The role of microtubules in cancer is well established and inhibitors of microtubule growth exhibit excellent anticancer properties [18, 19]. One of the earliest studies on the effects of eribulin on purified microtubules as well as in living cells showed that it inhibits microtubule growth by an end-poisoning mechanism [14]. This is in contrast to other anti-mitotic drugs such as vinblastine and paclitaxel that diminish the shortening and growth phases of microtubule dynamic instability. Eribulin does not cause the shortening of tubulin but it turns them into aggregates as observed through electron microscopy [14]. Eribulin significantly suppressed centromere dynamics in mitotic spindles at drug concentrations that arrested mitosis [15]. Results from binding studies on soluble tubulin suggested that eribulin had the ability to bind with high affinity to microtubule plus ends thereby it suppressed the dynamic instability [17]. Later studies involving X-ray crystallography eribulin was shown to bind to a site on beta-tubulin that is required for proto filament plus end elongation. In addition, single eribulin molecule was found to specifically interact with microtubule plus ends and is sufficient enough to either trigger a catastrophe or induce erratic microtubule growth [20]. Eribulin caused irreversible mitotic arrest unlike other microtubule inhibitors that leads to consistent inactivation of Bcl-2 and ended up with apoptosis [21, 22]. Molecular consequences of microtubule inhibition due to eribulin treatment are summarized in the Figure (Fig. 2).

Fig. 2.

Effects of microtubule inhibition due to eribulin treatment.

Recent evidences show that eribulin not only acts on the cancer cells but also shows its effects on the entire Tumor microenvironment (TME) [23, 24]. TME includes the entire cellular environment containing blood vessels, immune cells such as T cells, natural killer (NK) cells, activated fibroblasts and endothelial and mesenchymal cells along with the tumor cells [25, 26, 27]. The events orchestrated by all of these components of TME play a major role in tumor progression. Evidences indicate that effectiveness of the therapy depends on the capability of the anticancer agent to reduce the abnormalities in the TME [28]. Some of the alterations in the tumor environment due to eribulin treatment as observed until recently are discussed below. The different processes that are altered/ activated in the TME due to eribulin treatment is summarized in Fig. 3.

Fig. 3.

Effects of eribulin treatment on Tumor microenvironment (TME).

Besides the different mechanisms mentioned already, eribulin is shown to have specific inhibitory activity against TERT-RdRP as well [63]. TERT-RdRP is responsible for mitotic progression through the elevation of heterochromatin assembly and the maintenance of stem-cell property [64]. Eribulin-sensitive ovarian cancer cell lines expressed high levels of human TERT, and inhibition of the TERT expression reduced the eribulin sensitivity. Thus, suppression of the cancer cell growth by eribulin treatment was shown to be dependent on the TERT status [63]. The human TERT is shown to localize into mitotic spindles and centromeres during mitosis [65], Therefore it has been hypothesized that eribulin inhibited human TERT functions by interfering with the interaction between TERT and microtubules [63]. Subcutaneous U87MG brain tumor xenografts treated with eribulin was observed to have a decreased TERT-RdRP activity in a dose-dependent manner [66].

Several in vitro studies have accelerated the efforts to undertake the clinical use of Eribulin (Table 1, Ref. [6, 14, 21, 67, 68]). Eribulin treatment with wide varieties of human cancer cell lines such as breast, prostate, melanoma, colon, lung, ovaries, histiocytic lymphoma, pharyngeal squamous cell carcinoma and uterine sarcoma cell lines resulted in effective inhibition of cell growth even at very low concentrations (IC${}_{50}$ values in the range of 0.09–9.5 nM) [21, 69]. Anticancer activities of eribulin at sub nM range against several human STS cell lines such as, Ewing’s sarcoma, synovial sarcoma fibrosarcoma, leiomyosarcoma, liposarcoma and promyelocytic leukemia cell lines were reported [42]. Childhood cancer preclinical models were studied during Pediatric Preclinical Testing Program (PPTP) [70] in which eribulin demonstrated in vitro cytotoxic activity, with a median relative IC${}_{50}$ value of 0.27 nM [71]. Studies also showed that eribulin could inhibit the growth of the glioblastoma cell lines in vitro [68, 71].

