [101]. Studies have also shown that elevated AR expression in HER2-positive breast
cancer patients is associated with a poorer prognosis compared to HER2-negative cases [29].
BRCA1 and BRCA2
BReast CAncer genes-1/− 2 (BRCA1/2) are well-known tumor suppressor genes encoding proteins located at chromosome 17q and
13q, respectively. These genes play a crucial role in DNA repair via homologous recombination and maintenance of genomic stability,
making them key players in breast cancer susceptibility [61]. while primarily studied at the genetic level, their protein products can be
assessed in breast cancer tissue samples. Somatic mutations or epigenetic BRCA gene silencing can lead to BRCA protein deficiency in
sporadic breast cancers, a phenomenon known as BRCAness. This condition exhibits histopathological and molecular features similar
to BRCA1/2 germline mutation-related breast cancers. Several studies have associated BRCA1 protein expression with aggressive
phenotypes in breast cancer. Assessing alterations in BRCA1/2 protein expression levels, such as loss of expression or abnormal
localization, may offer additional diagnostic and prognostic insights. In a recent study involving 110 invasive breast carcinoma pa
tients, protein expression levels were evaluated to identify BRCA1 dysfunction using IHC, a promising and cost-effective technique for
detecting loss of BRCA1/2 protein expression in breast cancer tissue samples. The study observed altered BRCA1 protein expression in
19 % of the breast cancer cases, with loss of BRCA1 expression linked to aggressive breast cancer subtypes. Hence, the reduction of
BRCA1 protein expression holds promise as a potential protein biomarker for breast cancer diagnosis and prognosis [55].
Epithelial cell adhesion molecule
EpCAM, a type 1 transmembrane protein implicated in breast cancer progression, holds significant promise as a diagnostic and
therapeutic target. Studies have demonstrated its overexpression in diverse cancers, including breast cancer, suggesting its potential
utility in cancer treatment [133]. Detection of EpCAM in bodily fluids further underscores its importance as a diagnostic and prog
nostic marker across different cancers, including breast cancer [22,80]. Given its prevalence in TNBC, which is highly expressed in
36–38 % of cases, EpCAM emerges as a particularly promising target for diagnosis and therapy [121]. However, it is noteworthy that in
TNBC, EpCAM overexpression is linked to poor prognosis. To enhance the understanding and management of breast cancer, recent
research has focused on combining EpCAM analysis with other biomarkers, such as epithelial growth factor receptor (EGFR) and
HER-2, using advanced techniques like exchange-PAINT. This simultaneous analysis provides valuable insights into breast cancer
subtypes and guides personalized treatment decisions, marking a significant advancement in breast cancer stratification and therapy
[22]. Furthermore, EpCAM is not currently used routinely, and there is an ongoing clinical trial focusing on detecting EpCAM on
circulating tumor cells in blood for potential non-invasive diagnosis of breast cancer (ClinicalTrials.gov Identifier: NCT05658172).
Cyclin D1
Cyclin D1 is a key cell cycle regulator crucial for transitioning from G1 to the S phase. About 50 % of human breast cancers exhibit
high expression of Cyclin D1 [60]. Various research has linked the expression of cyclin D1 to tamoxifen resistance in breast cancer. As
such, Cyclin D1 has emerged as a potential biomarker of treatment response in an adjuvant setting [2]. In a study conducted on 1003
breast cancer patients where cyclin D1 expression was assessed using IHC, it was reported that Cyclin D1 was overexpressed in 59.4 %
of cases and correlates with breast cancer subsets that have favorable prognostic features. However, an inverse association was found
with TNBC [109]. The study also revealed that Cyclin D1 overexpression has a favorable impact on overall survival, and the patients
with cyclin D1 also have favorable recurrence-free survival. Thus, it denotes that cyclin D1 expression predicts favorable overall
survival, as shown in a recent study conducted on 100 cases of invasive breast cancer of NST. The study found that cyclin D1 over
expression was associated with favorable prognostic features, including ER and PR positivity and low-grade tumors. However, there is
no significant association with overall survival or DFS [18]. This aligns with the first study’s findings regarding the favorable prog
nostic features but differs in terms of the association with survival outcomes. On the other hand, a study observed that low cyclin D1
expression was linked to poor prognostic parameters such as advanced tumor stage, nodal stage, grade, and presence of metastasis
[49]. This contrasts with the findings of the other studies, suggesting a potential discrepancy in the prognostic value of cyclin D1
expression. This contradiction suggests that the prognostic value of Cyclin D1 expression may vary depending on factors such as tumor
subtype, patient characteristics, and detection methods [57]. Further research is needed to reconcile these discrepancies and clarify the
5
role of cyclin D1 expression as a prognostic biomarker in breast cancer.
Mammaglobin- A
Human mammaglobin -A is a secretory protein used as a biomarker for breast cancer diagnosis. Mammaglobin-A is one of the
proteins that differed in expression between breast cancer and normal breast. Mammaglobin-A is only produced by the mammary
gland in the normal breast. In breast cancer, it has been observed to be frequently upregulated, thus posing as a valuable therapeutic
target [117]. There has been a contrasting view concerning the effect of overexpression of mammaglobin-A on the aggressiveness of
breast cancer, with studies finding tumor progression, tumor suppression, or no effect. However, the preferential expression of
mammaglobin-A in breast epithelial cells made mammaglobin-A IHC a well-established technique for detecting metastatic breast
cancer. There is controversy on the data prevalence of mammaglobin-A expression in tumors, which may be a result of factors such as
different antibodies, immunostaining protocols, and thresholds for defining “positive” cases. Still, for breast cancer, the
mammaglobin-A positivity rates reported range from 59 to 100 % and 25–94 % in lobular breast carcinomas and invasive breast
carcinomas of NST, respectively [78]. In a recent study conducted using tissue microarray containing 16,328 samples from various
cancer types, it was reported that among 1139 evaluable invasive breast carcinomas of NST, reduced or absent mammaglobin-A
immunostaining was associated with a higher BRE grade, as well as loss of ER and PR expression and triple-negative status but not
with overall survival [45]. Suggesting that mammaglobin-A expression might not be a vital prognostic feature in breast cancer. In
contrast, about 11 different studies have previously investigated the prognostic significance of mammaglobin-A expression in a range
of 30 to 1017 breast cancer patients. Of these, three studies identified a negative prognosis associated with tumors exhibiting high
mammaglobin-A expression, while five studies reported a negative prognosis linked to tumors with low mammaglobin-A expression.
Additionally, three studies found no correlation between mammaglobin expression and patient outcomes. Researchers have investi
gated serum mammaglobin protein levels for diagnosis. Using specific monoclonal antibodies, they detected mammaglobin in 33 % of
serum samples from primary breast cancer and 44 % of serum samples from metastatic breast cancer, indicating its potential as a
diagnostic marker [78]. The development of alternative biomarkers, such as mammaglobin, highlights ongoing efforts to improve
diagnostic precision in breast cancer management [39].
Human epididymis protein 4
HE4 is greatly expressed in human epididymis epithelial cells [1]. HE4 is a secretory protein that plays physiological roles in cell
growth and differentiation. In recent times, there has been a rise in studies reporting overexpression of HE4 in different tumors, often
accompanied by increased levels of HE4 in the bloodstream. Studies have reported profoundly elevated levels of HE4 gene expression
in ovarian cancer and lung cancer, while breast cancer has moderate HE4 expression levels [50]. Studies have shown that HE4 plays a
crucial role in the diagnosis of different cancer types, including breast cancer, ovarian, and lung cancer [125]. Recent studies have also
indicated a significant increase in both HE4 mRNA and plasma expression of breast cancer patients in the early stage (stage I and II)
[79]. Hence, assessing plasma HE4 levels before surgery can distinguish patients and function as a serological marker for early
diagnosis of primary and recurring breast cancer [79,106]. Research findings have linked increased HE4 expression to poor prognosis
in breast cancer, and HE4 can also serve as a predictive biomarker for lymph node metastasis [1].
In summary, while protein biomarker development has made strides, delayed diagnoses remain a challenge for many breast cancer
patients. This underscores the importance of personalized medicine and the ongoing quest for even more robust biomarkers to improve
early detection and achieve better treatment outcomes.
Advances in protein biomarkers for point-of-care breast cancer diagnosis
The effectiveness of breast cancer treatment and the survival rates of patients hinge significantly on the timely and accurate disease
diagnosis. While mammography and ultrasonography are commonly employed clinical tests for breast cancer, they are limited by their
ability to detect tumors of certain sizes, often resulting in false positives or negatives. Additionally, in low-resource settings, access to
mammograms may be restricted, hindering effective breast cancer management [97]. Therefore, the importance of point-of-care
diagnosis cannot be overstated. Recent advancements in point-of-care diagnostic technology have introduced portable biosensors
that greatly enhance early breast cancer detection and monitoring. These biosensors provide non-invasive, cost-effective, and rapid
testing capabilities. For example, they have demonstrated the ability to detect protein biomarkers such as CA 15–3, CA27.29, HER-2,
and circulating tumor cells (CTCs) with high specificity and selectivity [62,135]. These innovations offer practical solutions for im
mediate implementation in clinical settings, especially in low-resource environments.
Point-of-care diagnosis refers to diagnostic tests conducted near the patient, providing clinically relevant information without
sending samples for processing and analysis in a clinical laboratory. This approach is powerful and accessible, as it is simple to use,
cost-effective, and does not require specialized equipment. Its primary advantage lies in its ability to deliver rapid results within
