Human epidermal growth factor receptor targeted inhibitors for the treatment of ovarian cancer

Introduction

Ovarian cancer is the eighth most lethal type of cancer in women worldwide, with over 184,000 fatalities reported in 2018¹. It has a poor prognosis and is usually diagnosed at late stage, due to lack of specific diagnostic biomarkers and relatively non-specific symptoms. Current diagnostic tests for ovarian cancer include the CA-125 blood test coupled with abdominal/pelvic ultrasound and computerized tomography (CT) scan. First line treatment of ovarian cancer is optimal debulking of macroscopic disease, generally followed by chemotherapy with carboplatin alone or in combination with paclitaxel^2,3.

The most common type of ovarian cancer is epithelial ovarian cancer (EOC), which is divided into 5 main subtypes, with differing histological, molecular, and genetic characteristics^3,4. The most common subtype is high grade serous ovarian cancer (HGSOC) which accounts for approximately 70% of the cases, with clear cell (10%), endometrioid (10%), mucinous (< 5%), and low grade serous cancer (< 5%) comprising the other significant subtypes^3,4. Recent investigations into the pathogenesis of ovarian cancer showed that it primarily originates from different parts of the female reproductive system and involves cellular migration to the ovaries. HGSOC is identified to originate from the distal fallopian tube, endometrioid and clear cell cancers arise from the endometrium, while low grade serous cancer might progress from serous cystadenoma and serous borderline tumors⁵.

Multiple drugs have been tested and approved for ovarian cancer although the response rate for second line therapy is only 10%–35% and different ovarian cancer subtypes respond differently to drug treatment. HGSOC patients usually respond well to initial platinum-based therapy, given their BRCA and p53 mutations⁷. However, these patients often present with resistance to initial therapy after a few months.

There is increasing interest in the potential use of targeted inhibitors for the treatment of ovarian cancer. This review seeks to overview the current clinical and preclinical status of human epidermal growth factor receptor (HER) targeted therapy in ovarian cancer, with special emphasis on tyrosine kinase inhibitors (TKIs).

Human epidermal growth factor receptors (HERs)

The HER family has been associated with the progression of several cancers including breast, lung and colon cancer⁸. In ovarian cancer, amplification, and/or high expression of epidermal growth factor receptor (EGFR), HER2 and HER3 receptors have been implicated in the progression and prognosis of the disease^9-12.

The HER family of receptors [also known as erythroblastic leukemia viral oncogene (erbB) family] are present on the cell surface as monomers, in the absence of ligand activation. There are four members in this family, EGFR (HER1/erbB1), HER2 (neu/erbB2), HER3 (erbB3) and HER4 (erbB4) (Figure 1). With the exception of HER2, ligands bind to their extracellular domain and form homo- or heterodimers with other members of the family, preferentially with HER2, since it has the most favorable kinase activity and exists in an activated form¹³. HER ligands are divided into three groups; those which bind specifically to EGFR (epidermal growth factor, amphiregulin and transforming growth factor-α), those conferring dual specificity to EGFR and HER4 (betacellulin, heparin-binding EGF, and epiregulin), and those which bind to HER3 and HER4 (neuregulins/heregulins)¹⁴.

Upon receptor dimerization, multiple downstream pathways are activated, which regulate cell proliferation, differentiation, angiogenesis, survival, and cellular metabolism amongst other functions. Heterodimerization allows for a myriad of phosphotyrosine residues to bind, which in turn increases the possibilities for signaling pathways¹⁵. These pathways (Figure 1) include the mitogen activated protein kinase (MAPK)/ERK pathway, which regulates the growth and development of cells, the signal transducer and activation of transcription (STAT) pathway, which governs cell proliferation and differentiation, the phosphoinosidyl-3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway, which regulates cell survival and metabolism, and the phospholipase Cγ (PLCγ) pathway, which controls calcium-dependent actions^13,16,17. In tumorigenesis, mutations within the components of these pathways can cause cancer cells to acquire certain aptitudes, including impartiality to proliferation signals, circumvention of apoptosis, insensitivity to growth inhibitory signals, augmented replicative potential and the capability to metastasize¹⁸.

EGFR (HER1) is a receptor tyrosine kinase comprising an extracellular ligand-binding domain of 622 residues, a 23 residue transmembrane domain, and a large, 522-residue intracellular domain¹⁵. EGFR is normally weakly expressed in the ovaries, however, several studies have found that EGFR is highly expressed in ovarian cancer. Immunohistochemical studies have indicated that 30%–70% of ovarian cancers have increased EGFR expression^19-21. High expression of EGFR is associated with poor, progression-free survival (PFS), advanced tumor grade, greater residual tumor mass and rapid proliferation^9,10,12. It has also been suggested that high EGFR expression in the tumor stroma is associated with aggressive clinical conditions and outcome and EGFR upregulation in fibroblasts is associated with growth and migratory abilities of ovarian cancer cells²².

