Cancer and Traditional Plant Knowledge, an Interesting Field to Explore: Data from the Catalan Linguistic Area

Cancer is the second cause of death in the world and is foreseen to be responsible for about 16 million deaths in 2040. Approximately, 60% of the drugs used to treat cancer are of natural origin. Besides the extensive use of some of these drugs in therapies, such as those derived from the genus Taxus, a significant number of plants have revealed themselves as useful against cancer in recent years. The field of ethnobotany focuses on documenting traditional knowledge associated with plants, constituting a starting point to uncover the potential of new plant-based drugs to treat or prevent, in this case, tumour diseases and side effects of chemotherapy and radiotherapy. From a series of extensive ethnobotanical prospections across the Catalan linguistic area (CLA), we have recorded uses for 41 taxa with antitumour effects. The two most quoted botanical families are Asteraceae and Ranunculaceae, and the most frequently reported species is Ranunculus parnassifolius, a high-mountain species, which is widely collected for this purpose. The reported species have been used to treat an important number of cancer types, focusing on preventive, palliative, and curative uses, as well as to deal with the side effects of conventional treatments. Comparing our results in CLA with previous data available in the most comprehensive databases of pharmacology and a review of cytotoxicity assays revealed that for the several species reported here, there was no previous evidence of traditional uses against cancer. Despite the need for further analyses to experimentally validate the information presented here, combining traditional uses and phylogenetically-informed strategies to phytochemical and pharmacological research would represent new avenues to establish more integrative approaches, hence improving the ability to select new candidate taxa in cancer research.


Introduction
Cancer is one of the leading causes of death worldwide, accounting for nearly 10 million deaths in 2020 [1]. There is about a 20% risk of developing a cancer in a lifetime and a 10% risk of dying from the disease; this means that one in five persons will suffer from some type of cancer in their lifetime and one in ten will, unfortunately, die from the disease [2]. The International Agency for Research on Cancer estimates an incidence of 30 million people and more than 16 million deaths directly linked to this illness by 2040 [3]. Breast cancer was amongst the most diagnosed cancer types in 2020, followed by lung, colon and rectum, prostate, skin (non-melanoma) and stomach cancer. By far, lung cancer, followed by liver and stomach cancer, represent the most deadly types of cancer [2]. The World Health Organization [1] suggests that between 30% and 50% of cancers could be avoided

General Data
Over the last 30 years of ethnobotanical prospection across the Catalan linguistic area, we have gathered information on folk plant uses related to cancer for 41 plant species, including references to curative, palliative or preventative purposes (Table 1). It is however, worth mentioning that in some cases, the informants use local euphemisms to describe the origin of the illness, such as "mal dolent" or "mal lleig", which literally translate to bad or ugly illness [30]. One explanation for this is that still today, it is hard for many people to use the word cancer, being considered to some extent a taboo disease (or forbidden word). This is especially common among the elderly, and is clearly associated with traditional knowledge. Certainly, fears behind the use of the word cancer could be most likely associated with the high rates of deaths attributed to this illness, prior to the improvement of detection and subsequent therapies of treatment.
A systematic review of the traditional uses of medicinal plants to treat cancer was published in 2021 [31]. The review reported a total of 948 plants species used against cancer around the world. Surprisingly, despite the large number of species reported in this former global review, 21 (out of the 41) taxa reported in our study were not included before in any compilation of uses, and also, a total of 37 taxa are quoted for the first time for the Iberian Peninsula and the Balearic Islands following this review publication. Altogether, these findings contribute significantly to improve our understanding of traditional plant uses related to cancer management, but also emphasise the power of in-depth surveys at regional and local levels to uncover new potential candidate species, and thus complement global initiatives in cancer research. Table 1. Plant taxa used against cancer in the Catalan linguistic area, grouped by curative, palliative, and preventive activities. Comparison of uses in the pharmacological comprehensive literature was extracted from: European Medicines Agency (EMA) monographs [32], Duke's CRC Handbook of Medicinal Herbs [33], and Fitoterapia.net webpage [34].