minutes to hours, facilitating the prompt initiation of treatment options and potentially improving patient outcomes. However, for
point-of-care testing to effectively diagnose breast cancer, the identification of suitable protein biomarkers is paramount. Moreover, a
variety of liquid biopsy biomarkers, including CTCs, circulating tumor proteins, and extracellular vesicles (EVs), have been deployed
for breast cancer diagnosis and prognosis. Liquid biopsies offer a non-invasive and convenient method for detecting and monitoring
breast cancer, providing real-time feedback. Identifying specific protein markers associated with breast cancer circulating tumor cells
could enable rapid point-of-care tests for early detection. Additionally, the analysis of individual proteins within EVs from breast
6
tumors may reveal unique signatures for breast cancer diagnosis. Furthermore, combining identified protein biomarkers from EVs has
the potential to enhance detection sensitivity.
While single protein biomarkers like CA 15–3 hold promise, recent advancements suggest that panels of protein biomarkers
identified from EVs may offer a more robust approach to breast cancer diagnosis. A study by Tian et al. [118] utilized a simple,
sensitive, and cost-effective thermophoretic aptasensor to identify seven serum EVs protein biomarkers (mucin-1, CA125, CEA, HER-2,
EGFR, EpCAM, prostate-specific membrane antigen (PSMA), and vascular endothelial growth factor (VEGF)) based on their expression
levels [118]. It was reported that the weighted sum of these seven identified biomarkers could differentiate patients with breast cancer
from healthy individuals and classify breast cancer types, including distinguishing metastatic from non-metastatic cancer. This sig
nifies the potential of using protein biomarker panels from EVs for improved detection and personalized treatment strategies.
Furthermore, recent research has explored the integration of proteomic analysis with other omics technologies. The study by [129]
combined transcriptomic and proteomic approaches to identify 18 protein biomarkers for each breast cancer stage [129]. This
innovative approach offers valuable insights not only for diagnosis but also for classification and potentially selecting optimal
treatment strategies.
These protein biomarkers promise greater diagnostic accuracy, less invasive sampling procedures, personalized treatment ap
proaches, and expedited diagnosis, ultimately improving patient care and outcomes. Without the prior identification of protein bio
markers, these benefits would remain elusive. Researchers can identify potential biomarker candidates by examining protein
expression patterns in healthy and cancerous tissues. These candidates then undergo rigorous testing to validate their effectiveness and
reliability in diagnosing breast cancer, thereby significantly contributing to improving the cure rate of breast cancer.
Proteomics techniques for protein biomarker detection
Proteomics, an ever-evolving field, explores gene expression patterns within the proteome, offering valuable insights into disease
biology. Modern medicine relies heavily on studying specific biomolecules, or biomarkers, to expedite diagnoses and inform treatment
decisions. Protein biomarkers, in particular, provide crucial information about disease states and can aid in early detection, screening
of high-risk individuals, and prediction of treatment outcomes [4,17]. In cancer research, proteomics has proven instrumental in
unraveling the complex molecular pathways driving tumor growth and evolution [66]. Researchers have developed protein panels
tailored for specific diseases by studying proteins of clinical relevance, aiding in early detection and personalized treatment strategies
[112]. Proteomic techniques are crucial in identifying and characterizing protein biomarkers (Fig. 1), particularly in breast cancer
research [88].
Proteomic techniques, such as mass spectrometry-based technologies and reverse phase protein array (RPPA), are commonly used
to measure protein levels [33]. Techniques like enzyme-linked immunosorbent assay (ELISA) and IHC are routinely used in clinical
settings for cancer diagnosis, prognosis, and treatment monitoring due to their reliability and sensitivity [71,95]. However, challenges
remain, including the scarcity of clinically explained proteomic data from large patient cohorts [84]. Despite these challenges, pro
teomics holds promise in advancing the understanding of disease mechanisms and identifying novel biomarkers for improved patient
care [108]. Its integration with other omics technologies, such as genomics and transcriptomics, offers a comprehensive approach to
unraveling the complexities of diseases like cancer [47,54,98].
Fig. 1. Visual depiction of the general overview of various proteomic techniques and the respective protein biomarkers of breast cancer identified
and validated by them.
7
Enzyme-Linked immunosorbent assays
ELISA is widely utilized in laboratory settings to analyze proteins in minute amounts quantitatively. It is an efficient tool for
biomarker verification, offering immediate detection capabilities [8]. However, ELISA’s limitations in multiplexing, which require
costly development and well-characterized antibodies, hinder its widespread use on a large scale. Nonetheless, ELISA remains a gold
standard for protein biomarkers detection in biological samples, with colorimetric or fluorescent readout signals commonly employed
in FDA-approved assays [93]. Ongoing advancements in cost-effective and label-free ELISA-based techniques hold promise for
enhancing early breast cancer diagnosis. For instance, recent innovations include the development of electrochemical immunosensors
for detecting and validating breast cancer protein biomarkers. Also, digital ELISA was developed to mitigate the challenges of
sensitivity in measuring the low concentration of proteins in biological samples and it is 1000-fold more sensitive than the conven
tional immunoassay method [68]. Yet, its sensitivity is still inadequate in quantifying proteins, thereby limiting the discovery of
biomarkers. However, another ultrasensitive protein detection approach, such as droplet digital ELISA (ddELISA), which uses digital
ELISA and droplet microfluidics, was developed for detecting low protein levels. This simple approach has maximum sensitivity,
surpassing digital ELISA using Single Molecule Arrays (Simoa), the current gold standard for ultrasensitive protein detection, by up to
25-fold. ddELISA is valuable for novel discovery of biomarkers for clinical applications [27]. A study reported an ultrasensitive
detection of protein biomarkers with the naked eye using DNA-programmed plasmonic ELISA. This technique is based on two
enzyme-free and isothermal nucleic acid amplification methods. The technique was able to detect as low as 1 pg mL-1 PSA target with
the naked eye, indicating that the method is potentially useful for early diagnostics and monitoring in less developed areas [23].
Western blots
Western blotting, also referred to as immunoblotting, is an analytical technique used to identify specific proteins within diverse
biological samples, including liquids, tissues, or cell homogenates [114]. First pioneered by Harry Towbin and colleagues in 1979,
Western blotting provides critical insights into protein phosphorylation states and altered protein forms, enabling qualitative or
quantitative analysis [115]. In breast cancer research, Western blot analysis has been instrumental in studying protein expression
changes associated with disease progression. For example, studies have used Western blotting to detect cyclin D1 expression levels in
breast cancer tissues, revealing a significant decrease in cyclin D1 expression with advancing disease stages [96]. Additionally, novel
techniques such as Kinase Activity-Tagged Western Blotting (KAT-WB) enable the assessment of multiple kinase signaling pathways
without using radioactive substances, offering valuable insights into cancer cell signaling mechanisms [38].
Reverse phase protein array
RPPA is a high throughput antibody-based proteomic technology capable of measuring thousands of proteins across numerous
samples, resulting in millions of data points, and is anticipated to play a central role in discovering and validating biomarkers and
therapeutic targets. RPPA is compatible with various protein samples, including cell lysates, plasma, various liquid biopsies, and fresh
and frozen tissues [26]. RPPA approach was introduced in 2001, and it involves immobilizing entire protein lysates onto a solid
support matrix. Specific antibody solutions are then applied to each array site independently, allowing for sensitive and simultaneous
identification of proteins in small sample volumes, such as biopsies. While RPPA requires highly specific antibodies, it enables the
measurement of phosphorylated proteins, facilitating the characterization of complete signaling pathways implicated in different
cancers, including breast cancer [123]. RPPA, known for its sensitivity and ability to quantify post-translationally modified proteins,
including phosphorylated ones, across numerous samples, requires minimal protein amounts. More so, RPPA identifies protein reg
ulators that are minute in abundance and are challenging to quantify by mass spectrometry profiling. Because of these qualities, RPPA
is used to validate potential protein biomarkers or protein pathways found by gene expression profiling or mass spectrometry [26].
With its expandable multiplex capability and extensive antibody libraries, RPPA is deemed ideal for clinical proteomics [123,128].
Immunohistochemistry
IHC serves as a standardized method in pathological diagnosis routines, facilitating the semiquantitative assessment of key bio
markers such as Estrogen Receptor 1 (ESR1), PR, and HER2 receptor levels in breast cancer [13]. This technique holds critical
importance in breast pathology and diagnosis, allowing for the distinction between malignant and non-malignant cells [24]. Through
IHC, researchers can investigate biomarker expression and tissue localization in cancer specimens [115]. Recent advancements in
microfluidic-based IHC have demonstrated clinically proven techniques that offer fast, accurate, and automated identification of breast
cancer compared to standard chromogenic staining methods. Additionally, IHC can be multiplexed using tissue microarrays, enabling
the simultaneous examination of multiple samples from different patients [81]. However, it is essential to note that while IHC is widely
utilized for protein analysis in cancer, it may have limitations in quantifying and detecting activated proteins, particularly those that
are phosphorylated, due to identification limits. Despite its limitations, IHC remains a cornerstone technique in breast cancer research,
facilitating the investigation of biomarker expression patterns and aiding in the discovery of novel biomarkers through tissue
microarrays [13].
8
E.E.J. Iweala et al.
9
Scientific African 25 (2024) e02308

