# Low risk of embryonic and other cancers in PIK3CA-related overgrowth spectrum: Impact on screening recommendations

Laurence Faivre, Jean-charles Crépin, Manon Réda, Sophie Nambot, Virginie Carmignac, Caroline Abadie, Tristan Mirault, Cécile Faure-conter, Juliette Mazereeuw-hautier, Aude Maza, et al.

► To cite this version:

Laurence Faivre, Jean-charles Crépin, Manon Réda, Sophie Nambot, Virginie Carmignac, et al. Low risk of embryonic and other cancers in PIK3CA-related overgrowth spectrum: Impact on screening recommendations. Clinical Genetics, 2023, 104, pp.554 - 563. 10.1111/cge.14410. hal-05290738

HAL Id: hal-05290738

https://hal.science/hal-05290738v1

Submitted on 30 Sep 2025

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# Low risk of embryonic and other cancers in PIK3CA-related overgrowth spectrum: Impact on screening recommendations

Laurence Faivre¹,² | Jean-Charles Crépin¹,³,⁴ | Manon Réda⁵ ID |

Sophie Nambot¹,²,⁵ ID | Virginie Carmignac¹,⁴ | Caroline Abadie⁶ |

Tristan Mirault⁷ ID | Cécile Faure-Conter⁸ | Juliette Mazereeuw-Hautier⁹ |

Aude Maza⁹ | Eve Puzenat¹⁰ | Marie-Agnès Collonge-Rame¹¹ |

Anne-Claire Bursztejn¹² | Christophe Philippe¹,¹³ | Christel Thauvin-Robinet¹,¹⁴ |

Martin Chevarin¹,¹³ | Claire Abasq-Thomas¹⁵ | Jeanne Amiel¹⁶ | Stéphanie Arpin¹⁷ |

Sébastien Barbarot¹⁸ | Geneviève Baujat¹⁶ | Didier Bessis¹⁹ | Emmanuelle Bourrat²⁰ |

Odile Boute²¹ | Nicolas Chassaing²² ID | Christine Coubes²³ | Bénédicte Demeer²⁴ ID |

Patrick Edery²⁵,²⁶ | Salima El Chehadeh²⁷ ID | Alice Goldenberg²⁸ |

Smail Hadj-Rabia²⁹ | Damien Haye¹⁷ | Bertrand Isidor³⁰ | Marie-Line Jacquemont³¹ |

Philippe Khau Van Kien³² ID | Didier Lacombe³³ | Daphné Lehalle¹ |

Laetitia Lambert³⁴ | Ludovic Martin³⁵ | Annabel Maruani³⁶ |

Fanny Morice-Picard³⁴,³⁷ | Florence Petit²¹ ID | Alice Phan³⁸ | Lucile Pinson²³ |

Massimiliano Rossi²⁵,²⁶ | Renaud Touraine³⁹ | Clémence Vanlerberghe²¹ ID |

Marie Vincent³⁰ | Catherine Vincent-Delorme²¹ | Sandra Whalen⁴⁰ |

Marjolaine Willems²³ ID | Nathalie Marle¹³ | Virginie Verkarre⁴¹ |

Christine Devalland⁴² | Mojgan Devouassoux-Shisheboran⁴³ | Marine Abad⁴⁴ |

Nathalie Rioux-Leclercq⁴⁵ | Bertille Bonniaud³ | Yannis Duffourd¹ |

Jehanne Martel⁴ | Christine Binquet⁴⁶ | Paul Kuentz¹,⁴⁷ | Pierre Vabres¹,³,⁴

## Correspondence

Laurence Faivre, Centre de Référence
Anomalies du Développement et Syndromes
Malformatifs, Hôpital d'Enfants, 14 rue
Gaffarel, 21079 Dijon cedex, France.

Email: laurence.faivre@chu-dijon.fr

Pierre Vabres, Centre de Référence MAGEC,
CHU Dijon, 14 rue Gaffarel, 21079 Dijon
cedex, France.

Email: pierre.vabres@chu-dijon.fr

## Funding information

Agence Nationale de la Recherche (ANR-
13-PDOC-0029), Grant/Award Number:
ANR-13-PDOC-0029; Dijon University
Hospital; European Reference network (ERN)

## Abstract

The PIK3CA-related overgrowth spectrum (PROS) encompasses various conditions caused by mosaic activating PIK3CA variants. PIK3CA somatic variants are also involved in various cancer types. Some generalized overgrowth syndromes are associated with an increased risk of Wilms tumor (WT). In PROS, abdominal ultrasound surveillance has been advocated to detect WT. We aimed to determine the risk of embryonic and other types of tumors in patients with PROS in order to evaluate surveillance relevance. We searched the clinical charts from 267 PROS patients for the diagnosis of cancer, and reviewed the medical literature for the risk of cancer. In our cohort, six patients developed a cancer (2.2%), and Kaplan Meier analyses
ITHACA and ERN SKIN; European Union through the FEDER programs; ISITE-BFC (PIA ANR); Programme Hospitalier de Recherche Clinique (PHRC) National, Grant/Award Number: NCT01950975

estimated cumulative probabilities of cancer occurrence at 45 years of age was 5.6% (95% CI = 1.35%-21.8%). The presence of the *PIK3CA* variant was only confirmed in two out of four tumor samples. In the literature and our cohort, six cases of Wilms tumor/nephrogenic rests (0.12%) and four cases of other cancers have been reported out of 483 proven *PIK3CA* patients, in particular the p.(His1047Leu/Arg) variant. The risk of WT in PROS being lower than 5%, this is insufficient evidence to recommend routine abdominal imaging. Long-term follow-up studies are needed to evaluate the risk of other cancer types, as well as the relationship with the extent of tissue mosaicism and the presence or not of the variant in the tumor samples.