Most Recurrent Taxa Used by Informants in Prospected Area
The most cited plant against cancer in our surveys across the area prospected was Ranunculus parnassifolius (Figure 1), a perennial herb belonging to the Ranunculaceae family, a botanical family widely studied over centuries in traditional ethnomedicine. In particular, this species grows in high-mountain screes in the Pyrenees and in other European mountain ranges. In the Catalan Pyrenees, this taxon is traditionally used for the treatment of "mal gra", a Catalan term that translates into "bad pimple", frequently used to designate some sort of skin cancer. It is usually prepared by combining it with chicken fat in the form of balm or unguent [35]. This plant is popularly very strongly associated with this specific use that it is, indeed, widely known by its local Catalan name "herba del mal gra" (i.e., "bad pimple herb") [30]. One of the negative consequences of this medicinal reputation is that, in recent years, this taxon has been intensively collected in the wild. Besides the impact of climate change in high mountain ecosystems, uncontrolled harvesting adds an extra level of threat to already damaged populations, many of which suffer the consequences of uncontrolled human activities, such as tourism and high-mountain sports, in these areas. Certainly, as already stated by Brower [36], unless action is taken, the rapid loss of biodiversity (to which we add the decline in traditional knowledge, particularly in heavilyindustrialised societies) can adversely affect future cancer plant-based drug discovery.
Besides R. parnassifolius, another four species belonging to the buttercup family (Ranunculaceae) were also recorded in our study (see Table 1 for details). This result is not surprising, based on the previous investigation by Hao et al. [37], who already pointed out that several phylogenetically-related genera within the family from China contain a series of phytometabolites (e.g., alkaloids, terpenoids, saponins and polysaccharides) with anti-cancer activity [38]. Altogether, this makes the species from the Ranunculaceae family reported here (and in particular R. parnassifolius) good potential candidates for future phytochemical and pharmacological investigation. chicken fat in the form of balm or unguent [35]. This plant is popularly very strongly associated with this specific use that it is, indeed, widely known by its local Catalan name "herba del mal gra" (i.e., "bad pimple herb") [30]. One of the negative consequences of this medicinal reputation is that, in recent years, this taxon has been intensively collected in the wild. Besides the impact of climate change in high mountain ecosystems, uncontrolled harvesting adds an extra level of threat to already damaged populations, many of which suffer the consequences of uncontrolled human activities, such as tourism and high-mountain sports, in these areas. Certainly, as already stated by Brower [36], unless action is taken, the rapid loss of biodiversity (to which we add the decline in traditional knowledge, particularly in heavily-industrialised societies) can adversely affect future cancer plant-based drug discovery. Besides R. parnassifolius, another four species belonging to the buttercup family (Ranunculaceae) were also recorded in our study (see Table 1 for details). This result is not surprising, based on the previous investigation by Hao et al. [37], who already pointed out that several phylogenetically-related genera within the family from China contain a series of phytometabolites (e.g., alkaloids, terpenoids, saponins and polysaccharides) with anti-cancer activity [38]. Altogether, this makes the species from the Ranunculaceae family reported here (and in particular R. parnassifolius) good potential candidates for future phytochemical and pharmacological investigation.
Another family that was recurrently cited among the informants is the sunflower family (i.e., Asteraceae). According to our results, five species were claimed to be used against cancer by locals ( Table 1). One of them is Silybum marianum, a very well-studied plant [39,40], which is indeed a great example to illustrate the importance of common plants for cancer research in particular, and for medicine in general. More specifically, S. marianum is known for its hepatoprotective properties, which has revealed promising results for cancer treatment in recent research [40]. In addition, other Asteraceae members were reported here with well-established traditional uses (Table 1). With such precedents, future analytical work should then focus on this group of candidate taxa for additional chemical exploration, and thus confirm whether any of these is of anticancer usefulness beyond the current knowledge. Among Rosaceae, four species in the family (Table 1) also play an important role in traditional medicine. Surprisingly, the utilised part of these species in the studied territories is not the fruit. In general, the presence of phytochemicals and antioxidants in Rosaceae fruits and their potential as cancer inhibitors are well known [41], but based on our results, investigating alternative tissues could indeed open new avenues in this field of research. Another family that was recurrently cited among the informants is the sunflower family (i.e., Asteraceae). According to our results, five species were claimed to be used against cancer by locals ( Table 1). One of them is Silybum marianum, a very well-studied plant [39,40], which is indeed a great example to illustrate the importance of common plants for cancer research in particular, and for medicine in general. More specifically, S. marianum is known for its hepatoprotective properties, which has revealed promising results for cancer treatment in recent research [40]. In addition, other Asteraceae members were reported here with well-established traditional uses (Table 1). With such precedents, future analytical work should then focus on this group of candidate taxa for additional chemical exploration, and thus confirm whether any of these is of anticancer usefulness beyond the current knowledge. Among Rosaceae, four species in the family (Table 1) also play an important role in traditional medicine. Surprisingly, the utilised part of these species in the studied territories is not the fruit. In general, the presence of phytochemicals and antioxidants in Rosaceae fruits and their potential as cancer inhibitors are well known [41], but based on our results, investigating alternative tissues could indeed open new avenues in this field of research.
Beyond the above-mentioned botanical families, other plant species were reported to fight against the more frequent cancers associated with high mortality rates in rural areas. Among them, Helleborus foetidus, Plantago lanceolata and Plantago major have been reported by our informants to treat skin carcinogenic injuries, Thymus vulgaris for throat-related forms of cancer, Crocus sativus for breast tumors, Anemone hepatica for liver, and Malva sylvestris for colorectal cancers (Table 1).