Eribulin treatment with human tumor xenograft models originated from breast, ovary, colon, lung, melanoma, pancreatic, and fibrosarcoma resulted in increased survival rates, tumor growth delays and reductions in size at dose levels below the maximum tolerated dose [6]. Eribulin showed in vivo anticancer efficacy in MDA-MB-435, COLO 205, and LOX xenograft models at much lower doses when compared with paclitaxel [6]. It has been proven that intermittent dosing was less toxic and more effective than daily dosing in preclinical models. Dose scheduling studies with eribulin on MDA-MB-435 breast, HT-1080 fibrosarcoma, U251, glioblastoma, SR-475 head and neck cancer, SK-LMS-1 leiomyosarcoma, NCI-H322M and NCI-H522 NSCLC, PANC-1 pancreatic cancer, and NCI-H82 small cell lung cancer (SCLC) models showed that maximal efficacy and minimal toxicity is achieved with moderate intermittent dosing [21]. Results from Pediatric preclinical testing program in childhood cancer models of different cancer histologies showed that eribulin exerted higher antineoplastic activity in ES xenografts than VCR [70, 71]. A latest report showed that eribulin penetrated the brain tumor tissue and prolonged the survival of mice by inhibiting the growth of glioblastoma cells transplanted subcutaneously or intracerebrally into mice [66].

Phase-1 trials are intended to identify the maximum tolerated dose, pharmacokinetics, toxicity profile and dose-limiting toxicities. First phase-1 trial of eribulin was conducted in patients with advanced solid tumors [72]. During such phase-1 trials, different dosing schedules were checked. Pharmacokinetic data showed linear, dose-proportional kinetics with a rapid distribution phase and a slower elimination phase [73]. As of now, 25 phase-1 studies are completed [74] in patients with different cancer types including advanced solid tumors, breast, ovary, head and neck, colon and bladder cancers. Eribulin has been well tolerated and has demonstrated good clinical response in breast, NSCLC, urothelial/bladder, and thyroid cancers. Febrile neutropenia, fatigue, anorexia, and peripheral neuropathy are some of the most common doses limiting toxicities reported [72, 73, 75, 76, 77, 78, 79]. The pharmacokinetic profile of eribulin showed an extensive volume of distribution, slow-to-moderate clearance, and slow elimination. Area under the plasma concentration versus time curve (AUC) and maximum plasma concentration (C max) has been reported to be directly proportional to the dose of eribulin (0.25–4 mg/m${}^{2)}$. It was also reported by several studies that metabolism and renal clearance contribute to the elimination of eribulin in a negligible way [80].

Reassuring tumor response data from the phase-1 trials led to the initiation of phase-2 studies in breast cancer, as well as in other solid tumor types. Phase-2 clinical trials gathered preliminary data about eribulin effectiveness on cancer patients compared to similar participants receiving a different treatment, Safety and adverse events were evaluated. As of now, 39 phase-2 studies on eribulin treatment are completed [74] in patients with cancer types including breast cancer, NSCLC, sarcoma, prostate, head and neck, urothelial, ovarian and pancreatic cancers [80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102]. In order to address the eribulin intolerance, a phase II, non-randomized, prospective study was conducted recently, which reportedthat a bi-weekly eribulin schedule (1.4 mg/m${}^2$ on days 1 and 15 of a 28-day cycle) is tolerable and has an almost equivalent efficacy in patients intolerant to the standard eribulin schedule [103].

Phase-3 trials of eribulin involved more number of participants, different populations and different dosages of the drug in combination with other drugs were tested to ensure safety and effectiveness. As of now, 6 phase-3 studies are completed [74] in patients with breast cancer, STS and NSCLC [10, 11, 104, 105, 106]. After getting FDA approval for marketing, two phase-4 trials (sponsored by Eisai) have been completed as of now, to obtain additional information on the safety, efficacy, and optimal use of eribulin by the study sponsor [107]. Very recent results favoring eribulin as a potent candidate for first-line treatment of patients with HER2-negative advanced breast cancer are obtained in a multicenter, prospective, post-marketing, observational study from Japan [108].

