The epidermal growth factor receptors as biological targets in penile cancer

Abstract

Penile cancer is a rare disease, with an incidence that is higher in less developed countries and is in the range of 1 – 10 per 100000 men worldwide. Early diagnosis is essential for cure, as 5 year cancer-specific survival is 90 – 100 % in patients with intraepithelial neoplasms and in those with low-grade superficial tumors without lymphovascular invasion, but it drops to 30% in men with multiple mobile or bilateral inguinal lymph nodes. The EGFR family plays a major role in penile cancer biology, with distinct receptors being involved in HPV-positive and -negative tumors. A number of anti-EGFR agents were used in penile cancer patients outside the context of a clinical trial, mainly as a salvage treatment after failure of first-line chemotherapy. A total of 28 patients received anti-EGFR monoclonal antibodies, with 50% of them showing a response to treatment, and a median PFS of ∼ 3 months. The rarity of the disease poses great challenge in terms of education and awareness of the general population, planning of preventive measures on a large scale, as well as conduction of prospective trials and approval of high-cost biological therapy.

Keywords:

• cetuximab
• EGFR
• panitumumab
• penile cancer

Penile cancer is a rare disease, with an incidence that is higher in less developed countries and is in the range of 1 – 10 per 100000 men worldwide [1]. Early diagnosis is essential for cure, as 5 year cancer-specific survival is 90 – 100 % in patients with intraepithelial neoplasms and in those with low-grade superficial tumors without lymphovascular invasion, but it drops to 30% in men with multiple mobile or bilateral inguinal lymph nodes [1]. In the case of recurrent/metastatic disease, the prognosis is poor, and the majority of patients die within a year [2]. In one retrospective study in 25 patients with advanced disease, combination of cisplatin plus 5-fluorouracil yielded a radiological response and a pain improvement in approximately one-third of evaluable patients [2], with a median overall survival of 8 months. Unfortunately, the effect of chemotherapy in terms of improvement of quality of life and palliation could not be assessed in this retrospective study [2]. Given the established pathogenic role of HPV infection [1], circumcision may decrease the rate of infection and therefore play a role in primary prevention of penile cancer, whereas awareness campaigns may favor early diagnosis and have an impact on mortality in high-incidence geographical areas. Recent advances in the biology of penile cancer allowed us to identify the human epidermal growth factor receptor family as a potential target of biological therapy. Members of this family, which is composed of EGFR, HER2, HER3, and HER4 transmembrane tyrosine kinase receptors, are activated via homo- or heterodimerization induced by extracellular ligand binding, which causes phosphorylation of tyrosine residues and signaling via a number of downstream pathways including the PI3K/Akt and Ras-Raf-MEK-ERK pathway [3]. Differently from EGFR, which is overexpressed in the majority of patients, its phosphorylated form was found to be present only in 25 – 50% of cases [3,4]. In a study of 148 samples of squamous penile carcinoma by Stankiewicz et al., the phosphorylated EGFR protein was expressed in 25% of cases, with a trend towards a more frequent expression in early-stage tumors. This result may explain the different incidence of phosphorylated EGFR found in a series of 30 patients with N0 – 1 penile cancer, with ∼ 50% of them showing positivity to phosphorylated cytosolic EGFR. Similar to other experiences showing a negative impact on prognosis in several solid tumors such as lung and head and neck cancer, cytosolic phosphorylated EGFR was associated in this study to an odds ratio for recurrence of 7.6 (95% CI = 1.49 to 50; p = 0.009) and to a hazard ratio for death of 9.0 (95% CI 1.0 – 100; p = 0.01) [4]. These results indicate that EGFR activation is an early event in the natural history of penile cancer and has a detrimental impact on prognosis. In more advanced stages, EGFR may not be mainly responsible for cancer progression, hence the potential of combination of anti-EGFR and chemotherapy agents. Of note, in penile carcinoma activation of EGFR is not associated to either gene amplification [3,4], or gene copy number gain [3]. We failed to identify any of the activating mutations in the tyrosine kinase domain of EGFR known to be implicated in lung cancer, that is EGFR E746 - A750-specific deletion in exon 19 and EGFR L858R-specific point mutation in exon 21 [5]. An analysis of 150 cases of penile squamous cell carcinoma did not detect a meaningful incidence of KRAS mutations at codons 12 and 13, and of BRAF mutation at codon 600 [6]. Whole genome sequencing of these genes is mandatory, as they may still be mutated in penile cancer, but mutations may be different than those of other solid tumors. Importantly, expression of RAS-association domain family 1A (RASSF1A) was found to be absent in > 95% of samples analyzed [6], which may play a fundamental role for RAS activation, rather than RAS mutation, especially in tumors with non-phosphorylated EGFR [6]. The RASSF1A protein is expressed in all nonmalignant epithelial cells and it exerts its tumor-suppressor activity via RAS-mediated apoptosis. Promoter CpG island hypermethylation in penile cancer is the major mechanism for RASSF1A inactivation, which appears to be closely related to RAS activation in human cancers. RASSF1A was frequently inactivated in tumor types without RAS gene mutations, such as small cell lung cancer, nasopharyngeal cancer and neuroendocrine pancreatic tumor, which implicates that RASSF1A inactivation may drive or facilitate RAS activation, even in the absence of RAS mutations [6,7]. Interestingly, distinct members of the EGFR family may be involved in HPV-positive and -negative tumors [3]. In the series by Stankiewicz et al., HER3 positivity was found in 95% of HPV-infected tumors and in 75% of HPV-negative tumors. Unlike EGFR, HER3 has an impaired kinase function but it can still generate downstream signals via heterodimerization with other member of the EGFR family. The phosphorylated EGFR protein was reported significantly less frequently in HPV-positive samples with respect to HPV-negative tumors (16 vs 40%, p = 0.01). These results are consistent with the negative prognostic impact of phosphorylated EGFR discussed above, as well as with the positive prognostic effect of HPV infection, and suggest distinct tumor biology in HPV-positive and -negative tumors. Interestingly, HER2 was not hyperexpressed in any sample, suggesting the lack of any pathogenic role in penile carcinoma. These molecular targets can be inhibited by a number of available agents, offering new possibility for treatment for a disease with limited therapeutic options. As an example, the lack of expression of the RASSF1A protein in the majority of cases of penile carcinoma offers the rationale for use of 5-aza-2’’-deoxycytidine, which was shown to inhibit retinoblastoma cells by reversing epigenetically silencing of the RASSF1A gene, either alone or in combination with EGFR-targeting agents. The use of dual targeting agents, such as the bispecific antibody against EGFR and HER3 called MEHD7945A, could be potentially effective in penile carcinoma. Such a compound inhibited the growth of human non-small cell lung cancer cell lines resistant to cetuximab and erlotinib and appeared to be safe in a phase I trial. Combination of MEHD7945A with chemotherapy was also feasible [8]. A number of anti-EGFR agents have been used in penile cancer patients outside the context of a clinical trial, mainly as a salvage treatment after failure of first-line chemotherapy [9-14]. Monoclonal antibodies appear to have promising efficacy, whereas currently available EGFR inhibitors such erlotinib and gefitinib seem to have no activity, which is likely to be related to the lack of EGFR mutating activation [11]. As summarized in Table 1, individual patient-level data were retrieved from published trials reporting on the use of monoclonal antibodies, which included cetuximab, panitumumab, and nimotuzumab. Of 28 patients (median age, 58 years old; interquartile range, 50 – 69), 15 had received therapy in the second-line setting, and 8 of them had received single-agent treatment. The most commonly used agent was cetuximab, which was administered to 24 patients. Overall, 50% of the patients showed a response to treatment, with a median PFS of ∼ 3 months (1.5 – 5.78). These results are comparable to that achieved with paclitaxel alone in the second-line setting [15]. Given the heterogeneity of data, any subgroup analysis would be biased and of little value.