KEYWORDS

cancer, PIK3CA, PIK3CA-related overgrowth spectrum (PROS), Wilms tumor

# 1 | INTRODUCTION

Overgrowth syndromes are a heterogeneous group of childhood disorders characterized by either generalized or localized tissue hypertrophy (limb or hemihypertrophy). Children with generalized overgrowth syndromes may have an increased risk of cancer, particularly embryonic malignancies such as Wilms tumor (WT), hepatoblastoma, and adrenocortical carcinoma.¹

Besides these well-defined overgrowth syndromes, the accurate clinical definition of localized overgrowth syndromes including seg-mental overgrowth or hemihypertrophy has remained elusive prior to identification of causative mosaic variants. Postzygotic variants in *PIK3CA* are now known to account for a wide spectrum of localized overgrowth syndromes.² The *PIK3CA*-related overgrowth spectrum (PROS) encompasses various conditions with localized overgrowth associated with postzygotic activating variants in *PIK3CA*. It includes mainly the CLOVES (Congenital Lipomatous Overgrowth, Vascular malformations, Epidermal nevi, Scoliosis/skeletal and spinal malforma-tion, MIM # 612918), KTS (Klippel-Trenaunay, MIM # 149000), and MCAP (Megalencephaly-CApillary malformation-Polymicrogyria, MIM # 602501) syndromes. PIK3CA encodes the catalytic p110 alpha subunit of phosphatidylinositol-3-kinase (PI3K). PI3K activation phos-phorylates and recruits a serine/threonine kinase (AKT), then activates the mammalian target of rapamycin (mTOR) complexes to regulate cell proliferation, apoptosis, angiogenesis, and growth. PI3K/AKT/mTOR signaling pathway is frequently upregulated in various solid cancers such as breast, lung, or colon cancer.³,⁴ More specifically, *PIK3CA* somatic variants occur at frequencies ranging from 1% to 46% in sev-eral common cancers,⁵ mainly in endometrial, breast, bladder, cervical, and colorectal cancers. In the latter, *PIK3CA* somatic variants are found with a high frequency: 24%-46% of endometrial cancers, 20%-32% of breast cancers, 20%-27% of bladder cancers, 14%-23% of cervical cancers, 13%-28% of colorectal cancers, and 12%-15% of head-and-neck cancers.⁵

Based on sporadically reported cases of WT and other types of cancer published in association with KTS before the genetic basis for PROS was known,¹,⁶ it has been postulated that patients with *PIK3CA*-related

disorders were at increased risk of cancer/WT. Two studies resulted in divergent conclusions, and should be interpreted with caution, as genetic test results were not available in all patients. Peterman et al, retrospectively reviewed patients with CLOVES syndrome in the Vascular Anomalies Center at Boston Children's Hospital between 1998 and 2016 to identify those who developed WT, together with a PubMed literature search.⁷ On a total of 122 patients (mean age 7.7 years, range 0-53 years), four developed WT, all diagnosed by 2 years of age. They calculated an incidence of WT in their CLOVES cohort of 3.3%, which was significantly higher than the incidence of WT in the general population (1/10 000; *p* < 0.001). They also found four additional CLOVES patients with WT in the literature. The authors concluded that patients with CLOVES syndrome have an increased risk of WT, and that quarterly abdominal ultrasonography until age 7 years should be considered (with screening being most beneficial for patients under age 3 years), given the benefits of early detection of WT and treatment. Later, Blatt, surveyed members of the KTS Support Group (KTSG) and reviewed PubMed for "Klippel-Trenaunay Syndrome" and "cancer."⁸ Individuals with cancer were reviewed for clinical confirmation of KTS, tumor type, location, and age at presentation. Of 223 KTSG respondents, 24 (10.8%) reported 23 cancers (diagnosed from 6 months to 68 years of age, median 41 years), including three who were younger than 18 years (two with WT). Nine of 23 cancers were basal cell carcinomas (4% of respondents). From 475 articles, authors identified 11 cancers in 10 individuals with KTS. Four of these occurred in children (WT *n* = 2; rhabdomyosarcoma *n* = 1; serous borderline tumor *n* = 1). Tumors in adults included skin squamous cell carcinoma (*n* = 2), basal cell carcinoma (*n* = 1), angiosarcoma (*n* = 1), Hodgkin's disease (*n* = 1), glioblastoma (*n* = 1), malignant hemangiopericytoma (*n* = 1). Authors concluded that the risk of embryonic cancer other than WT in children with KTS does not appear to be higher than in the general population, and WT incidence was under 5%, routine surveillance was not indicated according to the recommendation for best practices.⁹ In adults, they advised paying attention to skin in the area of malformations.

Given the uncertainty from these conflicting conclusions in the literature, and despite absence of definitive evidence, screening for WT remained indicated in international recommendation of follow-up
in PROS.²,¹⁰ Hence, in order to evaluate the relevance of current recommendations for surveillance in PROS, we have now added to the current literature a large series of patients with proven PIK3CA-related overgrowth to determine the risk of cancer, mainly pediatric given the median age of the cohort, with a focus on the risk of WT.

## 2 | PATIENTS AND METHODS

### 2.1 | Study subjects

Clinical data from patients with mosaic overgrowth syndromes and a documented postzygotic PIK3CA class IV and V variant (according to the ACMG guidelines) from the Mosaic Undiagnosed Skin Traits And Related Disorder (M.U.S.T.A.R.D.) cohort (NCT01950975) were retrospectively studied. Ethics committee approval has been obtained under the GAD collection. Molecular data of half of the cohort have previously been published.¹¹ DNA samples were obtained from affected tissue, mainly skin. Patient originated mainly from France.

### 2.2 | Ultra-deep targeted sequencing

Ultra-deep targeted sequencing of PIK3CA was performed as previously described in Kuentz et al, with an average sequencing depth of 1500×.¹¹

### 2.3 | Statistical analyses

The Kaplan-Meier method was used to estimate the cumulative probability of cancer.