Fig. 2. Schematic overview of proteomic techniques utilized for elucidation of protein biomarkers of breast cancer across various sample types like blood and tissue. The workflow encompasses protein
extraction, digestion for shotgun proteomics, LC-MS/MS, and bioinformatics analysis, then validation of targets and discovery of biomarkers.
Mass spectrometry
Mass spectrometry (MS) has undergone rapid advancements over the years, offering significant advantages in protein biomarker
analysis. Beyond its speed, MS-based technologies excel in de novo identification and simultaneous measurement of numerous pro
teins, making them invaluable tools in proteomic research [85]. Despite requiring substantial initial investment and time-consuming
sample preparation, MS-based techniques offer unparalleled insights into the proteomic landscape of diseases, including breast cancer
[103]. There are two main quantitative MS-based methodologies: directed (targeted) proteomics and shotgun or discovery proteomics.
Directed proteomics involves the quantification of predefined groups of ionic peptides, while targeted proteomics analyzes specific
groups of fragmented ions without identifying precursor ions [87,91]. In contrast, the shotgun approach relies on sequencing and
analyzing digested peptides from the entire proteome using liquid chromatography-mass spectrometry (LC-MS/MS) and automated
database searching. LC-MS/MS plays a crucial role in diagnostic procedures in laboratory medicine, particularly in the identification
and measurement of non-invasive biomarkers using bodily fluids to detect diseases. These methods are also instrumental in pin
pointing proteins present in proximal fluid and membranes, which are often glycosylated and released into the bloodstream of breast
cancer patients or emitted by various breast cancer cell lines. LC-MS/MS analysis can effectively track changes in the proteome of
nipple aspirate fluid, indicating breast cancer development. Additionally, urine also serves as a valuable non-invasive source for
potential biomarkers in breast cancer detection, with LC-MS/MS analysis detecting a range of overexpressed novel proteins [87].
Protein abundance is determined based on the signal of the detected peptides, typically normalized to the background signal of the
proteome or an internal standard. Shotgun proteomics, in particular, enables comprehensive characterization of protein level alter
ations associated with diseases and facilitates the discovery of novel biomarkers. With MS, researchers can study the human proteome
on a massive scale. Mass spectrometry-based techniques play a crucial role in protein biomarker detection in cancer samples, offering
unparalleled sensitivity and specificity for biomarker identification and quantification, as shown in (Fig. 2). Mass spectrometry is a
powerful tool for elucidating the proteomic signatures of breast cancer and holds promise for advancing personalized approaches to
diagnosis and treatment [25].
Challenges in protein biomarker detection and mitigation strategies
Despite their potential, protein biomarkers in breast cancer diagnosis and management face several challenges hindering their
widespread adoption and efficacy. Current protein biomarkers often lack consistent and unique expression patterns across all cancer
patients. Rather than universally applicable markers, proteins showing differential expression between normal and cancerous tissues
are detected [14]. While these proteins reflect similar cellular activities in both normal and cancer cells, their varying expression levels
and consistency pose challenges for reliable biomarker identification (Fig. 3). Additionally, proteins exclusively expressed in tissue
Fig. 3. Schematic representation of challenges and limitations associated with protein biomarkers, along with approaches for mitigation.
10
tumors may be present in limited quantities, limiting their utility as targeted biomarkers in serum samples from cancer patients [30].
Clinically relevant protein biomarkers are present at significantly lower levels than abundantly expressed proteins, complicating
their identification and study [16,113].
The pre-analytical handling and storage conditions of serum samples impact protein stability and biomarker detection. Variations
in temperature during sample storage can lead to protein degradation and alter biomarker profiles. Thawing of serum specimens may
result in partial protein degradation, affecting the reliability of proteomic profiling results [43,100,119]. Enzymatic degradation also
challenges serum or plasma proteomics, leading to biomarker loss [56].
Effective biomarkers for cancer diagnosis, prognosis, and therapy must demonstrate specificity to a particular cancer type and be
expressed exclusively in malignant tissues. However, many breast cancer biomarkers lack the required specificity, limiting their
clinical utility. Moreover, the high cost associated with biomarker discovery, validation, and implementation, coupled with the
invasiveness of some diagnostic tests, presents additional barriers to widespread adoption [16].
To mitigate these challenges, innovative strategies are being explored. These include using protease inhibitors during sample
processing to preserve protein integrity and applying advanced fractionation techniques to enhance the detection of specific bio
markers [35,42,74]. Addressing these technical hurdles is crucial for advancing protein biomarkers’ reliability and clinical utility in
breast cancer.
Clinical translation of protein biomarkers
Translating protein biomarker research findings into clinical practice requires careful consideration of several factors beyond the
inherent limitations of the biomarkers. A significant gap exists between the effort invested in discovering biomarkers and the number
of markers successfully integrated into clinical practice. One of the primary challenges in this translation is the lack of understanding
among some scientists involved in biomarker discovery regarding the analytical, diagnostic, and regulatory requirements for clinical
assays [102]. Bridging this knowledge gap is crucial for developing assays that can be effectively translated into clinical practice.
Once a promising biomarker is identified, developing a suitable assay with excellent analytical performance is imperative to ensure
diagnostic accuracy and enable its eventual integration into routine clinical practice [31]. Therefore, when designing a clinical assay, it
is essential to consider the nature and types of diverse clinical assays currently in use (enzymatic, spectrophotometry, immunoassays,
etc.). Analytical performance alone is insufficient for effective clinical adoption. Mistakes made during biomarker validation have been
reported as a significant reason promising biomarkers fail to make it into clinical practice.
Clinical validation is a challenging and costly process requiring meticulous planning at each step. [31]. To avoid resource waste, it
is crucial to establish a precise definition of how the tumor marker will be used in clinical practice to tailor the validation study
accordingly. This is important because the clinical effectiveness of a tumor marker can vary depending on factors like clinical settings,
disease types, and patient demographics. Assessing the clinical utility of a biomarker, which refers to the extent to which the test used
was able to enhance patient care and prognosis, also poses a challenge to its translation. Randomized controlled trials, considered the
least biased approach, are expensive, need many samples, and involve ethical issues, making them not always feasible [63].
Additionally, navigating the regulatory requirements of the targeted country adds another layer of complexity. For instance, the
