Multiple Hereditary Infundibulocystic Basal Cell Carcinoma Syndrome Associated with a Germline SUFU Mutation

Research

Case Report/Case Series Multiple Hereditary Infundibulocystic Basal Cell Carcinoma Syndrome Associated With a Germline SUFU Mutation

Joshua M. Schulman, MD; Dennis H. Oh, MD, PhD; J. Zachary Sanborn, PhD; Laura Pincus, MD; Timothy H. McCalmont, MD; Raymond J. Cho, MD, PhD

IMPORTANCE Multiple hereditary infundibulocystic basal cell carcinoma syndrome (MHIBCC) is a rare genodermatosis in which numerous indolent, well-differentiated basal cell carcinomas develop primarily on the face and genitals, without other features characteristic of basal cell nevus syndrome. The cause is unknown. The purpose of the study was to identify a genetic basis for the syndrome and a mechanism by which the associated tumors develop.

OBSERVATIONS Whole-exome sequencing of 5 tumors and a normal buccal mucosal sample from a patient with MHIBCC was performed. A conserved splice-site mutation in 1 copy of the suppressor of fused gene (SUFU) was identified in all tumor and normal tissue samples. Author Affiliations: Department of Additional distinct deletions of the trans SUFU allele were identified in all tumor samples, Dermatology, University of California, San Francisco (Schulman, Oh, Pincus, none of which were present in the normal sample. McCalmont, Cho); Department of Pathology, University of California, CONCLUSIONS AND RELEVANCE A germline SUFU mutation was present in a patient with San Francisco (Schulman, Pincus, MHIBCC, and additional acquired SUFU mutations underlie the development of McCalmont); Dermatology Research Unit, San Francisco Veterans Affairs infundibulocystic basal cell carcinomas. The downstream location of the SUFU gene within Medical Center, San Francisco, the sonic hedgehog pathway may explain why its loss is associated with relatively California (Oh); NantOmics, well-differentiated tumors and suggests that MHIBCC will not respond to therapeutic LLC, Santa Cruz, California (Sanborn). strategies, such as smoothened inhibitors, that target upstream components of this pathway. Corresponding Author: Raymond J. Cho, MD, PhD, Department of Dermatology, University of California, JAMA Dermatol. 2016;152(3):323-327. doi:10.1001/jamadermatol.2015.4233 San Francisco, 2340 Sutter St, Published online December 16, 2015. S431, San Francisco, CA 94115 ([email protected]).

asal cell carcinomas (BCCs) frequently arise in spo- cally or, intriguingly, in the setting of BCNS.5 Why infundibu- radic fashion, almost always in the context of long- locystic BCCs behave less aggressively than other BCC sub- B term exposure to UV radiation.1 They can develop in the types and why they occur in multiplicity in MHIBCC or in some setting of various genodermatoses as well, including basal cell patients with BCNS are not understood. We aimed to use whole- nevus syndrome (BCNS) (also known as Gorlin syndrome), exome sequencing to identify the genetic basis for MHIBCC and Bazex-Dupré-Christol syndrome, and Rombo syndrome, potential mechanisms for the development of infundibulo- among others.2 Of these, the genetics of BCNS are best under- cystic BCCs in that setting. stood, with most patients having an autosomal dominant inherited patched 1 (PTCH1) (OMIM 601309) gene mutation.3 The PTCH1 gene, a regulator of the sonic hedgehog pathway, is Report of a Case commonly mutated in sporadic BCCs also, although mutations in other components of this pathway have been impli- A woman in her 60s presented to our clinic for evaluation of cated in BCC tumorigenesis.1 multiple dome-shaped, skin-colored asymptomatic papules lo- In 1999, Requena and colleagues4 described a new geno- cated on the face and vulva (Figure 1A and B). These papules dermatosis characterized by numerous infundibulocystic BCCs first appeared around 50 years of age and gradually increased located mostly on the face, autosomal dominant inheritance, to approximately 60 lesions. Eight of the lesions underwent and an absence of palmar pits, jaw cysts, or other stigmata of biopsy, all of which showed buds and cords of basaloid cells BCNS; they termed the genodermatosis multiple hereditary enclosing small cystic spaces, consistent with infundibulo- infundibulocystic BCC syndrome (MHIBCC) (OMIM 604451). cystic BCC (Figure 1C). She had no palmar or plantar pits, and Infundibulocystic BCCs are a well-differentiated subtype of BCC radiographic studies did not reveal any jaw cysts. Therefore, composed of buds and cords of bland basaloid cells, often in we thought that MHIBCC represented the best diagnosis for association with cystic spaces that resemble the follicular in- her condition. The patient’s medical history was significant for fundibulum, which tend to behave in an indolent fashion. This a meningioma. Her family history was notable for a daughter uncommon subtype of BCC can also be encountered sporadi- with similar facial papules that had not undergone biopsy.