### 2.4 | Literature review

Various combinations of keywords were used in a PubMed search, cancer or tumor or Wilms tumor associated with PROS or CLOVES or Klippel-Trenaunay syndromes, with no limit of time. Only articles in English were included. Series of patients with PROS were gathered and the occurrence of cancer was noted. Isolated cases reporting cancer in a patient with PROS was also analyzed, although they may overestimate the risk of cancer. Presence or not of molecular confirmation was particularly studied, together with the clinical phenotype and the presence of cancer in an affected region.

## 3 | RESULTS

### 3.1 | Patients

Six of 267 patients from the M.U.S.T.A.R.D cohort had cancer (2.2%), five females and one male. Median age at last visit was 11.2 years [4 months; 68 years], with a sex ratio of 0.88.

The first patient (PED1276) was a 20-year-old female carrying the p.(His1047Leu) variant in her affected skin. She had KTS with right lower limb overgrowth, cutaneous capillary malformation, and lymphangiectasias. She has been monitored by annual abdominal ultrasound since birth. At 3-year old, an abdominal and pelvic magnetic resonance imaging (MRI), performed to assess extension of vascular malformations, revealed a left renal mass of 2 cm, initially considered as a vascular malformation. An annual US scan was performed for monitoring. At age 6 years, the left renal mass was no longer found in ultrasonography. Three years later, at 9-year old, US scan revealed a large left kidney mass with abdominal lymphadenopathy. Nephroblastoma stage III was diagnosed on renal biopsy. Clinical remission was obtained 1 year later after neo-adjuvant chemotherapy, nephrectomy, adjuvant chemotherapy, and radiotherapy. She was still in complete remission at last visit. Deep targeted sequencing failed to identify the mosaic variant p.(His1047Leu) in the renal tumor tissue.

The second patient (PED1405) was a 61-year-old female with CLOVES syndrome. She had abdominal lipomatous overgrowth, right hand macrodactyly and capillary malformation of upper limbs and trunk, harboring a p.(Glu453Lys) PIK3CA variant. Gastroscopy realized at 52 years old because of anemia and abdominal pain revealed a gastric adenocarcinoma with chronic atrophic gastritis due to *Helicobacter pylori*. Neoadjuvant chemotherapy and partial gastrectomy were performed, with a favorable outcome so far. She had no family history of gastric cancer.

The third patient (PED5337) was a 20-year-old female with multiple cutaneous vascular malformations, lipoma, and a lingual hemihyper-trophy, compatible with CLOVES syndrome. The p.(Glu542Lys) PIK3CA variant was detected on 20% of reads of a biopsy from an epidermal nevus of the neck. A juvenile granulosa tumor of the ovary (FIGO IC3) was diagnosed at 16-years old. Treatment included surgery with left annexectomy and adjuvant chemotherapy, with a favorable outcome so far. The PIK3CA variant could be identified in the ovarian tumor, with a 26.2%-46.5% mosaicism depending on the samples tested.

The fourth patient (PED6121) was a 61-year-old female with CLOVES syndrome associating patches of capillary malformation on her left lower limb and left flank since childhood, later associated with lymphedema. Affected skin biopsy showed the p.(Glu110del) PIK3CA variant. She had developed a myelodysplastic syndrome at 56-years old. She later presented a cutaneous basal cell carcinoma on the hypertrophic limb at 59-years old.

The fifth patient (PED5380) was a 41-year-old male with CLOVES syndrome. He presented at birth a lower limb overgrowth with extensive cutaneous capillary and lymphatic malformation, extending to his lower back and abdomen. Ultra-deep-sequencing on a left lumbar capillary malformation revealed the p.(Cys420Arg) PIK3CA variant. During investigations for rheumatoid purpura at 37-years old, a clear cell kidney cancer was diagnosed. Treatment involved radiofrequency of the kidney mass. He had no family history of clear cell kidney cancer. He smoked two packets of cigarettes per day. The p.(Cys420Arg) PIK3CA variant could be identified in the kidney cancer, with a <3% mosaicism.

The sixth patient (PED6941) was a 58-year-old female with KTS. She presented venous malformations of both legs and varicose
dilatations complicated with deep venous thrombosis and pulmonary embolism, requiring long-term anticoagulation. A right breast adenocarcinoma at 54-years old was treated by unilateral mastectomy with lymph node dissection, followed by hormone therapy. Follow-up at 57-years old was uneventful. Ultra-deep targeted sequencing on a skin biopsy of the left thigh detected the p.(Glu81Lys) PIK3CA variant at a mosaic state, but failed to identify the variant in the breast tumor tissue. She had a family history of breast cancer in her two sisters and her paternal grandmother that led to a gene-panel analysis of genetic predisposition to cancer that was negative.

## 3.2 | Statistical analyses

In order to have enough events and at least 25 people still at risk we estimated cumulative probabilities of cancer occurrence at 25 and 45 years of age. At 25 years of age, the probability of cancer occurrence in our population was 1.7% (95% confidence interval: 0.4%-6.8%). At age 45, the probability was 5.6% (95% CI = 1.35%-21.8%).

## 3.3 | Literature review

Tables 1 and 2 summarize the results of the literature review, first focusing on the risk of WT (Table 1), and second on other cancer types (Table 2), mentioning the sex and age of the patients, the

presence of molecular confirmation, and the presence of the cancer in an affected region.

# 4 | DISCUSSION

Here, we found a low frequency of cancer in the largest series of patients with a molecular confirmation of PIK3CA overgrowth disorder to date. Only six patients had cancer, including two pediatric and four adult cancers (2.2%). Kaplan-Meier analyses estimated cumulative probabilities of cancer occurrence at 25 years of age was 1.7% in our population (95% confidence interval: 0.4%-6.8%), and 5.6% at age 45 (95% CI = 1.35%-21.8%).