Table 1. Individual data of patients with penile cancer treated with anti-EGFR monoclonal antibodies in retrospective studies.

Ref.	Age	Number of previous chemotherapy regimens	Treatment	Response	Time to progression (in months)	Status
[11]	70	1	Cetuximab	No	1	1
[11]	69	1	Cetuximab	No	2.4	1
[11]	70	1	Cetuximab	No	1.9	1
[11]	61	1	Cetuximab	No	1.56	1
[11]	71	3	Cetuximab	No	2.1	1
[11]	46	1	Cetuximab	Yes	42	0
[11]	69	1	Cetuximab	Yes	9.3	1
[11]	72	1	Cetuximab + cisplatin	No	6.3	1
[11]	51	1	Cetuximab + cisplatin	No	3.8	1
[11]	43	1	Cetuximab + cisplatin	No	1.5	1
[11]	58	2	Cetuximab + cisplatin	No	0.3	1
[11]	58	2	Cetuximab + cisplatin	No	5.9	1
[11]	58	2	Cetuximab + cisplatin	No	1.1	1
[11]	67	2	Cetuximab + cisplatin	No	1	1
[11]	45	2	Cetuximab + cisplatin	No	0.7	1
[11]	71	2	Cetuximab + cisplatin	No	0.9	1
[11]	63	0	Cetuximab + cisplatin	Yes	4.5	1
[11]	62	1	Cetuximab + cisplatin	Yes	3.5	1
[11]	59	1	Cetuximab + cisplatin	Yes	3.1	1
[11]	45	4	Cetuximab + cisplatin	Yes	3.2	1
[12]	36	0	Cetuximab + TIP	Yes	2.8	1
[12]	59	1	Cetuximab + TIP	Yes	6.3	1
[12]	68	1	Cetuximab + TIP	Yes	2.4	1
[10]	75	1	Docetaxel + cetuximab	Yes	7	1
[14]	59	3	Nimotuzumab + paclitaxel	Yes	5.75	0
[13]	44	0	Nimotuzumab + paclitaxel + cisplatin	Yes	5.5	1
[9]	58	1	Panitumumab	Yes	3	0
[11]	43	2	Panitumumab + cisplatin	Yes	6	1

These data indicate that anti-EGFR monoclonal antibodies have some efficacy in penile carcinoma. The role of pEGFR as predictive of activity of anti-EGFR agents should be assessed. Anti-EGFR monoclonal antibodies could be used in addition to chemotherapy in the neoadjuvant setting to increase radiological responses and in the adjuvant setting to decrease the recurrence rate, as well as in the first-line setting in combination with chemotherapy or in more advanced lines of therapy as a single agent.

The rarity of the disease poses great challenge in terms of education and awareness of the general population, planning of preventive measures on a large scale, as well as conduction of prospective trials on biological therapy. International cooperation and management in referral centers are mandatory to tackle this terrible cancer, which carries an enormous social and psychological burden.

Acknowledgments

The authors wish to thank Antonella Buono from University of Salerno for the critical revision of the paper.

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.