HER2 is overexpressed in approximately 6%–30% of ovarian cancer patients^10,23 and is initially associated with DNA amplification and poor prognosis²⁴. Overexpression is often detected in the mucinous (19%)²⁵ and clear cell (14%)²⁶ subtypes. However, even some serous (3%) and endometrioid ovarian cancers (2%) have HER2 overexpression²⁷. Several studies have associated overexpression of HER2 with poor prognosis^10,28.

HER3 is expressed as a full-length receptor on the cell surface, in parallel with truncated intracellular isoforms. However, the activity of the latter is not well defined^29,30. HER3 lacks tyrosine kinase activity, hence it has to be transphosphorylated by other HER members to promote cell signaling¹⁵. HER3 is more frequently expressed in ovarian cancer (30%–80%) than EGFR and HER2³¹ and is more common amongst borderline and early-stage lesions³². Among the dimerization possibilities between these proteins, the most potent signaling complex is generated when HER3 heterodimerizes with HER2³³. Increased HER3 expression has been associated with poor clinical outcome and the average survival time for patients with low HER3 expression was 3.3 years, in contrast to 1.8 years for patients with high HER3 expression¹¹. Studies in various cancers have shown that when HER3 and MET are co-expressed, they are often associated with either response or resistance to therapy^34-36. Other studies have shown that high expression of HER3 might lead to HER3-PI3K-Akt signaling cascade in doxorubicin and cisplatin treated ovarian cancer, which often results in resistance to therapy^37,38.

HER4 is the least understood receptor of the HER family. It occurs as a spliced isoform, often being processed further by enzymes into a soluble intracellular domain, which can disperse to the cell cytoplasm or nucleus³⁹. In breast cancer, nuclear localization of the intracellular domain in combination with estrogen expression predicted worse clinical outcomes compared to membrane HER4 and estrogen⁴⁰. There are conflicting views about the expression of HER4 in ovarian cancer, with earlier reports suggesting either decreased or lack of expression of the receptor⁴¹, while more recent studies suggest an increased expression of HER4 in malignant tissues compared to normal tissues^31,42,43. Although the implication of HER4 expression in ovarian cancer is unclear, two studies found a possible correlation between HER4 expression and resistance of serous ovarian cancer to chemotherapy^42,44.

Monoclonal antibodies

HER-targeted monoclonal antibodies (mABs), such as trastuzumab (Herceptin^®) and pertuzumab (Perjeta^®) are recombinant humanized mABs, inhibiting HER2 extracellularly with differing modes of action (Figure 2). These agents have shown favorable results in HER2 positive cancers, especially HER2-positive breast cancer, where they are well established as standard therapy. More recently, trastuzumab-emtansine antibody-drug conjugate has been developed as another option for trastuzumab-resistant disease. Preclinical in vitro and in vivo studies and clinical trials have been focusing on the activity of these mABs in ovarian cancer, especially in selective subtypes, particularly mucinous cancers, which have HER2 amplification and overexpression^45,46.

TKIs

TKIs are small drug molecules that inhibit tyrosine kinases. Tyrosine kinases include the HER family, vascular endothelial growth factor receptors (VEGF), platelet-derived growth factor receptors (PDGFR), and also non-receptor tyrosine kinases BCR-ABL and KIT^68,69. Tyrosine kinases are enzymes that catalyze the transfer of phosphate from adenosine triphosphate (ATP) onto target proteins to elicit a response. There are three types of TKIs. Most small molecule TKIs are type I, which compete with ATP by binding to ATP binding sites on the active conformation of the receptors, thereby interfering with the action of tyrosine kinases⁶⁹. Type II TKIs bind to the inactive conformation of a kinase, while type III allosteric inhibitors bind to sites distant from the active site⁷⁰. To date, a few TKIs have been evaluated in ovarian cancer patients that are described below (Table 1, Figure 2). These include the first generation EGFR inhibitors, gefitinib and erlotinib, which have shown clinical efficacy against mutant EGFR lung cancer. Since, a resistance mutation develops frequently at T790M upon treatment, covalent irreversible second generation TKIs were developed. These consist of afatinib and neratinib that are active against this mutation. Other HER inhibitors were developed with broader inhibitory activity across multiple HER family members (pan-inhibitors) and these include lapatinib and canertinib as early developed inhibitors, followed by neratinib, sapitinib, and dacomitinib. Finally, multi-targeted TKIs that target the HER family among other targets (e.g., PDGFR, VEGFR, etc.) include vandetanib and leflunomide.

1.