We first focused on WT, as an excess of risk was discussed in the literature. WT (or nephroblastoma) has an incidence in the general population of 1 in 10 000.12 About 95% of children are diagnosed before the age of 10 years, with a median age at diagnosis of 3 years.13 Nephroblastomatosis is a precursor lesion to WT. It has been defined as the presence of diffuse or multiple nephrogenic rests, which are clonal foci of developmentally arrested embryonal renal cells rarely seen in normal kidneys but found in nearly 100% of patients with multifocal or bilateral WT.14 WT, together with nephroblastomatosis and nephrogenic rests, are among the most frequent neoplasia in various well-defined generalized overgrowth syndromes, in particular Beckwith-Wiedemann (BWS, MIM # 130650), but also at a lower risk in Sotos (MIM # 117550), Simpson-Golabi-Behmel (SGS,

<table><caption>TABLE 1 Wilms tumor or nephrogenic rests in cohorts of patients with suspected or proven PIK3CA-related overgrowth syndrome.</caption><thead><tr><th>Authors</th><th>Number of cases</th><th>Type of tumor</th><th>PIK3CA variant</th><th>Phenotype</th><th>Age at diagnosis</th></tr></thead><tbody><tr><td colspan="6"><em>Without genetic determination</em></td></tr><tr><td colspan="6"><em>Case reports</em></td></tr><tr><td>Mankad et al<sup>32</sup></td><td></td><td>Wilms tumor</td><td>Not genotyped</td><td>KTS</td><td>12 months</td></tr><tr><td>Pendergrass et al<sup>33</sup></td><td></td><td>Wilms tumor</td><td>Not genotyped</td><td>KTS</td><td>12 months</td></tr><tr><td>Ehrich et al<sup>34</sup></td><td></td><td>Wilms tumor</td><td>Not genotyped</td><td>KTS</td><td>12 months</td></tr><tr><td>Lapunzina et al<sup>1</sup></td><td></td><td>Wilms tumor</td><td>Not genotyped</td><td>MCAP</td><td>10 months</td></tr><tr><td colspan="6"><em>Series of patients</em></td></tr><tr><td>Greene et al<sup>20</sup></td><td>115</td><td>No tumor</td><td>-</td><td>-</td><td>-</td></tr><tr><td rowspan="2">Blatt et al<sup>8</sup></td><td rowspan="2">223</td><td>Wilms tumor</td><td>Not genotyped</td><td>KTS</td><td>6 months</td></tr><tr><td>Wilms tumor</td><td>Not genotyped</td><td>KTS</td><td>2 years</td></tr><tr><td colspan="6"><em>With genetic determination at least in some patients</em></td></tr><tr><td>Keppler Noreuil et al<sup>2</sup></td><td>35</td><td>Nephrogenic rests</td><td>p.His1047Leu</td><td>CLOVES</td><td>2 years</td></tr><tr><td>Mirzaa et al<sup>35</sup></td><td>86</td><td>No tumor</td><td>-</td><td>-</td><td>-</td></tr><tr><td>Gripp et al<sup>10</sup></td><td>73</td><td>Wilms tumor</td><td>p.His1047Arg</td><td>Hemihyperplasia</td><td>9 months</td></tr><tr><td>Postema et al<sup>36</sup></td><td>17</td><td>No tumor</td><td>-</td><td>-</td><td>-</td></tr><tr><td rowspan="4">Peterman et al<sup>7</sup></td><td rowspan="4">122</td><td>Wilms tumor</td><td>p.Asn345Lys</td><td>CLOVES</td><td>2 years</td></tr><tr><td>Wilms tumor</td><td>p.His1047Arg</td><td>CLOVES</td><td>11 months</td></tr><tr><td>Wilms tumor</td><td>Not genotyped</td><td>CLOVES</td><td>2 years</td></tr><tr><td>Wilms tumor</td><td>Not genotyped</td><td>CLOVES</td><td>16 months</td></tr><tr><td>Huchttagowder et al<sup>37</sup></td><td>14</td><td>Nephrogenic rests</td><td>p.His1047Arg</td><td>CLOVES</td><td>2 years</td></tr><tr><td>Our cohort</td><td>267</td><td>Wilms tumor</td><td>p.His1047Leu</td><td>CLOVES</td><td>9 years</td></tr></tbody></table>
<table><caption>TABLE 2 Other cancer types in patients with suspected or proven PIK3CA-related overgrowth syndrome.</caption><thead><tr><th>Authors/cohort</th><th>Sex</th><th>Age at tumor diagnosis</th><th>Phenotype</th><th>PIK3CA variant</th><th>Tumor/cancer type</th><th>Occurrence of cancer in affected region</th></tr></thead><tbody><tr><td>Howitz et al<sup>38</sup></td><td>M</td><td>8 years</td><td>KTS</td><td>NA</td><td>Astrocytoma</td><td>No</td></tr><tr><td>Wirtz et al<sup>39</sup></td><td>F</td><td>37 years</td><td>KTS</td><td>NA</td><td>Multiple myeloma</td><td>NA</td></tr><tr><td>Salman et al<sup>28</sup></td><td>F</td><td>38 years</td><td>KTS</td><td>NA</td><td>Basal and squamous cell carcinoma</td><td>Yes</td></tr><tr><td>Aygencel et al<sup>40</sup></td><td>M</td><td>31 years</td><td>KTS</td><td>NA</td><td>Hodgkin's disease</td><td>No</td></tr><tr><td>Lezama-del Valle et al<sup>41</sup></td><td>NA</td><td>NA</td><td>KTS</td><td>NA</td><td>Angiosarcoma</td><td>NA</td></tr><tr><td>Bujanda et al<sup>42</sup></td><td>F</td><td>38 years</td><td>KTS</td><td>NA</td><td>Esophagus carcinoma</td><td>No</td></tr><tr><td>De Simone et al<sup>43</sup></td><td>F</td><td>48 