FDA in the United States requires rigorous testing and evaluation before approving a new biomarker test for clinical use. These
regulations ensure the safety, efficacy, and reliability of the test for patients. Regulatory requirements can vary across different
countries. Finally, commercialization needs to be considered once a suitable assay format has been selected. The FDA considers
protein-based biomarker assays used for breast cancer diagnosis and management as medical devices and is bound by the same
regulatory principles as other medical devices [40]. This assay is regulated by In Vitro Diagnostics and Radiological Health in the
FDA’s Center for Devices and Radiological Health (CDRH). The FDA classifies medical devices into three classes (Class III, Class II, and
Class I) based on their intended use and risk to patients when incorrect results arise [28]. Biomarkers utilized for cancer diagnosis are
classified as Class III devices, while those used for cancer monitoring or prognosis are considered Class II [40]. This classification
reflects the varying risk levels associated with different diagnostic applications. In summary, translating protein biomarker research
findings into clinical practice requires a multi-faceted approach that considers analytical, diagnostic, regulatory, and commerciali
zation factors. Addressing these challenges is essential for the successful clinical translation of novel biomarkers, invariably leading to
improved patient care and outcomes.
Future direction and conclusion
Protein biomarkers are important in breast cancer diagnosis and therapy because they provide vital information about disease
progression and treatment response. Despite the challenges associated with biomarker identification and validation, proteomic ap
proaches continue to drive progress in personalized medicine. This review advances the knowledge concerning the biology of breast
cancer biology by clarifying the present landscape of protein biomarkers and highlighting ongoing research efforts. Future research
should address the current protein biomarker identification and validation hurdles. Integrating proteomic data with other omics
technologies, such as genomes and transcriptomics, could lead to a more thorough knowledge of breast cancer pathophysiology and
the unraveling of precise biomarkers. Furthermore, developing novel proteomic approaches with higher sensitivity and specificity is
critical for enhancing biomarker identification in serum and tissue. Collaborative efforts between researchers and clinicians are critical
for translating proteomic discoveries into clinical applications. Also, longitudinal studies with varied patient populations are required
to validate newly discovered biomarkers and evaluate their clinical value in real-world settings. More so, novel methodologies are
required to harness the full potential of protein biomarkers in improving early diagnosis, prognosis, and treatment results for breast
11
cancer patients.
CRediT authorship contribution statement
Emeka Eze Joshua Iweala: Conceptualization, Supervision. Doris Nnenna Amuji: Conceptualization, Writing – original draft,
Writing – review & editing, Visualization. Faith Chinasaokwu Nnaji: Writing – review & editing.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to
influence the work reported in this paper.
Acknowledgement
We thank the Covenant University Center for Research, Innovation, and Discovery (CUCRID), Nigeria, for the payment of the article
processing charges (APC) and the Covenant Applied Informatics and Communication Africa center of Excellence (CApIC-ACE) for their
support.
References
[1] M. A Abdelrazek, L. A Barakat, A. Nageb, R. Elbaz, A Abouzid, HE4 elevated levels correlated with breast cancer progression, J. Clin. Images. Med. Case Rep. 3
(3) (2022), https://doi.org/10.52768/2766-7820/1707.
[2] A.A. Abdelnaby, E.A.A. Azzam, A.T E.Al.Nashar, Immunohistochemical expression of cyclin D1 in breast carcinoma, Egypt. J. Hosp. Med. 87 (1) (2022)
2082–2090, https://doi.org/10.21608/ejhm.2022.232822.
[3] S. Afzal, M. Hassan, S. Ullah, H. Abbas, F. Tawakkal, M.A. Khan, Breast cancer; discovery of novel diagnostic biomarkers, drug resistance, and therapeutic
implications, Front. Mol. Biosci. 9 (2022) 783450, https://doi.org/10.3389/fmolb.2022.783450.
[4] A. Ahmad, M. Imran, H. Ahsan, Biomarkers as biomedical bioindicators: approaches and techniques for the detection, analysis, and validation of novel
biomarkers of diseases, Pharmaceutics. 15 (6) (2023) 1630, https://doi.org/10.3390/pharmaceutics15061630.
[5] M.M. Alasmari, A review of margetuximab-based therapies in patients with HER2-positive metastatic breast cancer, Cancers. 15 (1) (2022) 38, https://doi.
org/10.3390/cancers15010038.
[6] A. Alataki, L. Zabaglo, H. Tovey, A. Dodson, M. Dowsett, A simple digital image analysis system for automated Ki67 assessment in primary breast cancer,
Histopathology 79 (2) (2021) 200–209, https://doi.org/10.1111/his.14355.
[7] H.S.K. AL-Azzawi1, M.K. Rasheed2, M. AL-Naqqash3, CA 27-29: a valuable marker for breast cancer management in correlation with CA 15-3, Indian J.
Forensic Med. Toxicol. 14 (4) (2020) 1615–1621, https://doi.org/10.37506/ijfmt.v14i4.11773.
[8] R.A. Alharbi, Proteomics approach and techniques in identification of reliable biomarkers for diseases, Saudi. J. Biol. Sci. 27 (3) (2020) 968–974, https://doi.
org/10.1016/j.sjbs.2020.01.020.
[9] K.H. Allison, M.E.H. Hammond, M. Dowsett, S.E. McKernin, L.A. Carey, et al., Estrogen and progesterone receptor testing in breast cancer: american society of
clinical oncology/college of American pathologists guideline update, Arch. Pathol. Lab. Med. 144 (5) (2020) 545–563, https://doi.org/10.5858/arpa.2019-
0904-SA.
[10] F.O. Alshammari, A.O. Satari, A.S. Aljabali, Y.S. Al-mahdy, Y.J. Alabdallat, Y.M. Al-sarayra, M.A. Alkhojah, A.R.M. Alwardat, M. Haddad, S.A. Al-sarayreh, Y.
M. Al-saraireh, Glypican-3 differentiates intraductal carcinoma and paget’s disease from other types of breast cancer, Medicina 59 (1) (2022) 86, https://doi.
org/10.3390/medicina59010086.
[11] K. Asleh, G.L. Negri, S.E. Spencer Miko, S. Colborne, C.S. Hughes, X.Q. Wang, D. Gao, C.B. Gilks, S.K.L. Chia, T.O. Nielsen, G.B. Morin, Proteomic analysis of
archival breast cancer clinical specimens identifies biological subtypes with distinct survival outcomes, Nat. Commun. 13 (1) (2022) 896, https://doi.org/
10.1038/s41467-022-28524-0.
[12] T.N. Aung, B. Acs, J. Warrell, Y. Bai, P. Gaule, S. Martinez-Morilla, I. Vathiotis, S. Shafi, M. Moutafi, M. Gerstein, B. Freiberg, R. Fulton, D.L. Rimm, A new tool
for technical standardization of the Ki67 immunohistochemical assay, Modern Pathol. 34 (7) (2021) 1261–1270, https://doi.org/10.1038/s41379-021-00745-
6.
[13] S.S. Badve, Y. Gökmen-Polar, Protein profiling of breast cancer for treatment decision-making, Am. Soc. Clin. Oncol. Educat. Book (42) (2022) 73–81, https://
doi.org/10.1200/EDBK_351207.
[14] A.D. Barker, M.M. Alba, P. Mallick, D.B. Agus, J.S.H. Lee, An inflection point in cancer protein biomarkers: what was and what’s next, Mole. Cell. Proteom. 22
(7) (2023) 100569, https://doi.org/10.1016/j.mcpro.2023.100569.
[15] S.M. Bernhardt, P. Dasari, D.J. Glynn, A.R. Townsend, T.J. Price, W.V. Ingman, Comparison of hormone-induced mRNA and protein biomarker expression
changes in breast cancer cells, Breast Cancer Res. Treat. 187 (3) (2021) 681–693, https://doi.org/10.1007/s10549-021-06254-z.
[16] R. Bhawal, A.L. Oberg, S. Zhang, M. Kohli, Challenges and opportunities in clinical applications of blood-based proteomics in cancer, Cancers. 12 (9) (2020)
2428, https://doi.org/10.3390/cancers12092428.
[17] A. Bodaghi, N. Fattahi, A. Ramazani, Biomarkers: promising and valuable tools towards diagnosis, prognosis and treatment of Covid-19 and other diseases,