Wild and Cultivated Vegetables and Their Role against Cancer
In the area of preventive strategies against cancer by WHO [1], specialists pointed out the low consumption of fruit and vegetables as an important cancer risk factor. Even if the relationship between food and vegetable intake and cancer does not seem to be clear in most cases, their consumption is encouraged in trouble-preventive healthy diets [42,43]. In this respect, we want to emphasise the idea around the role of folk functional foods or nutraceuticals [44] as preventative and curative. For example, the species Brassica oleracea (wild cabbage and its common infraspecific categories including broccoli, cauliflower, kale, etc.), was widely reported as an effective source against stomach cancer by our informants. In addition, epidemiological studies highlight the positive effects of the ingestion of plants belonging to the genus Brassica as a cancer preventive [45], providing support to our reports.
In addition, four frequently cultivated vegetables (Allium cepa and Daucus carota subsp. sativus were specifically cited for the treatment of stomach cancer; Apium graveolens and Beta vulgaris), a minor or neglected crop (Helianthus tuberosus), and a wild plant often consumed as a vegetable (Urtica sp.) complete the set of folk functional foods potentially useful as antitumour sources (representing 17.5% of the cited species; Table 1). Altogether, these results show the importance of plants (both cultivated and their wild crop relatives), whose regular intake in diets can be beneficial and cancer preventive.

Plants for Dealing with Side Effects of Cancer Treatments
As highlighted in previous sections, this study provides compelling evidence regarding the importance of assessing traditional knowledge linked to plants as a possible source of new drugs. Whilst efforts need to focus on the prevention and/or cure of cancer, another important issue to tackle is the side effects of standard medical treatments, and investigation into how plants can contribute to the alleviation of the effects of antitumour therapies. Reducing (or minimising) the negative effects-e.g., anaemia, appetite loss, nausea and vomiting, general pain and distress, mouth infections, among others-of chemotherapy and radiotherapy treatments is one of the goals that is being pursued in this field, and plants are also present in this line of research [46]. Based on our research, the most common species reported with analgesic activity (i.e., pain relief) are Cannabis sativa and Papaver somniferum, whilst Plantago sempervirens and Santolina chamaecyparissus were used for mouth infections derived from therapies against cancer.