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Figure 1. Clinical and Histopathologic Features

A Multiple facial tumors B Close-up of one of the tumors

C Photomicrographs of patient’s tumors

A, Numerous small, domed, skin-colored papules are present on the central face and around the eyes. Similar papules were also noted on the vulva. B, The close-up appearance of one of the tumors. C, Photomicrographs of 2 infundibulocystic basal cell carcinomas from the patient showing buds and cords of basaloid cells arranged around small keratin-filled cysts; clefting between tumor aggregates and the surrounding stroma is focally evident (hematoxylin-eosin, original magnification ×40).

Owing to her increasing tumor burden, the patient elected to We prepared libraries for tumor and unaffected samples initiate therapy with the smoothened protein (SMO) inhibitor using a DNA library preparation kit (KAPA Hyper Prep; Kapa Bio- vismodegib; after 9 months of treatment, the size and num- systems) and used an exome sequencing system (HiSeq; Illu- ber of her tumors had not decreased, and therapy was discon- mina) with a reagent kit (Sure Select CRE; Agilent) to provide at tinued. The patient provided written informed consent, and least 200× coverage of the transcriptome. Sequencing data for our study was approved by the committee on human re- each sample was aligned by the Burrows-Wheeler transform al- search at the University of California, San Francisco. gorithm using default variables, duplicate marked by Samblaster Based on the autosomal dominant inheritance pattern of open-source software, and underwent insertion-deletion re- MHIBCC and the presence of tumors at multiple body sites, we alignment and base quality recalibration by GATK-Lite (version hypothesized that the patient could have a germline muta- 2.3; Broad Institute). Tumor vs matched normal tissue variant tion in a tumor suppressor gene with acquired second-hit mu- analysis was performed using an analysis pipeline (NantOmics tations in her tumors. Our approach therefore entailed sam- Contraster; Nanthealth) to determine somatic and germline pling unaffected (normal) buccal tissue to search for a mutation small variants and estimate relative coverage per exon, as pre- shared with the patient’s tumor samples, and we selected a total viously described.6 Small variants were annotated with base- of 5 tumors from the face (n = 3) and vulva (n = 2) for sequenc- level phastCons conservation scores (UCSC [University of Cali- ing to identify acquired mutations that, in the case of the vul- fornia, Santa Cruz] Genome Bioinformatics), population allele var tumors, would not be expected to be exclusively induced frequencies from the single-nucleotide polymorphism database by UV radiation. (dbSNP) (build 137), and their effect, if any, on genes.

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Figure 2. Allele Fraction Estimates of Chromosome 10

A Tumor 1 B Tumor 2

1.0 1.0

0.8 0.8

0.6 0.6

0.4 0.4

0.2 0.2 Relative Allele Proportion Relative Allele Proportion Relative

0 0 0 20 40 60 80 100 120 140 0 20 40 60 80 100 120 140 Chromosome 10 Position, Mb Chromosome 10 Position, Mb

C Tumor 3 D Tumor 4

1.0 1.0

0.8 0.8

0.6 0.6

0.4 0.4

0.2 0.2 Relative Allele Proportion Relative Allele Proportion Relative

0 0 0 20 40 60 80 100 120 140 0 20 40 60 80 100 120 140 Chromosome 10 Position, Mb Chromosome 10 Position, Mb

E Tumor 5

1.0

0.8

0.6

0.4

0.2 Relative Allele Proportion Relative

0 0 20 40 60 80 100 120 140 Chromosome 10 Position, Mb

Heterozygous single-nucleotide polymorphism database (dbSNP) loci are In addition, tumor 3 shows loss of the other copy of 10p. Mean allele fraction shown for all 5 tumors; the majority and minority alleles measured in tumor 1 are estimates are shown in red and gray lines. The position and allele fraction of the indicated by red and gray dots, respectively. Loss of whole chromosome 10 is SUFU splice site variant (10:104,377,045A>G) are indicated by the blue Xs. observed in tumors 1, 4, and 5. Tumors 2 and 3 show partial loss of 1 copy of 10q. Mb indicates megabase.