Heliyon. 9 (2) (2023) e13323, https://doi.org/10.1016/j.heliyon.2023.e13323.
[18] L. Bouzidi, S. Makni, J. Feki, R. Kallel, S. Graja, N. Gouiaa, T. Sellami-Boudawara, M. Mellouli, Immunohistochemical overexpression of cyclin D1 in tunisian
invasive breast carcinoma women, Arch. Iran. Med. 25 (4) (2022) 250–256, https://doi.org/10.34172/aim.2022.41.
[19] M.D. Cation, M.C. Ramos, Accelerating breast cancer biomarker discovery by mass spectrometry proteomics and current bioinformatics tools, Asian Pacific J.
Cancer Biol. 7 (3) (2022) 275–283, https://doi.org/10.31557/apjcb.2022.7.3.275-283.
[20] C. Chase Huizar, I. Raphael, T.G Forsthuber, Genomic, proteomic, and systems biology approaches in biomarker discovery for multiple sclerosis, Cell.
Immunol. 358 (2020) 104219, https://doi.org/10.1016/j.cellimm.2020.104219.
[21] R. Chen, C. Jiang, Q. Zhu, S. You, Y. Li, S. Li, L. Ding, H. Meng, Y. Yang, X. Zha, J. Wang, Combining the tumor abnormal protein test with tests for
carcinoembryonic antigens, cancer antigen 15–3, and/or cancer antigen 125 significantly increased their diagnostic sensitivity for breast cancer, Medicine 99
(29) (2020) e21231, https://doi.org/10.1097/MD.0000000000021231.
[22] Z. Chen, G. Yin, J. Wei, T. Qi, Z. Qian, Z. Wang, S. Zong, Y. Cui, Quantitative analysis of multiple breast cancer biomarkers using DNA-PAINT, Analyt. Methods
14 (37) (2022) 3671–3679, https://doi.org/10.1039/D2AY00670G.
[23] Y.H. Cheng, H. Tang, R.Q. Yu, J.H. Jiang, DNA-Programmed plasmonic ELISA for the ultrasensitive detection of protein biomarkers, Analyst 145 (14) (2020)
4860–4866, https://doi.org/10.1039/D0AN00656D.
[24] A. Cimino-Mathews, Novel uses of immunohistochemistry in breast pathology: interpretation and pitfalls, Modern Pathol. 34 (2021) 62–77, https://doi.org/
10.1038/s41379-020-00697-3.
12
[25] C.A. Ciocan-Cartita, A. Jurj, M. Buse, D. Gulei, C. Braicu, L. Raduly, R. Cojocneanu, L.L. Pruteanu, C.A. Iuga, O. Coza, I. Berindan-Neagoe, The relevance of
mass spectrometry analysis for personalized medicine through its successful application in cancer “Omics, Int. J. Mol. Sci. 20 (10) (2019) 2576, https://doi.
org/10.3390/ijms20102576.
[26] C. Coarfa, S.L. Grimm, K. Rajapakshe, D. Perera, H.Y. Lu, X. Wang, K.R. Christensen, Q. Mo, D.P. Edwards, S. Huang, Reverse-phase protein array: technology,
application, data processing, and integration, JBT 32 (1) (2021) 15–29, https://doi.org/10.7171/jbt.21-3202-001.
[27] L. Cohen, N. Cui, Y. Cai, P.M. Garden, X. Li, D.A. Weitz, D.R. Walt, Single molecule protein detection with attomolar sensitivity using droplet digital enzyme-
linked immunosorbent assay, ACS. Nano 14 (8) (2020) 9491–9501, https://doi.org/10.1021/acsnano.0c02378.
[28] L. Cooper, J. Chen, Changes in companion diagnostic labelling: implementation of FDA’s April 2020 guidance for industry for in vitro Cdx labeling for specific
oncology therapeutic groups, Ther. Innov. Regul. Sci. 56 (5) (2022) 689–697, https://doi.org/10.1007/s43441-022-00422-z.
[29] P. Cruz-Tapias, W. Rubiano, M. Rondón-Lagos, V.E. Villegas, N. Rangel, Intrinsic subtypes and androgen receptor gene expression in primary breast cancer. A
meta-analysis, Biology. 10 (9) (2021) 834, https://doi.org/10.3390/biology10090834.
[30] S. Das, M.K. Dey, R. Devireddy, M.R. Gartia, Biomarkers in cancer detection, diagnosis, and prognosis, Sensors 24 (1) (2023) 37, https://doi.org/10.3390/
s24010037.
[31] K.D. Davis, N. Aghaeepour, A.H. Ahn, M.S. Angst, D. Borsook, et al., Discovery and validation of biomarkers to aid the development of safe and effective pain
therapeutics: challenges and opportunities, Nat. Rev. Neurol. 16 (7) (2020) 381–400, https://doi.org/10.1038/s41582-020-0362-2.
[32] A. De Gregorio, T.W.P. Friedl, E. Hering, P. Widschwendter, N. De Gregorio, I. Bekes, W. Janni, D. Dayan, J.B. Huober, Ki67 as proliferative marker in patients
with early breast cancer and its association with clinicopathological factors, Oncology. 99 (12) (2021) 780–789, https://doi.org/10.1159/000517490.
[33] Z. Ding, N. Wang, N. Ji, Z.S. Chen, Proteomics technologies for cancer liquid biopsies, Mol. Cancer 21 (1) (2022) 53, https://doi.org/10.1186/s12943-022-
01526-8.
[34] T. Dokunmu, P. Obi, O. Fatiregun, O. Rotimi, S. Agodirin, S. Rotimi, Haptoglobin genotypes and malaria comorbidity in breast cancer and healthy Nigerian
women, Ann. Afr. Med. 21 (3) (2022) 231, https://doi.org/10.4103/1596-3519.356811.
[35] V.A. Duong, J.M. Park, H. Lee, Review of three-dimensional liquid chromatography platforms for bottom-up proteomics, Int. J. Mol. Sci. 21 (4) (2020) 1524,
https://doi.org/10.3390/ijms21041524.
[36] R. El Sayed, L. El Jamal, S. El Iskandarani, J. Kort, M. Abdel Salam, H Assi, Endocrine and targeted therapy for hormone-receptor-positive, HER2-negative
advanced breast cancer: insights to sequencing treatment and overcoming resistance based on clinical trials, Front. Oncol. 9 (2019) 510, https://doi.org/
10.3389/fonc.2019.00510.
[37] S. Eskandari, A. Baradaran, S. Ghorbani, P. Masoumzade, The correlation of estrogen and progesterone receptor expression with prognostic parameters in
women with breast cancer, Immunopathologia Persa. (2023) 1, https://doi.org/10.34172/ipp.2023.39486.
[38] M. Eto, S. Katsuki, Y. Tanaka, K. Takeya, Kinase activity-tagged western blotting assay, Biotechniques 68 (4) (2020) 211–213, https://doi.org/10.2144/btn-
2019-0136.
[39] M. Fatima, K.S.S. Sai Baba, N.N.R. Sreedevi, J.P. Kumar, G.S. Raju, S.G. Uppin, M.V. Bhaskar, S.A. Khan, K.M. Iyyapu, M. Noorjahan, Evaluation of serum
mammaglobin as an alternative biomarker in the diagnosis of breast tumors, J. Lab. Physicians 15 (01) (2023) 020–024, https://doi.org/10.1055/s-0042-
1747678.
[40] A.K. Füzéry, J. Levin, M.M. Chan, D.W. Chan, Translation of proteomic biomarkers into FDA approved cancer diagnostics: issues and challenges, Clin.
Proteomics. 10 (1) (2013) 13, https://doi.org/10.1186/1559-0275-10-13.
[41] L.H. Gam, Breast cancer and protein biomarkers, World J. Exp. Med. 2 (5) (2012) 0, https://doi.org/10.5493/wjem.v2.i5.86.
[42] P. Gamble, R. Jaroensri, H. Wang, F. Tan, M. Moran, et al., Determining breast cancer biomarker status and associated morphological features using deep
learning, Commun. Med. 1 (1) (2021) 14, https://doi.org/10.1038/s43856-021-00013-3.
[43] J. Gao, A. Ulvik, A. McCann, P.M. Ueland, K. Meyer, Microheterogeneity and preanalytical stability of protein biomarkers of inflammation and renal function,
Talanta 223 (2021) 121774, https://doi.org/10.1016/j.talanta.2020.121774.
[44] S. Giulianelli, C.A. Lamb, C. Lanari, Progesterone receptors in normal breast development and breast cancer, Essays Biochem. 65 (6) (2021) 951–969, https://
doi.org/10.1042/EBC20200163.
[45] N. Gorbokon, P. Timm, D. Dum, A. Menz, F. Büscheck, et al., Mammaglobin-a expression is highly specific for tumors derived from the breast, the female
genital tract, and the salivary gland, Diagnostics 13 (6) (2023) 1202, https://doi.org/10.3390/diagnostics13061202.
[46] T. Hanamura, J.L. Christenson, K.I. O’Neill, E. Rosas, N.S. Spoelstra, M.M. Williams, J.K Richer, Secreted indicators of androgen receptor activity in breast
cancer pre-clinical models, Breast Cancer Res. 23 (1) (2021) 102, https://doi.org/10.1186/s13058-021-01478-9.
[47] Y.J. Heo, C. Hwa, G.H. Lee, J.M. Park, J.Y. An, Integrative multi-omics approaches in cancer research: from biological networks to clinical subtypes, Mol. Cells
44 (7) (2021) 433–443, https://doi.org/10.14348/molcells.2021.0042.
[48] J.X. Hing, C.W. Mok, P.T. Tan, S.S. Sudhakar, C.M. Seah, W.P. Lee, S.M. Tan, Clinical utility of tumour marker velocity of cancer antigen 15–3 (CA 15–3) and
carcinoembryonic antigen (CEA) in breast cancer surveillance, The Breast 52 (2020) 95–101, https://doi.org/10.1016/j.breast.2020.05.005.