Clinical trials that gained approval for eribulin as a drug for cancer treatment are given in the table (Table 2, Ref. [10, 11, 93, 102, 104]). US Food and Drug Administration (FDA) approved eribulin in the year 2010 for the treatment of patients with metastatic breast cancer who were already pretreated with anthracycline and a taxane regimen. One of the pivotal phase-3 studies on eribulin treatment was Study 305/EMBRACE (Eisai Metastatic Breast Cancer Study Assessing Physician’s Choice vs E7389 Study). It was the first mono-therapy study of a cytotoxic agent to show increase of survival in patients with heavily pretreated metastatic breast cancer. Overall survival of eribulin (OS) was observed as 13.1 months versus treatment of physician’s choice (TPC) where the OS was 10.6 months. Survival benefit of 2.5 months was considered statistically significant. Neutropenia was the most common clinical grade 3 or 4 adverse event with eribulin (grade 3: 106 [21%] of 503 patients; grade 4: 121 [24%] of 503 patients) [10].

In another phase-3 trial (study 301), eribulin showed almost same efficacy as capecitabine while considering overall survival (OS) or progression free survival (PFS). The treatment had manageable safety profiles consistent with the known adverse effects; most adverse events were grade 1 or 2 [104]. As per the request from European Medicines Agency (EMA), pooled analysis of the above two phase-3 trials was carried out, to assess whether specific patient sub-groups, previously treated with an anthracycline and a taxane, benefited from eribulin. In this pooled analysis, serious adverse events were presented in similar magnitudes in the two groups (eribulin 21.1%; control 22.6%). Conclusively, 10.5% of patients in the eribulin group and 12% in the control group had an adverse event that led to discontinuation of the treatment and [109]. A subgroup analysis of 392 patients with human epidermal growth factor receptor 2 (HER2)-negative MBC showed a longer OS with eribulin (16.1 months) versus capecitabine (13.5 months) in this subgroup. The most frequent treatment-emergent adverse effect in the eribulin group of this study were neutropenia (53.3%), alopecia (34.8%), leukopenia (31.0%), peripheral neuropathy (pooled term, 23.9%), and anemia (21.2%) grade 3 or 4 neutropenia occurred in 43.5% of eribulin treated patients [110]. Very recent Post hoc, pooled exploratory subgroup analysis of EMBRACE and Study 301 revealed that eribulin provided a median OS benefit of 2.1 months over TPC/capecitabine in patients who received fewer prior regimens ($\le$3) for locally advanced/MBC. Although neutropenia and asthenia/fatigue rates were higher with eribulin treatment compared with control, the incidences of both were similar regardless of whether patients had $\le$3 or $>$3 prior regimens (neutropenia, 51.7% for both subgroups; asthenia/fatigue, 53.2% vs 55.1% for $\le$3 vs $>$3, respectively) [111].

Phase-3 study by Schoffski et al. [11] showed that better improvement of OS in the liposarcoma group compared to all sarcoma subtypes. The PFS was similar in both the study arms. Similar results were also observed in the MBC trial (study 301) with improvement in OS but without notable difference in the PFS (Table 2). The benefit of eribulin monotherapy based on improvement in OS led to approval by the FDA and EMA in 2016 for patients with unresectable (inoperable) or metastatic liposarcoma who had already undergone treatment with anthracyclines.