Pharmacological Activity Review
The official monographs of the European Medicines Agency (EMA) [32] and European Scientific Cooperative on Phytotherapy (ESCOP) [47] include just one out of the forty-one species reported in our study. According to the EMA records, the monograph report for Allium cepa provides details regarding the anticarcinogenic and antimutagenic activities associated with this species [32]. Consulting other sources of literature on phytotherapy made it possible to validate a total of 68.29% of the species reported here. In fact, Duke's CRC Handbook of Medicinal Herbs [33] was by far the most inclusive, systematic and detailed work analysed of any of the sources consulted (Table 1).
In addition, we carried out a review of the cytotoxicity tests against cancer cell lines for the plants reported by informants in our field surveys. So far, a total of 26 species have been the focus of different studies to test inhibition of cell growth in cancer cell lines (see Table 2 for details). Some of the species currently lacking any information regarding the inhibitory activity, such as Plantago major, Ranunculus bulbosus or Ranunculs parnassifolius, have been otherwise well studied from a genus level perspective, and the results obtained in related species could be similar for these taxa, although this would require future confirmation. In fact, the lack of cytotoxicity assays does not necessarily mean that some of these species are not indeed active against cancer. Other pharmacological activities, such as antioxidant capacity by scavenging reactive oxygen, are important in preventing potential damage to cellular components such as DNA, proteins and lipids. The oxidative damage can cause major problems, such as carcinogenesis [48]. Some of the species reported here are not studied against cancer cell lines such as Apium graveolens, Rubus ulmifolius or Sambucus nigra, but are well known for their antioxidant activity [49][50][51], and could be, therefore, good candidates to test in future assays.
In summary, our study provides relevant information on the traditional uses of plants against cancer across the Catalan linguistic area, contributing to the global understanding of ethnomedicine to mitigate the impact of an illness that kills nearly 10 million people per year. We want to stress, however, that our efforts to collate and make such data available should be paralleled by an increase in pharmacological studies to experimentally validate the data reported here, and also by subsequent analyses regarding the impact of these chemicals on cancer cell lines.   The essential oil causes a very strong inhibition (60%) of cell viability in PANC-1 cells, compared to 40% of reduction observed in U2OS cells at 10 µg/mL [93] Leaf EtOH -T47D The extract inhibited 75% of T47D cells at 200 µg/mL [94] Leaf CHCl 3 Polyphenol complex SH-SY5Y and SK-N-BE(2)-C The extract showed strong levels of cytotoxicity towards SH-SY5Y and SK-N-BE(2)-C cell lines at the highest tested dose level (125.0 µg/mL) [95] Verbena officinalis L.

Studied Area
The Catalan linguistic area constitutes a well-studied area from the following several perspectives: geographic [97], physiographic [98], floristic [99,100], vegetation [101], linguistic and cultural approach [102]. This territory, located in the eastern part of the Iberian Peninsula, also includes a northern Pyrenean portion, the Balearic Islands, and the city of L'Alguer on the island of Sardinia. Politically, this territory belongs to the following four states: Andorra (all the territory), France (Northern Catalonia or Eastern Pyrenees department), Italy (L'Alguer, Sardinia), and Spain (Balearic Islands, Carxe-a small area in Murcia, Catalonia, a portion of eastern Aragon, and Valencia) (Figure 2). It is home to around 14,000,000 people [103][104][105][106][107] and extends across 70,000 km 2 [100]. The orographic profile is quite diverse, from the Mediterranean Sea level to 3143 m a.s.l. at the summit of Pica d'Estats (Pyrenees). The landscape of the area of study is structured in several belts with distinct floristic and vegetation traits [99,100], harboring approximately 4300 autochthonous and 1200 allochthonous plant taxa, including species and subspecies [108].

Databasing and Data Selection
The information was collected through semi-structured ethnobotanical interviews [109], following the ethical principles of the International Society of Ethnobiology [110], and included in an open-access webpage (https://etnobotanica.iec.cat), which contains the ethnobotanical data for the Catalan linguistic area [111]. Herbarium vouchers were prepared for each species and are deposited in the herbarium BCN (Centre de Documentació de Biodiversitat Vegetal, Universitat de Barcelona). All the information available concerning plants used to treat, palliate or prevent cancer was retrieved from the open access webpage mentioned before. Data obtained from informants were compiled from interviews performed from 1990 to the present.
Bolòs et al.'s [100] was followed for taxonomic nomenclature, which is a specifically flora focused on the studied area. Plants of the World Online (https://powo.science.kew.org) was also consulted when exotic plants were involved. For family attribution, we followed the criteria stablished by the APG IV, the last Angiosperm Phylogeny Group's arrangement to date [112].