splice acceptor site of intron 6 of the suppressor of fused gene Results (SUFU [OMIM 607035]) (c.757-2A>G). This variant allele was also present on the same copy of chromosome 10 in the normal tis- We detected deletions of chromosome 10 in all tumor samples sue sample, consistent with a uniparental germline mutation. by drops in relative coverage and concordant allelic imbalances, Unlike in the tumors, the wild-type trans copy of the SUFU gene estimated by comparing reference and alternate allele sequenc- in the unaffected sample was preserved. ing depths in each tumor at heterozygous dbSNP loci in the unaffected sample (Figure 2). Based on the distinct locations and spans of the observed allelic imbalances, the losses on chromo- Discussion some 10 likely occurred independently in each tumor. We then examined the other copy of chromosome 10 in the tumors, in In the original report of MHIBCC,4 5 individuals from 2 fami- which we detected a mutation in all of the tumors at a conserved lies were identified with numerous infundibulocystic BCCs on

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photoexposed (face) and nonphotoexposed (trunk and geni- sequently, the association between a SUFU mutation and tal) sites. Genetic analysis of these individuals was per- MHIBCC in our patient can be considered a reproducible phe- formed to search for PTCH1 loss of heterozygosity in their tu- nomenon. mors, as would be expected in most cases of BCNS, but this Tumors that characterize MHIBCC behave more indo- aberration was not detected.4 In the present investigation, we lently than other subtypes of BCC. This behavior may reflect used whole-exome sequencing to conduct a much broader the relatively downstream position of SUFU protein in the sonic search for a genetic basis for MHIBCC. We identified a con- hedgehog pathway, such that SUFU mutations may be less dis- served SUFU splice-site mutation in normal and tumor tissue ruptive to the regulation of the pathway than mutations in up- samples from our patient, and we identified additional, dis- stream components, such as PTCH1 or SMO, might be. The tinct deletions affecting the other copy of SUFU in each of the downstream position of SUFU relative to SMO in the hedge- tumors but not in the normal buccal sample. No other muta- hog pathway also means that tumors with SUFU mutations may tions were present. Based on these results, we propose that be resistant to SMO inhibitors or other treatments targeting up- MHIBCC is caused by a germline SUFU mutation with indi- stream components of the pathway. This concept is sup- vidual infundibulocystic BCCs arising as a result of acquired ported by the observation that vismodegib resistance can be second-hit SUFU mutations. achieved through acquired copy number changes or muta- The SUFU protein is a component of the sonic hedgehog tions in SUFU, and it likely explains why our patient’s tumors pathway that acts as a tumor suppressor by binding to and were refractory to vismodegib treatment.15 modulating the function of the transcription factor Gli.7,8 Mu- tations of the SUFU gene have been studied mostly in the context of medulloblastoma, in which they have been implicated Conclusions in sporadic and familial cases, and in the context of familial multiple meningioma.7,9,10 Mutations in SUFU also have been A germline mutation in the tumor suppressor gene SUFU is as- identified in several patients who fulfill diagnostic criteria for sociated with MHIBCC, and the tumors that develop in this syn- BCNS but lack PTCH1 mutations.3,11,12 In each of these fami- drome are induced by additional acquired mutations in the trans lies with SUFU-associated BCNS, at least 1 family member de- copy of that gene. Mutations in SUFU are also found in some pa- veloped a medulloblastoma or meningioma; this is a much tients with BCNS, especially those with a predisposition to- higher rate than is seen in BCNS overall.11 Our patient also re- ward medulloblastoma or meningioma, and MHIBCC and SUFU- ported a remote history of a meningioma, further supporting associated BCNS may represent phenotypic variants of a this potentially significant association. common syndrome. A heightened awareness of the risk for me- Recently, Mann and colleagues13 described a patient with dulloblastoma or meningioma in patients with these syn- a germline SUFU splice-site mutation who had 2 children with dromes and their relatives may be warranted. Patients with these medulloblastoma; on physical examination, this patient had syndromes may not be responsive to vismodegib treatment or multiple skin-colored facial papules that histopathologically other SMO inhibitors. The downstream role of SUFU protein demonstrated basaloid proliferations with follicular differen- within the sonic hedgehog pathway also provides a potential tiation. We suspect that these facial lesions, characterized as explanation for the relatively indolent behavior of infundibu- basaloid follicular hamartomas, are actually infundibulocys- locystic BCCs that develop in patients with MHIBCC. Further tic BCCs (indeed, the 2 entities may be synonymous) and that investigation is warranted to determine whether the same this patient shares the same diagnosis as our patient.14 Con- mechanism underlies sporadic infundibulocystic BCCs as well.