years</td><td>KTS</td><td>NA</td><td>Squamous cell carcinoma</td><td>Yes</td></tr><tr><td>Fay et al<sup>44</sup></td><td>M</td><td>3 years</td><td>KTS</td><td>NA</td><td>Rhabdomyosarcoma</td><td>No</td></tr><tr><td>Chang et al<sup>45</sup></td><td>M</td><td>Adult-NA</td><td>KTS</td><td>NA</td><td>Esophagus carcinoma</td><td>No</td></tr><tr><td>Mathews et al<sup>46</sup></td><td>M</td><td>33 years</td><td>KTS</td><td>NA</td><td>Malignant hemangiopericytoma</td><td>No (sphenoid)</td></tr><tr><td>Stephens et al<sup>47</sup></td><td>F</td><td>3 years</td><td>KTS</td><td>NA</td><td>Serous borderline tumor</td><td>No</td></tr><tr><td>Karunamurthy et al<sup>48</sup></td><td>M</td><td>72 years</td><td>KTS</td><td>NA</td><td>Low grade angiosarcoma</td><td>Yes (scrotum)</td></tr><tr><td>Yilmaz et al<sup>49</sup></td><td>M</td><td>61 years</td><td>KTS</td><td>NA</td><td>Glioblastoma</td><td>No</td></tr><tr><td>Blatt et al, cancers and 2 WT (out of 2043 surveyed patients and 223 respondents)<sup>8</sup></td><td>M</td><td>20 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>Yes (on capillary malformation)</td></tr><tr><td></td><td>F</td><td>28 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>M</td><td>36 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>F</td><td>40 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>F</td><td>50 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>F</td><td>51 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>F</td><td>56 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>M</td><td>45 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>No</td></tr><tr><td></td><td>F</td><td>32 years</td><td>KTS</td><td>NA</td><td>Squamous cell carcinoma</td><td>No</td></tr><tr><td></td><td>M</td><td>48 years</td><td>KTS</td><td>NA</td><td>Squamous cell carcinoma</td><td>Yes (capillary and venous malformation on foot)</td></tr><tr><td></td><td>F</td><td>27 years</td><td>KTS</td><td>NA</td><td>Angiosarcoma</td><td>Yes (in area of capillary malformation)</td></tr><tr><td></td><td>F</td><td>19 years</td><td>KTS</td><td>NA</td><td>Hodgkin's disease</td><td>No</td></tr><tr><td></td><td>F</td><td>30-40 years</td><td>KTS</td><td>NA</td><td>Basal cell carcinomas</td><td>Yes (on capillary malformation)</td></tr><tr><td></td><td>F</td><td>After 40 years</td><td>KTS</td><td>NA</td><td>"Breast/lung cancer"</td><td>No</td></tr><tr><td></td><td>M</td><td>45 years</td><td>KTS</td><td>NA</td><td>Non Hodgkin lymphoma</td><td>No</td></tr><tr><td></td><td>M</td><td>68 years</td><td>KTS</td><td>NA</td><td>Bladder cancer</td><td>? (patient had pelvic malformations)</td></tr><tr><td></td><td>M</td><td>41 years</td><td>KTS</td><td>NA</td><td>Testicular cancer</td><td>No</td></tr><tr><td></td><td>F</td><td>29 years</td><td>KTS</td><td>NA</td><td>Uterine cancer</td><td>No</td></tr><tr><td></td><td>F</td><td>50 years</td><td>KTS</td><td>NA</td><td>Vaginal cancer</td><td>No</td></tr><tr><td></td><td>F</td><td>53 years</td><td>KTS</td><td>NA</td><td>Mucinous adenocarcinoma</td><td>No</td></tr><tr><td></td><td>M</td><td>64 years</td><td>KTS</td><td>NA</td><td>Leiomyosarcoma</td><td>Yes</td></tr><tr><td>Our cohort</td><td>F</td><td>16 years</td><td>CLOVES</td><td>p.Glu542Lys</td><td>Juvenile ovary granulosa tumor</td><td>No (variant found in tumoral tissue)</td></tr><tr><td></td><td>F</td><td>52 years</td><td>CLOVES</td><td>p.Glu453Lys</td><td>Gastric adenocarcinoma</td><td>Yes (variant could not be tested in tumoral tissue)</td></tr></tbody></table>
<table><caption>TABLE 2 (Continued)</caption><thead><tr><th>Authors/cohort</th><th>Sex</th><th>Age at tumor diagnosis</th><th>Phenotype</th><th>PIK3CA variant</th><th>Tumor/cancer type</th><th>Occurrence of cancer in affected region</th></tr></thead><tbody><tr><td></td><td>F</td><td>55 years</td><td>CLOVES</td><td>p.(Glu110del)</td><td>Basal cell carcinoma and myelodysplastic syndrome</td><td>Yes (BCC) (variant could not be tested in tumoral tissue)</td></tr><tr><td></td><td>M</td><td>38 years</td><td>CLOVES</td><td>p.(Cys420Arg)</td><td>Clear cell renal carcinoma</td><td>Yes (variant found in tumoral tissue)</td></tr><tr><td></td><td>F</td><td>54 years</td><td>KTS</td><td>p.(Glu81Lys)</td><td>Breast cancer</td><td>No (variant not found in tumoral tissue)</td></tr></tbody></table>

Abbreviations: CLOVES, congenital lipomatous overgrowth, vascular malformations; Epidermal nevi, scoliosis/skeletal and spinal malformation; F, female; KTS, Klippel-Trenaunay syndrome; M, male; MCAP, megalencephaly-capillary malformation-polymicrogyria; NA, not available.