[49] N.S. Holah, A.S. Hemida, Cyclin D1 and PSA act as good prognostic and clinicopathological indicators for breast cancer, J. Immun. Immunochem. 41 (1)
(2020) 28–44, https://doi.org/10.1080/15321819.2019.1677706.
[50] Z. Honarvar, B. Kalantari Khandani, M. Nazari, F. Karami Robati, Comparison of serum human epididymis protein 4 (HE4) levels in breast cancer patients and
healthy individuals, Indian J. Gynecol. Oncol. 19 (4) (2021) 1–5, https://doi.org/10.1007/s40944-021-00551-7.
[51] K.B. Horwitz, C.A. Sartorius, 90 YEARS OF PROGESTERONE: progesterone and progesterone receptors in breast cancer: past, present, future, J. Mol.
Endocrinol. 65 (1) (2020) T49–T63, https://doi.org/10.1530/JME-20-0104.
[52] H. Huebner, L. Häberle, V. Müller, I. Schrader, R. Lorenz, et al., MUC1 (CA27.29) before and after chemotherapy and prognosis in high-risk early breast cancer
patients, Cancers. 14 (7) (2022) 1721, https://doi.org/10.3390/cancers14071721.
[53] K.T. Hwang, Y.A. Kim, J. Kim, J.H. Park, I.S. Choi, K.R. Hwang, Y.J. Chai, J.H. Park, Influence of androgen receptor on the prognosis of breast cancer, J. Clin.
Med. 9 (4) (2020) 1083, https://doi.org/10.3390/jcm9041083.
[54] I. Isewon, E. Alagbe, S. Rotimi, J. Oyelade, A multi-omics classifier for prediction of androgen deprivation treatment response in prostate cancer patients, in:
2022 IEEE international conference on bioinformatics and biomedicine (BIBM), IEEE, Las Vegas, NV, USA, 2022, pp. 749–752, https://doi.org/10.1109/
BIBM55620.2022.9995633, 6 December 2022.
[55] R.M. Issac, P. Saldanha, J.M. Mathai, R. Mathews, B. Kumari, T. Chacko, Potential role of BRCA1 protein expression as a prognostic tissue biomarker in breast
carcinoma: an immunohistochemical and clinicopathologic study from South India, Oncol. Clin. Pract. 17 (5) (2021) 205–211, https://doi.org/10.5603/
OCP.2021.0031.
[56] A. Jafari, M. Farahani, M. Abdollahpour-Alitappeh, A. Manzari-Tavakoli, M. Yazdani, M. Rezaei-Tavirani, Unveiling diagnostic and therapeutic strategies for
cervical cancer: biomarker discovery through proteomics approaches and exploring the role of cervical cancer stem cells, Front. Oncol. 13 (2024) 1277772,
https://doi.org/10.3389/fonc.2023.1277772.
[57] S.A. Jeffreys, T.M. Becker, S. Khan, P. Soon, H. Neubauer, P. De Souza, B. Powter, Prognostic and predictive value of CCND1/cyclin D1 amplification in breast
cancer with a focus on postmenopausal patients: a systematic review and meta-analysis, Front. Endocrinol. 13 (2022) 895729, https://doi.org/10.3389/
fendo.2022.895729.
[58] S. Jeong, M.J. Park, W. Song, H.S. Kim, Current immunoassay methods and their applications to clinically used biomarkers of breast cancer, Clin. Biochem. 78
(2020) 43–57, https://doi.org/10.1016/j.clinbiochem.2020.01.009.
[59] W. Jiang, X.L. Xing, C. Zhang, L. Yi, W. Xu, J. Ou, N. Zhu, MET and FASN as prognostic biomarkers of triple negative breast cancer: a systematic evidence
landscape of clinical study, Front. Oncol. 11 (2021), https://doi.org/10.3389/fonc.2021.604801.
[60] X. Jiao, C. Xu, L. Tian, Z. Zhang, A.W. Ashton, Z. Li, R.G. Pestell, Abstract P3-10-03: enrichment of pro-oncogenic immune miRNAs in exosomes by cyclin D1,
promote cancer stem cell expansion, Cancer Res. 82 (4_Supplement) (2022) P3–10, https://doi.org/10.1158/1538-7445.SABCS21-P3-10-03.
13
[61] T.Y. Jin, K.S. Park, S.E. Nam, Y.B. Yoo, W.S. Park, I.J. Yun, BRCA1/2 serves as a biomarker for poor prognosis in breast carcinoma, Int. J. Mol. Sci. 23 (7)
(2022) 3754, https://doi.org/10.3390/ijms23073754.
[62] A. Joshi, G.K., A. Vishnu, T. Sakorikar, A.M. Kamal, J.S. Vaidya, H.J Pandya, Recent advances in biosensing approaches for point-of-care breast cancer
diagnostics: challenges and future prospects, Nanoscale Adv. 3 (19) (2021) 5542–5564, https://doi.org/10.1039/D1NA00453K.
[63] V. Kandi, S. Vadakedath, Clinical trials and clinical research: a comprehensive review, Cureus. (2023), https://doi.org/10.7759/cureus.35077.
[64] A. Kawiak, A. Kostecka, Regulation of Bcl-2 family proteins in estrogen receptor-positive breast cancer and their implications in endocrine therapy, Cancers.
(Basel) 14 (2) (2022) 279, https://doi.org/10.3390/cancers14020279.
[65] H. Kreipe, N. Harbeck, M. Christgen, Clinical validity and clinical utility of Ki67 in early breast cancer, Ther. Adv. Med. Oncol. 14 (2022) 175883592211227,
https://doi.org/10.1177/17588359221122725.
[66] Y.W. Kwon, H.S. Jo, S. Bae, Y. Seo, P. Song, M. Song, J.H. Yoon, Application of proteomics in cancer: recent trends and approaches for biomarkers discovery,
Front. Med. 8 (2021) 747333, https://doi.org/10.3389/fmed.2021.747333.
[67] Lakhani, J. & Harwani, D. (2021) In silico analysis and cluster validation of potential breast cancer nsSNPs of serological tumor marker CA27.29. doi:10.1101/2021.
01.19.427371.
[68] J. Lambert, A novel method of digital ELISA using single molecule amplification enables sub-attomolar detection limits for blood-based proteomics and
biomarker measurements, J. Immunol. 210 (1_Supplement) (2023) 251.13, https://doi.org/10.4049/jimmunol.210.Supp.251.13. -251.13.
[69] A.G. Lashen, M.S. Toss, N.P. Mongan, A.R. Green, E.A. Rakha, The clinical value of progesterone receptor expression in luminal breast cancer: a study of a large
cohort with long-term follow-up, Cancer 129 (8) (2023) 1183–1194, https://doi.org/10.1002/cncr.34655.
[70] E.G. Lee, D.E. Lee, J.H. Han, S. Lee, H.S. Kang, E.S. Lee, Y. Kwon, H.H. Kim, H.J. Chae, S.H. Sim, K.S. Lee, S.Y. Jung, Abstract P1-08-23: androgen receptor as
predictive marker for pathologic complete response in breast cancer with neoadjuvant chemotherapy, Cancer Res. 82 (4_Supplement) (2022) P1-08-23,
https://doi.org/10.1158/1538-7445.SABCS21-P1-08-23. -P1-08-23.
[71] J. Lengfeld, H. Zhang, S. Stoesz, R. Murali, F. Pass, M.I. Greene, P.N. Goel, P. Grover, Challenges in detection of serum oncoprotein: relevance to breast cancer
diagnostics, Breast Cancer 13 (2021) 575–593, https://doi.org/10.2147/BCTT.S331844.
[72] J. Li, X. Guan, Z. Fan, L.M. Ching, Y. Li, X. Wang, W.M. Cao, D.X. Liu, Non-invasive biomarkers for early detection of breast cancer, Cancers. 12 (10) (2020)
2767, https://doi.org/10.3390/cancers12102767.
[73] Y. Li, S. Jia, Y. Yao, Y. Zhou, S. Li, J. Li, Y. Liu, The novel role of cytomorphology from ultrasound-guided fine needle aspiration cytology in evaluating the
status of prognostic factors including estrogen receptor, progesterone receptor and HER2 in breast cancer G. Tuccari (ed.), Analyt. Cell. Pathol. 2022 (2022)
1–8, https://doi.org/10.1155/2022/6302751.
[74] Z. Li, Q. Wang, J. Mao, L. Zhang, W. Zhang, M. Ye, Selective enrichment of N-terminal proline peptides via hydrazide chemistry for proteomics analysis, Anal.
Chim. Acta 1142 (2021) 48–55, https://doi.org/10.1016/j.aca.2020.10.042.
[75] Z. Li, H. Wei, S. Li, P. Wu, X. Mao, The role of progesterone receptors in breast cancer. Drug design, development and therapy, Volume 16 (2022) 305–314,
https://doi.org/10.2147/DDDT.S336643.
[76] H. Liu, L. Ma, C. Li, B. Cao, Y. Jiang, L. Han, R. Xu, J. Lin, D. Zhang, The molecular mechanism of chronic stress affecting the occurrence and development of
breast cancer and potential drug therapy, Transl. Oncol. 15 (1) (2022) 101281, https://doi.org/10.1016/j.tranon.2021.101281.
[77] R. Maltoni, W. Balzi, T. Rossi, F. Fabbri, S. Bravaccini, M.T. Montella, I. Massa, L. Bertoni, F. Falcini, M. Altini, Appropriateness and economic analysis of
conventional circulating biomarkers assessment in early breast cancer: a real-world experience from the E.Pic.A study, Curr. Oncol. 29 (2) (2022) 433–438,
https://doi.org/10.3390/curroncol29020039.
[78] B. Milosevic, B. Stojanovic, A. Cvetkovic, I. Jovanovic, M. Spasic, M.D. Stojanovic, V. Stankovic, M. Sekulic, B.S. Stojanovic, N. Zdravkovic, M. Mitrovic,