ARTICLE INFORMATION Funding/Support: This study was supported by the REFERENCES Accepted for Publication: September 15, 2015. Well Aging Research Center, Samsung Advanced 1. Jayaraman SS, Rayhan DJ, Hazany S, Kolodney Institute of Technology, under the auspices of Sang MS. Mutational landscape of basal cell carcinomas Published Online: December 16, 2015. Chul Park, MD, PhD, and by grant K08 CA169865 doi:10.1001/jamadermatol.2015.4233. by whole-exome sequencing. J Invest Dermatol. from the National Cancer Institute, National 2014;134(1):213-220. Author Contributions: Drs Schulman and Cho had Institutes of Health (Dr Cho). 2. Castori M, Morrone A, Kanitakis J, Grammatico P. full access to all the data in the study and take Role of the Funder/Sponsor: The funding sources responsibility for the integrity of the data and the Genetic skin diseases predisposing to basal cell had no role in the design and conduct of the study; carcinoma. Eur J Dermatol. 2012;22(3):299-309. accuracy of the data analysis. collection, management, analysis, and Study concept and design: Schulman, Oh, Cho. interpretation of the data; preparation, review, or 3. Pastorino L, Ghiorzo P, Nasti S, et al. Acquisition, analysis, or interpretation of data: All approval of the manuscript; and decision to submit Identification of a SUFU germline mutation in a authors. the manuscript for publication. family with Gorlin syndrome. Am J Med Genet A. Drafting of the manuscript: Schulman, Oh, Sanborn, 2009;149A(7):1539-1543. Additional Contributions: We thank the patient for Pincus, Cho. 4. Requena L, Fariña MC, Robledo M, et al. Multiple Critical revision of the manuscript for important granting permission to publish this information. We are indebted to Jean Tang, MD, PhD, and Kavita hereditary infundibulocystic basal cell carcinomas: intellectual content: Schulman, Oh, Pincus, a genodermatosis different from nevoid basal cell McCalmont, Cho. Sarin, MD, PhD, Stanford University, for discussions regarding SUFU mutations in BCNS. NantOmics LLC carcinoma syndrome. Arch Dermatol. 1999;135(10): Statistical analysis: Sanborn. 1227-1235. Obtained funding: Cho. provided DNA preparation and DNA sequencing. Administrative, technical, or material support: Oh, No compensation was given for these 5. Walsh N, Ackerman AB. Infundibulocystic basal Pincus. contributions. cell carcinoma: a newly described variant. Mod Pathol. Study supervision: Oh, Pincus, Cho. 1990;3(5):599-608. Conflict of Interest Disclosures: None reported.

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6. Sanborn JZ, Salama SR, Grifford M, et al. Double younger than 3 years of age. J Clin Oncol. 2012;30 13. Mann K, Magee J, Guillaud-Bataille M, et al. minute chromosomes in glioblastoma multiforme (17):2087-2093. Multiple skin hamartomata: a possible novel clinical are revealed by precise reconstruction of oncogenic 10. Aavikko M, Li SP, Saarinen S, et al. Loss of SUFU presentation of SUFU neoplasia syndrome. Fam amplicons. Cancer Res. 2013;73(19):6036-6045. function in familial multiple meningioma. Am J Hum Cancer. 2015;14(1):151-155. 7. TaylorMD,LiuL,RaffelC,etal.Mutationsin Genet. 2012;91(3):520-526. 14. Honarpisheh H, Glusac EJ, Ko CJ. Cytokeratin SUFU predispose to medulloblastoma. Nat Genet. 11. Smith MJ, Beetz C, Williams SG, et al. Germline 20 expression in basaloid follicular hamartoma and 2002;31(3):306-310. mutations in SUFU cause Gorlin syndrome– infundibulocystic basal cell carcinoma. J Cutan Pathol. 8. Zhang Y, Fu L, Qi X, et al. Structural insight into associated childhood medulloblastoma and 2014;41(12):916-921. the mutual recognition and regulation between redefine the risk associated with PTCH1 mutations. 15. Sharpe HJ, Pau G, Dijkgraaf GJ, et al. Genomic suppressor of fused and Gli/Ci. Nat Commun.2013; J Clin Oncol. 2014;32(36):4155-4161. analysis of smoothened inhibitor resistance in basal 4:2608. 12. Kijima C, Miyashita T, Suzuki M, Oka H, Fujii K. cell carcinoma. Cancer Cell. 2015;27(3):327-341. 9. Brugières L, Remenieras A, Pierron G, et al. High Two cases of nevoid basal cell carcinoma syndrome frequency of germline SUFU mutations in children associated with meningioma caused by a PTCH1 or with desmoplastic/nodular medulloblastoma SUFU germline mutation. Fam Cancer. 2012;11(4): 565-570.