MIM # 312870), or Perlman syndromes (MIM # 267000). The prevalence of nephroblastoma in BWS have been estimated at 8.6%, but it has been showed that it was highly dependent of the molecular subtype of the variant, from 3.1% to 28.6%.¹⁵⁻¹⁸ Hence, guidelines have been formulated for the follow-up of these patients according to the molecular subtype of BWS. A semi-annual clinical examination and abdominal ultrasonography every 3 months up to the age of 8 years for screening of Wilms tumor have been recommended in high-risk patients.¹⁹ In our retrospective series of 267 patients with mosaic PIK3CA variants, one (0.04%) was diagnosed with WT. When adding previously published cases, six cases of WT (four cases) or nephrogenic rests (two cases) were found in a series of 483 PROS patients with proven PIK3CA mosaic variants (Table 1). Five out of the six cases had a variant located at codon 1047. Hence, the estimated risk of WT or nephrogenic rests in PROS is about 0.1%. A review of WT tumors in suspected KTS, CLOVES, and MCAP syndrome prior to identification of PIK3CA pathogenic variants in these conditions found three cases in KTS, and one in MCAP (Table 2). In a series of 115 KTS patients, no cases of WT were found. Conversely, none of 8614 WT patients in the US national database were reported to have KTS.²⁰ Hence, due to publication bias, the real incidence of WT in PROS might be even lower than calculated from the literature. Since the median age at last visit in our cohort was 11 years and 2 months, which is higher than the median age of onset of nephroblastoma in the general population—3 years and 2 months—and in patients with BWS (2 years),¹²,²¹ we believe that they are representative of the incidence of WT in proven PROS. WT has a good prognosis in the general population, with a 5-year survival of 85%.²² There is no clear evidence that screening reduces mortality or morbidity. The psychological issues and costs of WT repeated surveillance should be balanced with these unclear benefits. Pragmatic recommendations have been formulated for WT surveillance in children at risk, with a surveillance offered to children at >5% risk of WT. In that case, surveillance should be carried out by renal ultrasonography every 3-4 months, and should continue until 5 years of age in all conditions except BWS, SGS, and some familial WT pedigrees where surveillance is recommended until 7 or 8 years.⁹,¹⁹ Therefore, taken all these data in consideration, there is no sufficient evidence for iterative routine screening of WT in PROS. Literature data should continue to be analyzed in the future to

evaluate if some PROS patients may be particularly at risk for WT, depending on the location of overgrowth which could include the kidney region and the type of variants, with a particular emphasis on the p.(His1047Leu) and p.(His1047Arg) variants. Some authors have proposed the search for PIK3CA variants in urine cell-free (cf) DNA as a biomarker for nephroblastomatosis or WT in PROS patients.²³ Given the low frequency of WT and inconclusive evidence in favor of imaging screening, the systematic search for urine cfDNA as a candidate biomarker for nephroblastomatosis in PROS appears difficult to implement. Also, we failed to identify the mosaic variant p.(His1047Leu) found in affected skin in the renal tumor tissue from PED1276, although development of WT suggests that it is actually present.

Another case of tumor was found before adulthood in our series. Juvenile granulosa tumors are rare entities, representing less than 5% of ovarian tumors in children and adolescents, and developed from non-germinal ovarian cells. Their pathogenesis remains unclear.²⁴ They are considered to have low-to-moderate malignant potential, and in most cases, the tumor is unilateral. Treatment involves surgery and possibly chemotherapy. No other similar cases were found in the literature review.

Four additional cases of cancer have been reported in adults. A gastric adenocarcinoma was diagnosed in a 52-year-old patient with chronic atrophic gastritis due to Helicobacter pylori, a strong risk factor for gastric cancer. The role of the PIK3CA variant in this patient is unknown. The median age at diagnosis for gastric cancer is 71 years,²⁵ with earlier onset in cases with familial germline predisposition.²⁶ In gastric cancer, PIK3CA somatic variants, associated with poor survival, are uncommon.²⁷ The young age at diagnosis of clear cell renal carcinoma in a 37-year-old patient of our series with an involvement of the pelvic region is questionable, as the PI3K/AKT/mTOR signaling pathway serving an important role in the tumorigenesis. Another patient of our series has developed a basal cell carcinoma (BCC) in affected tissue. The question of an increased risk of skin cancer in affected regions has been discussed, with eight other cases with a BBC, but no molecular confirmation (Table 2).²⁸ BCC is the most common skin cancer worldwide, and a very common cancer in Caucasian populations (approximately 70 000 cases each year in France). BCC arises from uncontrolled growth of basal cells; ultraviolet rays, but also individual factors such as age, phenotype, immunosuppression, and
genetic predisposition contribute to the risk. Their development is almost exclusively local, and treatment usually involves surgery. In most cases, it did not occur in apparently affected region. The relationship with the *PIK3CA* variant is unknown. Finally, we found in our series one case of breast cancer, but at a common age, with a family history of breast cancer but no known genetic predisposition to cancer, and not located in an overgrowth area.

Interestingly, our literature review concerning other types of cancer than WT were all reported in KTS phenotypes with no molecular confirmation. One must note that cases from Blatt et al literature review included 23 self-reported cases from 223 respondents out of 2043 surveyed patients without medical direct confirmation, thus likely to result in bias.<fnref n="8" /> The low median age in our series and others cannot permit conclusions of an increased risk of adult-onset cancers. The probability of cancer occurrence of 1.7% at age 25 years (95% confidence interval: 0.4%–6.8%), and 5.6% at age 45 years (95% CI = 1.35%–21.8%) need to be confirmed with longer term follow-up studies. The apparently low rate of visceral cancers in PROS could be explained by the frequent absence of postzygotic pathogenic variant in visceral tissue due to mosaicism. In contrast, Cowden/PTEN hamartoma tumor syndrome (PHTS)—due to inactivating germline pathogenic variants in PTEN, a tumor suppressor gene which inhibits the PI3K pathway—is known to be associated with a higher risk for breast, thyroid, endometrial colorectal, kidney cancer, and melanoma.²⁹,³⁰ In this condition, unlike most cases of PROS, the germline pathogenic variant is present in all cells. Madsen et al mentioned that macroscopic overgrowth in PROS occurs in mesodermal and neuroectodermal derivatives, while *PIK3CA*-associated cancers most commonly arise in ectodermal or endodermal epithelia. Indeed, none of the cancer types enriched for *PIK3CA* variants (e.g., endometrial and breast) have been reported in PROS.³¹ Neuroectodermal and mesodermal cancers show very low occurrence, if any, of activating *PIK3CA* variants. It is also important to emphasize that when denoting a cancer-associated variant, a cancer “driver” implies that it confers a selective growth advantage at some point during cancer development, but is not sufficient for cancer initiation or development. *PIK3CA* variants could be weak drivers of malignancy, whatever tissue is affected.