Pharmacological Activity Review
In order to confidently assess how many of the species cited in our surveys (i.e., the Catalan linguistic area) had been previously studied in depth, a review of pharmacological activities was carried out. Monographs of the official sources, such as the European Medicines Agency (EMA) [32], the European Scientific Cooperative on Phytotherapy (ESCOP) [47], the encyclopedic bibliography on phytotherapy as Duke's CRC "Handbook of Medicinal Herbs" [33] and Fitoterapia.net webpage [34], were consulted.
In addition, an original publication search was carried out using four major online The orographic profile is quite diverse, from the Mediterranean Sea level to 3143 m a.s.l. at the summit of Pica d'Estats (Pyrenees). The landscape of the area of study is structured in several belts with distinct floristic and vegetation traits [99,100], harboring approximately 4300 autochthonous and 1200 allochthonous plant taxa, including species and subspecies [108].

Databasing and Data Selection
The information was collected through semi-structured ethnobotanical interviews [109], following the ethical principles of the International Society of Ethnobiology [110], and included in an open-access webpage (https://etnobotanica.iec.cat), which contains the ethnobotanical data for the Catalan linguistic area [111]. Herbarium vouchers were prepared for each species and are deposited in the herbarium BCN (Centre de Documentació de Biodiversitat Vegetal, Universitat de Barcelona). All the information available concerning plants used to treat, palliate or prevent cancer was retrieved from the open access webpage mentioned before. Data obtained from informants were compiled from interviews performed from 1990 to the present.
Bolòs et al.'s [100] was followed for taxonomic nomenclature, which is a specifically flora focused on the studied area. Plants of the World Online (https://powo.science.kew. org) was also consulted when exotic plants were involved. For family attribution, we followed the criteria stablished by the APG IV, the last Angiosperm Phylogeny Group's arrangement to date [112].

Pharmacological Activity Review
In order to confidently assess how many of the species cited in our surveys (i.e., the Catalan linguistic area) had been previously studied in depth, a review of pharmacological activities was carried out. Monographs of the official sources, such as the European Medicines Agency (EMA) [32], the European Scientific Cooperative on Phytotherapy (ESCOP) [47], the encyclopedic bibliography on phytotherapy as Duke's CRC "Handbook of Medicinal Herbs" [33] and Fitoterapia.net webpage [34], were consulted.
In addition, an original publication search was carried out using four major online databases for scientific bibliographic resources, namely PubMed, ScienceDirect, Scopus and Web of Science, using the following set of keywords: "scientific name" AND "cancer" AND "chemical compounds". The aim of this search was mainly to review the existence of cytotoxic activity tests in cancer cell lines involving the plant species of interest.

Conclusions
Ethnobotanical research, when integrated into new fields of study, such as molecular phylogenetics, phytochemistry and other "omic" disciplines, can become a powerful tool to experimentally validate traditional plant knowledge, as well as to predict promising new sources of plant-based drugs. This study represents a step forward in our understanding of folk medicine as a resource to obtain relevant information to treat symptoms related to cancer. We are, nonetheless, aware that there is still a long way to go to before this information can be used in oncological procedures. Plants with an ethnobotanical tradition deserve to be studied more thoroughly, as they provide potential candidate scenarios to fight against cancer and its associated side effects. It is, of course, important to continue to stay in alliance with conventional medicine, which already includes plant-based products. Based on this, ethnobotanical research should become a standard tool in pharmacological and medicinal research, as it could help to guide pathways for drug discovery, and play a significant factor when applying new solutions to the many rising challenges when fighting against diseases such as cancer.

Informed Consent Statement:
The studies involving human participants (in the present case not as patients, but as informants) were reviewed and approved. The information has been collected through semi-structured ethnobotanical interviews following the ethical principles of the International Society of Ethnobiology. The participants provided their informed consent to participate in this study.

Data Availability Statement:
The dataset analysed for this study are available in the manuscript, further inquiries can be directed to the corresponding authors.