NOTABLE NOTES

Enjoying Opera, Dermatology Style

Abigail L. Alexander, MS; Leonard J. Hoenig, MD

Opera has just about everything: great music, beautiful singing, AIDS, sought to lessen the stigma of the disease. The musical ends on talented performers, interesting plots, as well as a bit of medicine and a positive note as Mimi survives AIDS, surrounded by friends and dermatology thrown in. To enhance the appreciation of opera among hopeful for the future. physicians, we provide a discussion of several notable operatic Another disfiguring disease, syphilis, affects Dr Pangloss, a charac- works that explores the intriguing interface between opera and ter in Leonard Bernstein’s 1956 operetta Candide, based on the 1759 dermatology. satirical novel by Voltaire. Pangloss is described by Voltaire as having One of the most beloved of all operas is Giacomo Puccini’s La “the end of his nose eaten away”2 by syphilis and in the operetta he Bohème, which premiered in 1896. The plot centers around the lives wears a prosthetic nose. Tertiary syphilis can disfigure the nose in sev- of impoverished artists residing in Paris’ Latin Quarter during the eral ways. One clinical presentation was described by the noted 1830s and on the romance between Rodolfo, a poet, and Mimi, a French dermatologist Jean-Alfred Fournier (1832-1914). When seamstress. destruction of the inferior nasal cartilage occurs, the lower nose In the opening lines, the narrator explains that one of the reasons appears to be pushed into the intact upper nose much like the small Rodolfo finds Mimi so attractive is because of the “rosy hue to her clear tubes of opera glasses which retract into the larger ones.3 Fournier complexion that had the white velvety bloom of the camellia.”1 Sadly,this termed this finding “opera glass nose.” appearance was caused by tuberculosis, from which Mimi tragically dies Opera continues to inspire us with its beauty, which touches and during the final act. elevates the human soul. We hope that this brief dermatologic look at Pallor with red cheeks was often clinically seen in patients with tu- opera will enhance your appreciation of this magnificent form of berculosis. The pallor resulted from anemia, a common hematologic ab- musical art. normality in tuberculosis. The rosy cheeks were likely caused by facial flushing from fever. Author Affiliations: University of Miami School of Medicine, Miami, Florida (Alexander); private practice (Hoenig). In discussing La Bohème, it is almost impossible not to mention Rent, a rock opera written and composed by Jonathan Larsen as a modern- Corresponding Author: Leonard J. Hoenig, MD, 601 N Flamingo Rd, No. 201, day reinterpretation of Puccini’s work. Rent, like La Bohème, spotlights Pembroke Pines, FL 33028 ([email protected]). the plight of poor artists, this time living in Manhattan’s East Village dur- 1. La Bohème. Full text (4 Acts). Libretto by G. Giacosa and L. Illica. English version by W. Grist and P. Pinkerton. Archive. http://archive.org/stream ing 1989. Instead of tuberculosis, Mimi now has human immunodefi- /labohme4actsli00puccuoft/labohme4actsli00puccuoft_djvu.txt Accessed ciency virus (HIV) infection. June 7, 2015. In the 2005 Movie version of Rent, the character Angel suffers 2. Voltaire. Candide and Related Texts. Wootton D, trans. Indianapolis, IN: Hackett from Kaposi’s sarcoma and dies of AIDS. Kaposi’s sarcoma not only Publishing Co Inc; 2000:7. disfigured many AIDS patients but also stigmatized them as having 3. Renner WS. Tertiary syphilis of the nose and pharynx. N Y Med J Philadel Med HIV infection. Rent, through its humane portrayal of people with J. 1904;LXXIX(9):385-391.

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