Finally, we were able to test for the presence of the *PIK3CA* mosaic variant in the tumors of four of the six patients. In two cases, the variant was found in the tumor sample, but in the other two cases, ultra-deep sequencing did not evidence the *PIK3CA* variant in the tumor sample. The results were not correlated with the occurrence or not of cancer in the affected region (Table 2). These results suggest that the cancer reported in PROS patients may not always be related to their disease. In conclusion, a prospective international cohort should be set-up with long-term follow-up of PROS patients in order to be able to answer to the question of an increased cancer risk, besides the increased risk of WT. The increased risk of WT, however, is too low to justify screening. Testing for the presence of the variant in the tumor samples would also be required to try to assess the percentage of cancer that could potentially be attributed to the presence of the *PIK3CA* variant in the tumor tissue.

## AUTHOR CONTRIBUTIONS

Laurence Faivre, Jean-Charles Crépin, Christine Binquet, Paul Kuentz, Pierre Vabres: Contributions to conception and design. Laurence Faivre, Jean-Charles Crépin, Manon Réda, Sophie Nambot, Virginie Carmignac, Caroline Abadie, Tristan Mirault, Cécile Faure-Conter, Juliette Mazereeuw-Hautier, Aude Maza, Eve Puzenat, Marie-Agnès Collonge-Rame, Anne-Claire Bursztejn, Christel Thauvin-Robinet, Claire Abasq-Thomas, Jeanne Amiel, Stéphanie Arpin, Sébastien Barbarot, Geneviève Baujat, Didier Bessis, Emmanuelle Bourrat, Odile Boute, Nicolas Chassaing, Christine Coubes, Bénédicte Demeer, Patrick Edery, Salima El Chehadeh, Alice Goldenberg, Smail Hadj-Rabia, Damien Haye, Bertrand Isidor, Marie-Line Jacquemont, Philippe Khau Van Kien, Didier Lacombe, Daphné Lehalle, Laetitia Lambert, Ludovic Martin, Annabel Maruani, Fanny Morice-Picard, Florence Petit, Alice Phan, Lucile Pinson, Massimiliano Rossi, Renaud Touraine, Clémence Vanlerberghe, Marie Vincent, Catherine Vincent-Delorme, Sandra Whalen, Marjolaine Willems, Nathalie Marle, Virginie Verkarre, Christine Devalland, Mojgan Devouassoux-Shisheboran, Marine Abad, Yannis Duffourd, Jehanne Martel, Pierre Vabres: Acquisition of data. Laurence Faivre, Jean-Charles Crépin, Virginie Carmignac, Manon Réda, Christophe Philippe, Martin Chevarin, Christine Binquet, Paul Kuentz, Pierre Vabres: Analysis and interpretation of data. All authors: Involvement in drafting the manuscript or revising the manuscript, final approval of the version to be published.

## AFFILIATIONS

<footnote n="1">¹Equipe INSERM UMR1231, Génétique des Anomalies du Développement, FHU TRANSLAD, Université Bourgogne Franche-Comté, Dijon, France</footnote>

<footnote n="2">²Centre de Génétique, Centre de Référence Anomalies du Développement et Syndromes Malformatifs et FHU TRANSLAD, CHU Dijon Bourgogne, Dijon, France</footnote>

<footnote n="3">³Service de Dermatologie, CHU Dijon Bourgogne, Dijon, France</footnote>

<footnote n="4">⁴Centre de référence Maladies Rares Génétiques à Expression Cutanée (MAGEC), CHU Dijon, Dijon, France</footnote>

<footnote n="5">⁵Oncogénétique, Centre de lutte contre le cancer Georges François Leclerc, Dijon, France</footnote>

<footnote n="6">⁶Oncogénétique, CHU Rennes, Rennes, France</footnote>

<footnote n="7">⁷Université Paris Cité, PARCC INSERM U970, Centre de référence des maladies vasculaires rares, Hôpital européen Georges-Pompidou, Assistance Publique Hôpitaux de Paris, Paris, France</footnote>

<footnote n="8">⁸Institut d'hématologie et d'oncologie pédiatrique, Lyon, France</footnote>

<footnote n="9">⁹Service de Dermatologie, CHU Toulouse, Toulouse, France</footnote>

<footnote n="10">¹⁰Service de Dermatologie, CHU Besançon, Besançon, France</footnote>

<footnote n="11">¹¹Oncogénétique, CHU Besançon, Besançon, France</footnote>

<footnote n="12">¹²Service de Dermatologie, CHU de Nancy, Nancy, France</footnote>

<footnote n="13">¹³UF6254 Innovation en Diagnostic Génomique des Maladies Rares, Plate-forme de Biologie Hospitalo-Universitaire, CHU Dijon-Bourgogne, Dijon, France</footnote>

<footnote n="14">¹⁴Centre de Référence Déficiences Intellectuelles de Causes Rares, CHU Dijon Bourgogne, Dijon, France</footnote>

<footnote n="15">¹⁵Département de Pédiatrie et Génétique Médicale, CHU Brest Morvan, Brest, France</footnote>
¹⁶Service de Médecine Génomique des Maladies Rares et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, Hôpital Necker-Enfants Malades, AP-HP, Paris, France