HER-targeted TKIs evaluated in preclinical models of ovarian cancer

TKI	Chemical structure	Pharmacology	Clinical status	IC50: EGFR (nM)	IC50: HER2 (nM)	IC50: others (nM)	Reference
Afatinib (BIBW2992)		Potent and irreversible inhibitor of EGFR/HER2 including erlotinib-resistant EGFR T790 M71,72	Marketed for EGFR mutation positive lung cancer	0.5	14	-	73
Canertinib (CI-1033)		Irreversible non-selective EGFR family inhibitor, with an additional benefit of blocking mutant EGFRvIII74	Reached phase II; discontinued lately by Pfizer	1.8	11	HER4: 27	75
Dacomitinib (PF00299804)		Irreversible pan-HER inhibitor, especially EGFR	Phase III clinical trials	6.0	45.7	HER4: 73.7	75
Erlotinib (OSI-774)		Specific and reversible inhibitor of EGFR	Marketed for NCSLC and pancreatic cancer	0.5	512	HER4: 790	75
Gefitinib (ZD-1839)		Specific and reversible inhibitor of EGFR	Marketed for NCSLC	3.1	343	HER4: 476	75
Lapatinib (GW-572016)		Reversible and specific inhibitor to EGFR and HER276	Marketed for HER2 overexpressing breast cancer	10.8	9.2	HER3: 13, HER4: 367	77,78
Neratinib (HKI-272)		Potent irreversible, pan-HER (ie, HER 1, 2, and 4) TKI, with low molecular weight79	Marketed for adjuvant treatment of HER2 overexpressing breast cancer	92	59	KDR: 800, src: 1400	80
Sapitinib (AZD-8931)		Equipotent inhibitor of EGFR, HER2 and HER378	Phase II clinical trials	4	3	HER3: 4	78

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Biomarkers of sensitivity and resistance to TKIs in ovarian cancer

In breast and lung cancers, informative biomarkers of sensitivity to HER TKIs include overexpression of HER2 and mutation of EGFR. In breast cancer, HER2 overexpression is an effective biomarker of sensitivity to HER2-targeted TKIs, such as lapatinib. Preclinical studies of T-DM1 in ovarian cancers suggest that minimal expression of HER2 was essential for anti-tumorigenic properties of T-DM1 in model systems⁶⁶. Analysis of a series of ovarian cancer xenograft models demonstrated the curative potential of trastuzumab/pertuzumab combination in cancers with amplification and overexpression of HER2. Currently there is less information available regarding the association of HER2 expression levels and TKIs, in ovarian cancer^59,60. Interestingly, there is one case report of dramatic remission of a chemotherapy-resistant ovarian cancer to trastuzumab, which was HER2-negative suggesting that, the factors governing responsiveness in ovarian cancer might differ from those in breast cancer¹³⁰. The mutated form of EGFR with deletions in exon 19 indicates sensitivity to TKIs such as erlotinib and gefitinib, in NSCLC¹³¹. However, the importance of EGFR mutations in ovarian cancer is still not well researched since the occurrence of these mutations is much lower. As mentioned above, in a phase II trial of gefitinib in 27 ovarian cancer patients, the single patient showing response did contain an EGFR mutation (2235del15; E746-A750del) in the catalytic domain consistent with this molecular feature being an indicator of sensitivity⁸². This requires further validation in future studies.

The importance of mutations in HER2 and HER3 for sensitivity to pan-HER inhibitors, is under clinical investigation at present and a basket trial (SUMMIT) investigating neratinib treatment in multiple cancers with mutations, has been reported¹²⁴.

As observed with other chemotherapeutic drugs, resistance to TKIs is inevitable¹³². Mechanisms of resistance to EGFR-specific TKIs can include abnormalities in HERs, such as HER2 overexpression and mutations like EGFRvIII and HER2^L869R106,133 and secondary EGFR mutations in T790M¹³⁴, L747S, D761Y, and T854A¹³⁵. Downstream signaling pathways that are frequently modified include mutations in KRAS, BRAF, PIK3CA, and PTEN^136-138. Alternative pathways that can bypass control include aberrant activation of MET and HGF^139,140, modifications in VEGF receptors which trigger vascular permeability, in platelet-derived growth factors that regulate angiogenesis and in interleukin-6 that controls inflammatory processes^132-134. For other TKIs, overexpression of ABC resulting in low drug concentration in cells due to decreased uptake and increased efflux of the drug¹⁴¹ also contribute in resistance to therapy.

Future research

Conclusions

Ovarian cancer is a complex disease, with multiple molecular profiles. It frequently becomes resistant after initial therapy necessitating the development of new strategies.

The use of HER-targeted therapy continues to be assessed in this disease, since it might have value for selective patients and pre-clinical data supports the potential of this approach. Only a limited number of phase II trials have been completed in ovarian cancer and while response rates are low, there are frequent good percentages of stable disease. The pan-HER TKIs may have broader efficacy and utility than the early EGFR-targeted TKIs, which are dependent on the presence of mutations (are uncommon in ovarian cancer). Further biomarker studies are now required to help identify the most sensitive ovarian cancers and combination strategies require further development.

Acknowledgements

The work disclosed in this publication is partially funded by the Endeavour Scholarship Scheme (Malta). Scholarships are part-financed by the European Union–European Social Fund (ESF)–Operational Program II–Cohesion Policy 2014–2020 “Investing in human capital to create more opportunities and promote the well-being of society”.

Conflict of interest statement

No potential conflicts of interest are disclosed.