J. Stojanovic, D. Laketic, M. Vulovic, D. Cvetkovic, The enigma of mammaglobin: redefining the biomarker paradigm in breast carcinoma, Int. J. Mol. Sci. 24
(17) (2023) 13407, https://doi.org/10.3390/ijms241713407.
[79] N. Mirmohseni Namini, A. Abdollahi, M. Movahedi, A. Emami Razavi, R. Saghiri, HE4, A new potential tumor marker for early diagnosis and predicting of
breast cancer progression, Iran. J. Pathol. 16 (3) (2021) 284–296, https://doi.org/10.30699/ijp.2021.135323.2482.
[80] M.A. Mohtar, S.E. Syafruddin, S.N. Nasir, T.Y. Low, Revisiting the roles of pro-metastatic EpCAM in cancer, Biomolecules. 10 (2) (2020) 255, https://doi.org/
10.3390/biom10020255.
[81] H. Moses, M.H. Barcellos-Hoff, TGF- biology in mammary development and breast cancer, Cold. Spring. Harb. Perspect. Biol. 3 (1) (2011) a003277, https://
doi.org/10.1101/cshperspect.a003277. –a003277.
[82] G. Moustakas, S. Kampantais, A. Nikolaidou, I. Vakalopoulos, V. Tzioufa, G. Dimitriadis, HER-2 overexpression is a negative predictive factor for recurrence in
patients with non-muscle-invasive bladder cancer on intravesical therapy, J. Internat. Med. Res. 48 (1) (2020) 030006051989584, https://doi.org/10.1177/
0300060519895847.
[83] H. Muley, K. Valencia, J. Casas, B. Moreno, L. Botella, F. Lecanda, R. Fadó, N. Casals, Cpt1c downregulation causes plasma membrane remodelling and
anthracycline resistance in breast cancer, Int. J. Mol. Sci. 24 (2) (2023) 946, https://doi.org/10.3390/ijms24020946.
[84] F. Mundt, N.J.W. Albrechtsen, S.P. Mann, P. Treit, M. Ghodgaonkar-Steger, M. O’Flaherty, R. Raijmakers, J.A. Vizcaíno, A.J.R. Heck, M. Mann, Foresight in
clinical proteomics: current status, ethical considerations, and future perspectives, Open. Res. Eur. 3 (2023) 59, https://doi.org/10.12688/
openreseurope.15810.2.
[85] S. Na, E. Paek, Computational methods in mass spectrometry-based structural proteomics for studying protein structure, dynamics, and interactions, Comput.
Struct. Biotechnol. J. 18 (2020) 1391–1402, https://doi.org/10.1016/j.csbj.2020.06.002.
[86] Nan-Li, Z.D. Zhang, R.R. Li, J.Y. Chen, H.X. Huang, Y.W. Cheng, L.Y. Xu, E.M Li, The post-translational modification of Fascin: impact on cell biology and its
associations with inhibiting tumor metastasis, Amino Acids. 54 (12) (2022) 1541–1552, https://doi.org/10.1007/s00726-022-03193-x.
[87] A.N. Neagu, M. Jayathirtha, E. Baxter, M. Donnelly, B.A. Petre, C.C. Darie, Applications of tandem mass spectrometry (MS/MS) in protein analysis for
biomedical research, Molecules. 27 (8) (2022) 2411, https://doi.org/10.3390/molecules27082411.
[88] A.N. Neagu, D. Whitham, E. Buonanno, A. Jenkins, T. Alexa-Stratulat, B.I. Tamba, C.C. Darie, Proteomics and its applications in breast cancer, Am. J. Cancer
Res. 11 (9) (2021) 4006–4049.
[89] M.N. Nguyen, S. Akter, H. Akhter, S. Ansary, S. Han, Y. Shin, J. Ha, I. Kang, S.S. Kim, T.G. Choi, TOR1B: a predictor of bone metastasis in breast cancer
patients, Sci. Rep. 13 (1) (2023) 1495, https://doi.org/10.1038/s41598-023-28140-y.
[90] T.O. Nielsen, S.C.Y. Leung, D.L. Rimm, A. Dodson, B. Acs, et al., Assessment of Ki67 in breast cancer: updated recommendations from the international Ki67 in
breast cancer working group, JNCI 113 (7) (2021) 808–819, https://doi.org/10.1093/jnci/djaa201.
[91] Z. Noor, S.B. Ahn, M.S. Baker, S. Ranganathan, A. Mohamedali, Mass spectrometry–based protein identification in proteomics—A review, Brief. Bioinformatics
22 (2) (2021) 1620–1638, https://doi.org/10.1093/bib/bbz163.
[92] L.S. Normann, M.H. Haugen, V. Hongisto, M.R. Aure, S.K. Leivonen, V.N. Kristensen, A. Tahiri, O. Engebraaten, K.K. Sahlberg, G.M. Mælandsmo, High-
throughput screen in vitro identifies dasatinib as a candidate for combinatorial treatment with HER2-targeting drugs in breast cancer P.G. Petronini (ed.),
PLoS. One 18 (1) (2023) e0280507, https://doi.org/10.1371/journal.pone.0280507.
[93] Á. Ősz, A. Lánczky, B. Győrffy, Survival analysis in breast cancer using proteomic data from four independent datasets, Sci. Rep. 11 (1) (2021) 16787, https://
doi.org/10.1038/s41598-021-96340-5.
[94] J.H. Park, H.S. Han, S.D. Lim, W.Y. Kim, K.S. Park, Y.B. Yoo, S.E. Lee, W.S. Kim, Fatty acid synthetase expression in triple-negative breast cancer, J. Pathol.
Transl. Med. 56 (2) (2022) 73–80, https://doi.org/10.4132/jptm.2021.10.27.
[95] W.S. Phipps, M.R. Kilgore, J.J. Kennedy, J.R. Whiteaker, A.N. Hoofnagle, A.G. Paulovich, Clinical Proteomics for Solid Organ Tissues, Mole. Cell. Proteom. 22
(11) (2023) 100648, https://doi.org/10.1016/j.mcpro.2023.100648.
[96] S. Radenkovic, G. Konjevic, M. Nikitovic, S. Stojanovic Rundic, V. Plesinac Karapandzic, M. Milovic Kovacevic, V. Jurisic, Evaluation of Cyclin D1 expression
by western blotting methods and immunohistochemistry in breast cancer patients, J. B.U.ON. 26 (2) (2021) 475–482.
14
[97] T. Rajkumar, S. Amritha, V. Sridevi, G. Gopal, K. Sabitha, S. Shirley, R. Swaminathan, Identification and validation of plasma biomarkers for diagnosis of
breast cancer in South Asian women, Sci. Rep. 12 (1) (2022) 100, https://doi.org/10.1038/s41598-021-04176-w.
[98] L. Ramalhete, E. Vigia, R. Araújo, H.P. Marques, Proteomics-driven biomarkers in pancreatic cancer, Proteomes. 11 (3) (2023) 24, https://doi.org/10.3390/
proteomes11030024.
[99] T. Reinert, F. Cascelli, C.A.A.D. Resende, A.C. Gonçalves, V.S.P. Godo, C.H. Barrios, Clinical implication of low estrogen receptor (ER-low) expression in breast
cancer, Front. Endocrinol. 13 (2022) 1015388, https://doi.org/10.3389/fendo.2022.1015388.
[100] E. Revuelta-López, J. Barallat, A. Cserkóová, C. Gálvez-Montón, A.S. Jaffe, J.L. Januzzi, A. Bayes-Genis, Pre-analytical considerations in biomarker research:
focus on cardiovascular disease, Clin. Chem. Lab. Med. (CCLM) 59 (11) (2021) 1747–1760, https://doi.org/10.1515/cclm-2021-0377.
[101] N. Riaz, R. Idress, S. Habib, E.N. Lalani, Lack of androgen receptor expression selects for basal-like phenotype and is a predictor of poor clinical outcome in
non-metastatic triple negative breast cancer, Front. Oncol. 10 (2020) 1083, https://doi.org/10.3389/fonc.2020.01083.
[102] A. Root, P. Allen, P. Tempst, K. Yu, Protein biomarkers for early detection of pancreatic ductal adenocarcinoma: progress and challenges, Cancers. 10 (3)
(2018) 67, https://doi.org/10.3390/cancers10030067.
[103] R.J. Rotello, T.D. Veenstra, Mass spectrometry techniques: principles and practices for quantitative proteomics, Curr. Protein Pept. Sci. 22 (2) (2021) 121–133,
https://doi.org/10.2174/1389203721666200921153513.
[104] S.O. Rotimi, O.A. Rotimi, B. Salhia, A review of cancer genetics and genomics studies in Africa, Front. Oncol. 10 (2021) 606400, https://doi.org/10.3389/
fonc.2020.606400.
[105] C. Russo, L. Wyld, M. Da Costa Aubreu, C.S. Bury, C. Heaton, L.M. Cole, S. Francese, Non-invasive screening of breast cancer from fingertip smears—A proof of
concept study, Sci. Rep. 13 (1) (2023) 1868, https://doi.org/10.1038/s41598-023-29036-7.
[106] K.S.S. Sai Baba, M. Rehman, J. Pradeep Kumar, M. Fatima, G.S.N. Raju, S. Uppin, N. Mohammed, Serum human epididymis protein-4 (HE4) - a novel approach
to differentiate malignant frombenign breast tumors, Asian Pacific J. Cancer Prevent. 22 (8) (2021) 2507–2509, https://doi.org/10.31557/
APJCP.2021.22.8.2509.
[107] J. Schick, R.P. Ritchie, C. Restini, Breast Cancer Therapeutics and Biomarkers: past, Present, and Future Approaches, Breast Cancer 15 (2021)
117822342199585, https://doi.org/10.1177/1178223421995854.
[108] A. Schumacher-Schuh, A. Bieger, W.V. Borelli, M.K. Portley, P.S. Awad, S. Bandres-Ciga, Advances in proteomic and metabolomic profiling of
neurodegenerative diseases, Front. Neurol. 12 (2022) 792227, https://doi.org/10.3389/fneur.2021.792227.
[109] A.K. Siraj, S.K. Parvathareddy, P. Annaiyappanaidu, S.O. Ahmed, N. Siraj, A. Tulbah, F. Al-Dayel, D. Ajarim, K.S. Al-Kuraya, High expression of cyclin D1 is an