¹⁷Service de Génétique Clinique et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHRU de Tours, Tours, France

¹⁸Service de Dermatologie, CHU Nantes, Nantes, France

¹⁹Département de Dermatologie, CHRU de Montpellier, Montpellier, France

²⁰Service de dermatologie, centre de référence maladies génétiques à expression cutanée MAGEC, CHU St-Louis, Service de pédiatrie générale, CHU Robert Debré, Paris, France

²¹Service de Génétique Clinique et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU Lille, Lille, France

²²Service de Génétique Médicale et Centre de Compétence Anomalies du Développement et Syndromes Malformatifs, CHU Toulouse, Toulouse, France

²³Département de Génétique Médicale, Maladies rares et Médecine Personnalisée, et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHRU de Montpellier, Montpellier, France

²⁴Centre d'Activité de Génétique Clinique et Oncogénétique, CHU d'Amiens, Amiens, France

²⁵Service de génétique, Centre de Référence Anomalies du Développement, Hospices Civils de Lyon, Bron, France

²⁶INSERM U1028, CNRS UMR5292, Centre de Recherche en Neurosciences de Lyon, GENDEV Team, Université Claude Bernard Lyon 1, Bron, France

²⁷Service de Génétique Médicale, Centre de Référence Déficiences Intellectuelles de Causes Rares, Institut de Génétique Médicale d'Alsace (IGMA), CHRU de Strasbourg, Strasbourg, France

²⁸Service de Génétique et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU de Rouen et Centre Normand de Génomique Médicale et Médecine Personnalisée, Rouen, France

²⁹Service de Dermatologie et Centre de Référence des Maladies Rares Génétiques à Expression Cutanée (MAGEC), Université Paris Descartes-Sorbonne Paris Cité, Institut Imagine, Hôpital Universitaire Necker Enfants Malades, Paris, France

³⁰Service de Génétique Médicale et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU de Nantes, Nantes, France

³¹Unité de Génétique Médicale et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU de la Réunion, Saint-Pierre, France

³²Unité de Génétique Médicale et Cytogénétique, Centre de Compétence Anomalies du Développement et Syndromes Malformatifs, CHU de Nîmes, Nîmes, France

³³Service de Génétique Médicale et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU de Bordeaux, Bordeaux, France

³⁴Service de Génétique Clinique et Centre de Référence Anomalies du Développement et Syndromes Malformatifs, CHU de Nancy, Nancy, France

³⁵Service de Dermatologie, CHU d'Angers, Angers, France

³⁶Université de Tours, Service de Dermatologie, Tours, France

³⁷Service de Dermatologie, CHU de Bordeaux, Bordeaux, France

³⁸Service de Dermatologie, CHU de Lyon, Lyon, France

³⁹Service de Génétique Clinique et Centre de Compétence Anomalies du Développement et Syndromes Malformatifs, CHU de Saint-Etienne, Saint-Etienne, France

⁴⁰Unité Fonctionnelle de Génétique Clinique, Hôpital Armand-Trousseau, Paris, France

⁴¹Service d'Anatomie Pathologique, Hôpital Européen Georges Pompidou, Paris, France et INSERM UMR 970, Equipe 13, PARCC Université de Paris Cité, Paris, France

⁴²Service d'Anatomie Pathologique, Hôpital Nord Franche Comté, Trevenans, France

⁴³Service d'Anatomie Pathologique, Hospices Civils de Lyon, Lyon, France

⁴⁴Service d'Anatomie Pathologique, CHU Besançon, Besançon, France

⁴⁵Service d'anatomopathologie, CHU Rennes, Rennes, France

⁴⁶INSERM, Université de Bourgogne, CHU Dijon Bourgogne, CIC 1432, Module Épidémiologie Clinique, Dijon, France

⁴⁷Oncobiologie Génétique Bioinformatique, PCBio, CHU Besançon, Besançon, France

## ACKNOWLEDGMENTS

The authors thank the subjects and families involved in the study, their referring physicians, and M-CM France (http://m-cmfrance.com/). The authors also thank the University of Burgundy Centre de Calcul (CCuB) for technical support and management of the informatics platform. This work was funded by the Agence Nationale de la Recherche (ANR-13-PDOC-0029), the Programme Hospitalier de Recherche Clinique (PHRC) National (NCT01950975), and by grants from Dijon University Hospital, the ISITE-BFC (PIA ANR), and the European Union through the FEDER programs. Several authors are part of the European Reference network (ERN) ITHACA and ERN SKIN.

## CONFLICT OF INTEREST STATEMENT

The authors declare no conflict of interest.

## PEER REVIEW

The peer review history for this article is available at https://www.webofscience.com/api/gateway/wos/peer-review/10.1111/cge.14410.

## DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request.

## ORCID

Manon Réda https://orcid.org/0000-0003-3144-3990

Sophie Nambot https://orcid.org/0000-0002-9630-5049

Tristan Mirault https://orcid.org/0000-0001-8690-1093

Nicolas Chassaing https://orcid.org/0000-0003-0234-8820

Bénédicte Demeer https://orcid.org/0000-0001-5266-7710

Salima El Chehadeh https://orcid.org/0000-0003-1613-6570
Philippe Khau Van Kien https://orcid.org/0000-0001-7754-969X

Florence Petit https://orcid.org/0000-0002-1368-1023

Clémence Vanlerberghe https://orcid.org/0000-0003-3884-2054

Marjolaine Willems https://orcid.org/0000-0002-2959-0935

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**How to cite this article:** Faivre L, Crépin J-C, Réda M, et al. Low risk of embryonic and other cancers in PIK3CA-related overgrowth spectrum: Impact on screening recommendations. *Clinical Genetics*. 2023;104(5):554-563. doi:10.1111/cge.14410