Combination therapy is a treatment modality in which rational combinations of different anticancer agents are employed [112]. Synergistic effect of eribulin and other chemotherapeutic drugs are carried out to minimize the toxicities and to improve the drug sensitivity. Eribulin makes effective chemotherapeutic combinations with several cytotoxic and targeted inhibitors from widely differing mechanistic classes. The combination of S-1 (5-FU) and eribulin exerts a synergistic effect for TNBC cell lines through MET-induction by eribulin. Such combination therapy could be a potential treatment option for TNBC [113]. Several investigations were conducted in which, eribulin was combined with either one of the cytotoxic agents (capecitabine, carboplatin, cisplatin, doxorubicin, gemcitabine) or targeted agents (bevacizumab, BKM-120, E7449, erlotinib, everolimus, lenvatinib, palbociclib) in tumor xenograft models of breast cancer, melanoma, non-small cell lung cancer (NSCLC), and ovarian cancer. Eribulin showed positive combination activity with most of the agents with more efficacy than the individual agents [114]. Synergistic effects of eribulin on CDK inhibition [115], PIK3 inhibition [116, 117] and mTOR inhibition [118, 119] in TNBC and pAKT inhibition in STS [120] were carried out with corresponding inhibitors. In all these studies, positive synergistic effects were noticed. It is indeed a powerful strategy to increase the therapeutic index of the drug combination to improve drug efficiency even at the lowest doses on the aggressive or resistant cancer types. Recently several phase-1and phase-2 clinical trials were conducted with eribulin in combination with other chemotherapeutic agents to evaluate and improve the therapeutic efficiency [76, 77, 78, 79, 80, 82, 84, 100, 101, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132].

Resistance to chemotherapeutic drugs is a major obstacle to therapeutic success [133]. A common mechanism of drug resistance in various tumors is identified as over expression of a drug efflux pump such as P-glycoprotein (Pgp)/(ABCB1) [134]. Eribulin increased the mRNA and protein expression of P-glycoprotein in LS174T cells and MDR1 promoter activity in human breast cancer MCF7 cells thereby the drug efflux transporter P-glycoprotein is activated [135].

In a study involving 7 different eribulin resistant cell lines, ATP-binding cassette subfamily B member 1 (ABCB1) and subfamily C member 11 (ABCC11) were observed to be overexpressed and had an increased gene expression as well [136]. Both were responsible for the development of eribulin resistance in breast cancer cells regardless of the molecular subtype. Thus, it was suggested that ABCB1 and ABCC11 could be used as a biomarker for predicting the eribulin response in patients with breast cancer [136]. Studies have proven that therapeutic targeting of EMT may be beneficial in treatment-resistant cancer subtypes. Vimentin and claudin-1 are important markers of EMT pathway activation which were significantly up regulated in eribulin resistant cell lines [43]. Eribulin-resistant leiomyosarcoma cell lines were investigated to understand the mechanism of resistance [137]. Eribulin resistant cell lines express higher TUBB3 levels and grow more promptly than the parent cell line. Knockdown of TUBB3 by siRNA resulted in the inhibition of cell growth. Therefore, it could be predicted that if over expression of TUBB3 is observed for a type of cancer, eribulin would be suggestively be the drug of choice for treatment and alternative treatment approaches should be considered.

Further mechanistic understanding of Pgp activation, ABCB1 and ABCC11 activation, TUBB3 over expression might lead to novel therapeutic regimen to bridle eribulin resistance in the future.

Primary cultures moderately imitate the in situ microenvironment of the diseased condition and also preserve the crosstalk between malignant and healthy constituents. Such features are to be involved in the different responses to therapies and in all stages of the natural history of malignant tumors, i.e., carcinogenesis, migration, invasion and metastatic dissemination. Therefore, ex vivo models permit a more faithful reproduction of tumors and are a valid strategy especially for clinical and preclinical analyses [147]. Recently, efforts have been made to develop methods for tumor cell culture from biopsies and expansion of patient-derived circulating tumor cells CTCs) for individualized drug susceptibility [140, 141, 148]. A recent study by De et al. [149] had established an ex vivo 3D system for culturing CTCs derived from either primary or metastatic tumors that recapitulates their physiological characteristics. This scaffold imitates the architecture and stiffness of breast tumors and allows improved cell-matrix interactions. Patient-derived organoid models are also established as they offer spatial and temporal insights into cancer biology. These models conserve the heterogeneity of the primary tumor, which makes them more appropriate for identifying therapeutic targets and corroborating drug response [150]. The anticancer activity of eribulin on the patient-derived primary culture of a liposarcoma patient was assessed its antiproliferative effect by the arrest of cell motility and initiation of apoptosis [151, 152].