independent marker for favorable prognosis in middle eastern breast cancer, Onco Targets. Ther. 14 (2021) 3309–3318, https://doi.org/10.2147/OTT.
S309091.
[110] M.P. Sousa, A.M.L. Piloto, A.C. Pereira, F.C. Schmitt, R. Fernandes, F.T.C. Moreira, New quantum-dot-based fluorescent immunosensor for cancer biomarker
detection, Chemosensors 10 (12) (2022) 518, https://doi.org/10.3390/chemosensors10120518.
[111] R.C. Stein, A. Marshall, J. Bayani, A. Makris, I.R. MacPherson, L. Hughes-Davies, M. Sobol, T. Piper, G. Dotchin, H. Higgins, A. Shaaban, S.E. Pinder, J. Dunn,
J. Bartlett, OPTIMA Trial Management Group, Disparity between Ki67 measurements and tumor gene expression tests in patients with hormone-sensitive early
breast cancer from the OPTIMA preliminary trial, J. Clin. Oncol. 40 (16_suppl) (2022) 567, https://doi.org/10.1200/JCO.2022.40.16_suppl.567. –567.
[112] M. Su, Z. Zhang, L. Zhou, C. Han, C. Huang, E.C. Nice, Proteomics, personalized medicine and cancer, Cancers. (Basel) 13 (11) (2021) 2512, https://doi.org/
10.3390/cancers13112512.
[113] P. Subramanian, J.J. Jayapalan, P.S. Abdul-Rahman, Cancer proteomics: new horizons and insights into therapeutic applications, Curr. Proteomics. 18 (3)
(2021) 266–278, https://doi.org/10.2174/1570164617666200814153949.
[114] R. Sule, G. Rivera, A.V. Gomes, Western blotting (immunoblotting): history, theory, uses, protocol and problems, Biotechniques 75 (3) (2023) 99–114, https://
doi.org/10.2144/btn-2022-0034.
[115] S. Sundarraj, G. Rajagopal, B. Sundaramahalingam, M. Sundar, R. Thangam, Methods of protein detection in cancer for diagnosis, prognosis and therapy, in:
Y. Tutar, L. Tutar (Eds.), Biochemistry, IntechOpen, 2022, https://doi.org/10.5772/intechopen.101050.
[116] S.M. Swain, M. Shastry, E. Hamilton, Targeting HER2-positive breast cancer: advances and future directions, Nat. Rev. Drug Disc. 22 (2) (2023) 101–126,
https://doi.org/10.1038/s41573-022-00579-0.
[117] I.M. Talaat, M.Y. Hachim, I.Y. Hachim, R.A.E.R. Ibrahim, M.A.E.R. Ahmed, H.Y Tayel, Bone marrow mammaglobin-1 (SCGB2A2) immunohistochemistry
expression as a breast cancer specific marker for early detection of bone marrow micrometastases, Sci. Rep. 10 (1) (2020) 13061, https://doi.org/10.1038/
s41598-020-70012-2.
[118] F. Tian, S. Zhang, C. Liu, Z. Han, Y. Liu, J. Deng, Y. Li, X. Wu, L. Cai, L. Qin, Q. Chen, Y. Yuan, Y. Liu, Y. Cong, B. Ding, Z. Jiang, J. Sun, Protein analysis of
extracellular vesicles to monitor and predict therapeutic response in metastatic breast cancer, Nat. Commun. 12 (1) (2021) 2536, https://doi.org/10.1038/
s41467-021-22913-7.
[119] I.M.W. Verberk, E.O. Misdorp, J.M. Koelewijn, D. Shan, C. Lambrechts, L.M. Shaw, R.A. Rissman, K. Blennow, H. Zetterberg, A.J. Ball, R.M. Edelmayer, C.
E. Teunissen, A biorepository for the in-depth validation of pre-analytical sample handling effects on novel blood-based biomarkers for Alzheimer’s disease: the
first results: biomarkers (non-neuroimaging): method development and validation, Alzheimer’s Dementia 16 (S5) (2020) e045763, https://doi.org/10.1002/
alz.045763.
[120] F. Viehweger, L.M. Tinger, D. Dum, N. Gorbokon, A. Menz, et al., Diagnostic and prognostic impact of progesterone receptor immunohistochemistry: a study
evaluating more than 16,000 Tumors C. Ricciardelli (ed.), Analyt. Cell. Pathol. 2022 (2022) 1–15, https://doi.org/10.1155/2022/6412148.
[121] A. Vorobyeva, E. Bezverkhniaia, E. Konovalova, A. Schulga, J. Garousi, O. Vorontsova, A. Abouzayed, A. Orlova, S. Deyev, V. Tolmachev, Radionuclide
molecular imaging of EpCAM expression in triple-negative breast cancer using the scaffold protein DARPin Ec1, Molecules. 25 (20) (2020) 4719, https://doi.
org/10.3390/molecules25204719.
[122] I.A. Voutsadakis, Molecular alterations and putative therapeutic targeting of planar cell polarity proteins in breast cancer, J. Clin. Med. 12 (2) (2023) 411,
https://doi.org/10.3390/jcm12020411.
[123] N. Wang, L. Zhang, Q. Ying, Z. Song, A. Lu, A. Treumann, Z. Liu, T. Sun, Z. Ding, A reverse phase protein array based phospho-antibody characterization
approach and its applicability for clinical derived tissue specimens, Sci. Rep. 12 (1) (2022) 22373, https://doi.org/10.1038/s41598-022-26715-9.
[124] L. Wilkinson, T. Gathani, Understanding breast cancer as a global health concern, Br. J. Radiol. 95 (1130) (2022) 20211033, https://doi.org/10.1259/
bjr.20211033.
[125] H.J. Wu, P.Y. Chu, Recent discoveries of macromolecule- and cell-based biomarkers and therapeutic implications in breast cancer, Int. J. Mol. Sci. 22 (2)
(2021) 636, https://doi.org/10.3390/ijms22020636.
[126] Y. Wu, L. Li, D. Zhang, F. Ma, Prognostic value of the serum HER2 extracellular domain level in breast cancer: a systematic review and meta-analysis, Cancers.
14 (19) (2022) 4551, https://doi.org/10.3390/cancers14194551.
[127] S. Xia, Q. Lin, Estrogen receptor bio-activities determine clinical endocrine treatment options in estrogen receptor-positive breast cancer, Technol. Cancer Res.
Treat. 21 (2022) 153303382210903, https://doi.org/10.1177/15330338221090351.
[128] T. Yamada, Quantification of biomarker proteins using reverse-phase protein arrays, PROTEOMICS – Clin. Applicat. 14 (4) (2020) 1900120, https://doi.org/
10.1002/prca.201900120.
[129] H.P. Yao, X.M. Tong, M.H. Wang, Oncogenic mechanism-based pharmaceutical validation of therapeutics targeting MET receptor tyrosine kinase, Ther. Adv.
Med. Oncol. 13 (2021) 175883592110069, https://doi.org/10.1177/17588359211006957.
[130] C.P. You, M.H. Leung, W.C. Tsang, U.S. Khoo, H. Tsoi, Androgen receptor as an emerging feasible biomarker for breast cancer, Biomolecules. 12 (1) (2022) 72,
https://doi.org/10.3390/biom12010072.
[131] C.P. You, H. Tsoi, E.P.S. Man, M.H. Leung, U.S. Khoo, Modulating the activity of androgen receptor for treating breast cancer, Int. J. Mol. Sci. 23 (23) (2022)
15342, https://doi.org/10.3390/ijms232315342.
15
[132] A. Zhang, X. Wang, C. Fan, X. Mao, The role of Ki67 in evaluating neoadjuvant endocrine therapy of hormone receptor-positive breast cancer, Front.
Endocrinol. 12 (2021) 687244, https://doi.org/10.3389/fendo.2021.687244.
[133] D. Zhang, L. Yang, X. Liu, J. Gao, T. Liu, Q. Yan, X. Yang, Hypoxia modulates stem cell properties and induces EMT through N -glycosylation of EpCAM in
breast cancer cells, J. Cell. Physiol. 235 (4) (2020) 3626–3633, https://doi.org/10.1002/jcp.29252.
[134] H. Zhang, Y. Peng, Current biological, pathological and clinical landscape of HER2-low breast cancer, Cancers 15 (1) (2022) 126, https://doi.org/10.3390/
cancers15010126.
[135] M. Zubair, S. Wang, N. Ali, Advanced approaches to breast cancer classification and diagnosis, Front. Pharmacol. 11 (2021) 632079, https://doi.org/10.3389/
fphar.2020.632079.
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Scientific African 25 (2024) e02308

Contents lists available at ScienceDirect

Protein biomarkers for diagnosis of breast cancer

The current landscape of protein biomarkers

CA 27.29 Serum-based Prognosis, Treatment response Approved [52]

Human epidermal growth factor receptor 2

Proliferation index (Ki67)

Fatty acid synthase

BRCA1 and BRCA2

Epithelial cell adhesion molecule

role of cyclin D1 expression as a prognostic biomarker in breast cancer.

Human epididymis protein 4

Advances in protein biomarkers for point-of-care breast cancer diagnosis

Proteomics techniques for protein biomarker detection

Enzyme-Linked immunosorbent assays

Reverse phase protein array

Scientific African 25 (2024) e02308

Challenges in protein biomarker detection and mitigation strategies

Clinical translation of protein biomarkers

Future direction and conclusion

CRediT authorship contribution statement

Declaration of competing interest

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